Abstract
Present study was intended to the phenotypic and genetic categorization of the endangered catfish
Rita rita, development of sperm cryopreservation protocol and assessment of culture potentiality at
different stocking densities in raceway for ex-situ conservation. Landmark based morphometric and
meristic counts were employed to assess the status of the stocks of R. rita. Only one (caudal fin rays)
out of six meristic traits, 14 of 15 morphometric characters (except dorsal spine length), and 17 of 25
truss network distances showed a highly significant interstock variation (P<0.05, P<0.01 and P<0.001)
among the samples. Plotting discriminant functions and canonical analysis showed a clear separation
of the stocks for both morphometric and landmark measurements. Linear discriminant function
analysis for both the morphometric and landmark based measurements revealed equivalent overall
percentage of correctly classified fish into their original group as high as 78.5 and 78.0%, respectively.
The dendrogram based on morphometric and truss distances data placed the Old Brahmaputra
population in one cluster, and the Meghna and Jamuna populations in other cluster with a sub-cluster
of the Kangsa population. DNA microsatellite markers were also applied to evaluate the population
genetic structure of R. rita. Twelve microsatellite markers developed from other catfishes were tested
for PCR amplification of which five primers were amplified successfully. Four (except Cba06) of the
five amplified loci were found to be polymorphic (P95) in the four studied populations and a total of
46 alleles were recorded with 9 to 14 alleles per locus. The highest total number of alleles was
observed in both the Jamuna and Meghna (44) populations followed by the Old Brahmaputra (43)
and the Kangsa (41) population. The average observed heterozygosity (H,) value was the highest in
the Meghna population (0.7550) followed by the Jamuna (0.7000), the Old Brahmaputra (0.6050) and
the Kangsa (0.5700) populations. All the populations significantly deviated (P<0.001) from the HardyWeinberg Equilibrium for all the polymorphic loci. A comparatively highest population
differentiation (FsT- 0.0168) and lowest gene flow (N- 14.6175) were observed between the JamunaKangsa population pair; while the least FsT (0.0071) and uppermost N, (34.7476) were found between
the Jamuna-Meghna populations. The highest genetic distance (0.2563) was observed in the JamunaKangsa population pair compared to other population pairs. The UPGMA dendrogram based on
genetic distance produced a cluster with the Jamuna and the Meghna populations while the Old
Brahmaputra and the Kangsa populations remained in other cluster. An attempt was taken to
develop a cryopreservation protocol of sperm of R. rita. Milt was collected by crushing the testes and
suspended in extenders. Activation motility of sperm was improved from 48 to 128 mOsmol/kg, and
turned into stable condition by presenting complete activation at the osmolality of 128 mOsmol/kg
for all the tested extenders. The toxicity of three cryoprotectants- DMSO, methanol and ethanol each
with 5, 10 and 15% concentration to sperm was evaluated, and 10% concentration produced highest
sperm motility. Alsever's solution with 10% DMSO generated the significantly (P<0.05) highest
equilibration (89.67±5.51%) and post-thaw (79.33±5.13%) motility over other combinations of diluents.
For evaluating the effects of cryostorage, sperm was processed with Alsever's solution, egg-yolk
citrate and urea egg-yolk plus 10% DMSO and cryostored for six weeks. The sampling at seven days
interval showed a significantly (P<0.05) highest post-thaw motility of sperm processed with Alsever's
solution over egg-yolk citrate and urea egg-yolk. The catfish were reared in a raceway to observe the
culture potentiality at three different stocking densities (4, 8 and 16 juveniles/m²) under three
treatments (T1, T2 and T3, respectively) with three replications each. The mean initial length and
weight of Rita juveniles were 20.15±1.51 cm and 100.26±6.57 g, respectively. The fish were reared by
providing cleaned poultry viscera (containing 48.56% crude protein) at 5 - 2% of biomass once a day.
The fish were sampled once a month to observe the growth performance. The growth parameters
such as mean final length, relative length gain, mean body weight, relative weight gain and SGR were
significantly (P<0.05) higher in T1 compared to those of T2 and T3. The survival rate of the fish was
100% in all the treatments. The gross and net production of the fish were significantly (P<0.05) higher
in T3 (8.38±0.04 kg/m² and 6.78±0.004 kg/m²) than those of T2 (4.49±0.03 kg/m² and 3.69±0.003
kg/m²) and T1 (2.36±0.02 kg/m² and 1.96±0.002kg/m²). Though the growth performance of R. rita
was significantly influenced by the stocking density but the production findings suggest that this
species can be reared at the minimum density of 16 juveniles/m² in cemented raceway with suitable
environment and adequate supplemental feed. Mass seed production through induced breeding and
conservation of the stocks through proper management is suggested to save the endangered and
valuable Rita catfish from the threat of extinction.