Survival, male morphotypes, female and male

proportion, female reproductive status and tag loss in

crosses among three populations of freshwater

prawn Macrobrachium rosenbergii (de Man) in India

Bindu R Pillai1, Kanta D Mahapatra1, Raul W Ponzoni2, Lopamudra Sahoo1,†, P L Lalrinsanga1,

Wagdy Mekkawy2,3, Hooi Ling Khaw2, Nguyen H Nguyen2,*, Swagathika Mohanty1,

Sovan Sahu1 & Gunamaya Patra1

1Central Institute of Freshwater Aquaculture, Bhubaneswar, Odisha, India2WorldFish, Bayan Lepas, Penang, Malaysia3Animal Production Department, Faculty of Agriculture, Ain Shams University, Cairo, Egypt

Correspondence: B R Pillai, Principal Scientist, CIFA, Kausalyaganga, Bhubaneswar-751002, Odisha, India. E-mail: pillaibr@

hotmail.com

*Present address: N H Nguyen, School of Science, Education and Engineering, University of the Sunshine Coast, Maroochydore,

QLD 4558, Australia†Present address: L Sahoo, ICAR Research Complex for North East Region, Lembucherra, Tripura 799210, India

Abstract

The present study examined the variation in sur-

vival, proportion of male morphotypes, female and

male proportion, female reproductive status and tag

losses in nine crosses from a complete (3 9 3) dial-

lel mating of three populations of Macrobrachium

rosenbergii in India. The populations originated

from Gujarat (north-west), Kerala (south-west) and

Odisha (east), representing different agro-ecological

regions in India. Progeny from 60 families (4773

juveniles) were individually tagged and reared for

16–17 weeks in earthen ponds. Binary logistic

regression was used to analyse the data. Survival

rate was greatest (0.88) for Kerala 9 Odisha (KO;

the first letter represents the sire), whereas it was

lowest (0.75) for both Kerala 9 Kerala (KK) and

Gujarat 9 Kerala (GK). Significantly greater pro-

portions of large male morphotypes were observed

in KK cross and in combinations involving Kerala

population. The highest proportion of immature

females, as well as of non-berried females, was

observed in the Odisha 9 Odisha (OO) cross. Tag

loss was greatest for KK (0.35) and lowest for GG

(0.19). We found significant variations among

crosses for all studied traits suggesting the possibil-

ity of including them in genetic improvement pro-

grammes of this species.

Keywords: survival, male morphotype, diallel

cross, tag loss, Macrobrachium rosenbergii

Introduction

The giant freshwater prawn Macrobrachium rosen-

bergii (de Man) is an important species cultured

globally, with a production of 215 029 t worth

over a billion US dollars in 2010 (FAO 2012).

Aquaculture production of this species in India,

the third largest producer in 2005 with a produc-

tion of 42 800 t, started declining in 2006 and by

2010 it reduced drastically to 13 525 t (FAO

2012). The genetic deterioration of brood stock,

possibly due to unintentional inbreeding and nega-

tive selection was perceived as a major contribut-

ing factor to the decreased productivity. Similar

decline in productivity has been reported from

countries such as China, Taiwan and Thailand (De

Bruyn, Wilson & Mather 2004; New, Valenty,

Tidwell, D’Abramo & Kutty 2010).

Genetic improvement of cultured stocks through

selective breeding is an efficient way of increasing

productivity in farmed terrestrial animals and

plants, as well as in aquatic animal species (for

recent review see, Gjedrem, Robinson & Rye

2012). Favourable response to selective breeding

has been observed in a number of commercially

© 2014 John Wiley & Sons Ltd 1

Aquaculture Research, 2014, 1–12 doi:10.1111/are.12419

important finfish species such as salmon (Gjedrem

1997; Quinton, Mc Millan & Glebe 2005), tilapia

(Eknath, Bentsen, Ponzoni, Rye, Nguyen & Thode-

sen 2007; Ponzoni, Hamzah, Tan & Kamaruzz-

aman 2005) and rohu carp (Mahapatra, Jana,

Saha, Gjerde & Sarangi 2006). Selective breeding

programmes so far undertaken in commercially

important crustaceans including penaeids (Hetzel,

Crocos, Davis, Moore & Preston 2000; Argue,

Arce, Lotz & Moss 2002; Goyard, Patrois, Peignon,

Vanaa, Dufour, Viallon & Bedier 2002; Preston,

Crocos, Keys, Coman & Koeing 2004; de Donato,

Manrique, Ramirez, Mayer & Howell 2005; Gitte-

rle, Rye, Salte, Cock, Johansen, Lozano, Su�arez &

Gjerde 2005) and crayfish (Jones, Mc Phee & Rus-

coe 2000; Jerry, Purvis, Piper & Dennis 2005)

have also resulted in genetic gains in productivity.

Most aquaculture genetic improvement pro-

grammes have focused on growth traits. However,

there are other potentially important traits that

need to be considered as candidates for inclusion

in the breeding objective. Investigating the magni-

tude of genetic variation in such traits is impor-

tant in order to assess the feasibility of including

them in genetic improvement programmes. The

present investigation was therefore undertaken to

study the variation in important traits like sur-

vival, proportion of male morphotypes, female and

male proportion, female reproductive status as well

as tag loss among the nine crosses of three geo-

graphically distinct populations of M. rosenbergii

from India.

Material and methods

Sampling from different populations

A selective breeding programme aimed at increas-

ing the growth rate of giant freshwater prawn

(GFP) was initiated in 2007 in India, led by the

Central Institute of Freshwater Aquaculture (CIFA)

in collaboration with WorldFish, Malaysia. For the

establishment of the base population, different pop-

ulations of M. rosenbergii were sampled from three

locations in India, viz. Gujarat (north-west), Kerala

(south-west) and Odisha (East). These represent

different agro-ecological regions and are geograph-

ically distant from each other (Figure 1). Juvenile

Figure 1 Map of India showing

the collection sites for the popula-

tions of Macrobrachium rosenbergii

Gujarat, Kerala, Odisha.

© 2014 John Wiley & Sons Ltd, Aquaculture Research, 1–122

Selected traits in nine crosses of M. rosenbergii B R Pillai et al. Aquaculture Research, 2014, 1–12

prawns were collected from Narmada River, near

Bharuch, Gujarat (21º 420 N 72º 580 E) (2100

animals, average body weight (ABW): 0.30 g

(0.11–0.68 g), from the private hatcheries in Ker-

ala (9º 290 N 76º 190 E) [2000, ABW: 0.01 g

(0.008–0.02 g) and 200, ABW: 0.24 g (0.10–

0.55 g)], and from Brahmani River near Rajkani-

ka, Kendrapara, Odisha (20º 440 N 86º 420 E)

(900, ABW: 6.0 g (1.2–12.0 g). The juveniles

were transported to the institute farm and released

into indoor rearing facility separately. After

4 weeks, the three populations were tagged with

different coloured elastomer tags. Visible implant

elastomer tags (VIE) from Northwest Marine Tech-

nology, (NMT), USA, were used: green (Gujarat),

orange (Odisha) and red (Kerala). The tagged

prawns were then released into separate 0.04 ha

earthen ponds for grow-out. Prawns were fed com-

mercial freshwater prawn feed (Godrej Macro Gold,

30% crude protein, 5% lipid) twice daily at 10% of

biomass.

Family production and rearing

The experimental design consisted of a complete

3 9 3 diallel cross comprising three pure-bred mat-

ings (Kerala 9 Kerala (KK); Gujarat 9 Gujarat

(GG); Odisha 9 Odisha (OO)), and six crosses (Guja-

rat 9 Kerala (GK), the first letter representing the

sire; Gujarat 9 Odisha (GO); Kerala 9 Gujarat

(KG); Kerala 9 Odisha (KO); Odisha 9 Gujarat

(OG); Odisha 9 Kerala (OK)). The mating of the

brood stock was carried out in two batches. Table 1

shows the number of mating pairs for each popula-

tion and their combination. A total of 2233 juvenile

prawns (progeny of 27 sires and 30 dams) repre-

senting the three purebred and six crosses of the

three populations of GFP were tagged in the first

batch and another 2540 prawn juveniles (progeny

of 13 sires and 30 dams) representing six crosses

were tagged in the second batch.

For the production of healthy families, mature

female and male parents were carefully chosen to

maximize mating success. One male was stocked

in a mating tank (Ferrocement tanks

3 9 1.2 9 1.0 m) with four mature females (sex

ratio 1:4). Females carrying fertilized eggs (berried

females) were collected from the mating tank

within 24 h of spawning and maintained in sepa-

rate 500-L fibre glass tanks for 14–16 days until

the eggs had changed colour from bright orange

to grey. Grey-berried females were transferred to

the hatchery for egg hatching in 500-L circular

conical bottom fibreglass tanks. Larvae from indi-

vidual families from each population combination

were reared at a stocking density of 50/litre in

brackish water of 12 gL�1 salinity in the same

hatching containers. Clear water larval rearing

technique was employed that required water

exchange of 50% every second day. Larvae were

fed newly hatched brine shrimp nauplii for the first

10 days and a combination of brine shrimp nau-

plii and egg custard (egg, milk powder, mussel

flesh and cod liver oil and vitamin mineral mix).

One thousand postlarvae (PLs) were collected from

each family when approximately 90% of the lar-

vae metamorphosed to PL. They were reared at a

stocking density of 1 PL per litre for the first

20 days in the hatchery. Subsequently, PLs were

transferred to fine mesh hapas (2.5 9 191 m)

fixed in a 0.10 ha earthen pond for further

3 months rearing at a stocking density of 100/m2.

The PL were fed a 30% protein commercial prawn

pellet (Godrej Macro Gold) twice daily at 20% of

biomass.

Animal identification

Two tagging techniques were employed to identify

the animals either at the individual or at the

group level. Visible implant alpha numeric (VIA)

tags of standard size format (1.0 9 2.5 mm) from

Table 1 Numberofsires(s)anddams

(d)thatproducedprogenyforeachpop-

ulationcombinationinbatches1and2

(s1 9 d1)(s2 9 d2)a

Dams

Sires

Gujarat Kerala Odisha

Batch1

(s1 3 d1)

Batch2

(s2 3 d2)

Batch1

(s1 3 d1)

Batch2

(s2 3 d2)

Batch1

(s1 3 d1)

Batch2

(s2 3 d2)

Gujarat (5 9 5) (3 9 5) (3 9 3) (2 9 5) (2 9 2) (2 9 5)

Kerala (1 9 1) (2 9 5) (4 9 4) (2 9 5) (3 9 3) (0)*

Odisha (1 9 1) (2 9 5) (3 9 5) (0)* (5 9 6) (0)*

*No progeny produced.

© 2014 John Wiley & Sons Ltd, Aquaculture Research, 1–12 3

Aquaculture Research, 2014, 1–12 Selected traits in nine crosses of M. rosenbergii B R Pillai et al.

NMT, USA, with alpha numeric codes were used

for individual prawn identification. The VIE tags

mentioned earlier were used as means of identify-

ing prawns at a group level (nine codes, one for

each of the nine crosses using three colours and

three body locations). Purebreds were tagged on

both right and left sides with one colour whereas

crosses were tagged with one colour on one side

only. The VIA tags were inserted on the lateral

side of the second abdominal segment, whereas

the VIE tag was introduced into the ventral side of

the sixth abdominal segment. All prawns were

tagged with both types of tags.

Communal grow-out of tagged juveniles

The communal grow-out of tagged prawns was

carried out in earthen ponds of 0.04 ha

(400 m2) provided with continuous aeration

using an air blower. Two replicate ponds were

used for both batch 1 and batch 2. Stocking den-

sity was 2.8 and 3.2 individuals per square metre

of surface water area in batches 1 and 2 respec-

tively. About 130–140 days after hatching, the

prawns were tagged and stocked in the ponds.

Mean body weight at stocking for batch 1 was

1.3 g (0.46–6.3 g) whereas it was 2.1 g (0.30–

10.9 g) for batch 2. Prawns were fed twice daily

with a 30% protein commercial pellet (Godrej

Macrogold) at a feeding rate of 10% body weight.

Grow-out continued for 16–17 weeks for both

the batches. Water quality parameters (i.e. pH,

dissolved oxygen and total alkalinity) were mea-

sured weekly (once in the morning) whereas tem-

perature was measured twice daily. The ranges

for the measured water quality parameters during

the grow-out period were as follows: temperature

21.9 to 27.5°C; pH 7.5 to 8.4; dissolved oxygen

2.7 to 4.6 mg L�1; total alkalinity 85 to

121 mg L�1 as CaCO3.

Harvest and data recording

At harvest, surviving prawns were collected and

segregated sex-wise. The sex, body length and

bodyweight of each prawn were recorded. Only

those prawns that had a readable VIE tag could be

included in the analyses because otherwise the

effect of cross (the study of which was one of our

main aims) could not be fitted. The male prawns

were classified according to their morphological

appearance into Blue Claw (BC), Orange Claw (OC)

and Small (SM) as described by Kuris, Ra’anan,

Sagi and Cohen (1987). Large males without claws

were classified as no claw (NC) (Hulata, Karplus,

Wohlfarth & Halevy 1990). For the statistical

analysis BC, OC and NC were combined into a

large male category (LM) for comparison with SM.

Females were classified into four female maturity

stages: stage I (Immature, IM), stage II (early

maturity, EM), stage III (maturity, MA) and stage

IV (fully mature, FM) as per Rajyalakshmi (1961).

Ovaries in immature females show no visible sign

of ovarian maturation; the ovary is small and is

confined to the posterior region of the carapace. In

EM females, ovaries can be seen as a small orange

mass on the posterior region of the carapace. In

MA females, ovaries are larger and more intensely

coloured, occupying two-thirds of the carapace

cavity. In FM females, the ovary occupies the entire

carapace cavity. The number of females in EM and

MA categories was small. The reproductively active

females were combined with FM females and cate-

gorized as mature females. Females were further

classified into two categories: those carrying eggs

as berried (OB) females, and those not carrying

eggs as non-berried (NB) females. In this paper, the

maturity and berried status is generically referred

to as ‘female reproductive traits’. For the tag loss

analysis, two groups of surviving prawns at har-

vest were defined: a group that retained readable

VIA tags on their abdomen and another group that

either had lost the VIA tag or had an unreadable

VIA tag. The ‘tag loss’ trait referred to in this paper

is a combination of tag losses and unreadable tags.

Table 2 shows the total number of observations for

different traits and the proportion of the two cate-

gories defined for each trait.

Statistical analysis

A binary logistic regression model (LOGISTIC

REGRESSION procedure, SPSS (Statistical Package

for the Social Sciences) 2011) was fitted to the

data on survival to harvest, male morphotype,

female and male proportion, female reproductive

traits and tag loss.

The model fitted to survival, male morphotype,

female and male proportion, and female reproduc-

tive traits was:

logpijk

1� pijk

� �¼ lþ Ci þ BPj þ eijk

© 2014 John Wiley & Sons Ltd, Aquaculture Research, 1–124

Selected traits in nine crosses of M. rosenbergii B R Pillai et al. Aquaculture Research, 2014, 1–12

where, depending on the trait being analysed, p is

the probability of survival of large male type, of

female, of immature females or of non-berried

females, l is the overall mean, Ci is the fixed effect

of cross, BPj is the fixed batch by pond effect (four

levels) and eijk is the random residual error term.

In a preliminary analysis of male morphotype,

we tried to fit a multinomial logistic regression (for

more than two categories) but the analysis did not

converge. Some cells of the cross by batch by pond

table had very few or no observations for some of

the morphotype categories which might have been

the reason for lack of convergence.

The model fitted to tag loss was:

logpijkl

1� pijkl

� �¼ lþ Ci þ BPj þ Sk þ bðwÞi þ eijkl

where p is the probability of tag loss, l is the over-

all mean, Ci is the fixed effect of cross, BPj is the

fixed batch by pond effect, Sk is the fixed effect of

sex (two levels); Wl (harvest weight) was fitted as a

covariate, b is the regression coefficient of the logit

on Wl, and eijkl is the random residual error term.

All possible interactions between fixed effects

were initially tested in each model, and found to

be non-significant. The Bonferroni procedure was

used to test the differences among the levels of the

significant fixed effects.

The crosses were ranked based on the least squares

means for each trait with the addition of harvest

weight as reported by Pillai, Mahapatra, Ponzoni, Sa-

hoo, Lalrinsanga, Nguyen, Mohanty, Sahu, Vijayku-

mar, Khaw, Patra, Patnaik and Rath (2011). The

rank correlation among the traits was estimated as a

means of gaining some insight into the joint varia-

tion among the traits. We preferred rank correlation

over product moment correlation because it is a non-

parametric method that makes no assumptions

about the distribution of the variables among which

the correlation is being calculated. Note, however,

that the rank and product moment correlation val-

ues were in our case very similar.

Results and discussion

Estimates of fixed effects and covariates

Tables 3 depicts the significance of the fixed effects

and covariate fitted in the model and Table 4

shows the least squares means for cross, batch by

pond and sex. The effect of cross was highly signifi-

cant (P < 0.001) for all traits. The batch by pond

effect also was highly significant for all traits except

for the female berried status (P > 0.05). Regarding

VIA tag loss, the effect of sex was not significant

(P = 0.63). At harvest, 58% of the surviving

prawns of batch 1 retained readable VIA tags

whereas 79% did so for batch 2. Table 5 shows the

rank correlations among the least squares means

of the nine crosses for the different traits.

Survival

Survival rates for the nine crosses during the

16–17 weeks grow-out phase ranged from 75.8%

Table 2 Total number of observations, proportions and

codes of the categories defined for each trait

Trait Total Proportion (category = code)

Survival 4773 0.89* (alive = 1)

0.84† (alive = 1)

0.11* (dead = 0)

0.16† (dead = 0)

Male

morphotype

1352 0.51 (large = 1) 0.49 (small = 0)

Female and

male

proportions

3819 0.65 (female = 1) 0.35 (male = 0)

Female sexual

maturity

2437 0.82

(immature = 1)

0.18 (mature = 0)

Berried status 2437 0.86

(non-berried = 1)

0.14 (berried = 0)

VIA tag loss 4000 0.70 (with = 1) 0.30 (without = 0)

*All individuals.

†Individuals that lost VIE tag included in ‘dead’ category and

excluded in the calculations for all traits except survival.

Table 3 Fixed effects used in the analysis with the level

of significance for the studied traits

Trait Effect d.f. Wald v2 P > v2

Survival Cross 8 78.03 0.001

Batch by pond 3 33.94 0.001

Male morphotype Cross 8 124.88 0.001

Batch by pond 3 13.79 0.001

Female and male

proportions

Cross 8 128.06 0.001

Batch by pond 3 110.81 0.001

Female sexual

maturity

Cross 8 138.57 0.001

Batch by pond 3 23.12 0.001

Berried status Cross 8 327.51 0.001

Batch by pond 3 2.27 0.52

VIA tag loss Cross 8 45.85 0.001

Batch by pond 3 165.27 0.001

Sex 1 0.23 0.63

Harvest weight 1 108.70 0.001

© 2014 John Wiley & Sons Ltd, Aquaculture Research, 1–12 5

Aquaculture Research, 2014, 1–12 Selected traits in nine crosses of M. rosenbergii B R Pillai et al.

Table

4Least

squaresmeans�

SE*(inbrackets,totalnumber

ofobservationsinvolved

inthecalculationofeach

least

squaresmeans)

forthefixed

effectsfitted

intheanalysisof

survival,proportionsoflargemales,females,im

mature

females,non-berried

femalesandVIA

tagloss

Effect

Survivalrate

Proportionof

largemales

Proportionof

females

Proportionof

immature

females

Proportionof

non-berriedfemales

Proportionof

VIA

tagslost

Cross

GG

0.84�

0.02bc(618)

0.28�

0.04a(123)

0.79�

0.02a(506)

0.93�

0.01d(379)

0.95�

0.01cd(379)

0.19�

0.02a(530)

GK

0.75�

0.02a(678)

0.53�

0.04bc(197)

0.66�

0.02b(511)

0.84�

0.02bc(312)

0.86�

0.02b(312)

0.25�

0.02ab(535)

GO

0.77�

0.02ab(595)

0.29�

0.04a(176)

0.68�

0.02b(462)

0.80�

0.02b(280)

0.93�

0.02cd(280)

0.31�

0.02bc(484)

KG

0.87�

0.02c(563)

0.48�

0.03b(283)

0.50�

0.03c(490)

0.85�

0.02bc(205)

0.91�

0.02bc(205)

0.32�

0.02bc(513)

KK

0.75�

0.02a(847)

0.71�

0.03c(271)

0.61�

0.02b(631)

0.68�

0.03a(352)

0.51�

0.03a(352)

0.35�

0.02c(660)

KO

0.88�

0.02c(221)

0.72�

0.06c(58)

0.61�

0.04bc(196)

0.72�

0.04ab(136)

0.94�

0.02bcd(136)

0.34�

0.03bc(206)

OG

0.87�

0.02c(416)

0.28�

0.05a(98)

0.79�

0.02a(365)

0.92�

0.02

cd(265)

0.94�

0.01cd(265)

0.31�

0.03bc(382)

OK

0.87�

0.02c(412)

0.60�

0.06bc(70)

0.71�

0.03ab(344)

0.67�

0.03a(271)

0.87�

0.02bc(271)

0.33�

0.03bc(361)

OO

0.80�

0.02abc(423)

0.43�

0.06ab(77)

0.66�

0.03b(314)

0.97�

0.01d(237)

0.99�

0.01cd(237)

0.25�

0.02ab(329)

B9

P†

19

1‡

0.80�

0.01a(1100)

0.39�

0.04a(178)

0.79�

0.01a(852)

0.87�

0.01a(665)

0.93�

0.01(665)

0.45�

0.02a(892)

19

20.79�

0.01a(1133)

0.45�

0.04ab(225)

0.74�

0.02a(863)

0.89�

0.01a(629)

0.91�

0.01(629)

0.43�

0.02a(903)

29

30.85�

0.01b(1269)

0.50�

0.03ab(456)

0.56�

0.02b(1019)

0.85�

0.02ab(557)

0.91�

0.01(557)

0.19�

0.01b(1068)

29

40.87�

0.01b(1271)

0.57�

0.03b(494)

0.56�

0.02b(1085)

0.78�

0.02b(586)

0.90�

0.01(586)

0.17�

0.01b(1137)

Sex

Male

0.28�

0.01(1418)

Female

0.29�

0.01(2582)

*Least

squaresmeanswithacommonsuperscriptorwithnosuperscriptdonotdiffersignificantly(P

>0.05)from

each

other.

†B9

P:Batchbypond.

‡Batch1bypond1,andso

on.

© 2014 John Wiley & Sons Ltd, Aquaculture Research, 1–126

Selected traits in nine crosses of M. rosenbergii B R Pillai et al. Aquaculture Research, 2014, 1–12

to 88.5%. These values are consistent with those

reported for this species (New et al. 2010). In our

data, because we only recorded as surviving those

prawns that had a VIE tag, the number of missing

VIE tags is confounded with the number of dead

prawns within cross. Accordingly, our estimate of

survival is underestimated. Note, however, that

losses of VIE tags were relatively small (6.2% in

batch 1 and 3.7% in batch 2). We have no evi-

dence of differential VIE tag losses among crosses

so the differences between crosses in survival is

most likely unbiased. Among the three popula-

tions, Kerala population showed the lowest sur-

vival whereas Gujarat population showed the

highest survival. The KK, GK, GO and OO had sig-

nificantly lower survival compared to other

crosses. After a grow-out period of 8 months in

earthen ponds, Aflalo, Raju, Bommi, Verghese,

Samraj, Hulata, Ovadia and Sagi (2012) reported

survival rates of approximately 20% to 60%

among nine different crosses of three populations

of M. rosenbergii from India indicating among

cross variation for this trait. By contrast, in Viet-

nam, Thanh, Ponzoni, Nguyen, Vu, Barnes and

Mather (2009) did not find significant differences

in survival among nine crosses of three popula-

tions of M. rosenbergii during a 15-week grow-out

period in hapas. They report survival rates rang-

ing from 73.3 to 77.4%. Survival rates in our

study were similar to those of Thanh et al.

(2009). Note that the latter authors recorded sur-

vival rates in a grow-out period of approximately

the same duration as ours. However, unlike

Thanh et al. (2009) we found significant among

cross variation in survival rate. The rank correla-

tions between survival rate of the different crosses

and the other traits were low and statistically not

significant (Table 5).

Proportion of male morphotypes

Adult M. rosenbergii populations from culture

ponds exhibit a wide range of sizes, especially

among males. This variability poses a challenge to

increased profitability because prawn prices are

size dependent. Karplus and Sagi (2010) observed

that in male GFP populations live weights are usu-

ally bi-modal or multi-modal and positively

skewed, comprising a large fraction of small,

unmarketable individuals and a smaller fraction of

large prawns. Earlier workers found that prawn

size variation is the consequence of complex intra-

population interactions that result in a distribution

comprising three distinct male classes viz. SM, OC

and BC. These classes differ in their morphology,

physiology, behaviour and transform from SM to

OC and then to BC (Karplus & Sagi 2010). The BC

males are characterized by very long, robust sec-

ond walking legs that are deep blue, and are the

largest of the three morphotypes. They are aggres-

sive, territorial and dominant over the other two

morphotypes (Karplus, Malecha & Sagi 2000). A

second type of male, SM, with small claws, was

first described in pond populations (Fujimura &

Okamoto 1972). The SM are the smallest of the

three morphotypes and are slow growing and sub-

ordinate to both BC and OC. Sandifer and Smith

(1977) described a faster growing third male mor-

photype, OC, that also grows to a large size and

has long, second walking legs and orange claws.

A greater proportion of BC and OC morphotypes is

preferred under commercial grow-out because the

yield and the market value are greater. Our study

revealed a significant difference in the proportion

of male morphotypes among the crosses. The Ker-

ala population showed the greatest proportion of

large male morphotypes, whereas it was lowest in

Table 5 Rank correlations (probability of a greater value) among least squares means of the different crosses for the

different traits, including harvest weight as reported by Pillai et al. (2011)

Trait Survival

Proportion

of LM

Proportion

of females

Proportion

of IM females

Proportion

of NB females

Proportion of

VIA tags lost

Harvest weight �0.08 (0.84) 0.97 (0.0001) �0.68 (0.05) �0.75 (0.02) �0.63 (0.07) 0.74 (0.02)

Survival 0.07 (0.86) 0.03 (0.93) �0.03 (0.95) 0.40 (0.28) 0.26 (0.50)

Proportion of LM �0.68 (0.04) �0.74 (0.02) �0.55 (0.12) 0.75 (0.02)

Proportion of females 0.25 (0.51) 0.36 (0.33) �0.53 (0.14)

Proportion of IM females 0.74 (0.02) �0.80 (0.01)

Proportion of NB females �0.54 (0.14)

LM, Large males; IM females, Immature females; NB females, Non-berried females.

© 2014 John Wiley & Sons Ltd, Aquaculture Research, 1–12 7

Aquaculture Research, 2014, 1–12 Selected traits in nine crosses of M. rosenbergii B R Pillai et al.

the Gujarat population and intermediate in the

Odisha population. Among the crosses, KO and OK

had a significantly greater proportion of large male

morphotypes, whereas it was significantly lower in

OG and GO. Greater proportions of large male

morphotypes in KK, KO and OK would result in

greater production compared to the other crosses.

Among OO, OG, GG and GO the frequency of the

SM morphotype was significantly greater (>60%)

indicating poor yield from these crosses.

Earlier studies of the distribution of male mor-

photypes during grow-out culture of M. rosenbergii

indicate a density-dependent trend, with a larger

proportion of SM morphotype (and lower propor-

tion of BC and OC) at greater stocking densities

(D’Abramo, Heinen, Robinette & Collins 1989;

Karplus, Hulata, Wohlfarth & Halevy 1986; Ti-

dwell, Webster, Goodgame-Tiu & D’Abramo 1994;

Tidwell, Coyle & Schulmeister 1998). Karplus et al.

(1986) suggest that the changes in morphotype

frequencies with density reflect increased stunting

of prawn populations with increasing densities. In

our study, prawns from different crosses exhibited

differences even when communally reared in a

pond. Hence, the observed differences in the fre-

quency of male morphotypes among crosses could

not be due to density, which suggests that there

may be genetic variation in the potential for

expression of the various morphotypes. Aflalo et al.

(2012) examined the male morphotype distribu-

tion in different populations of M. rosenbergii in a

diallel cross experiment involving three popula-

tions. They reported differences in the frequency of

male morphotypes among different crosses, with a

greater proportion of large morphotypes (BC and

OC) in a pure-bred population from West Bengal,

as well as in its cross with Kerala population.

The rank correlations between the cross least

squares means of proportion of large males and

other traits showed a significant (positive) associa-

tion with harvest weight and VIA tag losses. By

contrast, there was a negative association with the

proportion of females relative to males, and of

immature and non-berried females.

Female and male proportions

The proportion of females was greatest in the Guj-

arat population and lowest in the Kerala popula-

tion. Overall, there were a greater proportion of

females in all crosses except in KG. The GG and

OG had significantly more females (79%) than the

other crosses. Earlier studies report greater propor-

tions of female M. rosenbergii in cultured popula-

tions from different geographical areas (Willis &

Berrigan 1977; Smith, Sandifer & Smith 1978;

Sandifer, Smith, Stokes & Jenkins 1982; D’Abra-

mo, Robinette, Heinen, Ra’anan & Cohen 1986;

Karplus et al. 1986; Jose, Joseph, Shyama, Mohan

& Nair 2007). In a diallel cross experiment involv-

ing three populations of M. rosenbergii in Vietnam,

Thanh et al. (2009) also report sex ratios that

were significantly different from 1:1, in favour of

females in all populations. Malecha, Nevin, Ha,

Barck, La Madrid-Rose, Masuno and Hedgecock

(1992) found a greater proportion of females in

the progeny of five crosses between normal males

(not sex reversed) and females. So far, there have

been no reports on the effect of density on female

to male ratio. Several authors attribute the preva-

lence of females in prawn populations to a greater

male mortality. New et al. (2010) notes that

prawns are vulnerable to limb damage and canni-

balism during moulting even under conditions of

excess food. Hence, the selective male mortality

hypothesis seems plausible under crowded pond

culture conditions. The greater proportion of

females in the present study could partly be attrib-

uted to greater male mortality. It would be inter-

esting to investigate as to what phase of the

development of males and females the sex imbal-

ance sets in (e.g. zygote, larvae, juvenile).

The most obvious feature emerging from the

rank correlations among the least squares means

for the crosses involving male and female propor-

tions was that a greater proportion of females were

associated with lower harvest weight (Table 5).

Female reproductive status

There were marked variations in the proportions

of immature females among the different crosses.

The pure-bred Odisha population had the highest

proportion of immature females (97%), followed by

the Gujarat population (93%). By contrast, the

pure-bred Kerala population had about 32% of

females having ovaries with developing oocytes.

Among crosses, OG, KG, GK and GO had signifi-

cantly greater proportions of immature females

whereas it was lower in OK and KO. The propor-

tion of non-berried females exhibited a similar

trend as that of immature females.

Generally, female growth in decapod crustaceans

slows down after they reach maturity because part

© 2014 John Wiley & Sons Ltd, Aquaculture Research, 1–128

Selected traits in nine crosses of M. rosenbergii B R Pillai et al. Aquaculture Research, 2014, 1–12

of the energy intake is diverted to egg production

and incubation (Hartnoll 1982). Freshwater

prawns can reproduce either continuously or peri-

odically, depending on their geographical location

(New et al. 2010). In tropical regions where the

water temperature remains above 24°C for most of

the year, reproduction occurs continuously (Ling

1969). By contrast, in monsoon regions, reproduc-

tion is seasonal (New et al. 2010). Ra’anan, Sagi,

Wax, Karplus, Hulata and Kuris (1991) report that

the growth rate of immature female M. rosenbergii

is relatively high, initially exponential, but after

sexual maturation, this rate reduces considerably

as the reproductive processes require significant

energy mobilization. Aflalo et al. (2012) reported

differences in reproductive performance among

crosses of M. rosenbergii in a diallel experiment

involving three populations from India. There was

a greater proportion of females in progressive repro-

ductive stages (bearing developed gonads or ber-

ried) in the pure-bred West Bengal population and

its crosses when it was used as a male parent. This

indicates the existence of population specific differ-

ences in reproductive activity of female M. rosen-

bergii. Populations showing fast rate of maturation

are not preferred for culture as this would retard

the growth rate and the presence of greater number

of egg carrying females further reduces the price.

The observed differences in proportion of reproduc-

tively active females among different populations of

M. rosenbergii may affect the yield and returns from

farming. It may however be noted that the rank

correlations between least squares means of the

crosses for harvest weight with the female repro-

ductive traits (proportion of immature and of non-

berried females) were negative (Table 5). This

indicates that across populations, heavier weight

was associated with a lower proportion of imma-

ture and non-berried females, although female sex-

ual maturation itself may slow down growth rate.

VIA tag loss

Two types of tags were used in the present study;

VIE as a batch tag to identify the crosses and VIA

tags for individual prawn identification. The losses

among the former were considerably lower than

the latter. Furthermore, all the retained VIE tags

were readable. Earlier studies on decapod crusta-

ceans including freshwater prawn on retention

and readability of VIE tags (Woods & James 2003;

Ashley & Jean-Paul 2006; Pillai, Rath & Sahu

2007) compare well with the results from our

study. By contrast, studies on retention and read-

ability of VIA tags report higher retention and

readability than what we experienced. Isely and

Stockett (2001) report 100% readability of VIA

tags in juvenile red swamp crayfish Procabarus

clarkii 150 days posttagging. Jerry, Stewart, Purvis

and Piper (2001) reported 96% readability in juve-

nile yabby (Cherax destructor). The lower readabil-

ity of VIA tag observed in our study could partly

be attributed to the pigmented and thicker shell

(exoskeleton) of the adult M. rosenbergii. Melaniza-

tion of tissues surrounding the tag could also have

resulted in decreased readability of VIA tags.

Under laboratory conditions, Pillai, Sahoo, Ma-

hapatra, Ponzoni, Sahu, Mohanty, Vijaykumar

and Sahu (2009) did not find any significant effect

of VIA tags on growth and survival of M. rosen-

bergii. However, our results indicated that higher

body weight was associated with greater losses of

VIA tags. This finding is important because it may

affect the family based selective breeding pro-

gramme for improved harvest weight of M. rosen-

bergii that requires individual prawn identification.

Greater tag losses among the heaviest individuals

may lead to lower selection intensity and reduced

selection response.

Note that the differences between crosses were

non-significant (results are not presented) when

the male morphotype was fitted as a fixed effect in

the statistical model used to analyse tag loss. The

effect of harvest weight as a covariate remained

significant but with lower odds ratio (2%) com-

pared with the analysis that did not include male

morphotype (3%). The BC morphotype had the

greatest tag loss (0.61) compared to the other mor-

photypes which ranged from 0.36 to 0.41. We also

found a significant difference in tag loss between

berried (0.31) and non-berried females (0.25). The

differences between crosses remained significant

when female maturity stage was included in the

model as a fixed effect. Fitting harvest weight as a

covariate did not affect the significance of the other

fixed effects. Table 5 shows that tag losses were

greater in the heavier crosses, whereas they were

lower among those with a greater proportion of

females, especially immature ones.

Conclusions

Evidence of variation among populations and

among crosses in survival rate, proportion of male

© 2014 John Wiley & Sons Ltd, Aquaculture Research, 1–12 9

Aquaculture Research, 2014, 1–12 Selected traits in nine crosses of M. rosenbergii B R Pillai et al.

morphotypes, female and male proportions and

state of reproductive maturity in females were

observed. These are biological attributes of impor-

tance in the production system. The presence of

variation among populations and crosses in

prawns bred and reared in a common environ-

ment suggests that the observed variation could

have a genetic basis. This variation could be

exploited in a synthetic population by including all

populations and crosses and selection practised in

ushering in genetic improvement. Greater VIA tag

losses among heavier animals (and among the lar-

ger male morphotypes in particular) are a concern

since they are bound to affect selection intensity

and response. Freshwater prawn has unique fea-

tures which are absent in other species. The pres-

ent study indicates two avenues that appear as

potentially useful to achieving large increases in

productivity: (1) an increase in the proportion of

males and (2) an increase in the frequency of the

larger male morphotypes.

Acknowledgments

This study was carried out at the Central Insti-

tute of Freshwater Aquaculture, Bhubaneswar

under a bilateral collaborative research project

between Indian Council of Agricultural Research

(ICAR), New Delhi, India and WorldFish, Malay-

sia. The authors express sincere gratitude to Dr.

S. Ayyappan, Secretary, Department of Agricul-

tural Research and Education and Director Gen-

eral, ICAR and former Deputy Director General

(Fisheries), ICAR for his keen interest and support

to this work. We are thankful to the Director,

CIFA for encouragement and for providing facili-

ties. Help received from the state fisheries depart-

ments of Gujarat, Kerala and Odisha during the

collection of different populations of M. rosenbegii

is gratefully acknowledged. This work is a contri-

bution to the CGIAR Research Program on Live-

stock and Fish.

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