Chile Wokshop, November 2003 Larval food (Artemia), larviculture and ecotoxicology group (UE partner 4) Members: Dr. Francisco Amat Dr. Francisco Hontoria Dr. Juan Carlos Navarro Dr. Inmaculada Varó Óscar Monroig Germán Medina Olga Ruiz Instituto de Acuicultura de Torre de la Sal Consejo Superior de Investigaciones Científicas 12595 Ribera de Cabanes (Castellón) SPAIN
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Chile Wokshop, November 2003Larval food (Artemia), larviculture and ecotoxicology group
(UE partner 4)
Members: Dr. Francisco AmatDr. Francisco HontoriaDr. Juan Carlos NavarroDr. Inmaculada VaróÓscar MonroigGermán MedinaOlga Ruiz
Instituto de Acuicultura de Torre de la SalConsejo Superior de Investigaciones Científicas12595 Ribera de Cabanes (Castellón)SPAIN
Brine shrimp Artemia franciscana as exotic invasive species in the Western Mediterranean and its dispersal by water birds
Brine shrimp in Argentina
Brine shrimp in Tunisia
Brine shrimp in India
Brine shrimp Artemia franciscana as exotic invasivespecies in the Western Mediterranean and its
dispersal by waterbirds
Francisco Amat, Francisco Hontoria, Olga RuizInstituto de Acuicultura de Torre de la Sal (CSIC). 12595 Ribera de Cabanes (Castellón). Spain.
Marta Sánchez, Andy Green, Jordi FiguerolaEstación Biológica de Doñana (CSIC). Avda. Mª Luisa, s/n. Pabellón del Perú. 40013 Sevilla. Spain.
Francisco HortasGrupo de Conservación de Humedales Costeros. Departamento de Biología. Facultad de Ciencias del Mar y Ambientales. Apdo. 40. 11510 Puerto Real (Cádiz). Spain.
Brine shrimp species autochthonous from the Western Mediterranean
Previous reports on Artemia franciscana presence in the Western Mediterranean
Hontoria et al. (1987) Ensayo de caracterización de cepasautóctonas de Artemia de Portugal. Seminario sobre Aquacultura. Publicaciones del Instituto de Ciencias Biomédicas. Porto. Portugal. 10 pp.
Narciso (1989) The brine shrimp Artemia sp.: an example of the danger of introduced species in aquaculture. ESA Special Publication, 10: 183-184.
Thiery and Robert (1992) Bisexual populations of the brine shrimp Artemia in Sète-Villeroy and Villeneuve salterns (Languedoc, France). Int. J. Salt Lake Res., 1: 47-63.
Amat et al. (1995) Biogeography of the genus Artemia (Crustacea, Branchiopoda, Anostraca) in Spain. Int. J. Salt Lake Res., 3: 175-190.
Materials and Methods
Artemia sources:• Artemia cyst bank available at the Instituto de Acuicultura de Torre
de la Sal: 130 cyst samples from Spain, Portugal, Italy, France, North Africa (1980-2003).
• Artemia cyst bank at the Artemia Reference Center, University of Ghent (Belgium).
• Artemia cyst samples recovered from waterbird faeces and pellets.
Specific adscription through hydrated cyst biometry.
Specific adscription through adult specimen morphometry (cultured under standard conditions). Multivariate discriminant analysis and morphological data base.
Specific adscription through PCR-RFLP analyses on part of the mitochondrial genome (INCO collaborative research CSIC-AUTH).
AVEIRO DistrictEsmolas salterns 1985 266 (**) ---- 263 A. parthenog. (d)Esmolas salterns 1991 249 ---- ---- A. franciscanaEsmolas salterns 1993 248 ---- ---- A. franciscana
Table 1. Artemia cysts samples available from Portugal, mean diameter of cysts and taxonomical adscription. Micro.: cysts diameter measured with micrometer eyepiece. C.C. cysts diameter measured with Coulter Counter. C.C.(*): idem (Hontoria, 1990).
(**) Vieira (1990). N.H: No hatching cysts. (d) A. parthenogenetica diploid.
Locality Samplingdate
Specific composition
SPAIN Huelva ProvinceEsteros Odiel saltern 06. 2002 A. parthenog. (d): 97 % A. parthenog. (t): 3 %
SPAIN CADIZ ProvinceN.S. del Rocío saltern 01. 2002 A. parthenog. (d): 98 % A. salina (bisex.) 2 %El Estanquillo saltern 01. 2002 A. franciscanaEl Pilar saltern 06. 2002 A. franciscanaSan Pascual saltern 02. 2003 A. franciscanaLa Dolores saltern 02. 2003 A. franciscana
MOROCCOLaguna Mar Chica saltern 06. 2000 A. parthenog. (d): 80 % A. franciscana: 20 %
FRANCESete-Listel saltern 05. 2002 A. franciscanaAigues Mortes saltern 06. 2002 A. parthenog. (d): 2 % A. franciscana: 98 %
Fos saltern 05. 2002 A. franciscana (p. s.)Pesquiers saltern 05. 2002 A. franciscana (p. s.)Hyère saltern 05. 2002 A. franciscana (p. s.)Thau Castelan saltern 05. 2002 A. franciscana (p. s.)
Table 2. Artemia populations obtained in the laboratory from cysts samples collected in SouthwesternSpain, North of Morocco and Southeastern France, and their relative specific composition wheredifferent species or strains did appear. A. parthenog. (d): Artemia parthenogenetica (diploid); (t): tetraploid; (p. s.): alcohol preserved original specimens.
Locality Samplingdate
Cyst diameter
Observations
VENETO ProvinceComacchio salterns 1985 278 N.H.
APULIA ProvinceMargherita di Savoia salterns 1988 258 A. parthenogenetica (d): 67 %
Table 3. Artemia cysts samples available from Italy. Mean diameter of cysts (µm) measured with micrometer eyepiece. Relative specific composition where different species or strains did appear. (d): Artemia parthenogenetica diploid, (t): tetraploid.
N. H. No hatching cysts. ARC 579: Artemia Reference Center cyst bank sample
Table 4. Number of samples (S) of faeces and pellets collected from different waterbirds and localities, and the quantities of Artemiacysts extracted and of viable nauplii hatched.
S with cysts S with only A.
franciscana
S with only A.
parthenog.1
S with both taxa
Total A. franciscana2
Total A.parthenog.1
Castro Marim
Redshank faeces 36 35 1 6 522 (267) 10
Redshank pellets 30 22 3 1 180 (96) 4
Godwit faeces 27 11 6 1 24 (14) 10
Cadiz Bay
Redshank pellets 25 0 14 1 1 (0) 40
Dunlin faeces 8 0 1 0 0 1
Table 5. Number of samples (S) of faeces and pellets collected from different waterbirds and localities that contained cysts of different Artemia taxa, and the numbers of adult Artemia reared after hatching cysts. Note that not all nauplii hatched from cysts survived to the adult stage that permitted species identification, thus the numbers of samples containing each Artemia taxon may be underestimated.
1 the diploid form, all individuals are females.2 all individuals were sexed, the number of males is given in parentheses
Nº of samples
Nº cysts per sample Nº nauplii per sample1 % cysts2
Table 6. Number of Artemia cysts collected from different waterbirds and localities, the numbers of viable nauplii produced and the proportion of cysts producing nauplii.
1 For samples with at least one cyst (see Table 4 for N).2 Mean ± s.e of % of cysts that produced nauplli from those sample with at least one cyst.
y = 59.1 - 6.97 x
1,5 2,0 2,5 3,0 3,5 4,0 4,5 5,0 5,5 6,0 6,5
loge (Nº cysts)
0
10
20
30
40
50
60
70
% c
ysts
hat
chin
g
Attributes of the invasive species specially present in Artemia franciscana
(Ehrlich, 1984; Lodge, 1993; McMahon, 2002)
Ability to function in a wide range of physical conditions. Muchgeneric variability or phenotypic plasticity: euryhaline (tolerance to different ionic brine composition) and eurythermal.
High competitive abilities: short generation time, rapid population growth, high fecundities, small offspring size, balanced ovovivi-parism/oviparism reproduction:
A. franciscana > A. parthenogenetica > A. salina
Associated with human activities: aquaculture and pet market.
High dispersal rate of the cysts: anemochory and zoochory.
Artemia in Argentina
Francisco Amat, Francisco Hontoria, Juan Carlos Navarro, Inmaculada Varó, Olga Ruiz, Germán MedinaInstituto de Acuicultura de Torre de la Sal (CSIC). 12595 Ribera de Cabanes (Castellón). Spain.
R. Graciela Cohen, Julieta GoenagaDepartamento de Biología. Facultad de Ciencias Exactas y Naturales. Universidad de Buenos Aires. Argentina.
Collaborative research ICI-AECI Spain-Argentina, and INCO CSIC-UBA.
Distribution of A. persimilis and A. franciscanapopulations in Argentina
Artemia franciscana is present North of 36º S and A. persimilis is present South of 37º S, and at 49º 18’ S in Puerto San Julián(Santa Cruz Province).
Geographical proximity with Chilean A. persimilis populations in Laguna Amarga (Torres del Paine National Park – 50º 29’ S) and Laguna de los Cisnes (Isla Grande, Tierra del Fuego – 53º 15’ S).
Morphometrics of cultured and preserved adult males and females obtained from original cysts, or collected in situ, allow to venture the presence of A. franciscana in seven localities, and A. persimilisin 25 localities.
FEMALES
First discriminant function
-6 -4 -2 0 2 4 6 8
Sec
ond
disc
rimin
ant f
unct
ion
-3
-2
-1
0
1
2
3A. franciscanaA. persimilis
Callaqueo
Palos Blancos
Piedras
Salina Chica
Colorada Chica Hidalgo
Pedro LuroCisnes
Laguna Amarga
Vidriera
El Inglés
GualichoSan Antonio
Algarrobo
Luzzetti
Winchel
Colorada Grande
Caleta Olivia
San Julián
Rada TillySalitral Negro La Pava
Villalonga
R3 km2035Cerro Pancho
Great Salt Lake
Pichilemu
Atacama
LlamaraLos Vilos
Manaure
San Francisco Bay
Cabo Rojo
AncashPiura
Yapes
Tunas
Mar ChiquitaGuantánamo
Oaxaca
San Quintín
Chimus
Las Cumaraguas
Hípico
Grossos
Coche
Yallahs
Yavaros
Salinas GrandesCelestun
Pampa de la Salina
Santiago del Estero
San Luís Potosí
A. franciscanaA. persimilis
First discriminant function
-8 -6 -4 -2 0 2 4 6
Sec
ond
disc
rimin
ant f
unct
ion
-3
-2
-1
0
1
2
3
MALES
Salinas Grandes
San Antonio
Algarrobo
Luzzetti
Piedras
Villalonga
Winchel
Great Salt LakeHidalgo
Grossos
Oaxaca
Cabo Rojo
San Francisco
Ancash
San Quintín
Chimus
Hípico
Guantánamo
Las Tunas
Coche
Las Cumaraguas
Mar Chiquita
Salina ChicaColorada Chica Callaqueo
Pedro Luro
Cisnes
YallahsYavaros
Celestun
Gualicho
Pampa de la SalinaEl Inglés
VidrieraPalos Blancos
Santiago del Estero
Colorada Grande
Rada Tilly
Salitral Negro
San Julián
Caleta Olivia
La Pava
Cerro Pancho
Laguna Amarga
Ruta 3 Km 2035
San Luís Potosí
Country/Province Population Coulter Counter Mean (in µm)
Standard Deviation
Microscope Mean (in µm)
Standard Deviation
Argentina Cordoba
(A. franciscana) Mar Chiquita 219.7 14.1 219.8 14.6 Salinas Grandes - - 229.4 12.6 Las Tunas 249.9 17.8 241.6 16.0
La Pampa (A. persimilis) Hidalgo 246.1 21.0 244.9 12.9
San Julián - - 237.1 15.6 Cerro Pancho - - El Caburé - - - -Chile
(A. persimilis) L. Amarga(PNTP) 250.9 18.7 - - L. Cisnes (TF) 237.5 21.3 - -*A.persimilis cyst diameter mean (microscope): 240.6µm (6.4); range 228.7-249.5µm. *A.persimilis cyst diameter mean (C.Counter): 237.6µm (7.6); range 227.9-250.9 µm.
Bioecological characterization (life-span) of Argentinean A. persimilis (Hidalgo) vs. A.
franciscana (Mar Chiquita)
Nauplii of both populations were reared to adulthood under different conditions of temperature and salinity.
All combinations of temperature (12, 21 and 28ºC) and salinity (30, 60, 90 and 120 gL-1) were assayed.
21ºC
SALINITY ‰
30 60 90 120
SUR
VIVA
L %
0
20
40
60
80
100
28ºC
SALINITY ‰
30 60 90 120
SUR
VIVA
L %
0
20
40
60
80
100
A. persimilis vs A. franciscana12ºC
SALINITY ‰
30 60 90 120
SUR
VIVA
L %
0
20
40
60
80
100
A. persimilisA. franciscana
FECUNDITYA.persimilis vs A.franciscana
12ºC
SALINITY
30 60 90 120
FEC
UN
DIT
Y
0
20
40
60
80
100
120
140
160
180
A.franciscanaA.persimilis
21ºC
SALINITY
30 60 90 120
FEC
UN
DIT
Y
0
20
40
60
80
100
120
140
160
180
28ºC
SALINITY
30 60 90 120
FEC
UN
DIT
Y
0
20
40
60
80
100
120
140
160
180
GROWTH RATE
21ºC
TIME
4 8 12 16 20 24 28 32 36 40 44 48 52 56 60
LEN
GTH
(mm
)
0
2
4
6
8
10
12
14
A. franciscana 12ºC
TIME
4 8 12 16 20 24 28 32 36 40 44 48 52 56 60
LEN
GTH
(mm
)
0
2
4
6
8
10
12
14 28ºC
TIME
4 8 12 16 20 24 28 32 36 40 44 48 52 56 60
LEN
GTH
(mm
)
0
2
4
6
8
10
12
14
30‰ 60‰ 90‰ 120‰
28ºC
TIME
4 8 12 16 20 24 28 32 36 40 44 48 52 56 60
LEN
GTH
(mm
)
0
2
4
6
8
10
12
14
30‰ 60‰ 90‰ 120‰
21ºC
TIME
4 8 12 16 20 24 28 32 36 40 44 48 52 56 60
LEN
GTH
(mm
)
0
2
4
6
8
10
12
14
A. persimilis12ºC
TIME
4 8 12 16 20 24 28 32 36 40 44 48 52 56 60
LEN
GTH
(mm
)
0
2
4
6
8
10
12
14
28ºC
TIME
4 8 12 16 20 24 28 32 36 40 44 48 52 56 60
% M
ATU
RE
FEM
ALES
0
20
40
60
80
100
21ºC
TIME
4 8 12 16 20 24 28 32 36 40 44 48 52 56 60
% M
ATU
RE
FEM
ALES
0
20
40
60
80
100
SEXUAL MATURITYA. persimilis 12ºC
TIME
4 8 12 16 20 24 28 32 36 40 44 48 52 56 60
% M
ATU
RE
FEM
ALES
0
20
40
60
80
100
SEXUAL MATURITYA. franciscana 12ºC
TIME
4 8 12 16 20 24 28 32 36 40 44 48 52 56 60
% M
ATU
RE
FEM
ALES
0
20
40
60
80
100
21ºC
TIME
4 8 12 16 20 24 28 32 36 40 44 48 52 56 60
% M
ATU
RE
FEM
ALES
0
20
40
60
80
100
28ºC
TIME
4 8 12 16 20 24 28 32 36 40 44 48 52 56 60
% M
ATU
RE
FEM
ALES
0
20
40
60
80
100
30‰60‰90‰120‰
A. persimilis always showed high mortality at low salinity (30 gL-1).
A. franciscana always showed low survival at the low temperature (12ºC), but survived better than A. persimilis at 21º and 28ºC.
A. franciscana always showed higher fecundity rates than A. persimilis.
A. persimilis showed higher growth rates than A. franciscana at all temperatures.
A. persimilis showed sexual maturity earlier than A. franciscana at 12ºC. For the other temperatures the sexual maturity occurred at the same time in both species.
In both species the increase of temperature is associated to an increase of survival and growth rates, and to a decrease of pre-reproductive period and maturity fulfilment.
OVOVIVIPARITY-OVIPARITYvs
Chl a and ORGANIC MATTER
TIME
nov dic ene feb mar abr may jun jul
% O
VOVI
VIPA
RIT
Y-VI
VIPA
RIT
Y
0
20
40
60
80
100
Chl
a ( µ
g/L)
0
50
100
150
200
250
300
0.00
0.01
0.02
0.03
0.04
0.05
OVOVIVIPARITYChl aORGANIC MATTER
OR
GA
NIC
MA
TTER
(mg/
ml)
O2 (mg/l)
T (ºC) S (‰)
mean (sd) max min mean (sd) max min mean (sd) max min
A prospection on the fatty acid composition of the total lipids from the following cyst samples is in progress: MAR CHIQUITA (CO, A. f) PALOS BLANCOS (BA, A. p) LAS TUNAS (CO, A. f) SALITRAL VIDRIERA (BA, A. p) HIDALGO (LP, A. p) EL ALGARROBO (BA, A. p) SALINAS CHICAS (LP, A. p) SALINA PIEDRAS (BA, A. p) CALLAQUEO (LP, A. p) SALINA EL INGLES (BA, A. p) COLORADA CHICA (LP, A. p) SAN ANTONIO (RN, A. p) COLORADA GRANDE (LP, A. p) EL GUALICHO (RN, A. p) SALITRAL NEGRO 2 (LP, A. p) CALETA OLIVIA (SC, A. p) SALITRAL NEGRO 3 (LP, A. p) SAN JULIAN (SC, A. p) Their characterization aims to establish the freshwater or marine type of these cysts according to:
• Taxonomic species: A. franciscana vs A. persimilis • Latitudinal distribution • Endorheic or litoral ecosystems
This is the first attempt to a broad PUFA profile characterization of cysts from different A. persimilis populations. CO: Province of Córdoba; LP: Province of La Pampa; BA: Province of Buenos Aires; RN: Province of Rio Negro; SC: Province of Santa Cruz. A. f : Artemia franciscana A. p : Artemia persimilis
Mutant depigmentedeyes, not sex linked, found in A. persimilis
Brine shrimp in Tunisia. INCO collaborative research CSIC-INAT
Three autochthonous Artemia populations from Tunisia: El Adhibet, Sijoumi and Sahline were characterised morphometrically as Artemia salina (= A. tunisiana).
Cyst and nauplius biometrics, as well as several life-span traits were established comparatively to other Mediterranean Artemia salina populations.
Brine shrimp in India. INCO collaborative research CSIC-IART
Six samples of Artemia cysts were afforded from India.
Puthalam cysts showed the exclusive presence of an autochthonous parthenogenetic strain (diploid or triploid?).
Thamarikulam cysts (suspected parthenogenetic) did not hatch.
Vembar, Veppelodai, Arasaradi and Theraspuran cysts, presumably a mixture of autochthonous parthenogenetic strain andintroduced American brine shrimp, showed the exclusive presence of A. franciscana (outcompeting exotic invasive species).