In the 1980's more emphasis on sturgeon aquaculture research. ⢠Established in 1988, first the sterlet was collected from the Hungarian rivers to supply the ...
EX-SITU CONSERVATION OF GENETIC DIVERSITY OF DANUBE STURGEONS IN HUNGARY 8th International Symposium on Sturgeons September 10th - 16th, 2017 - Vienna/Austria
Gyula Kovács, Gyöngyvér Fazekas, Vilmos Józsa and András Rónyai Photo by: Dr. Istvan Lehoczky PhD
National Agriculture Research and Innovation Centre Research Institute for Fisheries and Aquaculture H-5540, Szarvas, Anna liget 8.
BRIEF SITUATION OF DANUBE STURGEONS • IUCN Red List species
• Most of the sturgeons are endangered and threatened with extinction
NATURAL HABITATS • How to save them?
Sensitive habitats (spawning grounds, nursing and wintering habitats)
Anthropogenic effects on sturgeon populations and natural habitats (overfishing, poaching, extensive river regulations, establishment of hydro power plants, water pollution, etc.)
REHABILITATION OF NATURAL HABITATS
• Need to significantly decrease the anthropogenic effects • Rehabilitation of emphasized river sections (spawning grounds, nursing habitats, etc.)
• Solve the river connectivity problems (anadromous migratory species) • Slow process, by the completion of it the critically endangered species could extinct • What could be a prompt solution?
GENE CONSERVATION Key role in the rescue of Danube sturgeon species
Conservation
In vitro
In vivo
cryo-conserved materials
live animals
in situ
ex situ
in the natural habitat and production environment
in an artificial environment (zoo, research institute, etc.)
• • • •
juvenile restocking
• Shorter term -> Ex-situ gene conservation (conservation of genetic diversity) • Longer term -> In-situ gene conservation depends on the rehabilitation of natural habitats
• In vivo ex situ gene bank of NAIK HAKI
semen oocytes embryos somatic cells
ESTABLISHMENT OF EX-SITU STURGEON GENE BANK OF NAIK HAKI • In the 1980’s more emphasis on sturgeon aquaculture research • Established in 1988, first the sterlet was collected from the Hungarian rivers to supply the necessity of sterlet fingerlings in aquaculture research • Later the gene bank expanded with other species: Siberian sturgeon, Russian sturgeon, stellate sturgeon, great sturgeon (beluga) and paddle fish. • Gene conservation allowed the involvement in sterlet restocking programs • The Ministry of Agriculture supports its operation
Species
Origin
Quantity
Total weight (kg)
Average weight (kg)
Status
STOCKS IN THE GENE BANK
Danube, Tisza (Hungary)
198
505
2.6
Broodstock
Russia
19
190
10
Broodstock
USA
14
150
10.7
Broodstock
Danube, Tisza (Hungary)
492
185
0.4
Juvenile/pre adult
Danube (Romania)
3
47
16
Pre-adult
Caspian (Russia)
14
237
17
Pre-adult
Stellate sturgeon (Acipenser stellatus)
Danube (Romania)
4
16
4
Beluga sturgeon
Danube (Romania)
8
160
20
Pre-adult
Wild caught/ own 3rd and 4th generation
(Acipenser Baerii)
Russia
56
295
5.3
Pre-adult
Near to 500 juveniles
Paddlefish (Polyodon spathula)
USA/own
137
726
5.3
Pre-adult
from 4 different year classes
Sterlet (Acipenser
ruthenus)
Siberian Sturgeon
(Acipenser Baerii)
Paddlefish (Polyodon spathula) Sterlet (Acipenser
ruthenus)
Russian sturgeon
(Acipenser gueldenstaedtii)
Russian sturgeon
(Acipenser gueldenstaedtii)
(Huso huso)
Pre-adult
Siberian Sturgeon
The main focus is on the Sterlet Sex ratio: ~ 1 3 ♂ −
2
3
♀
MANAGEMENT OF THE GENE BANK Wild caught fish
Register of broodstock Tagging, fin clipping
Juvenile – pre-adult
Reproduction harvest in early spring selecting breeders
Hatching
Fingerling rearing
Larval rearing in RAS + weaning
BROODSTOCK COLLECTION, REGISTRATION AND HOLDING Classified fish from younger generation
Register of Broodstock
Wild caught fish
Tagging each specimen with PIT-tag
Collecting fin-clip samples for further DNA analysis
Feeding with artificial diet (crude protein 40-45%, crude fat 10-12%) with the feeding ratio 1-2%
Broodstock kept in 4-6 earthen ponds (total area 1.2-1.5 ha)
REPRODUCTION Selection of breeding candidates Choosing breeders in early spring harvest
Verification in the database
Number of breeders: 10 ♀ and 10 ♂ from different age grades
Sex Ratio: 50-50%
Using different breeders each year
Hormonal induction to trigger ovulation
1. Transportation breeders to hatchery Increase water temperature (16°C)
3.
2. Synthetic hormone, analogue of mammalian luteinizing hormone-releasing hormone
Milt collection
6. 4. Collection of eggs + fertilization
5.
Egg de-adhesion
Incubation of the eggs
LARVAL AND FINGERLING REARING Larval rearing in RAS
Fingerling rearing in RAS and pond condition (Floating tank system)
Feeding: natural food in the first 15-20 DPH
Feeding: formulated feed + natural food
Weaning to artificial diet (4 days)
Feeding ratio: 2-5 % of their body mass
Feeding ratio close to ad libitum Water temperature: 16-22 °C
Water temperature: around 20 °C/ depending on the weather Stocking: 20 kg/m3; 100-300 kg/ha
Stocking: 1000-2000 fry/m2
JUVENILE REARING UNTIL SEXUAL MATURITY • Rearing in pond conditions
• Feeding with artificial dry feed (crude protein 40-50% and crude fat 10-12 %)+ natural food produced by the pond
• Feeding ratio 1.5-2 % of the biomass (depending on the water temperature) • Systematic DO and water temperature
measurements • 3-5 sampling during growth season and annual harvests
GENOTYPING BROODSTOCK BY MICROSATELLITE MARKERS Genomic DNA purification from fin-clip sample
PCR amplification of microsatellite fragments
Fragment analysis of PCR products Genotypic data (loci and alleles)
TRACKING BREEDERS IN A DATABASE PIT-tag scanning at seasonal harvests (stock inventory)
Scanners connected to a computer and scanned code appears in our database (MS excel) as an individual • • • • •
Database contains • Information on their location Species (which pond) Year class • Body weight at the actual Place of origin harvest Sex • Data regarding the Genotype reproduction • Observations and notes
More precise broodstock management (growth, propagation plans, etc.)
Development a new software with special regards to ex-situ fish gene conservation that supports broodstock management decisions
UTILIZATION OF GENETIC RESOURCES OF THE GENE BANK BY RESTOCKING ACTIONS The gene bank of NAIK HAKI provided the genetic material for the National Restocking Programme on Sterlet in 2016 financed by the Ministry of Agriculture
18.000 sterlet fingerling was stocked to Danube and Tisza rivers at 6 locations Total weight of the restocked sterlet stock was 7 tons with 300 – 400 g average body weight Periodic monitoring of occurrence of the stocked fish
THANK YOU FOR YOUR ATTENTION!
Questions?
