Population genetic structure of freshwater mud eel, Monopterus cuchia (Hamilton, 1822) in Bangladesh

Abstract

n this study, population genetic structure of freshwater mud eel, Monopterus cuchia in Bangladesh was observed by RAPD and RFLP assay for finding a superior population with high genetic diversity. Some associated observations including rearing, breeding biology and artificial breeding has also been performed. First experiment was conducted to study the effect of different environments such as House tanks, Earthen ditches, Plastic tank and Back yard tanks on rearing and production performance of freshwater mud eel, M. cuchia where different types of biological food in different amounts were used, except the earthen ditch-2 (control). 40 fishes (approximately 1kg in weight) were stocked into each of the environment and the highest mean of final length (32.89±2.65423) and final weight (109.69±2.92777) were obtained in earthen ditch-1. The weight gain in length 17.78 ± 0.24 and in weight 85.07 ± 0.15 with 92.5% survival and production (4.07kg) were also highest in earthen ditch-1. On the other hand, the lowest mean growth considering final length (28.45±2.80092) and weight (67.24±3.79328) were recorded in house tank-2 while the weight gain in length (11.86 ± 0.26) and in weight (41.84 ± 0.59) were also observed in house tank-2 with the production of 2.02kg by 75% survival. Based on Pearson correlation coefficient no significant correlations were found between lengths and weights of fish in different culture regimes. In second study a successful protocol was developed to identify male and female M. cuchia through external morphology, internal anatomy and histological analysis. Furthermore, breeding biology of freshwater mud eel, M. cuchia was considered in terms of gonadosomatic index, length-weight relationship of gonad, ova diameter and fecundity. The ova diameter was recorded from 0.3 mm to 4.30 mm and the individual fecundity was recorded from 155 to 1495 while relative fecundity was found from 2.64 to 12.45. A peak of GSI was observed 2.14±0.2 in male and 5.1 ±1.09 in female. Artificial breeding of fish was practiced but unsuccessful spawning was recorded through different inducing agents in captive conditions. In this study, 16S rRNA and glutamine synthetase gene was used for identification of this fish and molecular standard was recorded 250bp length by 16S rRNA gene and 541bp length by the partial sequence of glutamine synthetage gene. In this study, genetic diversity of freshwater mud eel was studied by RAPD assay and a total of 735 bands with 228 polymorphic loci were detected while polymorphisms were revealed by all eight arbitrary primers. The highest and lowest number of bands was recorded 120 and 49. Thirteen different groups of inter individual pair wise similarity and 36 groups of genetic diversity were measured whereas the lowest and highest genetic distance was found 0.58 and 0.97 respectively while the average genetic diversity was found 0.8173716. The highest and lowest Nei’s genetic similarity values were recorded 0.60 and 0.06 respectively and linkage distance was computed between the ranges of 3.6 to 6.24 which were generated six clusters by 11 clades. Intra and inter population genetic diversity was analyzed by the same eight RAPD primers considering 2 populations (Tanguar Haor=P1 and Hakaluky Haor =P2). A total of 353 bands with 147 polymorphic loci were detected in P1 whereas a total 371 bands with 153 polymorphic loci were found in P2. Polymorphisms were revealed by all the primers in P1 and P2 while a single monomorphic locus were seen in P1 resulting 95% and 100% polymorphism showed in P 1 and P2 respectively. 9 and 13 groups of inter individual pair wise similarity were found in P1 and P2 respectively considering averages 6.46 and 6.08. The 27 and 29 different genetic distance values were recorded in P1 and P2 while the average genetic distances were calculated 0.80 in P1 and 0.816 in P2 respectively. Nei’s genetic similarity in P1 was ranged 0.11 - 0.58 with 34 diverse values and in P2 it was ranged between 0.06 – 0.54 with 36 different values, and in an average it were originated 0.323 in P1 and 0.301 in P2 respectively. A total 35 and 18 different linkage distances were estimated in P1 and P2 respectively whereas the average linkage distances were found 5.115 and 5.141 respectively in P1 and P2. A genetic relationship was made by using linkage distance where five clusters were created from 5 clades in both P1 and in P2. Based on allelic information, the frequency of q alleles were recorded highest in both the populations than the frequency of p alleles. Based on the intra-locus gene diversity the average highest gene diversity was observed in P2 in most of the primers. However, the average gene diversity was found highest 0.164635264 in P2 and lowest 0.150829501 in P1. Genetic diversity of glutamine synthetage gene was analyzed in this freshwater fish with two restriction enzymes of +CfrI and +Hpy178III while both the enzymes were digested the size of 541 bp and polymorphism was detected in terms of wild type homogygotes, polymorphic homozygote and heterozygosity. The genetic diversity was observed by using the RFLP band analysis and four different groups of individuals were identified with the p-values of 0, 0.033, 0.05 and 1 respectively whereas different distances were found among the groups. Intra and inter population genetic diversity of M. cuchia was analyzed by using same fragment of glutamine synthetage gene by the same enzymes considering the ecological habitats of Tanguar Haor (P1) and Hakaluki Haor (P2) resulting less polymorphism was observed in P1 than P2. Considering different genetic distances two groups of individuals were identified very close with 0.02 P-value in P1 and five groups were revealed in P2 from very close to large distances. The frequency of q alleles were recorded highest in both the populations than the frequency of p alleles except the enzyme +Hpy178III in P2 (0.433). However, intra-locus gene diversity was found highest in P2 by both the enzymes than P1. Inter population differentiation for one locus (gst = 0.5299) shows significant delineation between populations in allele frequencies and it can say that a high percentage of genetic distance is distributed among two populations. F-statistics was analyzed for observing the genetic structure of populations where degree of allele frequency (FIT = 0.7606) and differentiation of allele frequencies between two populations seems greater (FST = 0.1439), with only a moderate effect of random mating were observed within the populations (FIS = 0.7204). From the above records, polymorphism was found in glutamine synthetage gene of this fish and generally higher polymorphism was observed in P2 as well. Overall finding of this research is concluded that the earthen ditch was found one of the suitable culture environments of M. cuchia and live feed was recorded suitable for obtaining better growth. Breeding biology of this fish was considered with development of a successful protocol to identify male and female as prior knowledge for breeding program, and unsuccessful spawning was recorded through inducing agents. A molecular standard was recorded for species identification of this fish. Final observation in both cases of RAPD and RFLP study higher genetic diversity were observed in general as well as at population level though P2 was recorded stronger genetic status than P1. The research is indicating the good genetic status of this fish in Bangladesh particularly in the ecological habitats of Sylhet. e