Thereproductivebiologyandmovement ... final.pdf · supervisors: NedPankhurst, John Stevens andStewart Frusher. I will be forever grateful ... Thanks also to Colin andMichelle for

The reproductive biology and movement

patterns of the draughtboard shark,

(Cephaloscyllium laticeps): implications

for bycatch management

by

Cynthia Andrea Awruch

Submitted in fulfilment of requirements for the Degree of

Doctor of Philosophy

January 2007

Tasmanian Aquaculture and Fisheries Institute

School of Aquaculture

University of Tasmania, Australia

ii

Draughtboard shark, Cephaloscyllium laticeps

iii

DECLARATIONS

I hereby declare that this thesis is my own work except where due

acknowledgement is given, and that the material presented here has not been submitted

at another university for the award of any other degree diploma.

This thesis my be made available for loan and limited copying in accordance with

the Copyright Act 1968

Cynthia Andrea Awruch

January 2007

iv

ABSTRACT

The draughtboard shark (Cephaloscyllium laticeps) is the most common shark on

temperate reefs in southeastern Australia. In order to implement adequate management

plans its reproductive biology and movement patterns were studied.

Females developed a single external-type ovary with a maximum follicle diameter of 35

mm. Vitellogenesis commenced at 10 mm follicle diameter. The male reproductive tract

consisted of paired testis with spermatocysts undergoing diametric development.

The hormones testosterone, 17-β estradiol, progesterone and 11-ketotestosterone

(males only) were examined to determine their role in reproduction. Testosterone and

estradiol showed major changes during follicle development. Estradiol increased as the

follicle developed before declining as the follicle reached maturity. Testosterone

remained low during the first stages of follicular development and increased as the

follicle reached maturity. Progesterone showed a peak just prior to ovulation.

Testosterone was the only hormone that varied with maturity in males and no levels of

11-ketotestosteorne were detected.

Females were able to store sperm for at least 15 months and eggs were laid in pairs at

monthly intervals. Juveniles hatched after 12 months.

The size at maturity and seasonality of reproduction were estimated using reproductive

parameters obtained from dissected animals and from steroid hormones. The sizes at

onset of sexual maturity by both methods were similar. Females laid eggs throughout

the year with a peak in deposition between January and June. Elevated values of

testosterone and progesterone coincide with this period of egg deposition. Males

showed no seasonal pattern in reproduction although both testosterone and the amount

of sperm in the seminal vesicle were marginally higher in the first semester of the year.

Movement studies were undertaken using conventional and acoustic tagging. The area

of study included a marine reserve and the adjacent bays of southeast Tasmania. Both

v

methods demonstrate that the majority of sharks remained in the same region in which

they were tagged, although a few sharks moved large distances. Sharks were active

throughout the day and night with peak activity during dawn and dusk. This species

could remain stationary on the bottom for periods up to five days. No correlation was

found between activity and lunar patterns and both sexes showed similar activity

patterns

This study has provided the first information on reproduction and movement of

draughtboard shark and demonstrated the potential for hormones to provide

reproductive information necessary for management without the need to sacrifice the

shark.

vi

ACKNOWLEDGMENTS

I would like to start these acknowledgments by saying THANK YOU to my supervisors: Ned Pankhurst, John Stevens and Stewart Frusher. I will be forever grateful to them. I could not have done this study without their constant support; it has been a great pleasure and honour to work side by side with you all. I would specially like to say thanks to Ned for giving me the opportunity to come to Australia to complete this degree, for his patience and time in the laboratory, for discussing ideas and opening my mind. To John, for his support and advice since the very beginning and for showing me how to be a good scientist, but also, and more importantly, how to be a good person in science. And finally to Stewart, for absolutely everything, for spending endless hours with me helping with everything, discussing every detail, making me think, and for being always patience and open to see me. I would like to extent my thanks to Chris Carter, for his understanding and help. Thanks to Stewart’s family for accepting me in their house and for showing me such kindness. To the Tasmanian Aquaculture and Fisheries Institute, Marine Research Laboratories, and the School of Aquaculture for their funding support. I could not have done this research without the help of many commercial and recreational fishermen that collected samples and gave me tag returns. I would particularly like to thank Neville Perryman, for collecting so many sharks and for being so helpful. Bryan Hughes offered me great help by sharing his insight and knowledge when studying the gestation period of the sharks. To all the rock lobster section at Marine Research Laboratories, especially, Shane Fava, Craig McKinnon, Pip Cohen and Robbie Kilpatrick for their help on land and aboard the “Challenger”. A big thanks to Caleb Gardner for his kindness and constant support. I am very thankful to all the students at Marine Research Laboratories and members of the endocrine laboratory at the School of Aquaculture, Peter Lee, Hannah Woolcott and Quinn Fitzgibbon. Thanks to all of you for being always so helpful and for creating a fun research environment. Special thanks to Tobias Probst, for his constant help and for making me laugh so much! Matias Braccini and Javier Trovar-Avila, thanks for always being there and for our time together, it has been great have your friendship and your scientific advice. To Sarah Irvine for her constant support, for the fun at the conference, and for always cheering me up; to Michelle Treloar, for her constant encouragement, and to Cass Hunter for being such amazing friend through all these years. I feel so lucky for the times that we have shared. Jayson Semmens, Colin Simperdorfer and Michelle Heupel gave me helpful advice in the analysis of the tracking data. Thanks also to Colin and Michelle for their hospitality when I stayed in U.S.A. Julian Harrington offered me invaluable assistance in the use of the GIS software. Malcolm Haddon, Phil Ziegler, Al Hirst, Barry Bruce and Hugh Pederson all assisted with data analysis, for this I thank you.

vii

I could not have come to Australia without the invaluable help of my very good friends Cecilia Arighi and Sergio Schuchner. Without them, this PhD would be nothing but a dream. My stay in Australia has been a wonderful experience, I have come to know amazing people and I had the chance to make very good friends for life, thanks to all of you. How can I express my gratitude to my friends Louise Ward and Justin Ho? I will be forever grateful for their constant support, for always being there for me, and for making my life so much easier and happier in a new country. To my Argentinean friends, Lali, Marce and Ferchu, for, as usual, being such a good and important friends and for keeping our friendship without seeing me for all these years. To my uncle Juan, my aunty Raquel, my cousins, and specially my brother Alejandro, without your support and love I would not have done it, thank you.

TABLE OF CONTENTS

GENERAL INTRODUCTION ------------------------------------------------------------------------------ 9

CHAPTER ONE: 15

CHAPTER TWO: 18

2.1 INTRODUCTION 19

2.2 MATERIAL AND METHODS 25

2.2.1 Source of samples and data collection 25

2.2.2 Steroid hormone measurement 28

2.2.3 Classification of reproductive stage of the sharks 29

2.2.4 Data Analysis 34

2.3 RESULTS 35

2.3.1 Reproductive development 35

2.3.2 Embryo development 44

2.3.3 Endocrine correlates 46

2.3.4 Seasonality of reproduction 51

2.4 DISCUSSION 57

Females 57

Males 62

Seasonality of reproduction and egg laying behaviour 65

Incubation period 69

CHAPTER THREE: 74

3.1 INTRODUCTION 75

3.2 MATERIALS AND METHODS 78

3.2.1 Source of samples and data collection 78

3.3 RESULTS 83

3.3.1 Size at maturity of all sharks dissected 83

3.3.2 Sharks of known maturity stage (blood taken before dissected) 85

3.3.3 Sharks of unknown maturity 92

3.4 DISCUSSION 98

CHAPTER FOUR: 102

4.1 INTRODUCTION 103

4.2 MATERIAL AND METHODS 106

4.2.1 Acoustic tagging 106

4.2.2 Conventional tagging 115

4.3 RESULTS 118

4.3.1 Acoustic tagging 118

4.3.2 Conventional tagging 140

4.4 DISCUSSION 146

GENERAL CONCLUSIONS ----------------------------------------------------------------------------- 153

REFERENCES 159

9

General introduction


10

The practice of harvesting sharks has a long history. Although sharks were

considered edible prior the twentieth century, records associated with shark captures

were more likely to be related with rituals rather than eating habits (Budker, 1971;

Taylor et al., 2005). In the early part of the 1900s, sharks supported small regional

artisanal fisheries that targeted individual species for specific products, while towards

the 1920s the advance in fishing technology resulted in an increase in harvesting of

chondrichthyans globally (Budker, 1971; Taylor et al., 2005). The commercial

exploitation of sharks dates from the period between the two world wars (1930s to

1940s) when attention was drawn to the demand for shark liver oil stimulating a rapid

growth in shark fisheries (Budker, 1971; Taylor et al., 2005). Since the mid 1980s the

demand for shark products had greatly increased and by the late 1980s shark fisheries

were widespread. A further escalation in exploitation of sharks occurred in the mid-

1990s with the high price and increased demand for shark fins in Asian markets (Castro

and Brudek., 1999).

Currently, chondrichthyan populations around the world are harvested by

commercial, artisanal and recreational fisheries, and are mostly caught as bycatch

(discarded after capture) in the world’s fisheries which target teleost species (Walker,

1998; Stevens et al., 2000). As such, understanding incidental catch and mortality of

bycatch species is becoming an increasing requirement of future ecosystem management

plans (Hall et al., 2000; Pope et al., 2000). Furthermore, because most sharks exist at or

near the top of the food chain (Cortés, 1999; Heithaus, 2004), the removal of upper

trophic level predators from their ecosystem can have food web consequences. In

addition to their primary prey items, trophic cascades can have significant impacts on

non-prey species (Pauly et al., 1998; Stevens et al., 2000; Schindler et al., 2002; Scheffer et

al., 2005). With the fear of rapid depletion of world fish stocks because of over

exploitation, together with the reduction in sharks as top predators, the effects on

marine ecosystems of overfishing could result in an important loss of marine


11

biodiversity (Coleman and Williams, 2000; Mullon et al., 2005). Therefore, understanding

the role of the top predator sharks in the ecosystem are primary requirements for

management and conservation of the shark species but also to accurately address an

ecosystem based fisheries management framework.

It is increasingly being recognized that the life history characteristics of

chondrichthyans (long lived, slow growth and producing few offspring) make this group

a fragile marine resource that is vulnerable to exploitation (Walker, 1998; Dulvy et al.,

2003). Several species of sharks, rays and skates either targeted or caught as bycatch or

byproduct (retained after capture) have demonstrated substantial population declines

over the last 20 years (Pauly et al., 1998; Stevens et al., 2000; Graham et al., 2001; Baum et

al., 2003). By 2006, of the 547 chondrichthyan species listed in the International Union

of the Conservation of Nature’s (IUCN) Red List, 20% are threatened with extinction;

confirming that this taxonomic group is extremely vulnerable to overfishing and is

disappearing at an unprecedented rate across the world (IUCN, 2006). In recognition of

the expanding global catch of chondrichthyans and the potential negative impacts on

chondrichthyan populations, an International Plan of Action for the Conservation and

Management of Sharks (IPOA-Sharks) was adopted by the 23rd session of the United

Nations Food and Agriculture Organisation’s (UN FAO) Committee on Fisheries in

1999 (FAO, 1999; FAO, 2005).

Australia has an extremely rich chondrichthyan fauna with the most recent

taxonomic review estimating that of the 1025 species of chondrichthyans worldwide, at

least 297 species inhabit Australian waters. Of these species more than half (48% of

sharks, 73% of rays) are endemic to Australia (Last and Stevens 1994). In 2001 a Shark

Assessment Report, commissioned by the Department of Agriculture, Forestry and

Fisheries (DAFF), listed 5 species as protected, 6 species as endangered, 6 species as

vulnerable, 21 species as near threatened and 3 species as conservation dependent


12

(DAFF, 2001). As a member of the UN FAO, Australia committed to producing its

own National Plan of Action for the Management and Conservation of Sharks (referred

to as Shark-plan). The Shark-plan was endorsed by the Natural Resource Management

Ministerial Council on 16 April 2004 (DAFF, 2004). The Shark-plan recognises that

while Australia is not a major shark fishing nation, it is acknowledged that sharks are an

important part of the total quantity of Australia’s wild fish production and that

Australian vessels regularly take sharks as target and non-target catch (DAFF, 2004).

This thesis has focused on an endemic Australian shark species, the draughtboard

shark Cephaloscyllium laticeps (Duméril, 1853); that within the order Carcharhiniformes,

belongs to the Scyliorhinidae family. This family, the largest of the shark families,

includes up to17 genera and about 100 species. Particularly, 8 genera and 32 species are

found in Australia (Springer, 1979; Last and Stevens, 1994). The entire family lives in

marine habitats, feeding mainly on small fish and invertebrates. Most of scyliorhinids are

near-bottom dwellers in shallow waters, although a few genera include species that

occur along the continental slopes to depths exceeding 2000 m (Springer, 1979;

Compagno, 1984; Last and Stevens, 1994).

The draughtboard shark is the most common catshark in the coastal areas of

southern Australia, where it is a higher trophic level predator of temperate reefs. It is

particularly found inshore on the continental shelf of southern Australia from the

Recherche Archipielago (Western Australia) to Jervis Bay (New South Wales) down to

at least 60m (Last and Stevens, 1994). Common names for this species are: Australian

swell shark, sleepy joe, and nutcracker shark, and local synonymies are: Cephalloscylium

isabella laticeps and Cephalloscylium isabella nascione (Last and Stevens, 1994).

Draughtboard sharks form a significant component of the southeastern Australian

shark fishery where they are caught as accidental bycatch from rock lobster traps,

demersal trawls, long-lines and gillnets (Frusher and Gibson, 1999; Walker et al., 2005).


13

This species is usually returned to the water and fishing mortality is low due to its

resilience (Brickhill, 2001). In Bass Strait (southern Australia), there was a reduction of

approximately 54% in C. laticeps between 1973-1976 to 1998-2001 period. However, this

reduction has been attributed to commercial fishers avoiding fishing grounds where

these animals are abundant (Walker et al., 2005). There is currently no targeted

commercial fishery for draughtboard shark, although it has recently been marketed in

some areas of Tasmania, where there has been a trend for draughtboard sharks that

have been caught in commercial gillnets to be retained for local consumption as “flake”

(J. Lyle, TAFI Marine Research Laboratories, Hobart. pers. comm). Although caught as

a bycatch, draughtboard sharks are potentially vulnerable to population reduction

through fishing due to their high catchability in either pots or gillnets. Despite being a

common bycatch species, the lack of commercial value has resulted in this species not

being the subject of scientific study. Furthermore, assessing the potential impact of

fishing mortality of this top-level predator in a temperate reef ecosystem is currently

hindered by the poor knowledge of its biology.

The aims of this thesis were to address the biology, ecology and ecosystem role of

draughtboard sharks by studying their reproductive biology, movement patterns and

habitat utilisation. The results of this thesis will both, increasing the knowledge of

draughtboard sharks, but will also be essential to accurately addressing ecosystem based

fisheries management programs in Australian temperate reefs, where draughtboard

sharks share the habitat with other marine species of very high commercial value, such

as the rock lobster Jasus edwardsii (S. Frusher, TAFI Marine Research Laboratories,

Hobart. pers. comm). In addition, as this shark is, in general, not retained as a byproduct

there is a need for a non-destructive sampling methodology to study reproduction on

this species. In consequence, steroid hormones were used as a tool to assess

draughtboard sharks reproductive biology. These results will not only help to


14

understand reproduction of this species, but will also increase the knowledge of the

reproductive endocrinology of chondrichthyan species; an area within this marine group

that still remains highly unknown. Furthermore, because assessing and protecting

threatened species or species living in marine protected areas has become an important

part of global conservation activities (Powles et al., 2000; Blyth-Skyrme et al., 2006), there

is a need to find methodology of addressing the reproductive stage of chondrichthyans

species without killing the animals.

The thesis consists of the general introduction, four descriptive chapters and the

general conclusions. The next chapter, chapter one, describes the area of study. Chapter

two initially describes the reproductive characteristics and development of C. laticeps

obtained thorough anatomical and histological examination of the reproductive organs.

The second part of chapter two correlates reproductive hormones with basic

reproductive parameters to explore the role of steroid hormones on oviparous sharks.

Finally, in the last part of chapter two the seasonal reproductive cycle and embryo

development are described. Chapter three evaluates the use of reproductive hormones

as a non-destructive method to describe reproduction in sharks, and the hormones are

explored in the context of providing information required for fisheries management and

conservation. Chapter four explores the use of acoustic tagging techniques to assess

habitat utilisation and movement patterns and to compare this data with a conventional

tagging project that was also undertaken at the same time. Finally, in the general

conclusions, the information from the previous chapters was used to understand the

linkages between reproduction, movements and habitat selection, essential for

understanding the role of draughtboard sharks in the reef habitat and therefore address

future ecosystem management programs.

15

CHAPTER ONE:

Area of study

Chapter one – Area of study

16

Tasmania is an island located at 39-44°S and 144-149°E (Fig. 1.1 and 1.2a). Three

main bodies of water influence the island (Fig. 1.2b). The East Australian Current

(EAC) flows down the eastern seaboard to the southern tip of Tasmania where it

converges with colder subantarctic waters in a subtropical convergence zone (STC). The

Zeehan Current is an extension of the Leeuwin and flows down Tasmania’s west coast

and around southern Tasmania (Cresswell, 2000). Both the EAC and Zeehan Current

are nutrient poor water originating in sub-tropical regions. In contrast, the sub-antarctic

water mass is nutrient rich (Harris et al., 1987). Water temperatures in Tasmania range

from 10.7 to 18.6 (°C) between summers to winters (Cresswell, 2000).

The main marine habitat surrounding Tasmania is rocky reef formed of sandstone

and granite. While the reef supports a diverse and abundant fauna, seaweed and seagrass

is the predominant living flora (Edgar, 2001).

Fig. 1.1: Map of Australia. Tasmania is located in the south east of Australia.

AUSTRALIA

BASS STRAIT

TASMANIA

10°5'

43°60'

112°5' 154°

Chapter one – Area of study

17

Fig. 1.2: Map of Tasmania showing (a) the location of the main study site and (b) a satalite image demonstrating the main currents influencing Tasmania during autumn. The EAC and ZC extend further south during summer and retreat further north during winter. EAC: East Australian Current, ZC: Zeehan Current, STC: Subtropical Convergence, SAW: Subantarctic Water. NOAA 14 NLSSTC MOSAIC 11 MAR 1998 05 11Z-0653Z COPYRIGHT 1998 CSIRO.

BASS STRAIT

TASMANIA

b

EAC

STC

SAW

ZC

CRAYFISH POINT

RESERVE

TASMAN SEA

SOUTHERN OCEAN

40°00'

43°00'

145°00' 147°00' 148°00'

a

Bruny Island

Derwent Estuary

18

CHAPTER TWO:

Reproduction

Chapter two - Reproduction

19

2.1 INTRODUCTION

Reproduction involves one of the most important events in the life of any living

organism. The primary requirement for successful propagation of any species and their

individuals is the availability to reproduce. Understanding of the overall process of

reproduction requires knowledge of the morphology and physiology of the reproductive

tract, and reproductive strategies and cycles.

In life-history theory, reproductive strategy is defined as a complex mixture of

adapted characteristics designed by natural selection to solve ecological problems

(Stearns, 1976). A series of reproductive strategies has been developed by

chondrichthyan during their long evolutionary history. The general trend in

chondrichthyans reproductive evolution is a progression from oviparity to viviparity,

but within this there is still a great diversity of morphological and physiological

adaptations (Wourms, 1977; Carrier et al., 2004). These reproductive strategies are

expressed through reproductive cycles, which are regulated by a combination of physical

and biological variables to ensure that young fish are produced in the best environment

for their survival (Bromage et al., 2001; Pankhurst and Porter, 2003). Four categories of

reproductive cycles can be defined for chondrichthyan females: 1) species that are

reproductively active throughout the year, 2) species that are reproductively active

throughout the year, but exhibit seasonal periods where a greater proportion of

reproductive activity occurs, 3) species with a well defined seasonal cycle, where animals

are reproductively active for only a portion of the annual cycle, and 4) species that are

pregnant for approximately a full year, after which they spend a year or two non-

pregnant (Wourms, 1977; Hamlett and Koob, 1999; Koob and Callard, 1999). In

chondrichthyan males, sperm production can be either seasonal or occur throughout the

year and can be coupled or not with the mating period (Parsons and Grier, 1992).


20

Among oviparous elasmobranchs, both seasonal and non-seasonal reproductive

activity has been observed. For example, while a clear seasonal reproductive period for

both females and males has been reported in Hemyscyllium ocellatum (Heupel et al., 1999),

in Amblyraja radiata both sexes are reproductively active all year round (Sulikowski et al.,

2005a). Furthermore, male and females of the same species may differ fundamentally in

their reproductive tactics. In species such as Leucoraja ocellata, females are capable of

continuously laying eggs but show a seasonal peak in activity, while males are able to

continuously reproduce throughout the year (Sulikowski et al., 2004).

In the family Scyliorhinidae, species show single or multiple oviparity, as well as

aplacental yolk sac viviparity. Single oviparity, where only one egg case develops in each

uterus, has been recorded in the most primitive genera: Cephaloscyllium, Apristurus,

Scyliorhinus and some species of Galeus. Multiple oviparity, where several egg cases

develop in the uterus and the egg capsule is laid when the embryo reaches a certain

length, has been found in the genus Halaelurus and some species of Galeus (Nakaya,

1975; Springer, 1979; Compagno, 1984). Aplacental yolk sac viviparity, where embryos

are retained in the uterus during the entire period of development and depend solely on

yolk sac reserves, has been reported in some species of Galeus and in Cephalurus cephalus

(Nakaya, 1975; Springer, 1979).

Studies on reproduction of the scyliorhinids are limited to only a few species. In the

oviparous scyliorhinids, both sexes of Apristurus brunneus and Parmaturus xaniurus are

reproductively active throughout the year (Cross, 1988) whereas females of S. canicula

(Craik, 1978; Sumpter and Dodd, 1979) and Galeus melastomus (Costa et al., 2005)

although capable of producing eggs throughout the year, demonstrate seasonal periods

of reproductive activity. Embryo development in the scyliorhinid shark, Scyliorhinus retifer

(Castro et al., 1988) showed an incubation time of 256 days in captivity, while embryo

development in S. canicula (Capapé, 1977; Mellinger et al., 1986; Lechenault et al., 1993)


21

varied (in captivity) from 185 days in warm temperatures to 285 days in colder

temperatures.

Histological examinations of chondrichthyans show that follicular organization in

females is very similar to that of other vertebrate species (Fasano et al., 1989). However,

in chondrichthyan males, the testicular organization has been classified into three

different categories (radial, diametric and compound) (Pratt, 1988) depending on the

origin and propagation of the spermatocysts (the unit of structure and function of the

testis (Callard, 1991b)). The lamnid-alopiid testis type is radial and the testis is

comprised of lobes. The germinal zone is localized in the centre of each lobe and

development of spermatocysts proceeds radially from the germinal zone towards the

end of the lobes. The carcharhinid-sphyrnid testis is diametric; the development of the

spermatocysts proceeds from the germinal zone across the diameter of the testis. The

rajid testis is compound, combining both the radial and the diametric organization of

the testis (Pratt, 1988; Girard et al., 2000).

All scyliorhinid sharks share an external type of ovary (follicles are ovulated into the

body cavity where they reach the ostium (Pratt, 1988)). Ovarian follicles are embedded

under a single layer of generative tissue, and follicles are ovulated into the body cavity

(Pratt, 1988). Some species contain only one functional ovary, while in others both

ovaries are developed (Springer, 1979; Compagno, 1984; Cross, 1988). While the

anatomy of the scyliorhinid males have been less studied than that of females, it is

known that the entire family shares a pair of testes and accessory ducts, including

epididymis, efferent and deferent ducts and seminal vesicles (Springer, 1979; Dodd,

1983; Compagno, 1984). In the only two histological studies of the gonads of

scyliorhinids, Scyliorhinus retifer and S. canicula females had follicular organization similar

to other vertebrates, and males had a diametric type of testis (Dodd, 1983; Pratt, 1988).

Knowledge of vertebrate endocrinology is an essential component of understanding

reproductive processes, as reproductive hormones are involved as either triggers or


22

regulators of all aspects of reproduction. The brain-pituitary-gonadal axis is a cascade

system that regulates the entire reproductive process, promoting gametogenesis and

subsequent gamete maturation (Sherwood and Lovejoy, 1993; Gelsleichter, 2004;

Pankhurst, 2006). The release of gonadotropin releasing hormone (GnRH) by the brain

stimulates the production of the gonadotropins (GTH) from the pituitary gland. These

gonadotropins are released into the circulatory system, reaching the target cell where

they bind with membrane-bound receptors. This gonadotropin-receptor complex

triggers adenyl cyclase, to form cAMP (cyclic adenosine monophosphate), which in turn

activates protein kinases A, leading to the activation or de novo synthesis of steroid

synthesizing enzymes resulting in production of steroid hormones (Eckert, 1988). In

females, the ovary is the primary producer of steroid hormones, producing three main

gonadal steroids: testosterone (T), 17β-estradiol (E2) and progesterone (P4) (Koob and

Callard, 1991; Gelsleichter, 2004). Follicular estrogens are necessary for both hepatic

vitellogenin synthesis and reproductive tract development. Progesterone has

antagonistic actions with estrogens and is considered necessary for ovulation, the

continued maintenance of pregnancy, egg retention, and the simultaneous inhibition of

vitellogenin synthesis (Callard et al., 1991; Tricas et al., 2000; Gelsleichter, 2004). The

role that androgens play in female elasmobranchs is less clear. As in teleost fishes, T is

the precursor for biosynthesis of E2 (Tsang and Callard, 1982; Selcer and Leavitt, 1991;

Pankhurst et al., 1999; Tricas et al., 2000). While some authors have associated T only

with the follicular cycle of oviparous elasmobranchs (Sumpter and Dodd, 1979; Koob et

al., 1986), others have found high T concentrations during the egg retention and egg

laying process (Rasmussen et al., 1999; Sulikowski et al., 2004).

In vertebrate males, the testis is the principal source of reproductive hormones

(Fasano et al., 1989; Pankhurst, 2006) and although the presence of several gonadal

steroids has been reported in male elasmobranchs (Simpson et al., 1964; Callard, 1991a;

Manire et al., 1999), the function of most of these hormones remain uncertain.


23

Testosterone seems to be the primary androgen in elasmobranch males, and may play a

role in development and maturation of spermatocysts, and stimulation of the

development of secondary sex characteristics (Callard et al., 1985; Cuevas and Callard,

1992; Sourdaine and Garnier, 1993; Tricas et al., 2000; Gelsleichter, 2004). However,

other androgens such as dihydroxytestosterone (DHT), 11-ketotestoterone (11-KT),

and 11-ketoandrostenedione (11-KA) have also been reported to play a role in

spermatogenesis (Callard et al., 1989; Garnier et al., 1999; Manire et al., 1999). Although

male elasmobranchs do produce estrogens and P4 (Cuevas and Callard, 1992; Manire

and Rasmussen, 1997; Tricas et al., 2000) and both E2 and P4 receptors have been

identified in elasmobranch testes (Callard et al., 1985; Cuevas and Callard, 1992), their

roles in male reproduction remain unclear. Several authors (Callard, 1991a; Tricas et al.,

2000; Sulikowski et al., 2004) have reported elevated E2 levels with the middle stages of

spermatogenesis, while others (Manire and Rasmussen, 1997; Garnier et al., 1999) have

found no clear variation in E2 concentrations within the male reproductive cycle.

Male elasmobranch production of P4 has been associated with spermiogenesis and

spermiation (Gelsleichter, 2004), which suggests that P4 could act as a precursor for

androgen synthesis (Manire and Rasmussen, 1997; Gelsleichter, 2004). In contrast, P4

was found to peak independently of T in other shark species (Snelson et al., 1997;

Garnier et al., 1999; Gelsleichter, 2004). Studies on Squalus acanthias (Simpson et al., 1963;

Simpson et al., 1964; Callard, 1991a) have found that P4 may primarily be the substrate

for the production of T and other androgens.

The endocrinology of oviparous elasmobranchs has been reported in only a few

species such as Scyliorhinus canicula (Sumpter and Dodd, 1979), Leucoraja erinacea (Koob et

al., 1986), Raja eglanteria (Rasmussen et al., 1999), Hemiscyllium ocellatum (Heupel et al.,

1999) and Leucoraja ocellata (Sulikowski et al., 2004). Of these species, only S. canicula

belongs to the family Scyliorhinidae.


24

The aim of the present study was to investigate the reproductive biology of the

draughtboard shark, Cephalloscylium laticeps, a common species in southeast Australia. The

draughtboard shark belongs to the family Scyliorhinidae (Springer, 1979) and the only

information available states that the species is oviparous and that males reach maturity

at approximately 820 mm total length (Last and Stevens, 1994).

This study explores the anatomy and histology of the gonads, correlations of changes

in gonad condition with plasma levels of gonadal steroids, and assessment of the

seasonality of the reproductive cycle. The majority of the data were collected from

sharks immediately after capture from the wild, however some sharks were also kept in

captivity to address questions associated with embryo development and periodicity of

egg laying.


25

2.2 MATERIAL AND METHODS

2.2.1 SOURCE OF SAMPLES AND DATA COLLECTION

Draughtboard sharks were obtained from two different sources:

1) Commercial and research surveys

A total of 636 females and 468 males were collected throughout Tasmanian

coastal waters as bycatch from rock lobster trap, gillnet and hook fisheries between

June 2002 and April 2004 (Fig. 1.1 and 1.2, Table 2.1).

Date

Locations

Derwent Estuary East Coast South West Coast North West Coast

F M F M F M F M

Jun 2002 - - 3 2 - - - -

Jul 2002 5 25 - - - - - -

Aug 2002 - - - - - - - -

Sept 2002 1 4 - - - - - -

Oct 2002 1 - 16 18 - - - -

Nov 2002 - - - - 55 39 - -

Dec 2002 7 29 - - 34 17 - -

Jan 2003 - - - - 16 20 - -

Feb 2003 - - - - 39 26 - -

March 2003 - 2 39 46 25 24 - -

April 2003 - - - - 71 24 - -

May 2003 5 7 - - - - - -

Jun 2003 - 1 - - 22 21 - -

Jul 2003 - 2 - - 56 22 - -

Aug 2003 - - - - 27 13 - -

Sep 2003 - - - - - - - -

Oct 2003 - - 5 - 2 - - -

Nov 2003 - - - - - - - -

Dec 2003 - - - - - - - -

Jan 2004 4 4 - - - - - -

Feb 2004 6 40 4 - - - - -

March 2004 9 - 10 - 12 - - -

April 2004 1 - 33 27 19 7 106 16

Table 2.1: Number of female and male draughtboard sharks sampled, sorted by date and location of capture in Tasmania.


26

After capture sharks were euthanased by immersion in a benzocaine bath consisting of

0.5 l of benzocaine solution (40 g benzocaine. l ethanol-1) in 8 l of seawater, and the

following measurements were taken from both males and females: total length (mm

TL), total body weight (g TBW), liver weight (g), and stomach weight (g).

In addition, the following data were also recorded for males: calcification, rotation,

and length of the clasper (mm) (from the distal end of the metapterigyum to the tip),

testes weight (g) and weights (g) of the seminal vesicles: prior to and after expression of

any sperm. Ovary weight (g), oviducal gland width (mm) (at the widest part) and weight

(g), presence, condition and length of egg cases were recorded from females. For the

first 20 female sharks, the diameter (mm) of all follicles was measured, after which only

the largest 20 follicles were measured.

The following indices were calculated:

Gonadosomatic Index (GSI)= (Gonadal (testes or ovary) weight/Total weight)*100

Hepatosomatic Index (HSI)= (Liver weight/Total weight)*100

Where total weight= Total body weight – (Gonadal weight + Liver weight + Stomach

weight)

Proportion of sperm within seminal vesicle (PS) = [(weight of seminal vesicle – weight of

seminal vesicle after expression of any sperm)/ weight of seminal vesicle]*100

HISTOLOGY

For histological analysis, ovaries, testes and seminal vesicles were fixed in Bouin’s

solution, embedded in paraffin, sectioned at 7µm and stained with Haematoxylin-eosin.

Stained sections were examined and photographed under a light microscope (Leica

DMLB2 microscope with a Leica DFC 320 camera). Spermatogenic stages in the testis


27

were classified according to the descriptions of (Callard, 1991b) and (Parsons and Grier,

1992 ).

BLOOD SAMPLING

To correlate hormone levels with reproductive condition, blood samples (~3 ml)

from 118 females and 113 males, were collected by caudal venipuncture using pre-

heparinized syringes fitted with 22G needles. After extraction, blood samples were

placed on ice for 3-6 h and then centrifuged for 5 minutes at 8000 rpm. The plasma was

collected and stored at -15°C until thawed for analysis.

2) Captive sharks

Four female and three male sharks caught off Bruny Island (Southern Tasmania)

during January 2003 were held in captivity until December 2004 at Woodbridge Marine

Discovery Centre (southern Tasmania). A female shark caught at Bicheno (east coast of

Tasmania) in April 2004 was held in captivity until July 2005 at the Bicheno Aquarium

on the east coast of Tasmania. At Woodbridge, females and males were placed in a tank

of 7 m length, 3 m width and 1.2 m depth, supplied with water at ambient temperature

(10-11ºC winter and 16-17ºC summer) pumped from the sea. At Bicheno, the female

was held in a tank of 2.2 m length, 2.2 m width and 0.8 m depth, under static holding

conditions with a complete change of water once a month. The water temperature

ranged from 11-12ºC in winter to 21-22ºC in summer. Captive sharks were monitored

for the presence of egg laying and development of the embryos.


28

2.2.2 STEROID HORMONE MEASUREMENT

Levels of 17β-estradiol (E2), Progesterone (P4), Testosterone (T) for both females

and males, and 11-Ketotestosterone (11-KT) in males were measured by

radioimmunoassay (RIA). Plasma samples (200 µl) were extracted twice with ethyl

acetate (1 ml) and 100 µl aliquots were transferred to assay tubes for evaporation prior

to addition of an assay buffer. Assay reagents for E2, T and 11-KT were used as

described by Pankhurst and Carragher (1992). Progesterone was measured using

[1,2,6,7-3H] Progesterone supplied by Amerhsam Biosciences UK Ltd. The antibody is a

polyclonal full serum antibody raised in sheep and was donated by Dr Ken McNatty,

Wallaceville Animal Research Station, Upper Hutt, New Zealand. The assay protocol

used was as described by Pankhurst and Carragher (1992). Steroid assays were validated

by assessment of the slope of serial dilutions of extracted plasma against assay

standards. All samples diluted parallel to standard curves. Extraction efficiency was

determined from recovery of 3H– labelled steroid added to pooled aliquots of plasma.

Extraction efficiencies were 86, 74, 86 and 88% for T, E2, P4 and 11-KT respectively.

Each sample was analysed in duplicate and the assay values were corrected accordingly

to account for the extraction efficiency. The detection limit for all assays was 0.15 ng.ml-

1 plasma. Interassay variability was determined by repeat measurement of a pooled

internal standard and was 13 (9), 11 (9), and 9 (7) (%CV (n)) for T, E2 and P4

respectively. 11-Ketotestosterone data were measured in a single assay.


29

2.2.3 CLASSIFICATION OF REPRODUCTIVE STAGE OF THE SHARKS

Females

In this study the term “follicle” refers to the oocyte and surrounding theca and

granulosa layers prior to ovulation, and the term “ovum” refers to the oocyte after

ovulation.

The ovarian follicles were classified into four different stages based on follicle size,

colour, and histological characteristics as follows: previtellogenic (PV), early vitellogenic

(EV), vitellogenic (V), and mature (M) (Table 2.2, Fig. 2.1).

Based on follicle classification, oviducal gland condition and the presence of egg

cases in the uterus, females were then classified into five different reproductive stages as

shown in Table 2.3 and Fig. 2.2.

Males

Sperm was present in the seminal vesicles of individuals with uncalcified claspers

(Figure 3.4). As calcified claspers are required for copulation (Clark and Von Schmidt,

1965), the presence of sperm could not be used as a reliable indicator of functional

maturity. Therefore, clasper condition (determined by assessing the rigidity of the

clasper by hand) was used to decide the sexual stage of males. Males were classified as

juvenile, sub-adult and adult according to Table 2.4, and Fig. 2.5.

30

Follicle type

(See Fig. 2.1)

Maxim

um Follicle

Diameter (M

FD)

Colour

Histology

Follicular epithelium

Zone pellucida width

Yolk platelets

Previtellogenic (PV)

MF

D <

7 m

m

Ver

y w

hite

S

ingl

e ro

w o

f col

umna

r ce

lls

14-1

6 µm

N

o yo

lk p

late

lets

evi

dent

with

in t

he fo

llicl

e.

Early vitellogenic (EV)

7 m

m ≤

MF

D ≤

10 m

m

Slig

htly

yel

low

P

seud

ostr

atifi

ed

8-10

µm

In

divi

dual

yol

k pl

atel

ets

evid

ent

with

in th

e fo

llicl

e.

Vitellogenic (V)

10 m

m <

MF

D <

30

mm

Y

ello

w

Sin

gle

laye

r of

tall

colu

mna

r

cells

, sl

ight

ly lo

sing

the

pseu

dost

ratif

ied

appe

aran

ce

4-5

µm

Hig

h de

nsity

of y

olk

plat

elet

s w

ithin

the

folli

cle.

Mature (M

) *

MF

D ≥

30

mm

Y

ello

w

No

hist

olog

y du

e to

siz

e of

folli

cle

* 30 mm follicular diameter was the smallest follicle found within the initial egg encapsulation stage in pregnant animals.

Table 2.2: Classification of ovarian follicles based on size, colour and histological characteristics.


31

Female stages Type of MFD

(Refer Table 2.2)

Oviducal gland

Colour Width (mm) ± SE

Juvenile (J) PV Translucent 9 ± 0.5

Sub-adult (Sa) EV Pink 25 ± 0.6

Adult Stage 1 (As1) V Light red 33 ± 0.9

Adult Stage 2 (As2) V or M Dark red > 35

Adult pregnant (Ap) ** M Dark red > 35

Table 2.3: Classification of female sexual stages based on maximum follicular diameter (MFD), oviducal gland characteristics and presence of egg cases. PV: Previtellognic, EV: Early vitellogenic, V: Vitellogenic, M: Mature.

** Pregnant animals were defined by the presence of either partially formed egg cases in the oviducal gland or fully developed egg cases in the uterus.

Figure 2.1: Examples of follicle development (cross-sections stained with haematoxylin-eosin).

a) Presence of yolk inside the follicle. No yolk is present in PV follicle. Yolk started to be distinguished in EV

follicle, and follicle is filled with yolk in V follicle. b) Detail of the follicle walls.

PV: Previtellogenic, EV: early vitellogenic, V: vitellogenic. F: Follicle, ZP: zona pellucida, BM: basement

membrane, FE: follicular epithelium, YP: nucleus of yolk platelets, TH: theca, TI: Theca interna, TE: theca

externa. No histology of mature follicle was possible due to size of the follicle.

PV

V

EV

(b)

(a)

F

ZP

BM

FE

YP

TI

TE

50µm

F

ZP

BM

TH

FE

50µm F

ZP

BM

FE

TI

TE

YP

50µm

100µm

YP

100 µm

Follicle

YP

100µm

Detail of YP


32

For pregnant females, four stages of egg case development were identified (Fig. 2.3)

Stage 1 Stage 2 Stage 3 Stage 4

Oviducal gland

Tendrils

Anterior

Posterior

Egg case

Figure 2.3: Classification of the different egg case developmental stages in draughtboard shark.

Stage 1: Posterior coiled tendrils of the egg case are developed and protrude from the posterior part

of the oviducal gland.

Stage 2: Posterior half of the egg capsule is developed and protrudes from the oviducal gland.

Stage 3: The entire egg capsule is complete, but only the anterior coiled tendrils are still contained

within the oviducal gland. An ovum can be distinguished inside the egg case.

Stage 4: The egg case is free of the oviducal gland. An ovum can be distinguished inside the egg

case.

Figure 2.2: Examples of ovary conditions for the different sexual stages in female draughtboard sharks.

50mm

Juvenile Adult stage 1

Adult stage 2 Adult pregnant Sub-adult

50mm 50mm 50mm 50mm

Follicle

Follicle

Follicle

Follicle

Follicle


33

Male stages Clasper condition

Juvenile (J) Non-calcified

Sub-adult (Sa) Partially calcified

Adult (A) Fully calcified

Table 2.4: Classification of male sexual stages based on clasper calcification.

Figure 2.5: Macroscopic view of the development of the testes in

juvenile, sub-adult and adult draughtboard sharks. Criteria for

classification as given in Table 2.4

Juvenile Sub-adult Adult

50mm 50mm 50mm

Figure 2.4: Seminal vesicle of juvenile draughtboard shark (cross-section stained with haematoxylin -

eosin).

Spermatozoa

Epithelium

0.5 mm

50µm


34

2.2.4 DATA ANALYSIS

Plasma hormone level comparisons were analysed by one-way ANOVA and

subsequent Tukey’s multiple comparison tests (Quinn and Keough, 2002). For this and

all subsequent ANOVAS, residual plots were undertaken to assess the equality of

variances, and data was transformed (square root or logarithmic) where necessary. To

determine the relationship between adult and pregnant females a regression analysis was

made using SPSS. Assessment of reproductive seasonality of oviducal gland weight,

MFD, sperm accumulated in seminal vesicle and GSI in males, was made using one-way

ANOVA and Tukey’s multiple comparison tests. Unless otherwise noted, all data were

analysed using SPSS (SPSS® Base 10.0).

For the analyses of monthly variations in GSI and oviducal gland for females, the

sample size in the period March and April was considerably larger (n >120) than in the

other months. To ensure that sample sizes were not affecting the statistical results, the

ANOVA was repeated 15 times, randomly discarding a number of 50 animals each time.

The significance level was set at P=0.05 for all analysis.


35

2.3 RESULTS

2.3.1 REPRODUCTIVE DEVELOPMENT

Description of reproductive system

FEMALES

The reproductive system of Cephaloscyllium laticeps consisted of a single external

ovary, a single ostium, a pair of oviducts, oviducal glands and uteri (Fig. 2.6). The ovary,

embedded in the epigonal organ, was attached beneath the vertebral column to the

anterior-dorsal body by thin connective tissue. The smallest ovaries contained follicles

of less than 7 mm in diameter with non-discernible yolk. As the follicles grew by

acquisition of yolk, a group of 4-6 yellow follicles of similar size began to differentiate.

Follicles began vitellogenesis at about 10 mm diameter and reached the size for

ovulation at around 30 mm diameter. Follicles were ovulated in pairs and each ovum

enters a separate oviduct.

Macroscopic analysis was unable to separate atretic follicles from corpora lutea

although either or both were present in the ovary.

While the MFD of previtellogenic, early vitellogenic and vitellogenic follicles

generally correlated with juvenile, sub-adult, and adult stage 1 respectively; a small

proportion of vitellogenic follicles were also present in adult stage 2. Mature follicles

were present in both adult stage 2 and adult pregnant stages (Table 2.3 and Fig 2.7).

The relationship between maximum follicular diameter (MFD) and oviducal gland

width was sigmoideal (Fig. 2.8). The oviducal gland initially increased to about 30 mm

with small changes in MFD. Between 30-45 mm oviducal gland width, there was a

substantial change in MFD. The MFD remained high for the latter parts of the oviducal

gland growth (Fig. 2.8).


36

Figure 2.6: Reproductive system of female draughtboard shark. General view of reproductive organs. a) In situ. b) Dissected to show complete reproductive system. c) Cross-section of ovary from a juvenile shark (stained with haematoxylin-eosin).

1 mm

a

b

c

Oviducal gland

ostium

Uterus

Ovary

Cloaca

Egg case Follicle

Epigonal organ

Oviduct

Epigonal organ

Oviducal gland

Ovary


37

Figure 2.7: Size distribution of follicles in the different maturation stages of the draughtboard shark.

0

100

200

300

400

500

600Juvenile

05

101520

200300400500600

Sub-adult

05

101520

200300400500600

Adult stage 1

Follicle diameter

0-2 3-6 7-10 11-13 14-17 18-21 22-25 26-29 30-40

Nu

mb

er o

f fo

llic

les

05

101520

200300400500600

Adult stage 2

Adult pregnant-egg case stage 1



0-2 3-6 7-10 11-13 14-17 18-21 22-25 26-29 30-40



38

MALES

A pair of equally developed testes and genital ducts (epididymis, efferent ducts,

and seminal vesicles) constituted the macroscopic structure of the male reproductive

system. Testes, cylindrical in shape, were enveloped in a thin layer of epigonal organ

(Fig. 2.9). The proportion of epigonal organ attached to the testes decreased from

juvenile to adult (Fig. 2.10). Histological sections revealed a diametric spread of

spermatocyst development from the germinal zone to the efferent duct zone (Fig. 2.10).

Seven stages of spermatogenesis were distinguished (Fig. 2.11). Each of the three male

categories (juvenile, sub-adult, and adult) contained at least some spermatocysts with

Oviducal gland width (mm)

0 10 20 30 40 50 60 70

Ma

xim

um

fo

llic

ula

r d

iam

eter

(m

m)

0

5

10

15

20

25

30

35

40n=555

Figure 2.8: Relationship between maximum follicular diameter and oviducal gland width in the draughtboard shark.


39

spermatozoa although the proportion of spermatocysts containing spermatozoa

substantially increased from juveniles to adults.

Histological examination showed an inverse relationship between the different

spermatogenesis stages and male sexual categories. The proportion of early stages of

spermatogenesis (stage 3 and 4) decreased and the proportion of late stages (stages 6

and 7) increased with the transition from juvenile to adult categories (Table 2.5).

Testis

Epigonal organ

Deferent ducts

Zone of seminal vesicles

Epididymis

Figure 2.9: Reproductive system of male draughtboard shark.


40

Male sexual stages Proportion of spermatogenesis stages

Stage 1+2+3 Stage 4 Stage 5 Stage 6 Stage 7

Juvenile 0.76 0.12 0.06 0.03 0.03

Sub-adult 0.53 0.23 0.07 0.09 0.08

Adult 0.44 0.25 0.07 0.13 0.11

Table 2.5: Percentage of spermatogenesis stages in male draughtboard shark.

Figure 2.10: Histological sections of testes of draughtboard shark stained with haematoxylin-eosin. a: Juvenile, b: Sub-adult, c: Adult, d: detail of germinal zone, e: detail of efferent duct zone.

1mm a

Epigonal organ

Testis

Epigonal organ

Testis

Germinal zone

Efferent duct zone

1mm

c

1mm

Epigonal organ

Testis

b

Early stages of spermatocyst development

20µm

d

100µm

Efferent duct

Spermatozoa

e


41

Figure 2.11: Male testes - microscopic view. Cross-section of testis stained with haematoxylin-eosin. 1: Stage 1, spermatocysts containing spermatoblasts with spermatogonia. Sertoli cells can be seen in the interior of spermatocysts. 2: Stage 2, Sertoli cells (nuclei) are seen migrating from the interior to the periphery of spermatocysts. 3: Stage 3, spermatocysts contain primary spermatocytes. Sertoli cells have completed the migration to the basement membrane. 4: Stage 4, spermatocysts contain secondary spermatocytes. 5: Stage 5, spermatocysts contain spermatids with elliptical nucleus. 6: Stage 6, spermatozoa are developed. 7: Stage 7, head of spermatozoa tightly packed forming a spiral shape. A: detail of 1, B: detail of 2, C: detail of 3. SG: Spermatogonia, SPC: spermatocysts, blue arrows: show the basement membrane, red arrows: show the sertoli cell nuclei, SPM: spermatoblasts, SC1: primary spermatocytes, SC2: secondary spermatocytes, ST: spermatids, SP: spermatozoa.

100µm 7

SP

100µm

SP

6

100µm

4

SC2

50µm

3

SC1

50µm

2

SG

100µm

A SG

50µm B

SG

50µm

C

SC1

1

100µm

SG

SPC

100µm

ST

5

SPM


42

Female and male tissue indices

In females, GSI remained the same for juveniles and sub-adults, but increased

significantly in adult females. In contrast, HSI reached its peak in sub-adult animals (Fig.

2.12). The increase in GSI was most marked between adult stage 1 (As1) and adult stage

2 (As2), where there was a four-fold increase. The HSI steadily increased from juvenile

to As1, after which it significantly declined (ANOVA, P< 0.001). In males, GSI showed

a linear and significant increase from juveniles to adults (ANOVA, P< 0.001), and HSI

was not significantly different at any of the male sexual stages (Fig. 2.12).

The female ovulatory cycle

A general pattern was observed in follicle development in the ovary of female

draughtboard sharks. When the follicle size increased beyond 7 mm diameter, a group

of 4-6 follicles started to differentiate. These developed until they reached

approximately 30 mm diameter when they were ovulated. As these follicles developed, a

second, third and fourth group of 4-6 follicles also started to differentiate. Each of these

size class groups shared the same diameter, and the differences in diameter between

subsequent groups were about 4-5 mm. All pregnant females showed follicles of all size

classes indicating follicular development continued throughout the ovulatory, egg

retention and oviposition cycle (Fig. 2.7).

Observations from captive sharks demonstrated that eggs were released in pairs with

12-24 h intervals between each egg of the same pair. One female, maintained in a tank

by itself for 15 months, laid eggs at routine intervals of approximately 28 days. All the

eggs laid by this shark showed embryonic development, confirming that female

draughtboard sharks stored sperm for at least 15 months.


43

Figure 2.12: GSI and HSI for female and male stages in draughtboard shark. Values are mean + SE. For each index, different letters show significant differences between sexual stages. Juvenile (n= 317, 156) Sub-adult (n= 69, 36) for females and males respectively. Adult males (n=249), Adult stage 1 (n= 41), Adult stage 2 (n=103), Adult pregnant (n=92). GSI: Gonadosomatic Index, HSI: Hepatosomatic index.

Juvenile Sub-adult Adult stage 1 Adult stage 2 Adult pregnant

Ind

ex (

%)

0

5

10

15

20

GSIHSI

a

c

ab

c

aaa

b

cFemales

Sexual stages


Ind

ex (

%)

0

5

10

15

20

GSI

HSI

Males

a

bc


44

2.3.2 EMBRYO DEVELOPMENT

Observations from a single captive shark at Bicheno, showed that the female

swam in circles just before and during oviposition to enmesh the tendrils around any

object protruding from the substratum. Egg cases found by divers in the field (south

and east coast of Tasmania) have always been found attached to objects protruding

from the substratum. Observations from 20 eggs laid at Bicheno, showed that at early

stages of development, the embryo - connected through the yolk stalk to the external

yolk sac of 30 mm diameter - did not show any movement. When the embryo was about

two months old, and had grown to approximately 20 mm in TL, external gills were

visible. At this stage the embryo swam vigorously. The external gills reached their

greatest development at about four months, when the embryo was approximately 50-60

mm TL. At about five months old, the embryo skin developed pigmentation, the

external gills were reabsorbed, and the internal gills became functional. At this stage

mouth movements started. At about six months of age, there was a substantial increase

in total length (to about 120 mm), gill movements increased and the adult shape was

evident. At this stage, the external yolk sac became markedly reduced in size. At about

nine-ten months of age, the yolk stalk disappeared and the external vitelline sac was

about 3-5 mm diameter. Hatching in captivity occurred when the embryo was about 11-

12 months old and had reached 160-180 mm in TL (Fig. 2.13). One of the females kept

in captivity at Woodbridge, laid two eggs that were successfully hatched 13 months later.

The size of these two juveniles was 160 mm and 177 mm TL respectively.


45

Figure 2.13: Embryo development of draughtboard shark. A: Recently laid egg. B, C, D, E, F and G: 1, 2, 4, 5, 6 and 10 months after oviposition. H: Recently hatched embryo. SVE: External vitelline sac. Photos D, E and G by Bryan Hughes.

200mm

A

120mm

H

40mm

D

SVE

150mm

C

SVE

150mm

B SVE

150mm F

i

SVE

50mm E

SVE

60mm

G

SVE


46

2.3.3 ENDOCRINE CORRELATES

Females

Hormone levels varied among fish with different follicle types. Testosterone

began to increase in female sharks with vitellogenic follicles, prior to reaching its highest

levels in fish with mature follicles. Estradiol levels increased steadily with follicle

development reaching a peak in fish with vitellogenic follicles. The lowest levels of P4

were found in early vitellogenic females and the highest levels in sharks with mature

follicles. Similar plasma levels of P4 were found in fish with previtellogenic and

vitellogenic follicles (Fig. 2.14a). Assessment of steroid levels in adult animals showed

that both T and P4 levels were significantly higher in As2 and Ap (ANOVA, P< 0.001),

whereas E2 did not show a significant difference between any of the adult stages (Fig.

2.14b). To assess the possible role of steroids in pregnancy, plasma steroid

concentrations of pregnant females were plotted against the different egg case stages.

Non-pregnant animals that contained MFD ≥ 30 mm (type M) were classified as stage 0

and were included in the plot to compare the hormone levels prior to egg case

development. Due to the low number of pregnant females encountered during the

study, stages 1 and 2, and stages 3 and 4 were combined. Only P4 displayed a significant

change, reaching a peak of 8 ng.ml-1 during the early stages of egg case development

(ANOVA, P< 0.001) (Fig. 2.14c).

Plasma T levels were low in fish with oviducal glands of width < 50 mm and a

significant increase occurred as the oviducal gland reached its maximum size (ANOVA,

P< 0.001). Estradiol increased along with oviducal gland growth. Progesterone showed

a small increase in levels in the latter stages of oviducal gland growth before

substantially increasing in pregnant animals (Fig. 2.15).


47

Figure 2.14: Plasma levels of T, E2 and P4 in draughtboard shark for follicle types (a), for adult sexual stages (b), and for egg case development (c). Values are mean + SE for (a) and (b), and ± SE for (c). For each hormone, different letters show significant differences between follicles, sexual stages and egg cases respectively. PV: Follicle previtellogenic (n= 61), EV: Follicle early vitellogenic (n=10), V: Follicle vitellogenic (n-16), M: Follicle mature (n=27). As1: Adult stage 1 (n=12), As2: Adult stage 2 (n=11), Ap: Adult pregnant (n=20). Stage 0: non-pregnant animals with MFD type M (n=12). Stage 1 and 2 represent the development of the first half of the egg case (n=6) and stage 3 and 4 the completion of egg case development (n=14). T: testosterone, E2: 17β-estradiol, P4: progesterone.

Pla

sma

ste

roid

(n

g.m

l-1)

Sexual stagesAs1 As2 Ap

0

1

2

3

4

5

6

a

b b

A AA

a

b

b

PV EV V M

Follicle type

0

1

2

3

4

5

6T

E2

P4

a

c

b

aA

BB

C

a,b

c

b

a

Egg case stages

0

2

4

6

8

10

a

b

a

aa

A

A

B

a

0 1+2 3+4

T

E2

P4

(a)

(b)

(c)

Egg case stages


48

A diagrammatic summary of the ovulatory and hormonal cycle of the draughtboard

shark is presented in Fig. 2.16. Follicular development occurred in parallel with

ovulation, egg retention and oviposition. Testosterone remained low during the early

stages of follicle development and started to increase in sharks with late vitellogenic

follicles, reaching a maximum in sharks with mature follicles. Estradiol steadily increased

with follicle development up to the late vitellogenic stage. During the last stages of

follicle maturation E2 declined slightly, after which it remained relatively high.

Progesterone remained low until just prior to ovulation, when it rapidly increased. After

ovulation, P4 declined and then remained at a medium level.

Figure 2.15: Plasma steroid levels for oviducal gland width in draughtboard sharks. Values are means + SE. For each hormone, different letters show significant differences between oviducal gland widths. T: testosterone, E2: 17β-estradiol, P4: progesterone.

Oviducal gland width (mm)

Pla

sma

ster

oid

(n

g.m

l-1)

0

1

2

3

4

5

6TE2

P4

a aa

b b

a

b

c

b,cc

a

a,b

a

b

c

<10 ≥10 to ≤ 25 >25 to <50 ≥50 Pregnant

49

M

PV

V

E

V E

2

T

P4

Eg

g r

ete

nti

on

an

d O

vip

osi

tio

n

Ov

ula

tio

n a

nd

e

nca

psu

lati

on

ne

xt

cycl

e

Ov

ipo

siti

on

Fo

llic

ula

r p

has

e

CO

NT

INU

OU

S

20-2

8 D

AY

S I

N

CA

PT

IVIT

Y

30

mm

Fo

llic

le d

ev

elo

pm

en

t

Figure 2.16: A sum

mary of ovulatory and endocrine cycles in female draughtboard sharks. PV: Follicle previtellogenic, E

V: Follicle early vitellogenic, V

: Follicle

vitellogenic, M

: Follicle mature. T: testosterone, E

2: 17β-estradiol, P

4: progesterone.


50

Males

Plasma levels of 11-KT were undetectable (< 0.15 ng.ml-1) in all sharks.

Testosterone showed a significant increase from juveniles to adults (ANOVA, P<

0.001). Estradiol and P4 were detectable but displayed only minor and non-significant

differences between sexual stages (Fig. 2.17).

Figure 2.17: Relationship between plasma steroid levels and male sexual categories. Values are means + SE. Different letters show significant differences between sexual stages. Juvenile (n=54), Sub-adult (n=6), Adult (n=53). T: testosterone, E2: 17β-estradiol, P4: progesterone.

Sexual stages


Pla

sma

ster

oid

(n

g.m

l-1)

0

1

2

3

4

5

6

7

T

E2

P4

a

b

c


51

2.3.4 SEASONALITY OF REPRODUCTION

Females

Females of adult stage 1 formed a greater proportion of the catch in the later part

of the year, while pregnant females were more common in the first half of the year.

There was no seasonal trend in the intermediate stage (As2) (Fig. 2.18). A weak inverse

relationship was found between adult stage 1 and pregnant females (r2=0.54).

Concurrent with the apparent decline in pregnant females, and the increase in adult

stage 1 females throughout the year, there was a decline in both the MFD and oviducal

gland width (Fig. 2.19). Due to the small number of adult females present during most

surveys, samples were combined into bimonthly groups.

The low number of blood samples collected each month resulted in the data being

aggregated into periods of two or three months. In general, T and P4 showed a similar

trend to oviducal gland width and MFD with highest levels from the beginning to the

middle of the year and the lowest levels at the end of the year. E2 was highest at the start

of the year prior to falling to the lowest level in the March-April period (Fig. 2.20).


52

Figure 2.18: Percentage of adult female sexual stages per month in the draughtboard shark. No adult animals were captured in September. N values are the total numbers of sharks of all stages captured during that month. Ap: Adult pregnant, As2: Adult stage 2, As1: Adult stage 1.

Jan Feb Mar Apr May Jun Jul Aug Sep Oct Nov Dec0

20

40

60

80

100

Ap

Per

cen

tag

e o

f a

du

lt f

ema

les

0

20

40

60

80

100

As2

0

20

40

60

80

100

As1

11 15 34 117 3 9 17 5 7 18 7


53

Figure 2.19: Monthly variations of maximum follicular diameter (MFD) and oviducal gland weight in adult draughtboard shark females. Values are means ± SE. For each variable, different letters show significant difference. Numbers are sample sizes. Because of the low sample size for oviducal gland weight for Sep-Oct period, this period was excluded from the statistical analysis.

Months

Jan-Feb Mar-Apr May-Jun Jul-Aug Sep-Oct Nov-Dec

MF

D (

mm

)

15

20

25

30

35

40

45

50

55

60

Ov

idu

cal gla

nd

weig

ht (g

r)

0

10

20

30

40

50

60

70

80

90MFDOviducal gland weight

aa,b

b,c c c

a

a,b

bb

b

a,b

6121 12

2224

6

26 140

22

20

11

4


54

0

1

2

3

4

5

6

7

b

aa

a

P4

Ho

rmo

ne

lev

el (

ng

.ml-1

)

0

1

2

3

4

5

6

7

b

a

b,c

a,c

E2

Months

Jan-Feb Mar-Apr May-Jun-Jul Aug-Sep Oct-Nov-Dec0.0

0.5

1.0

1.5

2.0

13 11 8 10

a

b

a,b

a,bT

Figure 2.20: Mean grouped monthly plasma steroid levels for adult female draughtboard sharks. Values are mean ± SE. Different letters show significant differences between sample times. Numbers are sample sizes. T: testosterone, E2: 17β-estradiol, P4: progesterone.


55

Males

No significant monthly difference was found in the proportion of sperm in the

seminal vesicle (PS) and in the GSI throughout the year (ANOVA, P< 0.001). Due to

the small sample sizes, May and June, August to October, and November and

December samples were grouped (Fig. 2.21).

Average T values in males tended to decline throughout the year although the change

was not significant. Due to small monthly sample sizes, the samples were grouped into

periods of four months (Fig. 2.22).

Figure 2.21: Monthly variations of gonadosomatic index and proportion of sperm in the seminal vesicle of adult draughtboard shark males. Values are means ± SE. There were no significant differences (P> 0.05) between months. Numbers are sample sizes. GSI: Gonadosomatic index, PS: proportion of sperm in seminal vesicle.

Jan Feb Mar Apr May-Jun Jul Aug-Sep Oct-Nov Dec

PS

(%

)

0

10

20

30

40

50

60

GS

I (%)

0

2

4

6

8

10Proportion of spermGSI

17 48 20 37 14 12 11 8 25

Months


56

Figure 2.22: Monthly plasma testosterone (T) levels for adult male draughtboard sharks. Values are mean ± SE. Numbers are sample sizes. There were no significant differences (P> 0.05) between sampling times.

Months

Jan-Apr May-Aug Sep-Dec

Tes

tost

ero

ne

lev

els

(ng

.ml-1

)

3

4

5

6

7

8

16

29

21


57

2.4 DISCUSSION

FEMALES

The entire family Scyliorhinidae displays an external type of ovary (Pratt, 1988)

although the position of the ovary in the coelomic cavity varies among species.

Cephaloscyllium laticeps showed a single ovary located in the middle of the body cavity. In

contrast, several species (Apristurus brunneus, Halaelurus canescens and Scyliorhinus canicula)

have two ovaries present with only one being functional, and others (e.g. Cephalurus

cephalus) have two ovaries that are equally developed (Springer, 1979; Dodd, 1983; Cross,

1988; Balart et al., 2000). Although, the genus Cephaloscyllium is considered to occupy the

most primitive position within the family and Cephalurus is the most advanced group

(Nakaya, 1975; Springer, 1979; Dulvy and Reynolds, 1997), no correlations appear to

exist between reproductive mode and ovarian symmetry (Hamlett and Koob, 1999).

Vitellogenesis commenced in C. laticeps when follicles reach about 10 mm in

diameter. Similar to other elasmobranch species (Dodd, 1983; Storrie, 2004), the

follicular epithelium in C. laticeps began as a simple structure but as the follicle grew and

becomes vitellogenic, the epithelium became slightly pseudostratified. The

differentiation of the theca into internal and external layers in C. laticeps was also

reported in other elasmobranch species (Dodd, 1983; Prisco et al., 2002; Storrie, 2004).

However, in some other species such as Urolophus jamaicensis (Hamlett and Koob, 1999)

the theca remains undifferentiated.

Female draughtboard sharks were reproductively active all year round, showing a

constant overlap between follicular recruitment and development, and egg laying. As a

result, the ovary of adult sharks exhibited the full range of developing follicles in

addition to atretic follicles and corpora lutea from previously ovulated follicles. Hormone

levels were therefore a composite of different stages of follicle development.

Testosterone, E2 and P4 were found in all the different sexual stages although their


58

concentrations varied between these stages. Testosterone, along with E2, were found to

be the main steroids present during follicular development and both declined, although

not to baseline levels, during encapsulation and oviposition. In contrast, P4 was only

found in its highest concentrations during the ovulatory period.

Plasma E2 levels increased during follicle development of C. laticeps, reaching highest

concentrations (mean 4 ng.ml-1) in sharks with vitellogenic follicles, prior to declining

(mean 2.5 ng.ml-1) as follicles reached their maximum sizes. These results were also

correlated with the decrease in the HSI in the later stages of maturity. The substantial

increase in GSI between stages As1 and As2 and the corresponding decline in HSI

suggested that liver resources were being diverted to gonad development at this time.

The increase in E2 concentrations with the progression from previtellogenic to

vitellogenic follicles was related to the increase of steroidogenic activity, and stimulated

the liver to synthesize and release vitellogenin for its uptake by the developing follicle

(Dodd and Sumpter, 1984; Ho, 1987). Similar increases in E2 have been reported for

both oviparous and viviparous sharks (Manire et al., 1995; Heupel et al., 1999; Tricas et

al., 2002; Sulikowski et al., 2004). In vitro studies on oviparous sharks also found the

major production of E2 to be correlated with intermediate sized vitellogenic follicles

(Tsang and Callard, 1982; Koob and Callard, 1991; Callard et al., 1993).

Estradiol levels were found to decline in C. laticeps as follicles reached their maximum

size prior to ovulation, after which E2 levels rose to approximately equivalent levels of

mature follicles prior to egg case development (3-3.5 ng.ml-1). Although a decrease in E2

levels before ovulation was also reported in Leucoraja erinacea, the plasma steroid

concentrations in that species decreased to baseline levels during oviposition (Koob et

al., 1986). The present results also contrasted with seasonal oviparous sharks where E2

was at its highest concentration prior to ovulation (Sumpter and Dodd, 1979; Heupel et

al., 1999; Rasmussen et al., 1999). High levels of E2 before ovulation were also observed

in viviparous sharks (Manire et al., 1995; Snelson et al., 1997; Koob and Callard, 1999;


59

Tricas et al., 2000). The decline in E2 as follicles reach maximum size in C. laticeps may be

associated with changes in plasma levels of T and P4. Plasma T levels increased (mean

1.5 ng.ml-1) during the latter stages of maturation of the follicles of C. laticeps, suggesting

a down regulation of P450 aromatase activity (regulating conversion of T to E2), leading

to accumulation of T rather than its onward conversion to E2. Similar observations in T

levels have been found in several other species of shark (Dodd et al., 1983; Callard et al.,

1993; Manire et al., 1995). Moreover, in teleost fishes, most authors have found

maximum T concentrations just prior to ovulation or during final oocyte maturation

(reviewed in Pankhurst, 2006). Current results found that T levels remained high (mean

1.5 ng.ml-1) after ovulation in C. laticeps. Similar findings were reported for Leucoraja

ocellata and Raja eglanteria (Rasmussen et al., 1999; Sulikowski et al., 2004), but contrast

with the results of (Sumpter and Dodd, 1979; Koob et al., 1986) where T was only

elevated during follicle growth in Leucoraja erinacea and Scyliorhinus canicula. Both the

increase in E2 and the maintenance of high T concentrations after ovulation observed in

the present study, may have resulted from the continual recruitment of follicles into

vitellogenesis that occured during the reproductive process. This is similar to teleost

fishes with asynchronous gamete development where there is often no fall in plasma

steroids after ovulation, as there are further clutches of follicles undergoing

vitellogenesis (Pankhurst et al., 1999).

Several studies have also suggested additional roles that estradiol and androgens may

perform during the reproductive cycle of elasmobranchs. Estradiol has been suggested

to play a role in development and function of the reproductive tract (particularly the

oviducal gland). Gilmore (1983) and Koob and Callard (1991) suggested that endocrine

factors may induce egg capsule secretion, and Reese and Callard (1991) identified an E2

receptor in the oviduct of Leucoraja erinacea. As C. laticeps reproduced throughout the

year, the role E2 plays in the growth of the oviducal gland was difficult to determine

with E2 levels also being associated with recruitment of the next batch of maturing


60

follicles. A possible role of E2 in the storage of spermatozoa by the oviducal gland was

suggested by (Gelsleichter, 2004). This last author reported that females of Sphyrna tiburo

from populations exhibiting high rates of infertility, showed a reduced peak in

preovulatory E2 related with an apparent decline in the viability of stored spermatozoa

by the oviducal gland (Gelsleichter, 2004). As C. laticeps was able to store viable

spermatozoa, the high levels of E2 found in draughtboard sharks after ovulation may

indicate that E2 has also a role in the maintenance of spermatozoa. Estradiol has also

been associated with the expression of relaxin, the hormone required to enlarge the

cervix to allow egg passage during oviposition. Although relaxin has been identified in

the ovaries of several sharks and its effects are considered to be estrogen dependent

(Tsang and Callard, 1983; Callard et al., 1988; Koob and Callard, 1991), this hormone

and its interactions with E2 were not investigated in this study. There were, however, no

signs of a change in E2 in females in the present study with fully encapsulated ova that

would support this hypothesis.

Androgens may also be associated in the regulation of sperm storage by the oviducal

gland. Studies on Sphyrna tiburo females showed that T levels were highest for the 4-5

months between the mating and the ovulatory period suggesting that T is involved in

the regulation of sperm storage by the oviducal gland (Manire et al., 1995). As E2 was

correlated with oviducal gland function, it is possible that the oviducal gland’s ability to

store sperm is regulated by a combination of both E2 and T. Androgens might play a

role in encapsulation and oviposition, as levels of T increased by the onset of breeding

activity and remained high during egg laying in Raja eglanteria (Rasmussen et al., 1999),

and elevated T levels were found during egg case formation and oviposition in Leucoraja

ocellata (Sulikowski et al., 2004). This hypothesis is also supported with the finding in the

present study, where T levels remained high in C. laticeps. Further assessment of the

possible roles of both E2 and T in C. laticeps will require manipulative experiments.


61

Plasma levels of P4 remained low (mean < 1.5 ng.ml-1) during the follicular phase in C.

laticeps, but showed a marked peak (mean 8 ng.ml-1) in animals carrying partially formed

egg cases without an ovum, indicating that ovulation had not yet taken place (Fitz and

Daiber, 1963). A similar peak in P4 levels just before ovulation was observed in other

oviparous species (Koob et al., 1986; Heupel et al., 1999; Rasmussen et al., 1999). After

ovulation, P4 levels in C. laticeps fell, but still remained elevated during egg encapsulation

and oviposition.

Although studies on chondrichthyan endocrinology have advanced in the last few

years, there is still insufficient information to present a unified pattern of endocrine

control that encompasses all the oviparous species. Several studies have shown that the

same steroids hormones appear to behave differently in different oviparous species. For

example, while T concentrations were elevated during egg capsule formation and

oviposition in Raja eglanteria (Rasmussen et al., 1999) and Leucoraja ocellata (Sulikowski et

al., 2004), in Leucoraja erinacea (Koob et al., 1986) T production was reported to be very

low during the egg case production and oviposition. Although there is no explanation

for the differences in endocrine patterns among different species, it is important to note

that while some studies have been undertaken in the wild (Sulikowski et al., 2004;

Sulikowski et al., 2005a), others were done in captivity (Koob et al., 1986). Furthermore,

several authors have reported differences in the reproductive cycle between sharks held

in captivity in aquariums and sharks captured from the wild (Carrier et al., 1994; Heupel

et al., 1999). Heupel (1999) found Hemyscillum ocellatum to adjust its reproductive cycles

from a seasonal cycle in the wild to an annual cycle in captivity. Different factors such as

stress of capture or husbandry could produce differences in hormone levels or patterns

of secretion (Wardle, 1981; Cliff and Thurman, 1984). A decrease in steroid hormone

levels in response to confinement or acute stress were reported for the teleost fish

Onchorynchus nerka and Acanthopagrus butcheri (Haddy and Pankhurst, 1999; Kubokawa et

al., 1999). Sampling strategies, particularly the time of sampling in relation to the


62

reproductive cycle, could also account for differences in the reported roles of these

hormones. However, to date there is insufficient information to understand if the

differences in steroid hormone behaviour for oviparous species are a result of the

sampling methodology or are related to differences between species. Caution should be

applied when comparisons or generalization between oviparous species are made.

MALES

Male C. laticeps had two equally developed testes and reproductive ducts. The

origin and propagation of spermatocysts within the testes was characterised by a

diametric development, and was consistent with other species of carcharhiniformes that

have been studied (Pratt, 1988). In C. laticeps, the proportion of mature spermatocysts

and plasma T concentrations increased with sexual maturation. Similar results have been

found for both oviparous and viviparous elasmobranchs (Rasmussen and Gruber, 1993;

Heupel et al., 1999; Manire et al., 1999; Tricas et al., 2000; Gelsleichter et al., 2002;

Sulikowski et al., 2004). The results of the present study support the view of (Callard et

al., 1985; Sourdaine et al., 1990; Sourdaine and Garnier, 1993) that T plays a major role

in the regulation of testis development. In contrast, detectable levels of 11-KT were not

found in C. laticeps. Although, 11-KT is the main androgen reported for teleost fishes

(Pankhurst, 2006), and despite 11-KT being reported in both Sphyrna tiburo (< 2.21

ng.ml-1) (Manire et al., 1999) and Scyliorhinus canicula (< 0.27 ng.ml-1) (Garnier et al., 1999),

this hormone appears to play no role in C. laticeps.

In C. laticeps, clasper length and testis weight, along with the proportion of

spermatocysts containing spermatozoa, increased with increasing T concentrations.

Similar results have been reported for other elasmobranch species (Callard, 1991a;

Rasmussen and Murru, 1992; Sulikowski et al., 2005b). No androgen receptors have so

far been identified in male reproductive tracts (Callard, 1991b; Conrath and Musick,


63

2002), however, it is unlikely that androgens would not be involved in the development

of male reproductive organs. As androgens have such widespread actions as anabolic

agents in a wide range of vertebrate tissues (Eckert, 1988), it is likely that they would

exert similar effects on chondrichthyans. It also cannot be excluded that other

unmeasured androgens might also play a role in male reproductive development. As

both clasper length and testis weight increased simultaneously in C. laticeps with the

increase in T levels, it is suggested that T regulated both events.

Various authors (Heupel et al., 1999; Tricas et al., 2000) have associated T

concentrations with different stages of spermatogenesis suggesting the possible role of

T in regulating the final stages of sperm maturation. Plasma T concentrations increased

during the middle to late stages of spermatogenesis in some species such as Hemiscyllium

ocellatum (Heupel et al., 1999), Leucoraja ocellata (Sulikowski et al., 2004) and Dasyatis sabina

(Tricas et al., 2000). Cuevas and Callard (1992) reported that androgen receptors were

primarily localized in the early stages of spermatogenesis in the testis of Squalus acanthias,

suggesting that androgens may regulate the development of spermatogonia. Although

the proportion of spermatocysts containing spermatozoa increased from juvenile to

adult animals in parallel with the increase in T concentrations in C. laticeps, more

experimental studies need to be done to understand the role of T and possibly other

androgens, in regulating spermatogenesis.

Estradiol was present at very low plasma concentrations in male C. laticeps (< 0.20

ng.ml-1), similar to levels in S. canicula where plasma concentrations did not exceed 0.05

ng.ml-1 (Garnier et al., 1999). In males of the elasmobranch species Dasyatis sabina and

Sphyrna tiburo, relative changes in E2 levels have been reported throughout the year, but

the absolute value of E2 concentrations (< 0.27 and < 0.072 ng.ml-1 respectively) were

similar to, or lower than for C. laticeps (Manire and Rasmussen, 1997; Tricas et al., 2000).

Several studies have shown elevated E2 levels to be associated with the early to middle

stages of spermatogenesis (Manire and Rasmussen, 1997; Snelson et al., 1997; Tricas et


64

al., 2000). Furthermore, on the basis that estrogen and androgen receptors were found

to be higher in the regions of premeiotic stages of spermatogenesis, (Callard, 1991a)

suggested that intratesticular estrogens and androgens may cooperate in regulating the

early stages of spermatogenesis. Estradiol is also implicated in spermatogonial

proliferation in teleosts (Pankhurst, 2006). Gelsleichter (2004) proposed that circulating

levels of E2 might not reflect its rate of production or function in testis, if the effects of

this hormone were mainly paracrine. Furthermore, studies in S. canicula showed that

plasma E2 reaches maximum values of 0.05 ng.ml-1, while the maximum value in

testicular E2 was 0.50 ng.ml-1 (Garnier et al., 1999). It is not known whether E2 plays a

role in modulating spermatogenesis in C. laticeps; however, any such function is not

reflected in changing plasma levels of E2.

Detectable levels of P4 (~ 1 ng.ml-1) were found in male C. laticeps at all sexual

stages; however, there were no marked changes with changing sexual stage. This

contrasts with Sphyrna tiburo and Negaprion brevirostris where P4 increased with

testicular development (Rasmussen and Gruber, 1993; Manire and Rasmussen,

1997). Because P4 receptors were found to be higher in the post meiotic stage of

spermatogenesis, (Cuevas and Callard, 1992) suggested that P4 is primarily

associated with spermiogenesis and spermiation. Several authors (Manire and

Rasmussen, 1997; Gelsleichter, 2004) have identified P4 as a possible precursor of

androgen synthesis, while others reported that P4 peaks independently of T (Snelson

et al., 1997; Garnier et al., 1999; Gelsleichter, 2004). The role that P4 plays in male

C. laticeps remains unclear. As there was no increase in P4 with sexual development,

it is unlikely that plasma P4 was involved in spermatogenesis, with levels of P4

instead reflecting the rate at which it was being converted to downstream steroid

metabolites such as T.


65

SEASONALITY OF REPRODUCTION AND EGG LAYING BEHAVIOUR

The positive relationship between oviducal gland width and MFD suggests that

the draughtboard shark displayed a continuous breeding cycle, although within this

cycle there were peaks in both MFD and the oviducal gland weight between January

and June indicating that this was a preferred period for egg deposition. Elevated values

of T and P4 also coincided with this period. A greater proportion of adult stage 1

females were found towards the end of the year and these females contained ovarian

follicles in the advanced stages of vitellogenesis with corresponding elevated levels of

E2. This suggests that C. laticeps was reproductively active throughout the year with an

increase in mature egg production in the first half of the year (austral summer and

autumn).

This type of reproductive strategy, where animals are reproductively active

throughout the year but tend to exhibit one or two peaks in activity (Wourms, 1977),

has also been reported in other oviparous elasmobranchs including some scyliorhinid

sharks (Sumpter and Dodd, 1979; Cross, 1988; Richardson et al., 2000; Sulikowski et al.,

2004). In contrast, other oviparous species have well defined annual reproductive cycles

with reproduction restricted to a shorter period of the year. For example, in Hemiscyllium

ocellatum, females were found to lay eggs between August and January (Heupel et al.,

1999), and in Raja eglanteria between January and August (Rasmussen et al., 1999).

On the basis of the limited information available, there appear to be two basic

reproductive strategies in oviparous chondrichthyans. One group has a short incubation

period (less than 6 months), a limited time of sperm storage (at least 8 months), and a

shorter time between ovulation of each successive pair of eggs (less than about 1 week).

This group displays a seasonal reproductive cycle and tends to be found at lower

latitudes. In contrast, higher latitude species tend to be able to reproduce all year round,

have longer incubation periods (longer than 6 months), are able to store sperm for


66

longer period (at least two years) and oviposition of successive pairs of eggs occurs at

longer intervals (longer than one week) (Table 2.6). Similar results were found by

(McLaughlin and O'Gower, 1971) in their study on the genus Heterodontus, suggesting

that the breeding season is longer among cool water species. However, in

chondrichthyan males, there is no clear trend between the reproductive cycles and fish

from low or high latitudes. For C. laticeps males, the lack of marked annual changes in

GSI, proportion of spermatozoa in the seminal vesicles, or T levels would suggest that

males are able to produce spermatozoa all year round. Other high latitude oviparous

species such as Leucoraja ocellata and Amblyraja radiata, also continuously produce mature

spermatocysts throughout the year (Sulikowski et al., 2004; Sulikowski et al., 2005a). In

contrast, for morphological and hormone data some species from high and low latitudes

such as Hemiscyllium ocellatum and Scyliorhinus canicula show clear seasonality in their

reproductive cycles (Garnier et al., 1999; Heupel et al., 1999). Males of H. ocellatum were

reported to have red and swollen claspers (indicating the mating season) from July to

November, with a peak in androgen concentrations from July to October (Heupel et al.,

1999). In S. canicula males, both gonadal activity and T concentration were found to

peak in winter (Garnier et al., 1999; Henderson and Casey, 2001).

Among teleosts, fish at higher latitudes tend to have a markedly seasonal and

synchronised reproductive cycle within the population, while species at lower latitudes

are likely to display multiple spawning and less population synchrony (review in

(Pankhurst, 2006)). Teleost fishes tend to produce large numbers of eggs and larvae that

are dependent on environmentally driven phytoplankton and zooplankton production

cycles to survive. These cycles are more seasonal in higher latitudes. In contrast,

oviparous chondrichthyan young hatch as small juveniles and do not undertake a larval

phase that is dependent on seasonal planktonic production cycles. The fact that high

latitude elasmobranchs do not show seasonality suggests that the large size at hatching

uncouples juveniles from dependence on seasonal production cycles.


67

There are still significant gaps in understanding the process by which the

environmental signals are transmitted into the endocrine process that control

reproduction. However, in teleost fishes, there is evidence that the main factor driving

high latitude species is photoperiod, followed by temperature and social interactions,

while in low latitudes species this hierarchy may be inverted (Pankhurst and Porter,

2003).

In chondrichthyans, one possible explanation for seasonality among low latitude

species relates to sperm storage. In oviparous species found in warm waters the period

of sperm storage is usually coincident with the winter months (Rasmussen et al., 1999)

(Table 2.6). Furthermore, studies on androgen concentration and sperm production by

males from warm water species, showed that there is an inverse relationship between

androgen concentrations and temperature (Garnier et al., 1999; Heupel et al., 1999), and

that sperm production and its accumulation by the seminal vesicle is higher during the

winter months (Heupel et al., 1999). It could be hypothesised that the short periods of

sperm storage reported for elasmobranch females in lower latitudes may reflect the

inability of sperm to survive for long periods at elevated temperatures. An effect of this

type would truncate the period of reproduction among low latitudes species.

It is important to note that other selective forces such as juvenile mortality, growth

rate, size and age at maturity, offspring size, fecundity and longevity will influence the

differences in life history strategies of chondrichthyan species between high and low

latitudes (Stevens, 1999; Frisk et al., 2001; Cortés, 2004). However, the discussion of

these parameters is beyond the scope of the present study.

The relationship between female and male reproductive cycles can vary. While both

sexes of several species have a synchronized reproductive cycle (Heupel et al., 1999;

Kyne and Bennett, 2002; Sulikowski et al., 2004), there is an un-coupling of reproductive

activity in others (Ellis and Shackley, 1997; Henderson and Casey, 2001). In Scyliorhinus

canicula, the cycle of females and males was not synchronised with GSI peaking during


68

May in females, and November and December in males (Henderson and Casey, 2001).

In Hemyscillum ocellatum, females lay eggs from August to January and males have the

highest volume of sperm in the epididymis during August to November, indicating a

synchronous cycle (Heupel et al., 1999). For females that are able to store sperm (Pratt,

1993), there is no necessity to have a synchronous reproductive cycle between both

sexes. In the case of C. laticeps, females and males presented unsynchronised cycles.

Although females were able to reproduce all year round, a peak in egg deposition was

found between January to June. However, males did not show a peak in sperm

production at any time of the year.

Observation of C. laticeps held in captivity for 15 months, showed that females were

able to store sperm for this extended period. Although, migratory behaviour, peaks in

sexual aggregations and the benefit of different copulation and fertilization times have

all been hypothesized to explain sperm storage in vertebrates (Dodd et al., 1983;

Birkhead and Moller, 1993; Pratt, 1993; Conrath and Musick, 2002), there is limited

biological information available for C. laticeps to support any of these theories. However,

the presence of mature adult females and males throughout the year would suggest that

sperm storage was not associated with disaggregation of the sexes.

Studies on the mating behaviour of scyliorhinid sharks show that the male initially

bites the tail and then the pectoral axilla of the female (Castro et al., 1988). No mating

scars have been distinguished in any of the female C. laticeps sampled. However, the skin

of the draughtboard shark is very thick and this could prevent the damage that gives rise

to mating scars in other species. On the base of high levels of androgen during the

mating season, T has been associated with copulatory activity in elasmobranch species

(Rasmussen and Gruber, 1993; Heupel et al., 1999; Tricas et al., 2000). However, Crews

(1984) and Parsons and Grier (1992) suggest that a peak in testicular development or

circulating levels of gonadal hormones may not necessarily coincide with the peak in the

mating season. In species such us Mustelus griseus and Mustelus manazo there is a 6 month


69

delay between the peak in GSI and the mating season (Parsons and Grier, 1992).

Currently the mating season (if there is one) in C. laticeps is not known.

INCUBATION PERIOD

The different stages of embryonic development in C. laticeps were similar to those

reported for other scyliorhinids (Mellinger et al., 1986; Castro et al., 1988). In C. laticeps,

the incubation period in captivity was similar to the incubation time suggested by Castro

(1998) for Scyliorhinus retifer in the wild. In species from temperate waters such as

Leucoraja erinacea (Richards et al., 1963) and Raja clavata (Ellis and Shackely, 1995),

embryo development takes between six months to one year; while in species from warm

waters such as Raja eglanteria (Luer and Gilbert, 1985) and Hemiscyllium ocellatum (West

and Carter, 1990) the incubation period takes around four-five months (Table 2.6).

However, incubation time in captive Scyliorhinus canicula varied according to the

temperature of the aquarium, being shorter (180 days) for egg deposited in warm water

compared to those deposited in cold waters (285 days) (Capapé, 1977). For this species,

the increased water temperature was considered to increase the metabolic rate of

development of the embryo. For sharks with incubation times of less than 12 months,

the amount of time the egg is exposed to warmer (summer) or cooler (winter)

temperatures may account for variability in incubation times. For C. laticeps, with a

incubation time of approximately 12 months, temperature-generated variation in

incubation period was unlikely as each egg would appear to experience the same total

exposure to winter and summer temperatures, irrespective of the time of oviposition.

This assumes that temperature-dependent effects on development do not change with

stage of development.


70

In summary, female C. laticeps presented an external type ovary with follicles starting

vitellogenesis at 10 mm diameter and maturing at 30 mm. Testosterone and E2 played a

major role during the follicular phase, while P4 peaked during the ovulatory phase.

Females were reproductively active all year round with a seasonal period between

January to June where a greater proportion of eggs were laid. Deposition of the eggs

occurred once a month, and the incubation period is about 12 months. Male C. laticeps

presented diametric type testes. Testosterone played a major role changing according to

the sexual stage. Males were able to produce sperm all year round. The mating season

for this species (if there is one) remains to be determined.

71

Species name

Reproductive

activity

Females

Sperm

storage

Tim

e betw

een

laying

Tim

e betw

een

each egg

Incubation tim

e Latitude

Location

Authors

C. laticeps

Yea

r ro

und;

pea

k in

Ja

n -

June

A

t lea

st 1

5 m

onth

s 20

-28

days

12

-24

hrs

12 m

onth

s H

igh

Tas

man

ia

(Thi

s st

udy)

R. eglanteria *

Jan-

Aug

A

t lea

st 3

m

onth

s 4-

5 da

ys

Min

-hou

rs

3 m

onth

s Lo

w

Cap

tivity

(20

-22°

C)

(Lue

r an

d G

ilber

t, 19

85)

Y

ear

roun

d; p

eak

in

sprin

g N

o da

ta

No

data

N

o da

ta

3 m

onth

s H

igh

Del

awar

e B

ay, U

SA

(9°

C)

(Fitz

and

Dai

ber,

196

3)

R. clavata

Yea

r ro

und

No

data

N

o da

ta

No

data

4.

5-5.

5 m

onth

s (1

2-18

°C)

Hig

h P

lym

outh

, Eng

land

(C

lark

, 192

2)

N

o da

ta

At l

east

15

wee

ks

0-2

days

N

o da

ta

19 w

eeks

H

igh

Cap

tivity

(14

.9°C

) (E

llis

and

Sha

ckel

y, 1

995)

N

o da

ta

No

data

48

hrs

24

hrs

N

o da

ta

Hig

h C

aptiv

ity (

11-1

6°C

) (H

olde

n et

al.,

197

1)

Ja

n -S

ep

No

data

N

o da

ta

No

data

N

o da

ta

Hig

h S

outh

ern

Nor

th S

ea

(Hol

den,

197

5)

S. retifer

No

data

A

t lea

st 8

43

days

15

day

s F

rom

min

utes

up

to

8 da

ys

8.5

mon

ths

Hig

h C

aptiv

ity (

11.7

-12.

8°C

) (C

astr

o et

al.,

198

8)

A. brunneus

No

data

N

o da

ta

No

data

N

o da

ta

14 m

onth

s (1

0°C

) H

igh

Brit

ish

Col

umbi

a, C

anad

a (J

ones

and

Gee

n, 1

977)

P. xaniurus

Yea

r ro

und;

pea

k in

D

ec-M

ay

No

data

N

o da

ta

No

data

N

o da

ta

Hig

h C

alifo

rnia

, US

A

(Cro

ss, 1

988)

H. regani

Yea

r ro

und

No

data

N

o da

ta

No

data

N

o da

ta

Hig

h C

ape

Tow

n (9

8-51

5m)

(Ric

hard

son

et a

l., 2

000)

Table 2.6: Reproductive information on oviparous species. Data was not included in the table when it was considered unreliable due to sampling strategies or small sample size.

*Raja eglanteria lives either in both high and low latitudes.

** No data on water temperature was available; but most of the studies by Dodd were in captivity in cold water.

72

S. canicula

Yea

r ro

und

No

data

N

o da

ta

No

data

27

1-28

5 da

ys (

14-1

9°C

)

177-

180

days

(19

-24°

C)

Hig

h T

unis

ia

(Cap

apé,

197

7)

Y

ear

roun

d; p

eak

in

May

N

o da

ta

No

data

N

o da

ta

No

data

H

igh

Wes

t coa

st o

f Ire

land

(H

ende

rson

and

Cas

ey,

2001

)

N

o da

ta

Mor

e th

an 2

ye

ars

No

data

N

o da

ta

No

data

H

igh

No

data

but

pro

babl

y co

ld

wat

ers*

* (D

odd

et a

l., 1

983)

N

o da

ta

No

data

15 d

ays

No

data

N

o da

ta

Hig

h C

aptiv

ity (

14°C

) (M

elli

nger

, 198

3)

Y

ear

roun

d; p

eak

in

sprin

g an

d su

mm

er

No

data

N

o da

ta

No

data

N

o da

ta

Hig

h P

lym

outh

, Eng

land

(F

ord,

192

1)

10

mon

ths;

pea

k in

Ju

ne-J

uly

No

data

15

day

s to

on

e m

onth

N

o da

ta

5-6

mon

ths

(8.5

-18.

1°C

) H

igh

Brit

ish

wat

ers

(E

llis

and

Sha

ckle

y, 1

997)

N

o da

ta

No

data

N

o da

ta

No

data

17

0-20

0 da

ys

Hig

h C

aptiv

ity (

16°C

) (M

ellin

ger

et a

l., 1

986)

L. erinacea

Yea

r ro

und;

pea

k in

N

ov-J

an a

nd J

une-

Jul

No

data

7

days

(1

6.3-

20.3

C)

No

data

6-

8 m

onth

s H

igh

Con

nect

icut

and

Rho

de

Isla

nd

(Ric

hard

s et

al.,

196

3)

Y

ear

roun

d; p

eak

in

sprin

g an

d fa

ll N

o da

ta

No

data

N

o da

ta

6 m

onth

s H

igh

Del

awar

e B

ay, U

SA

(<

15°C

) (F

itz a

nd D

aibe

r, 1

963)

H. ocellatum

Aug

-Jan

N

o da

ta

No

data

N

o da

ta

No

data

Lo

w

Her

on Is

land

, Aus

tral

ia

(21-

28°C

) (H

eupe

l et a

l., 1

999)

Y

ear

roun

d N

o da

ta

No

data

N

o da

ta

4 m

onth

s Lo

w

Cap

tivity

(25

°C)

(Wes

t and

Car

ter,

199

0)

A. radiata

Yea

r ro

und;

pea

k in

S

epte

mbe

r N

o da

ta

No

data

N

o da

ta

No

data

H

igh

Wes

tern

Mai

ne, U

SA

(S

ulik

owsk

i et a

l., 2

005a

)

L. ocellata

Yea

r ro

und

peak

in

No

data

N

o da

ta

No

data

N

o da

ta

Hig

h W

este

rn M

aine

, US

A

(Sul

ikow

ski e

t al.,

200

4)

73

G. melastomus

Yea

r ro

und,

pea

k in

su

mm

er a

nd w

inte

r N

o da

ta

No

data

N

o da

ta

No

data

H

igh

Sou

th o

f P

ortu

gal

(Cos

ta e

t al.,

200

5)

P. extenta

Yea

r ro

und;

pea

k in

su

mm

er

No

data

N

o da

ta

No

data

N

o da

ta

Hig

h P

uert

o Q

uequ

én,

Arg

entin

a (B

racc

ini a

nd

Chi

aram

onte

, 200

2)

Y

ear

roun

d N

o da

ta

No

data

N

o da

ta

No

data

H

igh

Nor

ther

n co

ast o

f Sao

P

aulo

, Bra

zil

(Mar

tins

et a

l., 2

005)

R.maculata

Yea

r ro

und

No

data

N

o da

ta

No

data

5

mon

ths

(13-

18°C

) H

igh

Ply

mou

th, E

ngla

nd

(Cla

rk, 1

922)

R. brachyura

Yea

r ro

und

5-6

wee

ks

No

data

N

o da

ta

4 m

onth

s (9

-18°

C)

Hig

h P

lym

outh

, Eng

land

(C

lark

, 192

2)

R. microocellata

No

data

N

o da

ta

No

data

N

o da

ta

5-7

mon

ths

Hig

h C

aptiv

ity (

14.2

-16.

3°C

) (K

oop,

200

5)

H. portusjacksoni

Aug

and

Sep

. N

o da

ta

App

rox

13

days

N

o da

ta

No

data

Lo

w

Cen

tral

coa

st o

f New

S

outh

Wal

es, A

ustr

alia

(M

cLau

ghlin

and

O

'Gow

er, 1

971)

C. plagiosum

No

data

N

o da

ta

No

data

N

o da

ta

4 m

onth

s Lo

w

Cap

tivity

(24

-26°

C)

(Tul

lis a

nd P

eter

son,

20

00)

74

CHAPTER THREE:

Non-lethal assessment of reproductive parameters:

draughtboard shark - a case study

Chapter three - Non-lethal assessment of reproductive parameters

75

3.1 INTRODUCTION

The reduction and collapse of global fish stocks due to over exploitation is

increasing, with several species nearing extinction (Dulvy et al., 2003; Cortés, 2004;

Mullon et al., 2005). These declines have called for conservation strategies to be

developed for marine resources such as; implementing fisheries management policies,

establishing a global system of marine protected areas (MPAs) where fisheries are

restricted, or declaring some species as threatened or endangered where their capture is

prohibited. Currently, due to the potential for chondrichthyans to be strongly

susceptible to overfishing, the impact on fishing chondrichthyan species around the

world is the focus of considerable international concern (Stevens et al., 2000).

Chondrichthyan populations are harvested by commercial, artisanal, and recreational

fisheries (Bonfil, 1994; Walker, 1998) and while some species are the direct target of the

fishery others are taken as bycatch. It is commonly accepted that chondrichthyans have

slow growth, long life span, late sexual maturity, a low fecundity, long gestation period

and low natural mortality compared to teleost fish (Cortés, 2000; Stevens et al., 2000).

These life history strategies make this group very vulnerable to high levels of fishing

pressure and have led to a number of conservation and management strategies in an

attempt to protect chondrichthyan populations from decline (Simpendorfer and

Donohue, 1998; Stevens et al., 2000; Musick, 2004).

In order to manage chondrichthyan species, it is necessary to develop demographic

models that address their vulnerability to exploitation. Understanding their life history

strategies, particularly their reproductive cycles, is fundamental if species are to be

managed so that they reproduce to maintain appropriate population levels. Knowledge

of the size at which animals mature, is required to ensure that the species has sufficient

time to replace the stock prior to being harvested or impacted upon by fishing, and the

spatial and temporal timing of reproduction is therefore essential for sustainable


76

fisheries management to ensure that fishery activities are minimised during reproductive

periods.

Clasper calcification is the most common external method used to assess sexual

maturity in male chondrichthyans (Clark and Von Schmidt, 1965). However, not all

species (eg: seven gill sharks) alter the degree of calcification in their claspers as they

reach maturity, therefore the sacrifice of these males is necessary. In females, as

macroscopic examination of the ovaries from dissected animals is the only method to

assess sexual maturity, the sacrifice of females is always required. However, there are

many circumstances where killing the animal is inappropriate as in the case of

endangered or protected species, or species residing in MPAs. Similarly it may be

inappropriate to sacrifice bycatch species that would normally be returned to the water

alive. For studies on the reproductive biology and management of these species there is

a need to obtain data on reproduction without the requirement to kill the animal.

Furthermore, any investigation of the temporal and spatial timing of reproduction, for

both sexes, currently requires the examination of gonadal condition after dissection of

the animal.

Gonadal steroids, obtained from blood samples, could be used as endocrine markers

to determine the reproductive status of sharks without the need to kill and dissect the

shark. Only a few studies have compared the levels of plasma steroid hormones

between juvenile and adult chondrichthyans, and all of these suggest that hormones

could be used as an indicator of maturation status (Rasmussen and Gruber, 1990;

Rasmussen and Murru, 1992; Rasmussen and Gruber, 1993; Gelsleichter et al., 2002).

However, despite these results, only one study has linked plasma steroid hormones to

histological and morphological studies of the gonads to address size at onset of sexual

maturity (Sulikowski et al., 2005b).


77

This study has demonstrated that changes in plasma levels of reproductive hormones

are associated with maturation for both sexes in C. laticeps, and that reproductive

hormones reflect the temporal timing of reproduction (see chapter 3). This chapter

examines whether the endocrine markers, testosterone (T), 17β-estradiol (E2) and

progesterone (P4) could be used as an unambiguous indicator of sexual maturity in both

males (where gonadal sexual maturity might occur in advance of clasper calcification)

and females (where there are no external morphological markers of maturation), and

therefore eliminate the need for sacrificing sharks for subsequent macroscopic

examination of the gonads. The results from the assessment were then applied to

draughtboard sharks sourced from a marine protected area where only non-destructive

sampling methods are appropriate.


78

3.2 MATERIALS AND METHODS

3.2.1 SOURCE OF SAMPLES AND DATA COLLECTION

Draughtboard sharks were obtained from two different sources:

1) Commercial and research surveys: Animals from these surveys (see chapter 3,

section 3.2.1) were used to calculate size at maturity and to validate plasma steroid levels

against macroscopic examination of the gonads.

2) Surveys at a marine reserve: Eighty-two females and 54 males were caught between

May 2002 and May 2003 using rock lobster traps in the Crayfish Point Reserve in

southern Tasmania (Fig. 1.2a). Total length and total weight for each sex and clasper

length for males were recorded. Blood samples (as described in Chapter 2, section 2.21)

were taken prior to releasing the sharks.

For all sharks, steroid hormones were measured as described in section 2.2.2.

3.2.2 DATA ANALYSIS

To determine the maturity of sharks, from the marine reserve of unknown

maturation stage, that were released immediately after taking blood, plasma hormone

concentrations were compared with the hormone concentrations from sharks of known

maturity stages.


79

Size at maturity of sharks dissected

Reproductive stages of the sharks were described in section 2.2.3. For this chapter

adult females (As1, As2 and Ap) were combined into a single adult group. For both

sexes, juveniles and sub-adults were combined into a single group called juveniles.

To establish size at maturity of all sharks sampled in this study, oviducal gland width

(for females) and clasper length (for males) were compared to total length. Oviducal

gland width and clasper length were chosen as they were morphological parameters that

progressively grew with maturity, and were independent of the reproductive cycle. In

contrast, gonadal weight varied within mature animals depending on the cyclic

gametogenesis stage of the ovary or testis.

To determine the size at which 50% of the sharks were mature, animals were

grouped as either juvenile or adults. Sharks were grouped into 25 mm length-classes

ranging from 170 to 1020 mm. For dissected sharks, clasper calcification (males) and

macroscopic examination of the gonads (females) were used to distinguish between

juveniles and adults. A logistic regression was applied to each sex separately. The

proportion of adult animals (P) at 25 mm length class was obtained using the following

equation (Neter et al., 1990).

Equation 3.1

Where a and b are constants and x is the medium value of the length-class. Confidence

intervals around the logistic model were obtained by conducting 1000 simulations in a

bootstrapping routine where data were randomly sampled with replacement for each of

the 25 mm length classes (Turner et al., 2002). The middle 95% of the bootstrap

replicates constituted the confidence intervals. Values of P and the 95% confidence

limits were obtained from equation 3.1 using Excel (Microsoft® Excel 2000).

P=e (a+bx)/ [1+e (a+bx)]


80

Sharks of known maturation stage

LINEAR DISCRIMINANT PREDICTIVE MODEL (LDPM)

For both sexes, weighted averages of the predictive variables: total length (TL),

testosterone (T), 17β-estradiol (E2), and progesterone (P4) (for females) and clasper

length (CL), T, E2, and P4 (for males), were used to obtain discriminant function scores

(D) to distinguish juveniles from adult sharks. Discriminant function scores (D) were

calculated as follows:

D = Bo + B1X1 + B2X2 + …+ BiXi Equation 3.2

Where Xi is the value of each independent variable (i) and Bi is the coefficient estimated

from the data. From the discriminant scores it was possible to obtain the probability

that a shark was either a juvenile or adult. This probability P(Gi/D) was estimated by:

( ) ( )∑=

= g

i

GiPGiDP

GiPGiDPDGiP

1

/

)()/()/( Equation 3.3

Where P (Gi) is the prior probability and is an estimate of the likehood that a shark

belongs to a particular group (juveniles or adults). The prior probability was calculated

as the observed proportion of sharks in each group. The conditional probability P

(D/Gi) is the probability of obtaining a particular discriminant function value of (D) if

the shark belongs to a specific group. To calculate this probability, normal probability

theory (the D scores are normally distributed for each group) was assumed. Each shark

was known to belong to a particular group, and the conditional probability of the

observed (D) score given membership in the group was calculated.


81

The predictive function was built using Excel and SPSS (SPSS® Base 10.0).

MULTI-DIMENSIONAL SCALING (MDS)

For both sexes, a multidimensional scaling (MDS) ordination based on the

variables T and E2, (for females), and T and CL (for males) was used to separate

juveniles and adults using normalized Euclidean distances. Data were transformed when

necessary. To test the null hypothesis that there were no assemblage differences

between groups (juveniles and adults) in the spatial matrix, a one-way analysis of

similarities (ANOSIM) and a Pairwise test were performed. The MDS and ANOSIM

were performed using the Primer software package (Clarke and Gorley, 2001). Adults

were separated using a 95% cut off line. The line was calculated as the position on the

MDS ordination where 95% of adults were correctly classified.

The significance level was set at P=0.05 for all data analyses.

Size at maturity

To determine the size at which 50% of the sharks were mature, animals were

grouped as either juvenile or adult using LDPM and MDS analysis. Sizes at maturity

estimates were calculated for each method using equation 3.1.


82

Sharks of unknown maturation stage

To determine the size at maturity, sharks were classified as either juvenile or adult

based on their (D) scores using LDPM or on their MDS ordination. Size at 50%

maturity was calculated using equation 3.1 for both methods.

Reproductive cycle

For both sexes, differences in the proportion of adult sharks that came from the

marine reserve and from the rest of Tasmania were compared using a Chi-Square test

(Quinn and Keough, 2002).

Hormone comparisons were analysed by one-way ANOVA and Tukey’s multiple

comparison tests (Quinn and Keough, 2002). Residual plots were undertaken to assess

the equality of variances and data were transformed where necessary.

All data were analysed using SPSS and the significance level was set at P=0.05 for all

data analyses.


83

3.3 RESULTS

3.3.1 SIZE AT MATURITY OF ALL SHARKS DISSECTED

In females, oviducal gland width increased exponentially between 750-850 mm TL

(Fig. 3.1a). The largest juvenile female found was 850 mm TL and the smallest adult was

730 mm TL. For males, clasper length showed a steady increase as the animal grew until

715 mm TL (Fig 3.1b). From 700-780 mm TL, clasper length rapidly increased (Fig.

3.1b). The largest juvenile male recorded was 830 mm TL and the smallest adult male

was 725 mm TL. Size at 50% maturity of females was estimated at 815 mm TL (95%

confidence interval = 812.58 – 842.79, r2=0.80, n=609), and 761 mm TL (95%

confidence interval = 754.98 – 789.60, r2=0.84, n=462) for males (Fig. 3.1c).


84

Figure 3.1: Changes in (a) oviducal gland width (females) and (b) clasper length (males) with total length. Male claspers were classified as non-, partially and fully calcified for juveniles, sub-adults and adults respectively. Males mature at smaller sizes than females (c).

Ov

idu

cal

gla

nd

wid

th (

mm

)

0

10

20

30

40

50

60

70

a

n=609

Adult

JuvenileSub - adult

Cla

sper

len

gth

(m

m)

0

10

20

30

40

50

60

70

80

90

b

n=462

Adult

JuvenileSub - adult

Total length (mm)

100 200 300 400 500 600 700 800 900 1000

Pro

po

rtio

n m

atu

re

0.0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

1.0

c

Males (n=462)Females (n=609)


85

3.3.2 SHARKS OF KNOWN MATURITY STAGE (BLOOD TAKEN BEFORE DISSECTED)

Linear discriminant predictive model (LDPM)

FEMALES

Discriminant function analysis using TL, T, E2 and P4 showed significant

differences between juvenile and adult sharks (Wilk’s Lambda, χ2= 121.697, P< 0.001).

Both the standardized coefficient and the correlation of each variable with the

discriminant function showed that total length was the main variable to contribute to

the divergence between juveniles and adults. Testosterone and Estradiol contributed in

similar proportion while P4 did not explain any additional separation between groups

(Table 3.2). Progesterone was found to only play a major role in draughtboard sharks

during the ovulatory cycle (see chapter 2), and because its level only varied within adult

animals this hormone was unlikely to contribute to the separation between the two

groups. Therefore the model was rerun excluding P4. Discriminant function scores (D)

generated using TL, T and E2 were substituted into equation 3.2 as follows:

D= -6.919 + 0.008*TL + 0.90*T + 0.22*E2

Conditional probabilities under the discriminant scores (D) were generated for both

groups. The prior probability of any shark to be juvenile was 0.63 and to be adult was

0.37. The group to which a case belongs is based on its largest posterior probability.

From 118 females, 92% of cases were correctly classified.


86

MALES

Clasper calcification is traditionally used to determine maturity in male sharks,

however, the differences between partially and fully calcified claspers can be very

subjective. As the calcification of the clasper was related to clasper length (CL) (Fig.

3.2), clasper length was included to separate maturity stages in male sharks. Discriminant

function analysis combining CL, T, E2 and P4 showed significant differences between

juveniles and adults (Wilk’s Lambda, χ2= 41.377, P< 0.001). Clasper length and T were

the main contributors to the separation of juveniles and adults. Both E2 and P4 played a

minor role in the divergence of the two groups and were excluded from the analysis

(Table 3.2). Discriminant function scores (D) were generated using the following

equation:

D = -4.239 + 0.070*CL + 0.234*T

Variable Standardized

coefficient

Correlation with

discriminant function

Total length 0.61 0.90

Testosterone 0.42 0.80

Estradiol 0.40 0.71

Progesterone 0.16 0.58

Table 3.1: Standardized discriminant function coefficients and correlations of the discriminant linear function for draughtboard shark females. Total length showed the highest standardized coefficient and the highest relationship with the discriminant function.


87

Conditional probabilities under the discriminant scores were generated for juveniles and

adults. The prior probability was estimated as 0.55 and 0.45 for juveniles and adults

respectively. The group to which a case belongs was based on its largest posterior

probability. From 111 males, 99% of cases were correctly classified.

Figure 3.2: Relationship between clasper calcification and clasper length for draughtboard shark males.

Clasper length (mm)

0 10 20 30 40 50 60 70 80 90

Pro

po

rtio

n o

f m

ales

wit

h f

ull

y

calc

ifie

d c

lasp

ers

0.0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

1.0n=435


88

Multi-dimensional scaling (MDS)

FEMALES

A combination of T and E2 successfully separated the reproductive stages of

female sharks. Based on the discriminant function result of the contribution of P4 into

the separation of both groups, P4 was not included in the MDS analysis. The majority of

adult animals were on the left side of the ordination and the juveniles on the right side

(Stress=0) (Fig. 3.3). ANOSIM analysis showed that there were significant differences

between the reproductive stages (Global R=0.61, P< 0.001). A ‘95% cut off’ line for

adults resulted in 90% of the females correctly classified; eight juveniles were classified

as adults and five adults as juveniles (Fig. 3.3).

Variable Standardized

coefficient

Correlation with

discriminant function

Clasper length 0.89 0.96

Testosterone 0.59 0.78

Estradiol 0.16 0.12

Progesterone 0.07 0.27

Table 3.2: Standardized discriminant function coefficients and correlations of the discriminant linear function for draughtboard shark males. Clasper length showed the highest standardized coefficient and the highest relationship with the discriminant function.


89

MALES

Based on the results from the discriminant function analysis, where CL and T

played a major role in the separation between juveniles and adults, E2 and P4 were

excluded from the MDS analysis. A combination of CL and T separated adult male

sharks from most of the juveniles (Stress=0.01) and ANOSIM analysis demonstrated

that there were significant differences between the reproductive stages (Global R=0.70,

P< 0.001) (Fig. 3.4). Based on a ‘95% cut off’ lines of adults, 97% of the 111 males

sampled were correctly classified (Fig. 3.4).

Figure 3.3: Multi-dimensional scaling (MDS) of juvenile (white circles) and adult (black circles) draughtboard shark females of known maturity, using testosterone and 17β-estradiol. The vertical dashed line represents the “95% cut off” line whereby 95% of adults were to the left of this line.

Stress: 0

Adults Juveniles


90

Size at maturity

Hormone analysis was undertaken on 229 sharks that were also dissected. Size at

maturity was calculated for these sharks using equation 3.1 based on macroscopic

examination of the gonads (destructive sampling) and after classification of the sharks

into juveniles or adults using either LDPM or MDS analysis (non-destructive sampling).

For the MDS method, sharks on the left of the ‘95% cut off’ line were classified as

adults and sharks on the right of the ‘95% cut off’ line were classified as juveniles. All

three analyses resulted in a similar size at 50% maturity for both sexes, with females

within 1.8% and males within 0.4% of the estimated values from macroscopic

examination (Table 3.3).

Stress: 0.01

Juveniles Adults

Figure 3.4: Multi-dimensional scaling (MDS) of juvenile (white triangles) and adult (black triangles) draughtboard shark males of known maturity, using clasper length and testosterone. The vertical dashed line represents the “95% cut off” line whereby 95% of adults were to the left of this line.

91

50 % m

aturity

TL (mm)

Percentage

difference

r2

a and b values

95% Confidence interval

TL (mm)

n

Females (macroscopic)

814

- 0.

80

a =

-32

.16,

b =

0.0

4 79

8 –

830

118

Females (LDPM

analysis)

823

1.10

0.

77

a =

-59

.55,

b =

0.0

7 81

2 –

832

118

Females (MDS analysis)

829

1.84

0.

75

a =

-28

.54,

b =

0.0

3 81

1 –

848.

11

8

Males (macroscopic)

779

- 0.

80

a =

-47

.70,

b =

0.0

6 76

2 –

790

111

Males (LDPM

analysis)

776

-0.3

8 0.

80

a =

-63

.08,

b =

0.0

8 76

8 –

783

111

Males (MDS analysis)

782

0.25

0.

82

a =

-34

.23,

b =

0.0

4 76

0 –

802

111

Table 3.3. Com

parison of the size at 50%

maturity between destructive (visual examination) and non-destructive (LDPM

and MDS) methods for

female and male draughtboard sharks. Percentage differences in the size at maturity using the non-destructive method were compared with the

destructive method. LDPM: linear predictive discrim

inant m

odel, M

DS: M

ulti-dimensional scaling.


92

3.3.3 SHARKS OF UNKNOWN MATURITY

Sharks from the marine reserve were categorized as juveniles or adults based on

their posterior probabilities for the LDPM analysis. For the MDS ordination, the

unknown sharks were overlaid on the MDS plots for sharks of known maturity (Fig. 3.5

and 3.6). Sharks that fell to the left of the ‘95% cut off’ line for adults were classified as

adults and those on the right as juveniles.

Stress: 0

Juveniles Adults

Fig. 3.5: MDS ordination of draughtboard shark females (known and unknown maturity) using testosterone and 17β-estradiol. J (juveniles): white circles, A (adults): black circles, U (unknown): grey triangles. The vertical dashed line represents the “95% cut off” line whereby 95% of adults were to the left of this line.


93

Size at maturity

For both sexes, the LDPM and MDS analysis resulted in a similar size at 50%

maturity (Table 3.4.). There was no difference between the estimates of size at maturity

for female and male draughtboard sharks caught in the marine reserve compared to

those caught from the rest of Tasmania. The LDPM estimates of size at maturity were

closer to the macroscopic estimates for all values except males in the marine reserve.

Similarly, the confidence limits for the LDPM were narrower than the corresponding

MDS for all analyses except for males in the marine reserve. The techniques were

sensitive to sample sizes with the smaller sample sizes from the reserve population

resulting in larger increasing the 95% confidence limits (Fig. 3.7).

Stress: 0.01

Juveniles Adults

Figure 3.6: MDS ordination of draughtboard shark males (known and unknown maturity) using clasper length and testosterone. J (juveniles): white triangle, A (adults): black triangle, U (unknown): grey circles. The vertical dashed line represents the “95% cut off” line whereby 95% of adults were to the left of this line.


94

50 % maturity

ogive

TL (mm)

r2 a and b values

95% Confidence

interval

TL (mm)

n

Females (LDPM analysis) 818 0.74 a = -72.09, b = 0.09 803 – 855 82

Females (MDS analysis) 828 0.76 a = -9.44, b = 0.01 775 –870 82

Males (LDPM analysis) 768 0.83 a = -43.84, b = 0.06 745 – 803 54

Males (MDS analysis) 757 084 a = -25.73, b = 0.03 740 – 795 54

Table 3.4. Size at 50% maturity for female and male draughtboard sharks based on the hormone results of the linear discriminant predictive model (LDPM) and multi-dimensional scaling (MDS) analysis.

Figure 3.7: Comparison of 50% size at maturity and 95% confidence limits for female (circles) and male (triangles) draughtboard sharks caught in a marine reserve and from the rest of Tasmania using linear discriminant analysis (LDPM) and multi-dimensional scaling ordination (MDS).

Method

Siz

e at

50%

mat

uri

ty w

ith

95%

co

nfi

den

ce l

imit

s

720

740

760

780

800

820

840

860

880

Visual LDPM MDS MDSLDPM

Rest of Tasmania Marine Reserve

Visual LDPM MDS MDSLDPM

Rest of Tasmania Marine Reserve


95

Reproductive Seasonality

For both sexes the proportion of adult animals found in the marine reserve was

only slightly less than those found from the rest of Tasmania, although this difference

was non significant (Table 3.5).

Location Proportion adult animals

Females Males

Marine Reserve 0.29 0.48

Rest of Tasmania 0.38 0.54

To compare monthly variations of hormones between sharks obtained from the

marine reserve and the rest of Tasmania, sharks were grouped into three periods due to

the small sample sizes. For females, there were no significant differences between E2, or

P4 in sharks from the marine reserve compared with those from the rest of Tasmania. In

contrast, T levels were lower in sharks from the marine reserve than from the rest of

Tasmania in the March-May period (ANOVA, P< 0.001) (Fig. 3.8). The levels of T were

significantly lower for males in the marine reserve for the March-May period (ANOVA,

P< 0.001), although the gradual decline in T levels from January to December was

consistent in both the marine reserve and the rest of Tasmania (Fig. 3.9).

FIG. 3.5: Proportion of mature draughtboard sharks caught in lobster traps from a marine reserve and from the rest of Tasmania.


96

Figure 3.8: Seasonal variations in testosterone (T), 17β-estradiol (E2) and progesterone (P4) for adult female draughtboard sharks caught in a marine reserve () and the rest of Tasmania (�). Values are mean ± SE. Numbers are sample sizes. * Values are significant different.

Period

Ho

rmo

ne

lev

el (

ng

.ml-

1 )

0.0

0.5

1.0

1.5

2.0

14

6

4

T

13

11

10

1

2

3

4

5

6

7E2

0

1

2

3

4

5

6

7P4

Jan-Feb Mar-Apr-May Oct-Nov-Dec

*

*


97

Figure 3.9: Seasonal variations in testosterone for adult male draughtboard sharks caught in a

marine reserve (•) and the rest of Tasmania (▲). Values are mean ± SE. Numbers are sample sizes. * Values are significant different.

Period

Jan-Apr May-Aug Sep-Dec

Tes

tost

ero

ne

lev

els

(ng

.ml-1

)

0

1

2

3

4

5

6

7

8

11

9

12

16

29

21

*

*


98

3.4 DISCUSSION

Size at maturity obtained from blood samples was within 2% of the size at

maturity obtained from macroscopic examinations of gonads. For both sexes in C.

laticeps, the combination of external features (e.g. total length in females and clasper

length in males) and gonadal steroids can be used to obtain reproductive information

for management of sharks without having to sacrifice the animal.

From macroscopic examination of dissected animals it was clear that for

draughtboard sharks, maturity is strongly size dependent. Sharks larger than 860 and 870

mm TL (females and males respectively) were all adults and sharks below 750 and 710

mm TL (females and males respectively) were all juveniles.

For C. laticeps, both the linear discriminant predictive function and the multi-

dimensional scaling analysis provided objective methods to classify sharks as juveniles or

adults, and therefore address size at maturity and reproductive seasonality. Furthermore,

steroid hormones could determine the stage of maturity in the intermediate length size

classes where sharks could be either juveniles or adults. For these sharks neither total

length or clasper calcification could be used to determine the reproductive stage of the

animals, therefore hormones provided a mechanism for determining the reproductive

status of these sharks. The LDPM had narrower confidence limits and was, in general,

closer to the macroscopic estimates giving a more precise and accurate method than the

MDS, although no significant differences were found between values. The sample sizes

would suggest that for C. laticeps, it is necessary to have approximately 100 sharks with a

significant proportion in the critical region between 100% adults and 100% juveniles to

obtain an accurate estimate of size at maturity.

When selecting the hormones to use to separate juvenile or adult sharks,

understanding the role that each of the gonadal steroids play in shark reproduction is

important. Hormone analysis is relatively costly, thus knowledge of which hormones


99

contributed to the separation of the reproductive stages should enable costs to be

minimized. For draughtboard sharks there was a need to measure only two hormones, T

(for both sexes) and E2 (for females), to separate juveniles from adults. Testosterone

and E2 were found to be the principal hormones during the follicular phase of females,

while elevated plasma P4 was found primarily in the ovulatory phase (see chapter 2). As

P4 only varied in adult females and was dependent on the female ovulatory phase, it was

possible to find adult females with low or high levels of P4, whereas juvenile females

always had low levels of P4. Therefore, P4 was not a reliable discriminant factor for

separating juvenile and adult females. In males, only T showed a significant increase

from juvenile to adult animals (see chapter 2) and thus was the main contributor to the

separation. In C. laticeps males, clasper length and T contributed to the separation of

juveniles from adults. As the degree of calcification and size of claspers were external

features that could be readily assessed, clasper length and calcification will be the most

cost effective method of identifying the size at sexual maturity of C. laticeps males.

Steroid hormones were also important in providing data on seasonality of

reproduction. While clasper calcification can be used to address size at maturity in

several chondrichthyans species, dissection of these males is still required to understand

seasonality. Although C. laticeps was not found to have a defined seasonal reproductive

pattern (see chapter 2), variations in hormone levels followed similar trends in

reproductive activity obtained from macroscopic examination of the gonads. In

seasonally reproductive species such as Hemiscyllium ocellatum (Heupel et al., 1999), Raja

eglanteria (Rasmussen et al., 1999) and Dasyatis sabina (Tricas et al., 2000), strong

correlations in hormones and reproductive seasons have been reported.

A concern could be that the lower steroid plasma levels in seasonally reproductive

sharks captured during their non-reproductively active period could confound the

estimates of size at maturity as they could be classified as juveniles (ie. if the hormone


100

values fall to values equivalent of juveniles). To obtain size at maturity estimates it is

essential to sample animals during the reproductive period.

The population of C. laticeps from the Crayfish Point Reserve showed that the

proportion of adult sharks (for both sexes) was similar to the proportion in the rest of

Tasmania, suggesting that although this reserve would offer protection to adult animals,

this protection was not preferential. The similar seasonal trends in reproductive

hormones for females and males between the reserve population and the rest of

Tasmania is expected, as tagging studies (see chapter 4) demonstrated that this species

can move substantial distances and mix between regions. Although the seasonal sample

sizes were small, the similarity in trends between the dominant hormones and

macroscopic examination of the gonads in each sex demonstrated the potential of

hormones to define spatial and temporal variability in reproduction without the need to

sacrifice the sharks.

Different methods, such as ultrasonography and endoscopy, have been used to

assess gestation period and reproductive condition in females without killing the animal

(Carrier et al., 2003) (J. Daly, Melbourne aquarium, Melbourne. pers. comm.). To date no

estimates of size at sexual maturity or seasonality of reproduction have been reported

using these techniques. Ultrasonography or endoscopy also require substantial handling

and manipulation of the sharks, which could affect both the shark and its embryos

(Carrier et al., 2003). Obtaining a blood sample from the draughtboard shark for

estimating hormone concentration, involved a minimal handling time (2-3 minutes)

before the shark was returned to the water. The sample could be taken at sea in exposed

and rough conditions, making hormones a less invasive, quick technique.

Endocrine markers provide a non-destructive way to obtain information on somatic,

temporal and spatial reproductive parameters for management of sharks. Non-

destructive techniques are essential for sampling marine species on the world’s

threatened and endangered species lists, of which there are many chondrichthyans


101

(IUCN, 2006). Understanding the impact of fishing operations on bycatch is also

required for industry accreditation and meeting ecosystem based fishery management

objectives (Hall et al., 2000). In circumstances where the bycatch is not retained,

sacrificing the shark to obtain information on reproductive status would no longer be

required.

A general trend or common pattern in the MDS ordination or the (D) score values of

the LDPM analyses may also be found to distinguish juvenile and adult sharks for the

different reproductive modes (oviparity and viviparity). In this case, it would no longer

be necessary to sacrifice sharks that are not a target or by-product of fishing operations.

If the relationship between steroid hormones and reproduction is reproductive mode

specific or generic to all chondrichthyans, then validation for different species would

not be required and future reproductive needs (size at maturity, seasonal reproductive

activity) for management could be addressed non-destructively though blood sampling.

As hormones can also provide information on the seasonality of reproduction, they

have the potential to provide necessary information required for the conservation and

management of shark populations without the need to sacrifice the animal.

102

CHAPTER FOUR:

Movements, activity patterns and habitat utilisation

Chapter four - Movement

103

4.1 INTRODUCTION

Efforts to study the movement of fish at either the population or individual levels

have been in progress for over a century (Casey and Taniuchi, 1990; Kohler and Turner,

2001). Tagging is the most used method for studying fish movements, and provides

important information on life history and population dynamics (Hilborn, 1990; Eiler,

2000). Recently, the importance of incorporating fish behaviour and habitat utilisation

as components of fish movement studies has been recognized for marine management

and conservation programs (Shumway, 1999; Koehn, 2000).

Historically, migrations, movement patterns and habitat preferences of fishes were

determined by fishery dependent mark and recapture or visual (in situ) observations

(Gunn, 2000; Stevens, 2000; Lowe et al., 2003). Initially, mark-recapture studies used

conventional tags which are defined as those that can be identified visually without the

use of detection equipment (Kohler and Turner, 2001). Conventional tagging

experiments of cartilaginous fishes were first reported in the 1930s (McFarlane et al.,

1990; Kohler and Turner, 2001; Latour, 2004), and have subsequently continued to be

an important source of information for understanding chondrichthyan populations

(McFarlane et al., 1990; Hurst et al., 1999; Stevens, 2000).

Conventional mark-recapture studies rely on recaptures of the tag by a variety of

sampling gears used by either researchers or fishers. Bias associated with conventional

mark-recapture studies can occur when selectivity of sampling gears varies with habitat,

or habitat-specific movements alter catch rates, or fleet dynamics alter the probability of

recapture (Kohler and Turner, 2001; Simpendorfer and Heupel, 2004; Bolle et al., 2005).

The development of acoustic tracking technology in the 1960’s enabled detection of

the animal independent of the need to be recaptured. By either the use of hand held

detectors (active tracking) or moored ‘listening receivers’ (passive tracking) detailed

information on animal behaviour and movement can be obtained. Passive acoustic


104

monitoring technology allows movement patterns of multiple individuals tagged with

acoustic transmitters to be determined (Heupel and Hueter, 2001; Voegeli et al., 2001;

Heupel and Hueter, 2002). Hydrophone (listening) stations (receivers) record the date,

time and identity of an aquatic animal fitted with an acoustic transmitter swimming

within the detection range of the receiver (Voegeli et al., 2001). One of the most

promising current applications of acoustic tags to fishery management is elucidation of

home range area (Kramer and Chapman, 1999), and habitat utilisation (Sibert and

Nielsen, 2000). While there are many studies applying acoustic technology in sharks and

rays (Holland et al., 1999; Heupel and Hueter, 2001; Klimley et al., 2002; Nakano et al.,

2003; Garla et al., 2006)), only a few have addressed home range or habitat utilisation

(Morrissey and Gruber, 1993a; Morrissey and Gruber, 1993b; Heithaus et al., 2002;

Heupel et al., 2004; Duncan and Holland, 2006; Heupel et al., 2006b).

Although Cephaloscyllium laticeps is primarily caught as bycatch in rock lobster traps

and other inshore hook and gill net fisheries, there is concern that the small amount of

byproduct that is currently caught has the potential to expand (J. Lyle, TAFI Marine

research Laboratories, Hobart. pers. comm.). As a precautionary measure, Tasmania has

implemented a possession limit of two draughtboard sharks per person, or five sharks

per boat per day to constrain future catches.

Walker (2005) reported a 54% decline in draughtboard sharks caught in Bass Strait,

southern Australia between 1973-76 and 1999-2001. Although the cause for this decline

is uncertain, Walker suggested that it might be due to a change in fishing patterns in an

attempt to minimise bycatch of this species rather than a true decline in abundance due

to fishing.

Prior to considering any increased utilisation of this species it was important to

understand the mixing of populations between regions. Small-scale movement patterns

and habitat utilisation was also considered important to establish if sharks were more

vulnerable to capture at certain times of the day and on certain substrates. Knowledge


105

of the behaviour of draughtboard sharks can therefore be used to both increase

exploitation through targeted fishing or to minimise bycatch by avoidance.

Previous studies in other scyliorhinids showed that sharks are characterised as slow

swimmers (Springer, 1979; Compagno, 1984) and are often found resting in caves either

alone or in aggregations (Nelson and Johnson, 1970; Sims et al., 2005). Nelson and

Johnson (1970) reported nocturnal activity patterns for the scyliorhinids Heterodontus

francisci and Cephalloscylium ventriousus and Sims (2001) found differences in the day night

activity between males and females of Scyliorhinus canicula.

This study investigated the movement behaviour of the draughtboard shark using

conventional and acoustic tagging. Conventional tags were used to identify longer-term

movement (> 6 months) over larger geographic regions, while acoustic tagging

evaluated short-term movements (< 6 months) and habitat utilisation.


106

4.2 MATERIAL AND METHODS

4.2.1 ACOUSTIC TAGGING

Study site, acoustic receivers and transmitters

An array of 82 VR2 automated acoustic receivers (Vemco Ltd., Nova Scotia) were

deployed in October 2002 and retrieved in July 2003 in southeast Tasmania, Australia

(Fig. 4.1a). The sea floor in these areas consisted of sand, silt, seagrass and low profile

reef (Barrett et al., 2001; Jordan et al., 2001). Each receiver was secured to a vertical steel

post on a concrete mooring, approximately 1 m above the sea floor.

An extensive array of receivers was established as a series of acoustic ‘curtains’

separating the main bays and channels in southeast Tasmania (Fig. 4.1b). The depth of

receiver placement varied from 2 to 55 m. The distance between receivers was chosen

to ensure that detection distances had substantial overlap and varied from 720 to 930 m

depending on the habitat type. Receivers were positioned at the entrances of bays and

channels to ensure that no shark could move into or out of these areas without being

detected.

Within the extensive array, an intensive array was established at the Crayfish Point

Reserve (total area= 800 m2) and the adjacent areas of Alum Cliff and Taroona High

(Fig. 4.1 c). In the Crayfish Point Reserve, the sea floor includes a complex mix of sand,

silt, low and high profile reef (Barrett et al., 2001; Jordan et al., 2001). The complexity of

this habitat resulted in a reduction of the detection range for the acoustic receivers to a

minimum of 60 m (Semmens, unpublished data). Thus, the receivers were placed

approximately 100 m apart to provide sufficient overlap for determining position from

detection at multiple receivers. The receivers were placed in depths from 2 to 11 m. The

receivers that formed a small ‘curtain’ perpendicular to the shore at Alum Cliff and

Taroona High were 400-450 m apart.


107

The transmitters (V8SC-2H: Vemco, Nova Scotia), were cylindrical in shape, 30 mm

in length, 9 mm in diameter and weigh 3.1 g in water. The transmitters emit a 69 kHz

frequency “ping” code repeated after a random delay of 20 to 60 s. The battery life was

set at 180 days.

Alum Cliff

Taroona High

Crayfish Point

Reserve

c Figure 4.1: Acoustic receiver positions. a: Map of Tasmania showing out the southwest area. b: Receiver positions in the southwest area. Extensive curtains are labelled as: B: Lower mid-channel, C: Upper mid- channel, D: Upper channel, E: Upper Derwent, F: Lower Derwent, G: Storm Bay, H: Frederick Henry Bay, I: Norfolk Bay, J: Dunally, K: Eaglehawk Neck. c: Receivers position in Crayfish Point Reserve and adjacent areas. c: Receiver positions at the intensive array area established by the Crayfish Point Reserve, Alum Cliff and Taroona High.

Crayfish Point Reserve

B

D F

E

G

H

I J

C

b

K

a 147° 148°

43°

41°

Tasmania


108

Sampling methodology

Between January and March 2003, 25 (15 females, 9 males, 1 no sex recorded)

draughtboard sharks were caught in rock lobster traps. Fifteen sharks were sourced

from the Crayfish Point Reserve and 10 sharks from the east coast of Tasmania (42-

43°S, 147-148°E) (Fig. 2.1). All sharks were released in the Crayfish Point Reserve. Prior

to release, total length, total weight, and clasper length (for males) was recorded. Sharks

were fitted with the acoustic transmitters and injected with 25mg/kg of the antibiotic

tetracycline dissolved to saturation in seawater.

Initially, two sharks were internally tagged by inserting the tag into the body cavity.

After capture, these sharks were injected with a localised anaesthetic (Xylocaine 0.5%,

25 mg in 5 ml), and a 3-4 cm incision was made in the ventro-lateral region (the ventral

region was considered unsuitable as the sharks rest on the sea floor) towards the rear of

the stomach cavity. The transmitters were coated in 100% paraffin to prevent

transmitter rejection and to cover any sharp protrusion on the transmitter surface that

might irritate the shark (Heupel and Hueter, 2001). The transmitter was inserted and the

cavity closed using surgical glue (Indermil® Loctite Corporation, Dublin) and a

disposable skin stapler (Royal 35W, United State Surgical Corporation, Ltd). The sharks

were then held in captivity for one week prior to being released in the Crayfish Point

Reserve. However, the tagging wound was observed to re-open in several sharks,

probably due to the frenetic movements following release. Because of the uncertainty

associated with a partially open wound, the remaining 23 sharks were tagged externally.

For the external tagging, two 1.10 mm x 38 mm surgical needles were joined to the

distal end of the transmitters. The transmitter was attached to the base of the first dorsal

fin by the needles piercing through the fin. The needles passed through buttons on the

opposite side of the fin and were then crimped and the excess needle length removed

(Fig. 4.2).


109

Analysis of the data

Raw data collected by the receivers, including transmitter number, and time and

date of detection was downloaded (in March-April and July 2003) using the VR2 data

processing software (Vemco Ltd). Data from both arrays was analysed using ArcView

3.2 (ESRI 1999) with the Animal Movement Analyst Extension (AMAE) tool (Hooge

and Eichenlaub, 2000) and Microsoft Excel.

PERFORMANCE OF THE RECEIVERS WITHIN THE CRAYFISH POINT RESERVE

Receiver performance could be affected by the interference of acoustic noise (eg:

signals from motoring small vessels) in addition to obstruction and ‘bounce’ of acoustic

signals in association with different substrate types (eg: sand, high profile reef) and

vegetation (eg: density of seaweeds such as kelp). To evaluate the performance of the

receivers, data from sharks detected in the inner arrays of the Crayfish Point Reserve

and arrays outside the reserve (i.e. Alum Cliff or Taroona High) were compared. The

data were examined to determine if sharks detected inside, and subsequently outside the

reserve (or vice versa) were also detected by the outer ring of the Crayfish Point Reserve

receivers (Fig. 4.3). The performances of the receivers were then assessed by counting

Figure 4.2: External acoustic tag attachment on draughtboard sharks. a: Acoustic transmitter has been modified with the insertions of needles across both ends. b: The transmitter is inserted in the base of the first dorsal fin. c: The transmitter is attached to the dorsal fin and the residual needles are crimped. d: The attachment is complete.

a c b d


110

the times that the outer receivers detected the movement of 3 (1 internally and 2

externally tagged) sharks that moved, on 232 occasions, in and out of the inner area of

the Crayfish Point Reserve. Detection on the outer ring receivers indicated progress of

sharks from inner to outer regions and suggested good receiver performance.

DEFINITION OF MOVEMENT

For the intensive array, it was possible to obtain large datasets if the sharks

remained for periods of time within the reserve and adjacent areas. Due to the

complexity of the habitat, sharks could move by swimming between adjacent high

profile reefs (e.g. in narrow channels within the reef structure), over higher profile reefs

(e.g. elevated position in the water column) and over low profile reefs and other

Figure 4.3: The Crayfish Point Reserve was subdivided into four outside areas (1-4) and 1 inside area (inner). Sharks moving from the inner region of the reserve to either Alum Cliff or Taroona high (or vice versa) had to be detected by the outer ring receivers (areas 1, 2, 3 and 4) of the reserve.

4 3

1 2

Alum Cliff Receivers

Taroona High Receivers

Inner


1,2

2,3

3,4

4,1


111

habitats. Thus, the detection distance of the receivers changed as the habitat increases in

complexity (Fig. 4.4). Rocks and differing densities of macroalgae decreased the

detection distance and small movements of a shark, particularly vertically within the

water column, could result in detections by different receivers suggesting different

locations. Thus, movement in this study was defined as either movement from one area

to another non-adjacent area of the reserve (eg. from area 1 to 3 and from 2 to 4, or

non-adjacent joint areas from 1,2 to 3,4 and 4,1 to 2,3) or a consistent pattern of a new

set of receivers detecting a transmitter (see Fig. 4.3). For the extensive array, Alum Cliff

and Taroona High, movement was defined as when more than two non-adjacent

receivers of the curtain detected a shark.

Figure 4.4: Generalised representation of the reef habitat. The receiver (VR2) reception is permanently blocked when there are rocks between the sharks and the receivers, resulting in no detection (x). Intermittent blocking of the receiver reception (x√) occurs when there are algae between the sharks and the receivers. A receiver will also intermittently detect a shark, when the shark moves around a rock situated between the shark and the receiver (x√). When there is not obstruction between the shark and the receiver, sharks will be continuously detected (√).

Reef habitat

VR2

Rock Rock


112

DEFINITION OF STATIONARY OR MINOR MOVEMENT PERIODS

When the shark was detected by the same receiver or set of receivers for at least

60 minutes at intervals of 1 minute or less (60 + detections in an hour) the shark was

considered to be stationary. Often during this time it was possible for an additional

receiver to detect the shark, but it was unknown if the shark had moved slightly or the

detection was associated with movement of the habitat (eg. kelp) between the shark and

the receiver. In any case, stationary or minor movements indicated that the shark is not

actively swimming.

Differences in the number of sharks with stationary periods were tested by

Student t-test (Quinn and Keough, 2002). To determine if there was a difference with

size, sharks were classified as juveniles and adults according to 50% length at

maturity (see chapter 2). Sharks smaller or larger than 815 mm (for females) and 760

mm TL (for males) were considered as juveniles and adults respectively.

DEFINITION OF UNACCOUNTABLE TIME

There were times where an individual draughtboard shark could not be accounted

during tracking period. There were two possible reasons why the VR2 system did not

record the shark positions: 1) sharks were moving to areas outside the detection range

of the receivers, 2) sharks remained in areas were the VR2 system was obstructed by

external noise (eg. motor vessel) or marine substrate.

DEFINITION OF PRESENCE

As there were a large number of periods in the data set when individual sharks

could not be accounted for, the presence of a shark in any given area was defined as

when one or a group of receivers recorded at least 10 detections. This minimises the


113

number of false detections (single detection of any transmitter code) caused by external

noises, producing false readings of the transmitter identification number (tag) by the

receivers.

HABITAT UTILISATION

Habitat utilisation was determined as the 95% probability of a draughtboard shark

being found within a certain area calculated as the 95% kernel utilisation distribution

(KUD) (Worton, 1987) using the AMAE tool in Arcview. The spatial use of the habitat

through time was evaluated by examining the 95% KUD estimates for each shark per

month and by combining all months together. A student t-test was used to compare the

total number of animals that had stationary periods (see definition above) found in each

of the 5 areas of the Crayfish Point Reserve and the overlap areas (Fig 4.3) to determine

if sharks preferred specific areas of the Reserve.

HABITAT PREFERENCE

To evaluate the preferred habitats that sharks used to either move or have

stationary periods it was necessary to use separate approaches for movement and

stationary periods:

1. To determine if sharks preferred to move in any specific habitat, the proportion of

movements in each area was calculated, for each shark, as the number of

movements within an area divided by the total number of movements for that

shark. Differences in the proportion of movements in different areas were tested

using a Chi-Square test (Quinn and Keough, 2002).


114

To determine if there was a preference for certain boundary regions of the Crayfish

Point Reserve for access into and away from the reserve, the proportion of

movements in each area was compared. For this part of the analysis, area 1 of the

reserve includes the overlap between areas 1 and 4 (area 4,1), area 2 includes the

overlap between areas 1,2, area 3 includes the overlap between areas 2,3, and area 4

includes the overlaps between areas 3,4.

To determine if sharks preferred to spend stationary periods in any specific area, the

proportion of the hours that sharks were stationary in each area was calculated as

the number of hours that all sharks were stationary in each area, divided by the total

number of hours that sharks were stationary. Differences in these proportions were

tested using Chi-square. As this analysis was based on time, the same analysis was

undertaken based solely on the occurrence of a stationary period irrespective of the

length of the stationary period (providing it was greater than one hour). Thus, a

single stationary period of 20 hours provided a count of 1 whereas 3 separate one-

hour stationary periods separated by periods of movement provided a count of 3.

2. Habitat preference for both movement and stationary periods was determined as the

50% kernel utilisation distribution (KUD) using the AMAE tool in Arcview. The

50% contour was chosen to indicate the areas of greatest use.

DAY-NIGHT ACTIVITY

Day-night habitat utilisation and preferences were calculated using both the 95%

and 50% KUDs respectively.


115

SITE FIDELITY FOR THE CRAYFISH POINT RESERVE

To determine if sharks remained in, or dispersed away from the Crayfish Point

Reserve region, the number of days that each shark was detected in the Crayfish Point

Reserve was compared for each month after release and differences tested by a one way

ANOVA and Tukey’s multiple comparison tests (Quinn and Keough, 2002).

4.2.2 CONVENTIONAL TAGGING

Study site and sampling methodology

Between January 2000 and January 2005, sharks were tagged during routine

fishery dependent and independent rock lobster catch sampling trips around

southwestern and eastern Tasmania and in the Crayfish Point Reserve (Fig. 1.2a). Each

shark was tagged with a 35 mm yellow standard Rototag (Daltons, Henly-on-Thames,

England) externally attached to the second dorsal fin. For each shark, sex, total length

and clasper length (males) were recorded.

Analysis of the data

Differences in the proportion of sexes, sizes and time at liberty were tested by

Chi-square. The time at liberty was subdivided into 7 periods: 1) 0 to 6 months, 2) 7 to

11 months, 3) 1-2 years, 4) 2-3 years, 5) 3-4 years, 6) 4-5 years and 7) > 5 years. Size

frequency distributions were compared between sexes of released and tagged sharks

from different areas, using a randomisation procedure with the Kolmogorov-Smirnov

test statistic (D) with data pooled across sexes. Size frequency data from the two

distributions being compared were pooled and randomly reallocated to each original

distribution and the test statistic (D) recalculated. The procedure was repeated 1000


116

times and the test of significant difference between the two distributions made by

comparing the value of the observed test statistic to the distribution of D values

obtained by the randomisation procedure. Significant differences were identified when

less that 20 of the D values obtained from the randomisation procedure exceeded the

value of D from the original distributions (Haddon, 2001).

To calculate short and long term site fidelity for the Crayfish Point Reserve, data was

standardized to account for differing effort (number of trap lifts) undertaken in the

different surveys, by the following equation:

Where Pij is the proportion of sharks recapture in trip i that where tagged in trip j, where

j > i. Cj is the catch rate (number of sharks/trap) of sharks tagged during j, and was

calculated as Stj/Tj, where Stj is the number of sharks caught and tagged in trip j, and Tj

is the total number of traps set to capture the sharks in the trip j. The value Ti is the

total number of traps set to capture the sharks in trip i, Sri is the number of sharks

recaptured in trip i that were tagged in trip j, and Sti is the total number of sharks caught

in trip i.

To calculate the expected catchability the following assumptions were made:

1. Catch rate was a function of effort

2. The Crayfish Point Reserve had no finite carrying capacity

3. Tagged sharks were distributed randomly within the population

( )

=tj

tirii

SS

STjij CP


117

Differences in the proportion of sharks recaptured either per month or per year in the

Crayfish Point Reserve were tested using Chi-square test.

All statistical analyses, for both acoustic and conventional tagging, were carried out

using SPSS (SPSS® Base 10.0). The significance level was set at P=0.05 for all data

analysis.


118

4.3 RESULTS

4.3.1 ACOUSTIC TAGGING

Transmitter performances

Of the 25 sharks that were acoustically tagged, six transmitters did not start

working until one month after attachment (they were incorrectly set to start one month

after the battery was connected), one transmitter (#143) was never recorded and one

transmitter (#156), attached to a shark that was caught on the east coast of Tasmania,

was recorded only twice on the day of tagging. Transmitters #143 and #156 were

excluded from the analysis.

Performances of the receivers within the Crayfish Point Reserve

The performances of the receivers was considered to be high as 97% (± 0.43 SE)

and 86% (± 4.44 SE) of sharks that moved between the inner region of the Crayfish

Point Reserve and Alum Cliff or Taroona High respectively were detected (Table 4.1).

Number of sharks

Inner CPR-Alum Cliff Inner CPR-Taroona High

Total number of movements

Percentage of movement

detected by the outer CPR receivers

Total number of movements

Percentage of movements detected by

the outer CPR receivers

2 sharks (externally tagged) 92 97% (n=89) 67 88% (n=59)

1 shark (internally tagged) 49 96% (n=47) 24 79% (n=19)

Total 141 97% (n=136) 91 86% (n=78)

Table 4.1: Movements of three draughtboard sharks between the inner areas of the Crayfish Point Reserve (CPR) and Alum Cliff or Taroona High.


119

Movement patterns

For the 17 sharks where transmitters were working on release, 10 were recorded

at the marine reserve on the same day of released, six within the first week of being

released and one more than a week after release (Fig. 4.5). Presence of sharks was

detected in the entire intensive array and some areas of the extensive array (the River

Derwent, the Upper Channel and Storm Bay (Fig. 4.6)).

Figure 4.5: Number of draughtboard sharks recorded at the Crayfish Point Reserve (CPR) soon after being released.

Days between release and first detection

Nu

mb

er o

f sh

ark

s re

cord

ed a

t th

e C

PR

0

2

4

6

8

10

12

1 2 3 4 5 6 7 > 70

n=17


120

Sharks showed two typical patterns of movement. The majority of the sharks (n=20)

were never recorded beyond the Derwent River during the survey period (Fig. 4.7a).

However, three females moved away from the intensive array and beyond the Derwent

River (Fig. 4.7b).

Figure 4.6: Presence of draughtboard sharks in the study area (red triangles). D: Upper channel, E: Upper Derwent, F: Lower Derwent, G: Storm Bay. The intensive array is the area formed by Crayfish Point Reserve, Alum Cliff and Taroona High.

G

Intensive array

D

E

F

147.38° 147.88°

43°

42.75°


121

The minimum time that sharks were recorded before leaving the extensive array was

11 days and the maximum overall time was 188 days. The last records for the 23 sharks

were evenly split between the intensive array (n=11) and the extensive array (n=12)

(Table 4.2). Although the sharks that were translocated from the east coast of Tasmania

tended to leave the reserve earlier than those captured from the Crayfish Point Reserve

(Fig. 4.8), all of the nine sharks sourced from the east coast remained within the

Derwent River region during the study period.

Neither of the two sharks that were internally tagged demonstrated any behaviour

that would suggest an impact of tagging different to the fin-tagged sharks. One of the

sharks moved away from the Crayfish Point Reserve and out of Storm Bay whereas the

other was last located, at the end of the study, within the reserve.

Figure 4.7: Examples of the two typical movement patterns of the draughtboard sharks. a) Shark 146 was never recorded beyond the Derwent River, moving between the intensive array, the Derwent River (F) and the Upper mid-channel (D). b) Shark 121 left the Crayfish Point Reserve (CPR) soon after it was tagged travelling through the Lower Derwent (F) to Storm Bay (G). Blue arrows show the movement of the sharks.

F D

Initial recorded position of shark

#146

Final recorded position of shark #146

CPR

a

Final recorded position of shark

#121

G

CPR

Initial recorded position of shark

#121

F

b 147.38°

43°

43°

147.50° 147.88° 147.62°

42.75°

43.6°

12

2

Shark ID

Sex

Size

Source

Date of

tagging

Date of last

record

Number of

hits

Zone of last

record

164

F

920

EC

18

/03/

03

29/0

3/03

31

3 F

16

3 **

F

76

0 E

C

26/0

4/03

4/

05/0

3 7

AC

16

2 **

M

76

0 E

C

26/0

4/03

29

/04/

03

20

TH

16

1 **

M

88

0 E

C

26/0

4/03

28

/05/

03

137

F

160

F

820

CP

R

18/0

2/03

17

/03/

03

56

G

159

M

715

CP

R

13/0

2/03

9/

05/0

3 25

53

AC

15

8 F

62

0 C

PR

20

/02/

03

6/05

/03

93

B

157

M

530

CP

R

20/0

2/03

22

/03/

03

1286

F

15

6

M

820

EC

26

/03/

03

26/0

3/03

2

- 15

5 F

60

0 C

PR

11

/02/

03

20/0

3/03

14

58

CP

R

154

F

820

CP

R

18/0

2/03

01

/07/

03

2702

A

C

153

F

630

CP

R

18/0

2/03

22

/03/

03

146

AC

15

2 F

75

0 E

C

25/0

3/03

4/

04/0

3 88

A

C

151

M

770

CP

R

18/0

2/03

8/

06/0

3 36

16

AC

14

9 F

83

0 C

PR

12

/02/

03

30/0

3/03

14

846

E

148

M

610

CP

R

19/0

2/03

22

/03/

03

677

TH

14

7 M

87

0 C

PR

12

/02/

03

16/0

5/03

85

41

TH

14

6 **

M

75

0 E

C

26/0

4/03

7/

07/0

3 41

5 F

14

5 **

M

66

0 E

C

26/0

4/03

21

/05/

03

1758

F

14

4 **

F

65

0 E

C

26/0

4/03

22

/05/

03

235

F

143

F

88

0 C

PR

13

/02/

03

MIS

SIN

G

- -

141

F

870

EC

18

/03/

03

25/0

4/03

19

03

F

140

F

770

CP

R

18/0

2/03

06

/07/

03

9685

A

C

121

* F

83

0 C

PR

16

/01/

03

21/0

2/03

20

G

11

6 *

F

770

CP

R

16/0

1/03

22

/07/

03

1313

C

PR

Table 4.2: Summary data for draughtboard sharks tracked in the southeast region of Tasmania. All sharks were tagged and released

at the Crayfish Point Reserve. Ten sharks were caught on the east coast of Tasmania and translocated to the Crayfish Point Reserve

(CPR), the other 15 were caught at the CPR. Two sharks were internally tagged, these sharks are indicated with a single asteric (*).

One transmitter did not work (shark ID 143). AC: Alum Cliff, B: Lower mid channel, E

: Upper Derwent River, E

C: East coast of

Tasmania, F

: Lower Derwent River, G

: Storm Bay, T

H: Taroona High. (**) For 6 sharks, transmitters started working one month

after insertion; the date of tagging was used as the date of the first record. Sharks with less than two hits (#156, #143) were

excluded from the analysis.


123

Movement behaviour

Two types of movement behaviour were recorded; sharks that only showed

movement and sharks that alternated movement with stationary periods. Nine sharks

showed continuous movement and 12 sharks alternated between movement and

stationary periods (Fig. 4.9, Table 4.3). Movement behaviour could not be determined

for two sharks due to a low number of detections. All sharks were out of the range of

detection for several days (Table 4.3). Within the total number of days between the first

record and the end of the study period, the percentage of days that sharks were detected

averaged 17 % (± 4.11% SE) with 1% and 69% as the minimum and maximum (Table

4.3). The average time spent by sharks having stationary periods was eight hours per day

(± 1 hr SE) with one shark spending a continuous period of five consecutive days

stationary. There was no difference in the average time spent in stationary periods for

Months after being released

0 1 2 3 4 5 6

Pro

po

rtio

n o

f sh

ark

s a

t th

e C

PR

0.0

0.2

0.4

0.6

0.8

1.0

Figure 4.8: Monthly proportion of draughtboard sharks at the Crayfish Point Reserve (CPR). 13 sharks were sourced from the CPR (black bars) and 10 sharks were translocated from the east coast of Tasmania (grey bars).


124

either sex (seven females and five males) or between juveniles (n=7) and adults (n=5).

No correlation existed between the movements of the draughtboard sharks and lunar

phases (Fig. 4.9).

Figure 4.9: Examples of movement behaviour of draughtboard sharks. Nine sharks showed only movement records (eg: shark # 146), while 12 sharks alternated between movements and stationary periods (eg: shark #147 and #140). The figure indicates both movement and stationary behaviour simultaneously as these behaviours could occur on the same day” was added in the figure legend. Dashed lines represent the initial and the last record of the sharks. Black and grey regions represent movement and stationary periods, respectively. Lunar phases are shown at the top of the graph.

Sh

ark

# 1

40

Sh

ark

# 1

47

19/0

1

26/0

1

2/02

9/02

16/0

2

23/0

2

2/03

9/03

16/0

3

23/0

3

30/0

3

6/04

13/0

4

20/0

4

27/0

4

4/05

11/0

5

18/0

5

25/0

5

1/06

8/06

15/0

6

22/0

6

29/0

6

6/07

13/0

7

20/0

7

Sh

ark

# 1

46

Day


12

5

Shark ID

Number of

detections

Movement

Stationary

Uncertain

Number of

days between

first and last

record

Number of

days recorded

betw

een first

and last record

Number of days

betw

een last

record and the end

of the study period

Percentage of days

within the study period

that shark was recorded *

164

271

18

2 25

1 12

4

115

3 16

3 7

7 9

4 79

5

162

16

16

4 2

84

2 16

1 10

4 3

0 10

1 32

3

55

3 16

0 44

6

0 38

28

4

127

3 15

9 22

19

410

659

1150

86

40

74

25

15

8 89

6

0 83

76

4

77

3 15

7 10

63

2 81

4 24

7 31

10

12

2 8

155

1215

26

92

7 26

2 38

8

124

6 15

4 23

72

26

1622

72

4 13

4 55

21

35

15

3 10

0 4

0 96

33

2

122

1 15

2 69

10

0

59

11

8 10

9 7

151

3233

26

1 36

2 26

10

111

69

44

45

149

1225

6 20

6 96

32

2418

47

33

11

4 21

14

8 58

6 48

84

44

4 32

5

121

3 14

7 30

66

299

2767

0

94

63

67

58

146

346

8 0

338

72

10

15

12

145

1474

46

8 0

1006

25

6

62

7 14

4 21

2 9

129

74

26

4 61

5

141

1610

45

9 30

2 84

9 39

16

88

18

14

0 84

40

40

5702

26

98

139

67

16

43

121

17

6 0

11

37

9 15

1 5

116

1155

12

0 41

3 62

2 18

2

125

69

Table 4.3: Type of movement of draughtboard sharks. The total num

ber of records were classified as movement, stationary period, or uncertain. Sharks showed days

when they were not recorded. * The percentage of days that had shark records was calculated assuming that all sharks were alive at the end of the study period.


126

Habitat utilisation

To determine habitat utilisation, the analysis was restricted to sharks with ≥ 20

movements. Sharks (n=12) with less than 20 detections, had insufficient information to

demonstrate movement behaviour (Fig. 4.10)

For the 11 sharks with ≥ 20 movements, movements were recorded in all areas of

the intensive array, the Upper and Lower Derwent River and the Upper Channel (Fig.

4.11a and Fig. 4.11b). The majority of the sharks remained within the Derwent River

region (Fig. 4.11a). Nine of the 11 draughtboard sharks had ≥ 20 movements in the

Crayfish Point Reserve. These sharks utilised all regions of the reserve, with a minimum

Figure 4.10: The distribution of movements for the 23 acoustically tagged draughtboard sharks.

Number of movements

Nu

mb

er o

f sh

ark

s

0

2

4

6

8

10

12

14

0-20 21-40 41-60 61-80 81-100 > 100


127

of five sharks being recorded in each region and a maximum of nine sharks being

recorded in region three. No region was visited by all 11 sharks (Fig. 4.11a).

The majority of the sharks (n ≥ 8) were detected as having stationary periods in the

Crayfish Point Reserve and in the Derwent River array, while less sharks (n ≤ 3) were

found to have minimal movements around Alum Cliff and Taroona High (Fig. 4.12a

and Fig. 4.12b). As the Derwent River, Alum Cliff and Taroona High have substrates of

sand with small areas of reef (Table 4.4), it is uncertain if these sharks were in stationary

Nu

mb

er o

f sh

ark

s d

etec

ted

TH DerwentCPR AC

Areas

0

3

6

9

12

15

Up CH

Movements

Areas of Crayfish Point Reserve

1 2 3 4 Inner

Movements

1,2 2,3 3,4 4,1

(a)

Shark 116

CPR

Figure 4.11: a) Number of draughtboard sharks detected in the different study regions estimated by 95 % KUD contours. Sharks (n=11) with at least 20 recorded movements were used to calculate habitat utilisation of movement. CPR: Crayfish Point Reserve. b) Examples of habitat utilisation distribution for draughtboard sharks estimated by 95% KUD contours.

Shark 164

CPR

(b)


128

periods on the sand which surrounds these receiver arrays, were in stationary periods on

the small reef areas, or are utilising the cement tyre that was used to support the

listening stations. In comparison to the broad areas of the Crayfish Point Reserve

moved over by the draughtboard sharks, sharks showed a preference for areas within

the reserve to have stationary periods. The majority of the sharks were detected in area 1

and its overlap with area 2, followed by a significant decline (t-test, P<0.001) in the

other areas of the reserve used for stationary periods (Fig. 4.12a).

Area Substrate type

Area 1 of CPR Mostly high profile reef and some sand and hard sand

Area 2 of CPR Mostly silty sand and some patchy reef

Area 3 of CPR Mostly different profile sands and some patchy reef

Area 4 of CPR Different profile reefs (low, patchy and high)

Inner area of CPR Reefs and some sand and hard sand

Alum Cliff Sand and small areas of reef

Taroona High Sand, silty sand, and small areas of reef

Derwent Silty sand, sand and small patchy reef

Table 4.4: Type of marine substrate for the intensive and extensive array. CPR: Crayfish Point Reserve.


129

Based on both movement and stationary periods, all monitored sharks showed

similar habitat utilisation either by month or when all months were combined (Fig. 4.13

a and b).

Figure 4.12: a) Number of draughtboard sharks having stationary periods in the different study regions. Stationary periods were recorded for 12 sharks. b) Examples of habitat utilisation during stationary periods of draughtboard sharks estimated by 95% KUD contours. CPR: Crayfish Point Reserve.

Nu

mb

er o

f sh

ark

s d

etec

ted

0

3

6

9

12

15 Stationary periods Stationary periods

TH DerwentCPR AC

Areas

Up CH


1 2 3 4 Inner 1,2 2,3 3,4 4,1

(a)

(b) Shark 148

CPR

Shark 141

CPR


130

Figure 4.13: Examples of individual and combined monthly habitat utilisation for (a) movements and (b) stationary periods of draughtboard sharks using 95% KUD contours. CPR: Crayfish Point Reserve.

(a)

(b)

Movement April

CPR

Shark 116 Movement May Shark 116

CPR

Movement June Shark 116

CPR

Stationary March Shark 147

CPR

Stationary April Shark 147

CPR

Stationary combined months Shark 147

CPR

Movement combined months Shark 116

CPR

Movement July Shark 116

CPR

Movement March Shark 116

CPR


131

Habitat preference

Of the total shark movements (n= 2594), 6% occurred when sharks moved

between the outside and the inside of Crayfish Point Reserve. When moving between

Alum Cliff and Crayfish Point Reserve, draughtboard sharks used the region of the

reserve (area 1) adjacent to Alum Cliff on 80% of occasions. In contrast, sharks that

moved between Taroona High and Crayfish Point Reserve used the closest area of the

reserve (area 4) equally (43%) to the furthest area of the Crayfish Point Reserve (area 1).

Sharks moving between the Derwent River and Crayfish Point Reserve used the closest

area of the reserve (area 3) and the adjacent area (area 1) in equal proportions (38%).

Area 1 was the preferred region for sharks to enter or leave the Crayfish Point Reserve

(Fig. 4.14).

Despite the higher number of receivers in the Crayfish Point Reserve, the greatest

number of movements were recorded by the six receivers at the Alum Cliff. These

receivers recorded a significantly greater number of detections than all the other sites

(Chi-square, P< 0.001) (Fig 4.15a). Although not significantly different, the two

receivers at the Taroona High also had a higher number of movements compared to the

Crayfish Point Reserve. The draughtboard sharks tended to move along the shore rather

than spend time moving within the reserve. To determine if there was a preference for

sharks to move around the Alum Cliff region compared to the Taroona High region, the

proportion of detections at Taroona High were compared with the proportion of

detections at the two inner (closer to shore) receivers of the six at Alum Cliff. These two

receivers are at approximately the same distance from the Crayfish Point Reserve and

cover the same detection radius as the Taroona High receivers. The two receivers at the

Alum Cliff detected a significantly greater proportion of shark movements (0.30 ± 0.12


132

SE) than those at Taroona High (0.10 ± 0.05 SE) (Chi-square, P< 0.001), indicating that

tagged sharks dispersed out of the Derwent River rather than further up the river.

Within the Crayfish Point Reserve the two sides of the reserve closest to the Alum

Cliff (areas 1 and 2) detected a greater proportion of movements than those facing

Taroona High (Fig. 4.15b). At a finer scale, sharks showed a higher proportion of

movements in shallower regions (the overlaps areas 3,4 and 4,1) rather than the deeper

region (the overlaps areas 1,2 and 2,3) (Chi-square, P< 0.001). Different areas of the

Crayfish Point Reserve overlapped within a 50% KUD for each shark, two sharks were

detected only in area 1, two sharks used the composite region of areas 1, 2 and the inner

region of the reserve, two shark used only area 1 and 3, and three sharks used the entire

reserve (Table 4.5 and Fig. 4.16). All nine sharks monitored were detected in area 1

during this study.


133

Areas

Pro

po

rtio

n o

f m

ov

emen

ts

0.0

0.2

0.4

0.6

0.8

1.0

AC TH D UCCPR

a,b

a

c

bb

Figure 4.15: Area preferences for draughtboard sharks. a) Intensive and extensive array. b) Areas of the Crayfish Point Reserve (CPR). Values are mean + SE of the proportion of movements of each shark within specific regions. AC: Alum Cliff, TH: Taroona High, D: Upper and Lower Derwent, UC: Upper Channel. Different letter shows significant differences between movements and different areas.


Pro

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ts

0.00

0.05

0.10

0.15

0.20

1 32 4 1,2 2,3 3,4 4,1i

aa

aa a

a,b a

b,cc

(a)

(b)


134

In contrast to shark movements within the Crayfish Point Reserve, the areas that

overlapped within the 50% KUD contours for each shark indicated that sharks had

stronger preferences for areas in which to have stationary periods (Table 4.6, Fig. 4.17).

Area 1 and its overlap with area 4 has 66% of the 33 recorded stationary periods and

92% of the 674 hours spent stationary were recorded in these two areas (Fig. 4.18 a and

b). These areas are the only two areas characterised by a high profile reef (Table 4.4),

indicating preferences for this reef type. Although areas 2 and the overlapping area 3,4

had a high proportion of movements they recorded a low number of stationary periods

indicating that they were only transit zones to other areas of the Crayfish Point Reserve.

Areas Number of sharks whose 50% KUD overlapped in

each area Area 1 and the joint area of 2 and 4 2

Areas 1, 2 and inner area 2

Area 1 and 3 2

The whole CPR 3

Table 4.5: Areas within the Crayfish Point Reserve (CPR) that overlapped with the estimated 50% KUD of individual draughtboard sharks.

Figure 4. 16: Example of habitat utilisation for draughtboard sharks determined by 50% KUD contours. CPR: Crayfish Point Reserve.

Shark 140

CPR

Shark 149

CPR


135

However, areas 1 and the overlapping area 1,4 were preferred for both visiting and

having stationary periods.

Areas Number of sharks whose 50% KUD overlapped

in each area Area 1 and the joint area of 2 and 4 7

Areas 1, 2 and inner area

Area 1 and 3

Inner area 2

The whole CPR

Table 4.6: Areas within the Crayfish Point Reserve (CPR) that overlapped with the estimated 50% KUD of individual draughtboard sharks during their stationary periods.

Figure 4. 17: Example of habitat utilisation for draughtboard shark stationary periods determined by 50% KUD contours. CPR: Crayfish Point Reserve.

Shark 154

CPR

Shark 155

CPR


136

Pro

po

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f h

ou

rs i

n s

tati

on

ary

per

iod

s

0.0

0.1

0.2

0.3

0.4

0.5

0.6n=674


Nu

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f st

atio

nar

y p

erio

ds

0

2

4

6

8

10

12

14

16

18

20

1 2 3 4 Inner 1,2 2,3 3,4 4,1

Figure 4.18: Habitat preferences for draughtboard shark stationary periods. a) Proportion of hours that draughtboard sharks spent in stationary periods in each area. b) Number of stationary periods spent in each area.

(a)

(b)


137

Day-night activity, habitat utilisation, and preference

Twelve draughtboard sharks were excluded from the day-night activity analysis

because they had less than 20 movements (see Fig. 4.10). Of the remaining 12 animals,

nine moved mostly at night, while the other three moved predominantly during the day

(Fig. 4.19). Similar patterns were found irrespective of whether the sharks were inside or

outside of the Crayfish Point Reserve. No sharks had an even distribution of

movements over a 24 hr period.

The majority of the sharks (n=8) showed similar 95% and 50% KUD distributions

between day and night suggesting no change in diel activity patterns either for habitat

utilisation and preferences (Fig. 4.20 a and c). The remaining four sharks showed

distinct diel patterns, although the areas used still overlapped (Fig. 4.20 b and d). Three

sharks increased their 50% KUDs during the night moving out of the Crayfish Point

Reserve and one shark increased its 50% KUD during the day.


138

Figure 4.19: Example of daily movement activity of draughtboard sharks. Two distinctive movement patterns were found, 3 sharks moved more during the day (shark 147) and 9 sharks moved more at night (shark 116).

0.0

0.1

0.2Inside the Crayfish Point Reserve Shark 147

Pro

po

rtio

n o

f m

ov

emen

t

0.0

0.1

0.2Outside the Crayfish Point Reserve Shark 147

Time (hr)

0.0

0.1

0.2Outside the Crayfish Point Reserve Shark 116

0.0

0.1

0.2 Inside the Crayfish Point Reserve Shark 116

17 19 20 21 22 23180 1 2 3 4 5 6 7 8 9 11 12 13 14 15 1610


139

Site fidelity for the Crayfish Point Reserve

Sharks tended to leave the Crayfish Point Reserve within the first month of being

released. The average number of days spent at the reserve significantly decreased from 6

days (± 2 days), just after being released, to 1 day (± 1 day) after 4 to 5 months (t-test,

P< 0.001) (Fig. 4.21).

Figure 4.20: Example of day and night habitat utilisation (95% KUDs, a and b) and habitat preference (50% KUDs, c and d) for draughtboard sharks. Grey and white spaces represent the day and night movements respectively. CPR: Crayfish Point Reserve.

Shark 147 a

CPR

Shark 149 b

CPR

c Shark 147

CPR

Shark 149 d

CPR


140

4.3.2 CONVENTIONAL TAGGING

Between January 2000 and January 2005, 1234 draughtboard sharks were tagged

in southwest and eastern Tasmania and the Crayfish Point Reserve. The Crayfish Point

Reserve showed the highest recapture rate, 36% of 364 sharks tagged, followed by

eastern and southwestern areas where the recapture rate was 9% (sharks tagged n=398)

and 3% (sharks tagged n=472) respectively (Table 4.7). However, the fishing effort by

researchers in the reserve was higher than for southwest and eastern Tasmania. The

maximum time at liberty ranged from one month to up to five years in the Crayfish

Point Reserve, from one month up to four years in both eastern and southern Tasmania

Figure 4.21: Average number of days spent at the Crayfish Point Reserve for the draughtboard sharks. Values are means (± SE). Different letters show significant differences.

Months at the Crayfish Point Reserve

1 2 3 4 5

Nu

mb

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f d

ays

at t

he

Cra

yfi

sh P

oin

t R

eser

ve

0

2

4

6

8

10

a

b b

a,b

a,b

n=17


141

(Table 4.7). There were no significant differences in the proportion of recaptures (for

either sex or size) with time (chi-square, P< 0.001).

Distances travelled

The majority of the draughtboard sharks were recaptured in the vicinity of where

they were released (Table 4.8). The majority of the draughtboard sharks travelled a

maximum distance of up to 10 km over the study period. However, for 4%, 7% and

18% of the sharks tagged in the Crayfish Point Reserve and on the east and southwest

coast, the maximum distances travelled were 75, 250, and 300 km respectively (Fig.

4.22). No relationship between time at liberty and distances travelled was found.

Area Tagged (n)

Recaptures (n)

Recaptures (%)

Maximum time at liberty (years)

Crayfish Point Reserve 364 132 36.3 5

Southwestern of Tasmania 472 17 3.6 4

Eastern of Tasmania 398 37 9.3 4

Area Recaptures in same area (n)

Recaptures in different areas (n)

Crayfish Point Reserve 118 14

Southwestern Tasmania 15 2

Eastern Tasmania 34 3

Table 4.8: Recapture areas for conventionally tagged draughtboard sharks.

Table 4.7: Summary of draughtboard sharks conventional tagging. Sharks were tagged in southwestern and eastern Tasmania and in the Crayfish Point Reserve.


142

Southwest

East


Figure 4.22: Examples of maximum reported distances travelled for draughtboard sharks. Tagged in southwestern Tasmania. Tagged in eastern Tasmania. Tagged in Crayfish Point Reserve. The lines represent the shortest possible route between the release and the recapture position.


143

Length-frequency composition of tagged and recaptured sharks

The size composition of female and male tagged draughtboard sharks was similar

in the southwest area and the Crayfish Point Reserve, but was significantly different

from eastern Tasmania which had a larger number of smaller males (Kolmogorov-

Smirnov, P< 0.005) (Fig. 4.23). There were insufficient recaptures to compare the size

of the sharks that were recaptured for the southwest and east coast populations. For the

Crayfish Point Reserve, the size of recaptured sharks was not significantly different

from the initial population that was tagged (Fig. 4.23).

Figure 4.23: Length-frequency composition of tagged and recaptured female (black bars) and male (grey bars) draughtboard sharks in different regions of Tasmania.

Total length (mm)

0 200 400 600 800 1000 12000.0

0.1

0.2

0.3

0.4

0.5

0 200 400 600 800 1000 1200

Pro

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s

0.0

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0.2

0.3

0.4

0.5

0.0

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0.2

0.3

0.4

0.5

East of TasmaniaTags

Females (n=292)Males (n=105)

Southwest of TasmaniaTags



Crayfish Point ReserveTags


Crayfish Point ReserveRecaptures

East of TasmaniaRecaptures


Southwest of TasmaniaRecaptures



144

Short and long term site fidelity for the Crayfish Point Reserve

Sharks were found up to five years after release at the Crayfish Point Reserve.

Around 2% (proportion 0.02) of sharks were subsequently recaptured each month

during the first 11 months of release (Fig. 4.24). Sharks showed a gradual dispersion

away from the Crayfish Point Reserve over the six years since tagging began (Fig. 4.25).

Figure 4.24: Monthly proportion of recaptured draughtboard sharks at the Crayfish Point Reserve. Numbers are mean + SE. Numbers above the bars are total number of recaptures. No significant differences were found between months (Chi-square test).

Months after being released

Pro

po

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f re

cap

ture

d s

har

ks

0.00

0.05

0.10

0.15

0.20

0.25

0.30

1 2-3 4-5 6-7 8-9 10-11

n=109


145

Figure 4.25: Yearly proportion of recaptured draughtboard sharks at the Crayfish Point Reserve. Numbers are mean ± SE. Numbers above the bars are total number of recaptures. No significant differences between years were found (Chi-square test).

Year after being released

Pro

po

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f re

cap

ture

d s

har

ks

0.00

0.05

0.10

0.15

0.20

9

1-2 2-3 3-4 4-5 5-6

7

21

8 2


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4.4 DISCUSSION

A few studies have been reported using passive acoustic receivers to understand

shark movements in complex reef habitats (Chapman et al., 2005; Garla et al., 2006).

However, this was the first study using passive acoustic technology to address bottom

dwelling sharks moving within a complex environment. In addition to external noise

(produced by shipping traffic, waves, currents and vegetation) that can provide mixed

signals to the receivers (Clements et al., 2005; Heupel et al., 2006a), the physical nature of

the habitat (rocky reef) could also reduce the distance over which signals can be reliably

detected. Draughtboard sharks were routinely observed by divers to rest in depressions

in the reef and under ledges, which could result in missing or uncertain records.

Consequently, it was difficult to know if sharks were resting in the reef area without

being detected or had left the area. In this study, the combination of a complex habitat

and the movement dynamics of the bottom dwelling draughtboard shark created

uncertainty in positioning an animal. Despite these shortcomings a greater

understanding of behaviour of this species was developed, including small-scale reef

utilisation preferences. By determining the accuracy of the outer ring of receivers in the

Crayfish Point Reserve and finding that the probability of a shark being detected was

high (86 to 97%), there was greater confidence in correlating detections to behaviour.

In this study, only two sharks were internally tagged with acoustic transmitters. Initial

trials with surgical implantation in aquaria studies were problematic with the sharks

opening the insertion wound when they flexed as they were released back into the water.

The attachment of external transmitters required less handling of the shark and did not

involve local anaesthesia, surgery and recovery. However, external tags were susceptible

to fouling which can lead to a reaction at the attachment site. Skin damage caused by

abrasion of a fouled conventional tag was occasionally observed in draughtboard sharks.

Moreover, sharks could rub the external transmitters against the reef or other sharks,


147

and the tags could also increase the probability of entanglement in nets and lines.

McKibben and Nelson (1986) suggested that the behaviour of three grey reef sharks,

Carcharhinus amblyrhynchos, was altered by the continual irritation of the dorsal fin where

the transmitter was attached. As no draughtboard sharks have been resighted carrying

the external transmitters, the impact of the external acoustic tag on the sharks remains

uncertain. The two sharks with internally placed tags did not appear to behave any

differently to the externally tagged sharks during the period of the study. Similar

conclusions on the behaviour of externally and internally tagged sharks were reported

for other species such as: Galeocerdo cuvier (Holland et al., 1999), Negaprion brevirostris

(Gruber et al., 1988) and Scyliorhinus canicula (Sims et al., 2001). From this study it could

be suggested that although the external tagging procedure was quicker, the uncertainty

of possible damage on draughtboard sharks due to the external tags is an important

factor that could affect future shark behaviour. Further research is required to determine

the benefits of either internally or externally tagging draughtboard sharks.

While it was difficult to determine the impact of in situ tagging on draughtboard

sharks, 41% of individuals were not recorded in the Crayfish Point Reserve on the day

of release. This suggests that they moved to regions of the reef where they were not

detected. As all sharks were released towards the middle of the reserve it was unlikely

that they could have swam beyond the detection limits of the outer Crayfish Point

Reserve receivers before transmitting their first signal (ie. Maximum time between

‘pings’ was 60 seconds). It was also unlikely that all these sharks they would have swam

through the outer ring of receivers without being detected given the greater than 86%

chance of detection. It was possible that tagging affected the initial behaviour of the

sharks causing them to rest and recover from capture and tagging in regions of the reef

(eg. caves) where signals could not be detected.

Sharks tagged in this study were either, captured and released in Crayfish Point

Reserve or captured on the east coast of Tasmania and released in the Crayfish Point


148

Reserve. The majority of the draughtboard sharks that were translocated from the east

coast did not stay within the release area for longer than one month, after which they

left the intensive array to be finally recorded around the lower section of the Derwent

River. While no significant difference could be detected with the small number of sharks

tagged, there were indications that sharks did show some site fidelity as the sharks

sourced from the Crayfish Point Reserve were sighted back in the reserve on more

occasions and tended to disperse less widely than those sourced from the east coast.

The rapid decrease in acoustically tagged sharks recorded in the Crayfish Point

Reserve was similar to the conventional tag data from the reserve where the majority of

sharks were not seen after tagging. The acoustic data suggests that these sharks were still

in the general vicinity of the Derwent River but only revisited the Crayfish Point

Reserve at decreasing intervals as time increased. As with the large distances recorded

for conventional tag recaptures, two of the acoustically tagged sharks moved out of the

Derwent River and associated Storm Bay. Similarly, (McLaughlin and O'Gower, 1971)

found that the demersal shark Heterodontus portusjacksoni undertook both short

movements around its reef habitats and occasional long (hundred of kilometres)

movements. Only short-term movements were reported in the scyliorhinidae Scyliorhinus

canicula (Rodriguez-Cabello et al., 1998; Sims et al., 2001).

Draughtboard sharks of both sexes and all sizes used the complete habitat within the

boundaries formed by the intensive array, the Derwent River and the Upper Channel.

Within the Derwent region, the Crayfish Point Reserve appeared to be towards the limit

of draughtboard shark habitat as a greater proportion of sharks moved between the

Reserve and the mouth of the Derwent rather than moving in the other direction.

Within the small region occupied by the Crayfish Point Reserve, draughtboard sharks

did not use the reserve in a random manner. The great number of movements detected

on the Derwent mouth side of the Reserve (area 1) would be expected as a result of the

greater movement in this direction (as noted above). However, the high use of area 1 by


149

sharks entering and exiting the reserve from the Taroona High site suggest that sharks

were actively using this area as a region to enter and exit the Crayfish Point Reserve.

The main difference between area 1 and the rest of the Crayfish Point Reserve was the

increased presence of higher profile reef. As higher profile reef would be expected to

interfere with acoustic signal transmission, detections in this region were possibly under

represented.

Cooper (1978) and Simpendorfer and Heupel (2004) recommended that the

temporal pattern of spatial occupation is crucial for determining whether an animal

randomly visits habitat or the habitat is the area usually occupied by it (home range). In

species such as neonate blacktip sharks Carcharhinus limbatus (Heupel et al., 2004), the

sixgill shark Hexanchus grisesus (Dunbranck and Zielinski, 2003), and the temperate rocky

reef teleost Cheilodactylus fuscus (Lowry and Suthers, 1998) changes over time of the home

range or seasonal variations in habitat use were reported. These seasonal movements

were related to survival strategies, feeding activity and reproductive behaviour. In

contrast, and similar to other species such as juveniles of Carcharhinus perezi (Garla et al.,

2006), the coral reef fish Plectomorus leopardus (Zeller, 1997) and the snapper Pagrus auratus

(Parsons et al., 2003), draughtboard sharks showed no temporal patterns of habitat

utilisation throughout the study period.

Draughtboard sharks showed a preference for crepuscular and night time activity in

comparison to moving during the day. Similarly, theses same activity periods have been

reported for other bottom dwelling shark species in their natural environment, such as

the angel shark Squatina californica (Standora and Nelson, 1977), the horn shark

Heterodontus francisci (Nelson and Johnson, 1970), the scyliorhinids Scyliorhinus canicula

(Sims et al., 2001), and Cephaloscyllium ventriosum (Nelson and Johnson, 1970). Movements

in draughtboard sharks were probably associated with feeding activity, as the main

dietary items are nocturnally active animals such as octopus (Octopus maorum), squids,

southern rock lobster (Jasus edwardsii) and crabs (Awruch, personal observation).


150

Although night time activity was most common, several sharks also moved during

the day. This has also been observed for other bottom dwelling species such as

Heterodontus portujacksoni (McLaughlin and O'Gower, 1971), Heterodontus francisci and

Cephalloscyllium ventriosum (Nelson and Johnson, 1970) which were all found to feed

mainly at night with a small number of observations of day time feeding. Although

day/night differences in habitat utilisation are common among chondrichthyans

(Gruber et al., 1988; Holland et al., 1993; Sims et al., 2001; West and Stevens, 2001; Sims,

2003) for the majority of the draughtboard sharks there were limited differences

between the areas utilised during the day and night. This suggests that the sharks had

established feeding areas or recognised certain habitat types (eg: high profile reef) as a

more productive region to locate food.

Within the Crayfish Point Reserve, draughtboard sharks preferred high profile reef

habitat to have stationary periods. In addition, divers have reported draughtboard sharks

resting in rocky crevices by themselves or in groups. Similar activity has been reported

for H. portusjacksoni (McLaughlin and O'Gower, 1971) and other species of scyliorhinids

such as C. ventriosum, S. canicula, S. stellaris and C. ventriosum (Nelson and Johnson, 1970;

Sims et al., 2001; Sims et al., 2005). Although periods of inactivity have been reported in

other species, such as H. francisci, C. ventriosum, S. stellaris and S. canicula (Nelson and

Johnson, 1970; Sims et al., 2001; Sims et al., 2005), this was the first time that a

continuous period of five days in stationary behaviour has been documented for any

species. Avoidance of predators, thermoregulation, sexual behaviour and digestion have

all been suggested as reasons for periods of inactivity among benthic sharks, especially

within the Scyliorhinids (Economakis and Lobel, 1998; Sims et al., 2001; Sims, 2003;

Sims et al., 2005). (Sims et al., 2001) reported different sexual aggregation behaviours in

S. canicula, where the resting periods in males occur on gravel substratum in deep water,

while females rest in caves or under rocks in shallow water. In other species such as S.

stellaris, no sexual segregation was reported with both sexes found to rest in a rocky


151

habitat (Sims et al., 2005). In this study, there was no correlation between periods of

inactivity and either physical parameters such as lunar phase, diel cycle, months or tides

or between biological parameters such as sex, reproductive condition, or size. It is

therefore most likely that the reason for extended periods of inactivity was due to

digestion of prey. Awruch (unplub. data) found that large prey items (eg: 4 kg octopus)

were often present in the stomach of the sharks and these would be expected to take a

considerable period to digest. Observations in captivity found that draughtboard sharks

swallowed rather than chewed food items, and recreational fishers report that a problem

with catching draughtboard sharks is that they swallow the baited hooks. It is postulated

that the long stationary periods that were found in this study were associated with

sharks digesting large prey items.

The recapture of the majority of the draughtboard sharks in the vicinity of where

they were released was most likely a function of the research design as few (16.2%) were

returned by non-researchers. Research surveys in the Crayfish Point Reserve and the

east and southwest coasts revisited the same sites. Thus it is reasonable to expect that

the majority of the recaptures would come from these surveys. The higher number of

research recaptures in the Crayfish Point Reserve (89%) could be associated with the

increased and more frequent sampling undertaken in this region. In contrast, surveys in

southwest and eastern Tasmania occurred once a year at similar periods and for the

same duration.

The lack of tag reporting by fishers using traps or nets clearly highlights the problems

associated with gathering data on by-catch species. Although there is greater recognition

that fisheries are to be managed under the principles of ecosystem based fisheries

management, and that recording and reporting of by-catch is important, the reporting of

recaptured tagged animals that are returned to the sea ( ie. no commercial value) remains

problematic. During this study, the tagging program was published in fishing industry

magazines and explained through talks given to both gillnet and trap fishers. Fishers


152

were familiar with reporting tags as many of the Tasmanian target species have been the

subject of tagging studies. Despite this lack of reporting, conventional tag returns have

indicated a degree of mixing over larger ranges. Large distance movements were

recorded between eastern and western Tasmania and between southern and northern

Tasmania. The conventional tag returns have also demonstrated longer-term site

affinities with several sharks being recaptured in the same location up to five years after

tagging. Similarly, long-term site fidelity or philopatric behaviour (animals returning to a

specific location) has been recorded for other species of sharks. The horn shark H.

portujacsoni, the dogfish S. canicula and the hammerhead shark Sphyrna tiburo were

reported to return to a specific location after periods of absence that can be measured in

months or years (Rodriguez-Cabello et al., 1998; Sims et al., 2001; Heupel et al., 2006b).

In summary, although short-term, the acoustic tag data has revealed information on

specific habitat preferences, movements, activity and stationary patterns about this

species that were previously unknown. Together with the conventional tagging data,

there was confirmation that while mixing between broad regions does occur, the general

pattern of movement was of limited dispersion within Tasmania’s major coastal regions.

153

General conclusions

General conclusions

154

The aim of the present study was to investigate the biology and ecology of the

draughtboard shark, Cephaloscyllium laticeps; a common predator of rocky reef ecosystems

in southeastern Australia. This thesis has focused on two important components:

reproduction, including endocrine control, and the study of movement, activity patterns

and habitat utilisation.

In chapter two, the reproductive biology of Cephaloscyllium laticeps was described in

detail. As reproduction is one of the most important events in the life cycle of any living

organism, being the primary requirement for successful propagation, understanding the

reproductive process was considered the first necessary component for scientific

investigation in this species. Sexually mature draughtboard shark females were found

throughout the year with a slightly higher proportion of pregnant females in the first six

months of the year. Sperm production in males was also higher early in the year with a

subsequent slight decline later in the year, evident from both macroscopic examination

and steroid hormones levels. Females laid two eggs at monthly intervals and embryo

development took approximately one year. Together with the results of other studies,

two basic reproductive strategies in oviparous chondrichthyans are apparent. Species

from higher latitudes, such as the draughtboard shark, tend to reproduce all year round

and have longer periods of egg incubation, sperm storage and time between oviposition

of successive pairs of eggs. In contrast, species from lower latitudes tend to have

distinctive reproductive seasons and have relatively shorter periods for incubation,

sperm storage and oviposition.

In addition to macroscopic examination of gonadal stages, the role of steroid hormones

in sexual maturation and egg development was explored, in view of their actions as

triggers or regulators of all aspects of reproduction. In the present study, the steroid

hormones testosterone (T) and 17β-estradiol (E2) played a major role during the

General conclusions

155

follicular phase of draughtboard shark females, while progesterone (P4) was primarily

involved with the ovulatory phase. These results follow the general pattern in T and E2

during follicle maturation reported in other oviparous species. However there appears to

be no constant pattern during the latter stages of the reproductive cycle, with a diverse

behaviour of these hormones in different oviparous species (Sumpter and Dodd, 1979;

Koob et al., 1986; Heupel et al., 1999; Sulikowski et al., 2004). In contrast, and in

accordance with previous studies (Koob et al., 1986; Heupel et al., 1999; Koob and

Callard, 1999), the results of this work showed a clear role of P4 during ovulation and

oviposition. In draughtboard males, T was the main steroid hormone produced during

sexual development. The results of the present study supported the view that T played a

major role in the regulation of testis development and in the final stages of sperm

maturation, as was suggested by various authors (Callard et al., 1985; Sourdaine et al.,

1990; Sourdaine and Garnier, 1993; Heupel et al., 1999; Tricas et al., 2000).

Although studies on chondrichthyan endocrinology have advanced in the last few years

(reviewed in Gelsleichter, 2004), the information is still very limited and insufficient to

completely understand the endocrine control of reproduction in all the oviparous

species in this group of fish. However, results from the present study provided new

information and an improved understanding of endocrine control of reproduction and

reproductive strategies in oviparous chondrichthyans.

Based on the positive correlation between sexual maturity and steroid hormone

levels described in chapter two, chapter three explored the potential of steroid hormone

measurements as a non-destructive technique to assess reproduction for applied

fisheries research. Hormone measurements were found to produce almost identical

results in estimating size at maturity and elucidating the reproductive cycle as

macroscopic examination of gonadal stages from dissected sharks. Although only

validated for Cephaloscyllium laticeps, it is possible that this technique will be widely

General conclusions

156

applicable for describing size at maturity and reproductive seasonality in different

chondrichthyan species. Positive correlations found by other authors between

macroscopic observations and steroid hormones for different species and different

reproductive modes (Rasmussen and Gruber, 1990; Tricas et al., 2000; Sulikowski et al.,

2004), suggest that there is the potential to apply these methods to all chondrichthyans.

While validation of the steroid hormone levels for different chondrichthyans will require

sacrificing a small number of individuals, the hormone levels may be sufficiently

consistent between species or reproductive groups to minimise the need for validating

each species.

Whether studying chondrichthyans bycatch for ecosystem based fisheries

management or managing vulnerable and endangered species, the need for reproductive

data to ensure that populations contribute to future generations is essential (Hall et al.,

2000; Walker et al., 2005). Therefore, non-lethal sampling for biological assessment will

be increasingly important in the management of vulnerable chondrichthyan populations.

These results were the first to demonstrate the potential use of steroid hormones for

applied fisheries management and open the way for hormone measurements to become

a significant scientific tool for non-destructive sampling in chondrichthyans.

In chapter four, a combination of conventional and acoustic tagging studies were

used to understand the movements, activity patterns, and habitat used by draughtboard

sharks. As a bottom dwelling species it was not surprising to find that sharks alternated

between swimming and stationary periods, although the finding that a shark could be

stationary for up to five continuous days has never been previously reported. Although

the majority of the sharks tended to move at night (probably related to movements of

their main prey items), they also make opportunistic movements at other times.

Sharks were found to use all regions of the Crayfish Point Reserve during the six

month study, with preferred regions for stationary periods as well as entering and exiting

General conclusions

157

the reserve. The preference for sharks having stationary periods in high profile reef

areas suggested that the sharks may seek caves as refuge area for predator avoidance.

Although information on draughtboard shark predators has not been reported, this

species has been found in the stomach of seven-gill sharks, Notorynchus cepedianus (Last,

P., CSIRO Marine and Atmospheric Research, Hobart. pers. comm.). Both the

conventional and acoustic tagging data showed a preference for sharks to remain in the

general vicinity of tagging. Gradual dispersion rather than established migratory routes

appeared to be the general movement pattern, however, recaptures from conventionally

tagged sharks did demonstrate that this species is capable of travelling relatively long

distances.

The results of this work have increased the understanding of the behaviour of

bottom dwelling sharks, particularly within the Scyliorhinidae family. Although, a few

studies have described the movement patterns and habitat utilisation of scyliorhinids

(Nelson and Johnson, 1970; Sims et al., 1993; Sims et al., 2001; Sims et al., 2005), no

information on longer-term movements (>6 months) using acoustic technology have

been previously reported. In addition, this was the first work using listening stations to

understand movement behaviour in bottom dwelling sharks in complex reef habitats,

and it was the first study using passive tracking within the scyliorhinids.

With the move to ecosystem based fisheries management it is important to consider

the sustainability of catches of major bycatch species. Fundamental to management of

bycatch will be the need to ensure that populations can be sustained through adequate

reproduction and available habitat. The results of the present study provided important

information on the reproductive cycle, movements, habitat requirements, and activity

patterns of draughtboard sharks. This will allow development of management plans that

consider the requirement of this temperate, rocky reef predator.

General conclusions

158

Marine protected areas (MPA’s) or fishery closures have been reported as an effective

spatial tool for fisheries management (Jamieson and Levings, 2001; Stevens, 2002;

Baelde, 2005; Blyth-Skyrme et al., 2006). Although sharks are usually highly mobile

animals which often have an extensive distribution (Stevens, 2002), MPA’s can still play

a useful role in their management and conservation; as closed areas effectively reduce

fishing mortality protecting parts of the population. However, as draughtboard sharks

showed no indication of distinctive reproductive seasons or areas, and no strong site

fidelity; the implementation of shark refuge areas is unlikely to be particularly effective

in protecting draughtboard sharks. Instead, a minimum legal size above the 50% size at

maturity that enables sharks to reproduce for several years before being harvested

should be implemented. Limiting catches between January and June, the time of peak

egg deposition, should also be considered.

In summary, this study has markedly increased the knowledge of the biology and

ecology of the draughtboard shark. With new requirements to address bycatch in

integrated ecosystem based management programs, together with the ecosystem

consequences of removing upper trophic level predators, it is important to conserve

draughtboard sharks to have a healthy southern Australian reef ecosystem. As well as

providing important life-history information on this species, this work has been some of

the first to investigate hormonal control of reproduction in chondrichthyans in the wild.

This study has also pioneered the use of hormone measurements as a non-lethal

sampling tool for elasmobranch reproductive studies, which has important conservation

implications for protected and endangered species.

159

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