Selective breeding of RotifersImproved hatchery and grow-out technology for grouper aquaculture in the
Asia-Pacific region (FIS/97/73)
Dr Inneke Dr Inneke RumenganRumengan; Mr ; Mr Stenly WullurStenly WullurSam Sam Ratulangi Ratulangi UniversityUniversity
Dr Richard KnuckeyDr Richard KnuckeyQueensland Department of Primary IndustriesQueensland Department of Primary Industries
Rotifers are traditional aquaculture food for fish Rotifers are traditional aquaculture food for fish larvaelarvae
Grouper larvae and rotifersGrouper larvae and rotifers
Rotifers <100 µm best for 1Rotifers <100 µm best for 1stst feeding larvaefeeding larvaeScreening rotifers is wastefulScreening rotifers is wastefulSelect for rotifer strain with smaller average lengthSelect for rotifer strain with smaller average length
123 ± 8123 ± 8147 ± 11147 ± 11SSSS--typetype
133 ± 10133 ± 10176 ± 16176 ± 16SS--typetype
186 ± 13186 ± 13219 ± 13219 ± 13LL--typetype
Width (µm)Width (µm)Length (µm)Length (µm)RotiferRotifer
First feeding grouper larvae have a mouth gape large enough only for SS-type rotifers
RotifersRotifers
A normal population of A normal population of rotifers has rotifers has reproducing females of reproducing females of a range of sizesa range of sizes
Apply selection pressures to get a population of rotifers Apply selection pressures to get a population of rotifers with smaller average body lengthwith smaller average body length
•• FirstlyFirstly-- Characterise rotifer strainCharacterise rotifer strain•• SecondlySecondly-- Manipulate culture methods to select for Manipulate culture methods to select for
smaller rotifersmaller rotifer
90 100 110 120 130 140 150 160 170 180
35
30
25
20
15
10
5
0
Rotifer ResearchRotifer Research
Characterise rotifer strainCharacterise rotifer strain•• Growth rateGrowth rate
-- Relationship between salinity growth rate and Relationship between salinity growth rate and reproductive sizereproductive size
•• HeritabilityHeritability-- Size of Size of amicticamictic & resting eggs and resulting rotifer& resting eggs and resulting rotifer-- Size of mother and resulting offspringSize of mother and resulting offspring
Manipulation of rotiferManipulation of rotifer•• Effect of diet particle size on rotifer sizeEffect of diet particle size on rotifer size•• Mutation by UV lightMutation by UV light
Clonal stock
30 eggs in each well
• Different salinities : 5, 20, 30 ppt• N. oculata 3 x 106 cells/ml• Observation at 1 hour interval for 24 hours
1 h 2 h 3 h
24 h
B
A
C
Size measurements
A = Lorica lengthB = WidthC = Anterior width
Characterise Characterise -- development ratedevelopment rate
Lower salinity = faster developmentLower salinity = faster development
Characterise Characterise -- development ratedevelopment rate
100
120
140
160
180
0 2 4 6 8 10 12 14 16 18 20 22 24Age post-hatch (h)
Loric
a le
ngth
(µm
)
5 ppt
20 ppt
30 ppt
Reproductive females appear before maximum size Reproductive females appear before maximum size is reachedis reached
•• 5ppt = 6 h; 20 ppt = 12 h; 30 ppt = 20 h5ppt = 6 h; 20 ppt = 12 h; 30 ppt = 20 h
Characterise Characterise -- heritability heritability
Amictic Amictic eggseggs•• Relationship between:Relationship between:
•• rotifer and egg sizerotifer and egg size•• egg size and offspringegg size and offspring
Resting eggsResting eggs•• Relationship between small Relationship between small
resting egg, hatched rotifer resting egg, hatched rotifer and resulting offspringand resulting offspring
Characterise Characterise -- heritabilityheritability
Poor correlation between the size of parent rotifer and Poor correlation between the size of parent rotifer and theirtheir amicticamictic eggegg
50
60
70
80
90
100
90 110 130 150 170 190Lorica length (µm)
egg
size
(µm
)
Rotifer length vs Amictic egg size
Freq
uenc
y
0
4
8
12
16
20
110 120 130 140 150 160 170 180
Lorica length (µm)
Average = 147 µm
Offspring of rotifer hatched from small amictic egg
Offspring from rotifer hatched from small Offspring from rotifer hatched from small amicticamictic egg egg are not smaller than initial populationare not smaller than initial population
Characterise Characterise -- heritabilityheritability
Offspring from small resting egg and small Offspring from small resting egg and small parent are not smaller than initial populationparent are not smaller than initial population
Female parent
Offspring
Avg = 148 µm0
2
4
6
8
10
12
90 100 110 120 130 140 150 160 170
Lorica length (µm)
Freq
uenc
ySmall resting eggs
Hatched population
Select small reproductive female
Characterise Characterise -- SummarySummary
Distribution skewed toward Distribution skewed toward small rotifer because small rotifer because rotifers continue to increase rotifers continue to increase in size after becoming in size after becoming reproductive and not due to reproductive and not due to genetic variation. genetic variation. 90 100 110 120 130 140 150 160 170 180
35
30
25
20
15
10
5
0
Rotifer size is only weakly Rotifer size is only weakly related to parent, related to parent, amictic amictic or or resting egg size.resting egg size.
Clonal stock1st laid eggs
P • Different diets : - Stichococcus sp. (1 pg/cell)
(30 ppt) - N. oculata (10 pg/cell)- Chlorella like (10 pg/cell)- Tetraselmis sp. (170 pg/cell)
• Observe at 12 h interval until all P dead• Life table computation
A
BC
3 days old rotifers
F1 A = Lorica lengthB = WidthC = Anterior width
Manipulation Manipulation -- Diet particle sizeDiet particle size
Manipulation Manipulation -- Diet particle sizeDiet particle size
Size distribution of reproductive females is effected by Size distribution of reproductive females is effected by algal dietalgal dietThe diet with the smallest particle size (The diet with the smallest particle size (StichococcusStichococcus) ) produced a population with the highest percentage of small produced a population with the highest percentage of small (<150 µm) rotifers(<150 µm) rotifers
<150 µm
52%
151-180 µm
48%
Stichococcus sp. (n = 52)
72% 28%
Chlorella like (n = 40)
6%
94%
Tetraselmis sp. (n = 16)
51%
30%19%
Nannochloropsis oculata (n = 57)
>180 µmLorica length
Rotifer stock
Eggs collected
UV treatments : 30 & 60 min
UV lamp
B
A
C
UV treated eggs
Clonal culture
A = Lorica lengthB = WidthC = Anterior width
Manipulation Manipulation –– UV exposureUV exposure
Manipulation Manipulation –– UV exposureUV exposure
Some clonal lines of Some clonal lines of rotifers from UV rotifers from UV treatment have smaller treatment have smaller size than original populationsize than original populationSome of this difference Some of this difference may be due to a reduced may be due to a reduced growth rategrowth rateLongLong--term stability of the term stability of the size trait is not yet size trait is not yet establishedestablished
0
10
20
30
40
50
110 120 130 140 150Lorica length (µm)
160
0
10
20
30
40
50
110 120 130 140 150 160 170
Size distribution of two UV treated clonal lines
ConclusionsConclusions
Two methods have shown potential to Two methods have shown potential to select for smaller rotifers in an SSselect for smaller rotifers in an SS--rotifer populationrotifer population
•• DietDiet-- very small particle size of diet very small particle size of diet produces a rotifer population with more produces a rotifer population with more small individualssmall individuals
•• MutationMutation-- UV exposure produces UV exposure produces population with a smaller average body population with a smaller average body sizesize
