A review of endoparasitic acarines of Malaysia with ...msptm.org/files/01_-_22_M_Nadchatram.pdf · A review of endoparasitic acarines of Malaysia with special reference to novel endoparasitism

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Tropical Biomedicine 23(1): 1–22 (2006)

A review of endoparasitic acarines of Malaysia with special

reference to novel endoparasitism of mites in amphibious

sea snakes and supplementary notes on ecology of chiggers

M. NadchatramFormerly Entomologist and Founding Head, Division of Acarology (1967-1982), Institute for Medical Research,Kuala Lumpur, Malaysia; Senior Teaching Fellow, Department of Zoology, National University of Singapore,Kent Ridge, Singapore (1983-1988).

Abstract. Some 2,000 species of mites of the family Trombiculidae are known in the world.The 6-legged larvae are mostly ectoparasites of reptiles, birds, mammals and invertebrates.Their 8-legged active nymphs and adults are free-living predators. In the Asia-Pacific region,a few species in various genera are vectors of scrub typhus and scrub-itch. In this a paper, avery bizarre trombiculid species, Vatacarus ipoides Southcott 1957, endoparasitic in thetrachea of the amphibious sea snake, Laticauda colubrina (Schenider) is re-described basedmostly on new-born larvae reared in the laboratory. Life history study of the mite producedvery novel and interesting results. A brief account of the life-cycle was presented at the firstlaboratory demonstration of the Malaysian Society of Parasitology and Tropical MedicineMeeting by Nadchatram and Audy (1965). The life history is illustrated and described here ingreater detail. The active nymphal, and the akinetic teleiophane stages are bypassed, whichis unusual in the life-cycle of the family Trombiculidae. Also, the larva is the only stage inthe life-cycle that feeds. The sexes are predetermined in the larval neosomatic stage and giverise to small males and bigger females. Having obtained adults of the species, by rearing, itis deemed unnecessary for the original proposal by Southcott to erect a new family,Vatacaridae, because the adults share all the attributes of the family Trombiculidae. The maleand female obtained through laboratory rearing are illustrated for the first time. Relationshipof V. ipoides with Laticauda snakes show close host-specificity, in a group of acarines thatare generally habitat-specific. Possible explanations for their association are discussed. Theunusual morphology and the formation of new structures during an instar is of ontogeneticand evolutionary importance. The hypertrophic larvae are superficially vermiform, rather thantypically acarine in shape. This, and other biological features, necessitated the proposal ofnew morphological terms, and they are discussed here.

INTRODUCTION

An essential component of investigationsof tropical or arthropod-borne endemicinfections is the systematic collection ofthe incriminating agents to investigatetheir biomedical and scientific importanceon a broad basis. International cooperationbetween scientists at the IMR and scientistswith common interest from variousuniversities and institutions in America,Australia, Europe and elsewhere, and thefree exchange of material, made it possibleto better understand the Malaysian

organisms of diverse forms and life-styles,and their medical or potential medicalimportance.

For example, in scrub typhus research,the study of parasites on a broad basis alsocontributed to our knowledge of ecologyand host-parasite-habitat relationships,which in turn, helped in the use of animalsand their ectoparasites as ecological“labels” or markers to monitor the effectsof deforestation on the natural environ-ment of the Malaysian countryside (Audy,1948; 1954; 1956a; 1956b; 1958; 1960).

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As I look back over some 35 years ofmy happy and fulfilling association withthe Institute for Medical Research throughbiomedical studies of ticks and mites, Irealize how much I owed it to others andhow dependant I was on the ideas andachievements of other learned peers andcolleagues, nationally and internationally.Working in a research laboratory in aspecialized field in a somewhat isolatedenvironment for many years, the realiza-tion that we must always be reminded thatthe vigour of a research organization mustfinally depend on the range and degree ofits contacts with the outside world forfruitful endeavours in the best interest ofscience and the nation, especially in a fieldof study which is unique to the country.Our vigorous activities and collaborativeparticipation gained international popu-larity, and many overseas scientistsinterested in ticks and mites for theirscientific and public health importancespent days, weeks, months and, in somecases more than a year through grants theyapplied for and received from varioussources.

Participation by international re-searchers proved to be of mutual benefit.Some 500 parasitic and free-living mites,and ticks were discovered and describedin collaboration with overseas specialists,and their bionomics clarified through fieldinvestigations and life history studies inour laboratory. These Malaysian investi-gations also contributed to the discoveryof other parasites, both ecto- and endo-parasites, published elsewhere by variousspecialists. There are many new emergingand re-emerging diseases lurking on thedoor steps of the rainforest ecosystem richin the diversity of biosystems. Under-standing the natural history of infectiousand vector-transmitted diseases must be anon-going programme in our country andin any other country like ours in thetropical rainforest ecosystem, rich in faunaand flora. Early in my years I was made tolearn to be broad based in investigationsof parasites, without being rigidly directedby knowledge or suspicion of their abilityto transmit disease. Understanding the

parasites in relationship with the animalhosts has proven equally important inscience.

Known Malaysian endoparasitic mites

of terrestrial birds and animals

Instances of the broad based investigationsof parasites in Malaysia were the dis-coveries of the numerous endoparasiticmites that were found living naturally inthe lungs of monkeys, stomachs of fruitbats, as well as the nasal and respiratorypassages of birds and ground mammals.Though these studies were incidental tothe investigation of the vectors andpotential vectors of scrub typhus, the dis-covery broadened our knowledge of acarineparasitism. Therefore, the opportunity istaken to briefly summarise endoparasitismof terrestrial reptiles, birds, mammals andinvertebrates recorded in Malaysia. Withthe first discovery of the intranasal biotopeof chiggers in Malaysia in 1952, a total of86 species in 16 genera of chigger mitesinfesting intranasal passages of vertebrateswere recorded in a world review up to 1970,and of these 14 were Malaysian species(Nadchatram, 1970a). Over a 40-yearperiod many more species representingmany other families of mites were found invarious biotopes, especially those parasiticin birds by non-trombiculid families ofmites, e.g. Rhinonyssidae, Ereynetidae,Cytoditidae and Turbinoptidae (Nadchatramet al, 1964). Subsequently, additionalspecies of Rhinonyssidae were found onmigratory birds in Malaysia (IMR AnnualReport, 1977: p. 23). The monkey lungmite, Pneu-monyssus simicola Banks wasfound naturally infecting the lungs of theSilver Leaf Monkey, Presbytis cristatus

(Stiller et al., 1974 and IMR Annual Report,1974: p. 85). It is noteworthy that thenymphal stage of the mite was not foundfrom among the thousands of specimenscollected. Many other species of monkeysand tree shrews were also examined forlung mites, with negative results. Fain(1969) discusses the adaptation ofparasitism of mites on various animalhosts. The lungs of the colubrid landsnake, Natrix chrysarga was found

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infected with Entonyssus asiaticus (Fain,1961; IMR Annual Report 1975: pp.34-35;Stiller et al., 1977). More than a hundredspecimens were collected from 4 of 14snakes of N. chrysarga. Bats are also notspared by endoparasitic mites. Thestomaches of fruit bats were infected withGastronyssus bakeri Fain (Fain, 1955;Stiller 1977b; IMR Annual Report 1975: p.35). Of 52 fruit bats collected, 36.5% oftwo species of fruit bats were foundharbouring the mites in their stomach.

Another group of mites that are veryspecialized and restricted in their biotopeare demodicid mites of the speciesDemodex intermedius Lukoschus et al.that were found in the Meibomian glandsof the eye lids of the tree-shrew, Tupaia

glis in Bukit Fraser, Malaysia (Lukoschuset al., 1984). All stages of the vermiformmites were found and described, followedby a discussion of its taxonomy andpathology. Meibomian glands associatedwith the eyes are multi-alveolar glandswith holocrine secretion of a fatty paste-like sebum. Large amounts of this sebum,essential for the normal function of thecornea, are spread on the cornea by themovements of the eye-lids. All stages ofthe life-cycle were found in the glands ofthe upper and lower eye-lids of both eyes.The mites were found in 4 of 17 T. glis

examined and D. sabani in several speciesof rodents (Desch et al., 1984). The genusDemodex is found in humans throughoutthe world living in all stages of the cyclein facial pores and hair follicles of the eye-brows as observed by the author onhimself. Two species of human Demodex

are known – D. folliculorum (Simon) andD. brevis (Akbulatova). It is also worthy ofnote that many different groups ofarthropods are hosts to endoparasiticmites (Fain and Lukoschus, 1983).

Endoparasitism in amphibious sea

snake

The main object of this paper is thedescription of an extremely classicalexample of endoparasitism of sea snakes.The biology of the mites of the familyTrombiculidae found in the trachea of theAmphibious Sea Snake, Laticauda colubrina

(Schenider) (plate I) is described in moredetail from the earlier synoptic descriptionby Nadchatram and Audy (1965) whichwas presented as a laboratory demonstra-tion, organized by the then MalaysianSociety of Parasitologists, as the MalaysianSociety of Parasitology and TropicalMedicine was known then.

The family Trombiculidae is one of thelargest families in the order Acari, withsome 2,000 known species in the world.

Plate I. Banded Amphibious Sea Snake, Laticauda colubrina. The oar-like tail is clearly seen, upperright. (After Ditmars, 1962).

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The 6-legged larvae are mostly ecto-parasites of reptiles, birds, mammals andinvertebrates. Members of this family arealso the important vectors of scrub typhus.In the Asia- Pacific region, includingMalaysia, at least a dozen species of larvalmites in various genera of this family areeither vectors of scrub typhus or thecausative agents of scrub-itch. The 8-leggedactive nymphs and adults are free-livingpredators. The Institute for MedicalResearch, Kuala Lumpur, Malaysia has acolourful background in investigations onvarious aspects of scrub typhus research,since 1925.

In the study of the life history ofVatacarus ipoides, carried out by theauthor at the Institute for MedicalResearch, Kuala Lumpur, it was found thattwo stages in the life-cycle were bypassed,the larvae developed from a singlemoulting stage into adults without feeding,a very unusual phenomenon. The by-passing of the active nymphal stage andthe quiescent teleiophane stage (anexample of tachygenesis) is unique amongmites of the family Trombiculidae. Therevelation of this unusual phenomenon in1960 prompted much excitement. Theunique morphology and the formation ofnew structures during an instar wassufficiently novel and of sufficient onto-genetic and evolutionary importance todeserve new descriptive terms as stated byAudy. Consequently, Audy et al (1963)proposed new terms to define the newstructures in V. ipoides and a number ofother species of trombiculid mites. Thiswas followed by a review of examples ofexternal transformations in a variety ofinsects, acarines and crustaceans (Audy et

al 1972). However, in this paper only themorphological structures and biologicalfeatures that apply to Vatacarus ipoides

are discussed. The terms and definitions asapplied to this species and presented hereare from that of Audy et al. 1963 and 1972,as defined below:

Neosomy (new term, n. Greek neo –new, soma – body), meaning theprocess of new formation of structuresor tissues in a single active instar of an

arthropod, at present recorded only forcertain groups of trombiculid larvaeand represented by development ofnew integument material;

Neosome is a new derivative term: thedeveloped product of neosomy to bedistinguished from earlier stages of theinstar, as in plate II.

Neosomule is a new external structureresulting from neosomy

Neosomatic structures or organs:

new descriptive term. Distinct struc-tures of adaptive value and usually oftaxonomic importance.

The bizarre mites found in the respiratorysystem of amphibious sea snakes of thegenus Laticauda, are capable of increasinggreatly in size to a worm-like or maggot-like creature with neosomatic structures.The term ipomorphy was proposed bySouthcott (1957) to denote such amodification, in groups of animals notnormally so.

In this paper, the larva of V. ipoides isre-described and illustrated based for themost part on unengorged or newly hatchedlarvae obtained from rearing the mites inthe laboratory, for the first time. Descrip-tion of neosomatic or engorged larvaefollows the description, with slight mo-dification, by Southcott, 1957. The adultmale and female are also illustrated for thefirst time, obtained from the mites rearedin the laboratory, to exemplify the featuresof an adult trombiculid mite of normalcontour that emerged from the hyper-trophic larvae bearing corniculate papillae,defined as neosomules and are vermiformrather than typically of the oval or roundshape of larval mites of the familyTrombiculidae. This led Southcott topropose a new family, Vatacaridae toaccommodate the genus and species.However, laboratory rearing of this mitehas shown that the adult mites have all thecharacteristic morphological features ofthe family Trombiculidae, and of normaltrombiculid form, hence there is no validreason to erect a new family, Vatacaridae.However, consideration may be given for

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subfamily ranking of this bizarre creaturein the larval stage. However, the taxonomictreatment is best left to the activelyinterested acarologists.

TAXONOMY

Revision of the genus Vatacarus

Southcott, 1957

(based on larval characters)A mite of the family Trombiculidae,

found in the respiratory system of the genusLaticauda, maggot-like in appearance inthe engorged state (neosome), capable ofincreasing greatly in size, with conicularprojections or neosomules developing onlyin the engorged specimens. Urstigmatapresent. Trachea, though not at all seen inwhole mounted specimens, are apparent inlarval pelts of V. ipoides. Cheliceral fangsrecurved, hinged. Palpal tibial claw 2-pronged. Palpal tarsal formula (PTF) 7BS.Eyes 2 + 2, visible only in unengorgedlarva, not discernable in engorged speci-mens. Scutum trapezoidal, shoulderspresent. 2 anterolateral setae (AL) placed

below anterior margin, 1 anteromedianseta (AM) and 2 posterior lateral setae (PL)situated well above line of posteriormargin. A pair of filiform sensillae. Genuof leg III multisetose. Empodia of tarsus Ito III setiform.

Re-description of Vatacarus ipoides

Southcott

(mostly based on new-born larva, figs. 1-8)

(Terminology in description followsthat of Nadchatram and Dohany (1974).The re-description is based on unengorged(UL) larvae reared in the laboratory for thefirst time, and from neosomatic or fullyengorged larvae collected from the host.(The identity of the mite species wasconfirmed by Dr. Southcott).

Unfed larva 0.45 x 0.21 mm, partiallyfed larva (PEL) elongated, length 2.94 mm;width across idiosoma 1.2 mm, widthacross hysterosoma, or posterior idiosome0.97 mm; engorged larva or neosome 4.0 -4.8 mm x 2.0 - 2.3 mm. Colour in life of ULand PEL yellow to light orange, EL brightorange.

Plate II. A, Scanning electron micrograph of ventral view of entire engorged larva or neosome ofV. ipoides. B, posterior ¾ of the neosome or larva of V. ipoides showing ambulacral papillae(neosomules).

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Figure 1–8. Unengorged Larva of Vatacarus ipoides Southcott:– 1, dorsal and ventral aspects ofidiosome; 2, scutum; 3,4, dorsal and ventral aspects of gnathosome; 5,6,7, distal segments of legI, leg II and leg III; 8, humeral, dorsal and ventral setae.

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UL and PEL free of conical projectionsor neosomes of idiosome. UL broadly oval,PEL idiosome swollen, hysterosomagreatly elongate. EL elongate, conicularprojections or neosomes more prominentdorsolaterally and posteriorly. Eyes 2 + 2,placed on ocular plate; anterior eye roundand oculate, posterior eye ovoid andindistinct; ocular plate situated laterally inthe posterior half of scutum, the midline ofplate being in line with PL base. (Eyes notapparent in engorged specimens, butvisible in UL and PEL, less so in the latter,and not easily discernable in old mounts.)Eyes, in live UL and PEL visible as redspots under low magnification. Gnatho-

some: Fairly heavily chitinised, chelicera(37 um), flexed with pointed apex, a small

distal denticle and a big hook-like ventraldenticle. Galeal seta (15 um) nude, fineand tapering. A pair of coxal setae withadpressed ciliations. Palpal formula b/b/bNN + 7BS (PTF); palpal setae weak,femoral and dorsotibial setae more deve-loped than others and have swollen bases;dorso-tibial seta stoutest; lateral seta andventral seta of tibia normally ciliated, thelatter two nude and tapering. Palpal tarsuswith a long, blunt subterminala, a sub-apical sensory seta, and 3 fairly long, and4 short setae with short indistinct barbulesand appearing almost nude. Claw 20 umwith 2 strong prongs, ventral stronglyrecurved with pointed apex, dorsal clawless recurved and blunt. Scutum: (also seescanning electron micrograph, plate III)

Figure 9–16. Neosomy in Vatacarus ipoides Southcott, larva (9–10) new born, lateral and dorsalview; (11–13), progressive intermediate degrees of enlargement, lateral view; (14–16), neosome,ventral, dorsal and lateral view (after Audy et al., 1963).

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trapezoidal, anterior width greater thanposterior, anterior shoulders not evenlyrounded, lateral margins sinuous, posteriormargin slightly concave medially; posteriormargin more distinct than the other scutalmargins, but not chitinised. Punctationsclose-set and evenly distributed. Scutalsetae with slightly expanded bases, buttapering distally, with minute barbs.PL>AM>AL, the difference in measure-ments being gradual. Sensillae long andfiliform (85 um).

Standard measurements (in um) ofscutum, mean of 4 unfed larvae:

AW A-AL PW P-PL SB ASB PSB AP AM AL PL SENS96 14 82 36 59 49 69 32 52 48 65 85

Dorsal setae (56 um), slightly swollen atbase, and tapering distally. DS arranged insomewhat transverse rows of 2. 6. 4. 6. .6.4. 2. 2 (0) in unfed larva. Ventral setae (VS)2 pairs ciliated sternal setae, plus 40- 44true ventral setae 32 um long. Both DS andVS are inserted on neosomules or conicalprojections in engorged larvae.

Legs: 7-segmented and fairly sclero-tized. Tarsal claws I-III stout; empodialong, thin and delicate, and easily brokenin engorged specimens. Coxae I and IIwell separated in engorged neosomes andpartially engorged larvae, but less so in thelatter. Coxae I and II almost fused in ULs.Ordinary setae have very short adpressed

Plate III. Electron micrograph of scutum of engorged larva.

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ciliations and therefore appear nude.Coxae I to III unisetose, the setae beingslightly swollen at base. Sensory andbarbed setae as follows: Leg I: tarsus +pretarsus 95 x 41, tarsala elongate andblunt (44), microtarsala situated at tip oftarsala; a pretarsala, a long subterminala,a small tapering parasubterminala placedclose behind subterminala, and 20-22barbed setae. 2 tibialae, distal one stout,blunt and striated, proximal slender andpointed; a fairly long microtibiala placedbelow and in line with distal tibiala, 6barbed setae; 6 tapering genualae (variesfrom 5-7 in some specimens), l micro-genuala and 4 ciliated setae; remaining 3basal segments with 5, 1,1 ciliated setae.The seta on basifemur nude and pointed.Coxa I 76 x 56. Leg II : tarsus + pretarsus75 x 40 um, a short cigar-shaped striatedtarsala (26 um), a microtarsala abovetarsala plus 15-16 ordinary setae; 2 tibialae,the distal striated and blunt, proximaltapering plus 5 ordinary setae; 3 subequal,tapering genualae and 3 ordinary setae;other 3 basal segments with 4, 1, 1 ordinarysetae. Coxa II: 80 x 84 um. Leg III : Tarsus+ pretarsus 82 x 40 um. One indistinctmastitarala and 13 ordinary seate; apointed tibiae + 3 ordinary setae; other 3basal segments with 3, 2, 1 ordinary setae.Coxa III 86 x 52 um.

Remarks on taxonomy: It isinteresting to note that ordinary or non-sensory setae are of two kinds - setaewithout basal expansion and those withswollen bases as in the genus Babiangia -complex. Radiating lines around scutumand coxae I and II were observed in someengorged larvae, but not so in UL or PEL.Evidence of trachea was found in larvalpelts, but not on larval specimens. It is alsonoteworthy that variations in numeroustaxonomic characters were observed whichis either related to sexual differences orother intraspecific variations.

Southcott in his description of V.

ipoides stated that the empodia tended tobe retroflexed, and had figured it so. Hehad, since, examined some of the speci-mens we sent him, and has noted (pers.commun.) that the empodia are setiform.

Material examined: The mites werefound in the air-sacs (tracheae) of theAmphibious Sea Snake, Laticauda colu-

brina (Schneider), found in low tide in themangrove tree holes and in rock holes ina tidal reef, near Pulau Sudong, about 5miles off Singapore by M. Nadchatram inOctober, 1959.

General remarks on slide preparation

Making slide preparations of the neosomesof Vatacarus caused considerable tech-nical problems at first due to their largesize and cylindrical shape. The cuticle ofthe species in the engorged state was fatty,leathery, and thick-walled. Potassiumhydroxide was an unsatisfactory clearingagent, because specimens cleared in itdisintegrated when mounted. At first it waspossible to mount neosome Vatacarus onlyafter dissecting it into several parts, amethod that is still useful for detailedstudy of certain taxonomical characters.Subsequently, a somewhat satisfactorymethod for making whole mounts wasdeveloped. After a few tiny holes made inthe posterior end, the specimen wasplaced between 2 slides, and gradualpressure was applied and the glass slideswere tied together with a string. Some ofthe liquid fatty content oozed out throughthe artificial pores on pressure. The speci-men was then placed in a petri dish whileit was still pressed between the slides,flooded in lactic-glycerine. After a weekthe specimen was transferred to the PVAmounting medium. The method was notideal, but it was quite satisfactory. (Thesenotes were made in the early 1960s, whenthe choice mounting medium was PVA orPolyvinal Alcohol. The medium of choicenowadays is Hoyer’s medium. This mediumis found to be satisfactory because age-oldmounts are easily recoverable and re-mounted.)

The adult male and female V. ipoides

Synopsis of the descriptions of the adultmale and female Vatacarus ipoides

Southcott (Fig. 17-28, male; Fig. 29- 40,female).

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Figure 17–28. Male Vatacarus ipoides Southcott:– 17, dorsal and ventral aspects of idiosome; 18,crista and epistome; 19,20, dorsal and ventral aspects of gnathosome; 21, male genitalia;22,23,24,25, distal segments of legs I, II, III, IV; 26, coxae I and II; 27, coxae III and IV; 28, dorsaland ventral idiosomal setae.

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Figure 29–40. Female Vatacarus ipoides Southcott:– 29, dorsal and ventral aspects of idiosome;20, crista and epistome; 31,32, dorsal and ventral aspects of gnathosome; 33, female genitalia;34,35,36,37, distal segments of legs I, II, III, IV; 38, coxae I and II; 39, coxae III and IV; 40, dorsaland ventral idosomal setae.

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(Some drastic changes and movementshad taken place in the acarology labora-tory a few years ago after my retirement,when the slide collections of these andother material were either lost or mis-placed. Therefore, the synopsis presentedhere is based on the illustrations only.However, it is felt justified to publish theillustrations to show the typical contour ofthe slide preparation of the adult mites,which were directly reared in the labora-tory by the author. The publication of theillustrations would also be of some help tofuture workers. Also, the illustrations wereprepared in Japan, at the U.S. Army 406laboratory, at considerable expense andlabour and would prove it to be wastefulif they are not published).

The adult mites emerged directly fromthe neosomatic larvae. They were orangecoloured in life. Live males were slightlysmaller and slender.

The adults are of the general facies oftrombiculid mites, with the usual scutumor crista and epistome, the genitalia ofmale and female show 3 pairs of genitaldiscs (nymphs only have two pairs). Thegnathosome and legs are also of normalcontour. However, the dorsal aspect oftibia of the gnathosome bears 3 pairs ofspur-like setae just proximal of the claw, asin other trombiculid mites. The male wasmore elongate and smaller than the female,the idiosome of male more slender. Male2.0 to 2.5 mm, with propodosoma 1.2 mm,hysterosoma 1.7 mm. The constriction ofthe idiosome is characteristic for mites ofthe family Trombiculidae, the constrictionof both male and female is between thirdand fourth pair of legs. Idiosome of female3.0 to 3.5 mm with hysterosoma wider thanpropodosoma. As in other trombiculidmites it has more setae of the ordinarybarbed type to give the adult a velvetyappearance.

BIOLOGY

The life history of Vatacarus ipoides

The amphibious sea snakes, Laticauda

colubrina located near Pulau Sudong,

Singapore were seen in most of the treeholes by J.R. Audy, and rock holes in thetidal reef at low tide by M.N. As many as 5or 6 in each of the many holes or crevicesin the coral reef were found. A total of 18snakes were collected and transported inmetal tanks to the Institute for MedicalResearch, Kuala Lumpur, where the livesnakes were kept in tanks filled with seawater. The snakes were dissected fromtime to time, to recover the mites alive.The trachea or air-sac was slit open toexpose the entire windpipe. All of the 18snakes examined were infested with mites,which were subsequently identified as V.

ipoides, and confirmed by Dr Southcott.The older and larger snakes harboured themost number of mites, 60 being the largestnumber found in a single big snake. Thebigger, maggot-like mites or neosomeswere confined to the upper region of thetrachea, a few were found dispersed in theposterior portion of the trachea, but nonewere found in the lung-sac. Almost allmites found were fully fed. Lungs ofamphibious sea snakes are much largerthan their land based relatives. Mostchigger mites increase their volume some20 x with engorgement, Vatacarus larvaeincrease 1,500 x or more. The legs,functional in the unfed stage, are useless,relatively minute appendages of theneosome. The cornicular projections orneosomules (plate II) have an obviousfunction enabling the larval mites to movein the mucus secretions in the manner ofmaggots or worms (plate IV A). The pro-gressive degrees of enlargement fromnew-born to neosomatic larvae is asillustrated in figures (Fig. 9–16).

The engorged or maggot-like mites inbatches of 15 x 4, were reared in porouspots made of unglazed clay, about 1½” inheight and 1¾” wide (Nadchatram,1968).The pots were boiled several times toremove soluble minerals before use. Apinch of sterile soil was introduced tosimulate natural conditions. A glass coverwas placed over the pot to contain themites within the pot. The ambient tem-perature of the area housing the breedingunit was min. 25.5ºC, max. 30ºC, with RH

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63 - 87 %. The larvae were reared to adults,and F1 eggs and larvae were obtained.

The neosomatic larvae were sluggishand moved very slowly or not at all. Twosizes of neosomatic larvae, some smallerthan the others, were seen. In the formercase it was possible to determine if themite was alive by the contraction andexpansion of the body. On the third dayafter being placed in the breeding pot thelarvae became quiescent, without visiblemovement. From this stage onwards thecornicular projections or neosomules(plate II) gradually disappeared until thebody was smooth-surfaced, and resembledan inflated, elongate balloon, or shapedlike a Havana cigar (plate IV D). Thischange was completed by the end of thefifth day, and 9 days later, the adults of twosizes emerged. The adults were orangecoloured, the contour was typically oftrombiculid form, with medial body con-striction. It was at first thought that thepost-larval stages were nymphs, but wassubsequently established (more from theirbehaviour than anything else) that theywere, in actual fact, sexually separableadults. Only half of the larvae emerged asadults. Of the others, many were attackedby fungi, and a few failed to metamorphosefor unknown reasons.

The development of adults directlyfrom larvae is an extreme example oftachygenesis. Both sexes of adults wereobtained, and it is highly probable that thesmaller, neosomatic larvae gave rise to themales, and the bigger ones to females,suggesting that sex differentiation ispredetermined, hence the sexes were easilyseparable, (plate IV E) unlike in otherspecies of trombiculid adults, whichneeded to be turned-over for a ventralview, to examine the genitalia to determinethe sex. The males in this species wereabout 2.5 to 3 mm, which are approximatelythe size of Blankaartia acuscutellaris, ascrub-itch chigger, and bright orange incolour. The females were 1.5 times biggerthan the males, and lighter orange. Theratio of males to females was 3 to 2.

Both males and females were normallyactive. One day after the first batch of

adults emerged, a few tiny mould-likestructures were seen here and there in apot containing both sexes. The head of thismould-like structure (sperm-sac) wassmall and round and was supported on avery thin stalk. (Some of the stalks werewithout the head, suggesting that thesperm-sac were taken by the female) Thehead of the sperm-sac capsule was pallidand translucent. On careful examination itwas established that the structures were,indeed, spermatophores with the spermpackets. Adults, both male and female, thatemerged subsequently were segregated bykeeping them in separate pots for twodays, but spermatophores were not seen inthe pot containing the males. However,within about an hour after the males andfemales were brought together, three stalksof spermatophore with sperm packets orsacs were seen. This showed that the maleswere stimulated to deposit the sperm onlyin the presence of the “fairer sex”. Thoughit was not observed by us, it is most likelythat the females take up the sperm throughthe genitalia, leaving only the empty packetand stalk as demonstrated by the femaleBlankaartia acuscutellaris. This uniquemethod of insemination and fertilisation ofthe female trombiculid mites was describedby Lipovsky et al. (1957). Whether or nota ritualistic dance precedes spermatophoredeposition, as in some other groups ofacarines, is not known. However, the modeof transfer of the male sperm packet to thefemale further supports the fact that Vata-

carus is a member of the family Trombi-culidae. Two virgin females were isolatedimmediately upon emergence, but they didnot produce eggs till the day they died 30and 36 days later. From these observationsit is perceived that parthenogenesis doesnot occur in this species. Also, the adultmites seem to have a much shorter lifespan (6 to 8 weeks) compared with adultsof Leptotrombidium deliense or Blan-

kaartia acuscutellaris whose life spanunder laboratory conditions is about ayear, or longer.

Eggs were seen 12 days following thedeposition of spermatophores, and onefemale continued to oviposit for 24 days.

15

Eggs were laid singly at long intervals, thenumber per day ranging from 1 to 7. Thenumber of eggs laid by a single female was52, the average number of eggs laid varied.To observe the number of eggs laid, andthe duration of egg laying, 3 fertile femaleswere placed in 3 separate pots. In pot # 1:46 eggs were laid in 17 days; the femaledied on the 27th day; highest number ofeggs laid was 7, on the 9th day. In pot #2:52 eggs were laid in 22 days, the highestnumber laid was 6, on the 15th day, thefemale died on 43rd day. In pot #3: 42 eggswere laid in 24 days; the female died on30th day, highest number of eggs laid was6, on 7th day. The eggs were orange, roundand a little bigger than those of Lepto-

trombidium deliense, the vector of scrubtyphus. Deutovums were seen 7 days afterovipositon, and larvae hatched 16 dayslater.

The newly emerged larvae were appro-ximately 2 x the size of newly hatchedlarvae of L. deliense, and the idiosomeunder low-power magnification was clearlydivided into a broad anterior half and anarrow posterior half. On hatching, theunfed larvae crawled up the pot and restedin small clusters underneath the glasscover, as new born larvae of L. deliense

and B. acuscutellaris did. The larvae in theunfed state were yellowish-orange andquite active, and lived for a maximum of

18 days. Adult females lived for 8 weeks,and the males for 5- 6 weeks. During theperiod under observation cannibalismamong the adults did not occur.

Four of the host snakes were kept alivein sea water. One of them died 3 weeksafter capture. Six engorged larval miteswere recovered from the tank that held thesnake, and 20 more from the dead snake.The last 3 snakes died 64 days after theircapture. A total of 57 engorged larvae werefound in the tracheae. In all the 18 snakesexamined, almost all the larvae wereengorged; only about 10 partially engorgedlarvae were found among the severalhundreds that were recovered from thesnakes. Not a single unengorged larva wasfound. The feeding time of the larval miteis unknown. It is suspected that thefeeding time is short, because almost allthe mites found in the snakes were mostlyfully engorged. The life-cycles of Blan-

kaartia acuscutellaris, a species with atypically normal trombiculid life cycle, andV. ipoides are presented in tabular form,for comparison

The normal life-cycle of mites of thefamily Trombiculidae consists of the egg,deutovum, active parasitic larva, akineticnymphophane, active free-living nymph,akinetic teleiophane, and finally the free-living adult. In Vacacarus, the activenymphal and the akinetic teleiophane

Table 1. Life cycle of V. ipoides compared with B. acuscutellaris

Stages of life-cycle Blankaartia acuscutellaris Vatacarus ipoides

No. of days No. of Days00l

Engorged larvae placed in breeding pot 0 0

Larvae quiescent in 2-3 “Pupal stage” 8

Nypmphs emerged (fed on culicine eggs) 6-7 lacking

Nymphs quiescent in 8-9 lacking

Adults emerged in 8-9 9-10

Spermatophore deposited in 1-2 1

Eggs laid after 12-13 10-12

Deutovum developed in 5 7

Eggs hatched in 10-12 14

Development period of F1 from EL to UL 48-56 48-52

16

stages are absent. However, the totalnumber of days required to complete thelife-cycle is not significantly different.

The breeding experiment has esta-blished a few interesting biological facts,as listed below:

(1) The protonymphal and active nymphalstages are bypassed, the neosomaticlarvae giving rise to adults of differentsexes, after a single moulting stage.This is an example of tachygenesis andis unique, and unusual for the familyTrombiculidae.

(2) The larva is the only stage in the life-cycle that fed, the free-living adults didnot feed, but produced eggs. (Normal-ly, free-living adults feed on otherarthropods or their eggs beforeproducing eggs.)

(3) The sexes of the adults were easilydistinguished, the male being smallerthan the female, the sex difference isprobably predetermined in the em-bryonic stage, but quite evident in theneosomatic larval stage. (Sexes ofnormal adults are of similar size.)

(4) The male deposited sperm packetsonly in the presence of the oppositesex.

(5) Cannibalism was not seen to occuramong adults.

(6) Newly emerged larvae, male andfemale adults had short life span. Forexample, ectoparasitic unfed larvaelived for up to 18 weeks, not 18 days,free-living adults lived for over a year,not for 6 to 8 weeks.

(7) Normal chiggers increased theirvolume with engorgement some 20 x,but Vatacarus larvae increased 1,500 xor more.

(8) When the new born larva enter therespiratory track and begins to feed,the mite is a completely new creaturecompared with other ectoparasitictrombiculid mites larvae.

The following freshwater snakes wereexamined for tracheal mites but were

found uninfected: 12 Dog-Faced Watersnakes, Cerberus honchos, a commonspecies in fresh and tidal waters, especial-ly in muddy creeks near the coast. 20 Puff-Faced Water Snake, Homalopsis buccata,common in freshwater ponds and swamps,both near the coast and far inland; and 5Elephant’s Trunk Snakes, Acrochordus

javanicus, common in tidal streams.In addition, 39 specimens of true sea

snakes of 6 different species held incaptivity by the Penang Hospital forresearch on sea snake anti venom, wereexamined for mites in the trachea, withnegative results.

RESULTS AND DISCUSSION

Relationship of V. ipoides with the host,

L. colubrina – a possible explanation

The habitat and behaviour of the Amphi-bious Sea Snake is based on Sharma(1973), Tweedie (1983), Ditmars (1962),and from my observations and inferences.

The Amphibious Sea Snake, Laticauda

colubrina, is the only local species foundin the tidal reefs and mangrove tree holesof Singapore, and possibly the west coastof Peninsular Malaysia. It has broadabdominal plates like terrestrial snakes,although the tail is wide, flat and paddle-like. It is said to be encountered on thebeach where it seeks shelter under bouldersor in rocky niches. It also lays its eggs onland, unlike other sea-snakes which areviviparous and entirely aquatic. This speciesalso frequents mangroves, and off the near-by island of Pulau Sudong, where the authorcollected as many as 5-6 snakes from asingle rock hole in the tidal reef. Althoughconsidered potentially dangerous, thisspecies has never been known to attackanyone in the sea in this area. It appearsto be quite nonaggressive and makes noattempt to bite even when molested.According to Tweedie (1983) the sea snakesare common and found in numbers on theshore on small islands, hiding in crevicesor under stones, off Singapore. The landarea near Raffles Lighthouse had beenreported as a nesting site. Species of

17

Laticauda are known for their wide distri-bution across the Pacific and IndianOceans. They are associated with marinewaters and rocky areas or coral along theseacoast. Nadchatram and Radovsky(1971) described a second species ofVatacarus from L. colubrina from Lan Yu,a small island of unknown name, 70 km SEof Taiwan. The endoparasitic mites aresupposedly common wherever the snakesare found. In 1980, a collection was re-ceived from Professor A. Fain of Belgium.The collection of mites were from the lungsacs of L. colubrina and L. laticaudata

found in New Guinea and Caledonia andpreserved since 1878. The species wasclose to Vatacarus, and described asIguanacarus alexfaini Nadchatram 1980.

The sea snakes have a cylindrical body,with a laterally compressed, paddle-liketail. A salt gland under the tongue givesamphibious snakes the ability to expelexcess salt absorbed from the marineenvironment. Whether or not this bio-logical feature would be a factor which islikely to facilitate endoparasitism of theamphibious sea snakes is not known. Thegenus Laticauda appears to have evolvedfrom an elapid stem much later than theother sea-snakes. In their natural habitat,the snakes feed primarily on eels, but willoccasionally prey on small fishes. Whenfeeding they have been seen to trap smallfish in rock crevices with coils of the bodyand then seize them. Laticauda species.

(there are 3 known species, colubrina,

semifasciata, and laticaudata) are theonly sea snakes known to lay eggs on land,either on the sand or under it. Courtshipand mating occur on land. The breedinghabitat must be protected sufficiently toprovide shade and shelter. Some form ofbiological mechanism must trigger the fedlarvae to leave the hosts when the snakescome ashore for the mites to continue theirfree-living phase. Snakes of Laticauda

species spend much time on land and,therefore, are the only ones which couldhave any kind of “den” relationship thatcould be utilised by the mite parasite forspreading from snake to snake. Also,because of the gregarious behaviour of

these snakes, they must have recognisedsites to which they come for egg laying andother reasons.

There has to be a very closerelationship between the snake and theparasites. Large populations of the snakesare said to be found on relatively smallislands during the breeding season. Therock holes and the holes in mangrove treesare only the resting sites during low tide.These holes were examined, but did notyield the free-living adults of the mite. Alsoall the larvae found in the tracheae of thesnakes were mostly fully engorged, but didnot detach, only waiting for the snakes togo ashore. The sea snakes leave the oceanfor land at about 10 day intervals, usuallyat night, to digest food, engage incourtship, lay eggs, and slough skin. Thisis when the engorged or replete larvaeleave the host to continue with theprogression of their life cycle. In thebreeding season, the movement of thesnakes from sea to land is said to be morefrequent.

Tick Infestation

It is ecologically noteworthy that a speciesof tick, Amblyomma nitidum Hirst andHirst, 1910 was found on at least threeseparate occasions by Audy et al. (1960).A female adult, nymphs and a few larvaewere collected, which supports the “den”behaviour of the amphibious sea snake onland Audy et al. Warburton (1933) describedA. laticaudae from the shed skin of L.

colubrina held at the Raffles Museum,Singapore. Some doubt has been cast onthe status of the Amblyomma speciesinfesting this species of sea snake (Petney& Keirans, 1995). They concluded that bothnitidum and laticaudae are valid speciesbased on hypostomal dentition of malespecimens. A. laticaudae possess 3/3dentition, whereas A. nitidum has 4/4dentition in the male. For a species of seasnake that lives and breeds in completeecological isolation from other species ofsnakes, and where the ticks are identicalin other taxonomic ways, it is very unlikelythat on the bases of hypostomal dentition

18

the two forms can be regarded as separatespecies. I support, as Audy et al did, Kohls’contention that the ticks represent a singlespecies. Kohls, in Audy et al (1960) hadcompared the Singapore specimens withthe descriptions of A. nitidum and A.

laticaudae and had observed the differencein hypostomal dentition in the males, andhad suggested that this could be due tovariation. Kohls (1957) concluded thatlaticaudae is a junior synonym of A.

nitidum. From the literature reviewed, A.

nitidum is host-specific to amphibious seasnakes.

Habitat and Ecological Niche of Chiggers

With reference to the behaviour of thelarval trombiculids mites, Audy (1956a)and other observers Nadchatram &Dohany, (1974) and Traub & Wisseman(1974) have noted that trombiculid mitesgenerally show little or no host-specificity,but rather a varying degree of habitat-specificity, with a few notable exceptions,for example, in birds, a few genera ofchiggers are primary parasites showinghost preference. Like-wise with reptilesand bats in Malaysia. Years of studies inMalaysia have borne it out. Thoughhabitat-specificity may give an appearanceof host-specificity in many genera oftrombiculid mites, it is, however, a wellestablished fact that all the mite vectors ofscrub typhus and the mites causing scrub-itch, which are red or orange in colour, arehabitat-specific (Audy, 1968). It is nowclear, based on Malaysian studies of host-parasite relationship of the thousands ofreptiles, birds, bats and other mammals,that certain groups of mites pertainparticularly to reptiles (e.g. the generaEltonella, Babiangia and Fon-secia), andothers almost exclusively to groundmammals (e.g. the genera Gahrlie-pia,

Walchia, and Helenicula). Certain groups,especially closely related to their hostsin one way or another, show signs ofdeveloping true host-specificity on birds ofsome genera (e.g. Neoschoengatia, Tori-

trombicula and Odontacarus, and on bats(e.g. Chiroptella, Trombigastia, andRiedlinia).

Based on approximately 35 years offield and laboratory investigations carriedout at the Institute for Medical Research,Kuala Lumpur, Malaysia, 148 species oftrombiculid mites or chiggers were re-corded. Taxonomical, biological andecological investigations were conductedon most of the species, with reference tounderstanding their epidemiologicalimportance. On the basis of these studiesan ecological classification was conceivedand published by Nadchatram (1970b). Theclassification divided the Malaysianchigger mites into seven different eco-logical categories. Alland (1967) statedthat all scientific theories have three thingsin common. They are simple; they explaina great deal; they can be tested througheither observation or experimentation, orboth. According to him, Darwin’s theory ofevolution is one such elegant theory. Witha few basic assumptions, Darwin’s theoryattempts to explain the development anddiversification of life. Nadchatram (1970)following Darwin’s concept endeavouredto illustrate the diversification of thetrombiculid mites or chiggers to betterunderstand their epidemiological im-portance or significance. The followingcriteria were used to arrive at the eco-logical classification.

A. Colour of chiggers; all chiggers aresoft bodied, the exoskeleton sclerotizedonly in certain regions, their colour rangefrom white or pallid, to yellow, orange tored. The colour is mostly influenced by thehabitat; B. natural host preference ofchiggers based on the examination ofthousands of reptiles, birds and mammals;C. natural habitat preference of new bornchiggers; D. breeding experiments usinga standard rearing technique; E. mor-phological and taxonomical characteristicsof the chigger species.

A major attribute of this classificationis its provision of an ecological frameworkwithin which the present wealth of chigger-related information may be considered. Itwas shown that all the orange and redchiggers living on the ground-surface wererelatively highly adaptable to fluctuatingenvironmental conditions, and parasitic on

19

all animals, including man. The sensillaewere either filiform or filamentous. Thetarsus of the palp always has a basal tarsalaand a number of barbed setae, varyingfrom 3 to 7, and with or without nudesubterminala(s). This is expressed as apalpal tarsus formula (PTF). For detailedex-planation see Nadchatram & Dohany,1974. For the purpose of this discussionthe PTF determines the ecological affinityof the chiggers, so that 7B or 7BS isassociated with orange or red chiggers thatinfest a variety of species of host animalsand are tolerant to changing environmentalconditions, whereas species of chiggerswith 3B and 4B are white or pale coloured,restricted in their host and habitat andsensitive to changing environmentalconditions. The orange or red ones werechiggers of Eco-logical Group I, the groupthat included all the species of chiggermites of epidemio-logical importance.Conversely, those species of chiggers ofEcological Group II were pallid or white,or pale yellow, and were sensitive tochanging environmental conditions andthe free-living post-larval stages werecryptozoic and lived in ground burrows ofrodents. Sensillae were club-shaped orglobose and PFT 3B, 4B, 4BS or 5B. Of theother groups i.e. Ecological Group III toVII, the determining factors were based onhabitat and host. Group VI is discussedhere because it involves the two speciesfound hypodermal in frogs, and endo-parasitic in sea snakes, respectively. Bothspecies, Vercammenia hendricksoni Audyand Nadchatram and V. ipoides wereorange in colour, suspected to be living inclose association in the same habitat as thehosts. Whether or not they are adaptableto fluctuating environmental conditions willnot be known until the nest sites areinvestigated to determine the behaviour ofthe adult mites. The PTF is 7BS in larvalV. ipoides and 7B in Ve. hendricksoni asin other surface dwelling chiggers. Asmentioned earlier the larvae and adultsof both species were orange coloured.However, based on the know-ledge gainedfrom our laboratory studies of V. ipoides

and the morphological modifica-tions seen

in the larval stage, and based on theabbreviated life-cycle and short life span,it is sufficiently evident to postulate thatV. ipoides is a host-specific parasite, andthat the adults are surface dwelling withinthe confines of the snake’s breeding sites.There is an extremely close relation-shipbetween the parasite and host, because ofthe clustering behaviour of the larvalmites and the “den” relationship of the seasnakes.

Acknowledgements. The support of theDirectorate, I.M.R., Malaysia is gratefullyacknowledged. I am grateful to a numberof friends and colleagues. First of all, Iwish to thank Professor Yong Hoi Sen,Univ. Malaysia for his editorial review ofthe first draft of this paper. I gratefullyacknowledge the arrangements andlogistic support, and many other courtesiesprovided by Dr John R. Hendrickson,formerly Professor and Head of Zoology,University of Malaya, Singapore in 1959,which made it possible for both Dr J. RalphAudy and me to visit, collect and transportthe snakes to Kuala Lumpur. My thanks aredue to Dr. Lim Boo Liat, formerly Head,Medical Zoology Division, I.M.R., KualaLumpur for providing the fresh watersnakes. The close collaboration thatprevailed between his Division and theDivision of Acarology made it possible forthe thousands of animals collected by theDivision of Medical Zoology to be madeaccessible to my Division for study overmany years. Dr R.H. Reid, formerly Headof the Sea Snake Anti-venom ResearchCenter, General Hospital, Penang forpermission to examine sea snakes from hiscollection in Penang. Grateful thanks areextended to Captain Murdoch, formerly ofthe U.S. Army Medical General Laboratory(406), Tokyo, Japan for the arrangementsmade to illustrate the larval and adultmites by Mr Sonobe. The scanning electronmicro-graphs were made at the HooperFounda-tion in the early 1960s. For the useof the micrographs in this paper, I amgrateful to the Directorate of theFoundation. My grateful thanks are due MrLee Woon Kai of the Entomology Unit for

20

scanning the plates and illustrations usedin this publication at very short notice, andwith excellent results.

Most of all, my mentor, formerly Head,Division of Virus Research and MedicalZoology, I.M.R. (1950-59), and later Director,Hooper Foundation, University of CaliforniaMedical Centre (1959-74), the late ProfessorJ. Ralph Audy, was instrumental in mydevelopment as a research worker. Heworked very hard to insure that myachievements were recognized. It was DrAudy who inspired me in this, and manyother research programmes with provisionof funds from the U.S. International Centerfor Medical Research and Training, SanFrancisco, through the Hooper Founda-tion, attached to Institute for MedicalResearch, Kuala Lumpur, to recruittechnical staff and for other laboratoryneeds. This paper was to have been writtenjointly much earlier, but sadly it did notmaterialize for various reasons. It is withmuch pride and humility that I dedicatethis publication in fond memory of thisinspiring genius, the late Professor (Dr.) J.Ralph Audy. His significant contributionsto Malaysian biomedical science ispublished in the form of an obituary(Malayan Nature Journal 28: 51-56, 1974.

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