Dr. Naresh S. Nagpure

The present study explored the induced genotoxicity (DNA damage) due to organophosphate pesticide profenofos (PFF) after in vivo exposure in freshwater fish Channa punctatus by the use of Comet assay and Random amplified polymorphic DNA (RAPD). The fish specimens were exposed to sub-lethal concentration of 1.16 ppb (50% of LC) in a semi-static system and the DNA damage was assessed in exposed and control fish. The DNA damage was measured in erythrocytes as the percentage of DNA damage in Comet tails and RAPD technique using oligonucleotide primers of fish specimens exposed to the sublethal concentrations of PFF. The most informative primers in terms of variation in RAPD profile were found to be OPA-01, OPA-03, OPB-02, OPB-01 and OPA-13. Appearance/disappearance of bands and increase/decrease in the band intensity were evident in the RAPD profile of fish specimens exposed to PFF as compared to the control. Findings from the present study suggest that the potential impacts of assessme...

The walking catfish Clarias magur (Hamilton, 1822) (magur) is an important catfish species inhabiting the Indian subcontinent. It is considered as a highly nutritious food fish and has the capability to walk to some distance, and survive a considerable period without water. Assembly, scaffolding and several rounds of iterations resulted in 3,484 scaffolds covering ∼94% of estimated genome with 9.88 Mb largest scaffold, and N50 1.31 Mb. The genome possessed 23,748 predicted protein encoding genes with annotation of 19,279 orthologous genes. A total of 166 orthologous groups represented by 222 genes were found to be unique for this species. The Computational Analysis of gene Family Evolution (CAFE) analysis revealed expansion of 207 gene families and 100 gene families have rapidly evolved. Genes specific to important environmental and terrestrial adaptation, viz. urea cycle, vision, locomotion, olfactory and vomeronasal receptors, immune system, anti-microbial properties, mucus, therm...

Danio rerio, zebrafish, has been widely used as a non-mammalian vertebrate model organism in various studies. The present research describes to develop and characterize a new cell line from a wild strain Indian zebrafish native to Brahmaputra River, Assam, India. The new cell line designated as DRCF was developed from the caudal fin of D. rerio. The cell line was successfully subcultured up to 31 passages. Growth studies revealed that cell growth of DRCF was optimal at 28 °C in L-15 medium supplemented with 20% FBS. Molecular characterization of the DRCF cell line using mitochondrial genes namely cytochrome oxidase subunit I gene (COI) and 16S rRNA authenticated the true origin of the cell line. The chromosome analysis of the DRCF cell line expressed its 50 diploid chromosome number of D. rerio. The immunocytochemical characterization of the cell line exhibited its fibroblastic morphology. The expression of the green fluorescent protein (GFP) following transfection revealed the suitability of the cell line for transfection studies.

Cyprinion semiplotum (order: Cypriniformes; family: Cyprinidae) is endemic to Southern Asia and has limited distribution in rivers of Eastern Himalayan region. It has an important role in ecology and is an important source of nutritious food and livelihood to the local people. In the recent past, the population of this species has declined due to various anthropogenic factors, and consequently, it has been categorized under vulnerable species. Genetic stock structure information would be useful to formulate appropriate management and conservation measures. The present study is carried out to characterize the Cyprinion semiplotum stocks collected from selected rivers in the state of Arunachal Pradesh, India, using mitochondrial ATPase 6/8 gene. A total of 14 haplotypes were found with nucleotide diversity in the range of 0.0013–0.0028. The result of AMOVA analysis also revealed that 99.44% of the genetic variation contained within stocks/populations and 0.56% occurred among populations/stocks. The results of the present study indicated that C. semiplotum stocks in north eastern part of India have low genetic differentiation and may be considered as a single panmictic population.

Malathion [S-(1,2-dicarboethoxyethyl) O, O-dimethyl phosphorodithioate] is a widely used organophosphorus insecticide throughout the world. However, limited efforts have made to study its genotoxic effect in different fish tissues. The present investigation was aimed to assess the genotoxic potential of the pesticide to the freshwater teleost fish Channa punctatus at sublethal concentrations using the micronucleus test and comet assay. Initially, the 96-h LC50 value of commercial-grade malathion (50% EC) was determined as 5.93 ppm in a semistatic system. Based on LC50, three test concentrations (viz. sublethal I, sublethal II, and sublethal III) were determined to be 1.48, 0.74, and 0.59 ppm, respectively, and the fish specimens were exposed to these concentrations. Tissue samplings were done on days 0, 1, 3, 7, 15, 22 and 29 of malathion exposure for assessment of the induction of micronuclei (MN) frequency and DNA damage. The MN formation in the peripheral blood cells was found to be significantly higher (p < 0.05) in the treated specimens at all sampling intervals compared to the control. The MN frequency reached maximum on days 3 and 7 at sublethal I and II concentrations, respectively, followed by a nonlinear decline with the progression of the experiment. Similarly, significant effects (p < 0.05) of both concentration and time of exposure were observed on DNA damage in the gill, kidney, and lymphocytes. All of the tissues exhibited a concentration-dependent increase in DNA damage up to day 3, followed by a nonlinear decrease with the duration of exposure. A comparison of the extent of DNA damage among the tissues showed the sensitivity of gill tissue to malathion.

River pollution due to rapid industrialization and anthropogenic activities adversely affects the aquatic organisms, especially fish. Here, we assessed the genotoxicity, mutagenicity and bioaccumulative aspects of tannery effluents in freshwater murrel, Channa punctatus, an inhabitant of river Ganges. Test specimens were collected from three different polluted sites of the river within and nearby Kanpur area during different seasons and blood samples of these specimens were processed for comet assay and micronucleus test as genotoxicity biomarkers. A significantly (P < 0.05) higher micronuclei induction, nuclear abnormalities and % tail DNA was observed in the specimens collected from the polluted sites. Bioaccumulation studies in the muscle (1.202 μg/g) and gill tissues (< 0.300 μg/g) of the specimens revealed the concentration of chromium (core component of tanning industry) above the maximum permissible limits as prescribed by World Health Organization (WHO). The findings of the present analysis indicated contamination of river Ganges with tannery effluents which induce genotoxicity in fish with seasonal variation.

The results of the Cytogenetic studies carried out in two marine ornamental fishes, Cl~aetodon rollare and Stegastes insularis, occurring in the south-eastem Arabian Sea, by Giemsa staining and nuckolar organizer regions (Ag-NOR) staining are reported. The diploid chromosome number (2n) in C. collare and S. insularis was found to be 48. Based on the karyomorphology, the karyotype formula was derived as 48t (FN=48) and 14m + 24 sm + 6st + 4 t (FN=86), respectively for these two species. Silver staining revealed presence of one pair of NORs in both the species. This is the first description of karyotypes in these species using conventional staining and chromosome banding techniques. The rich fish diversity of India is reflected in its large number of documented 2163 fish species of which 1370 (63%) are from the marine ecosystem. So far, marine fishes have not received much deserved attention towards their genetic characterization and only about 40 of them have been cytogenetically cha...

DNA markers are being increasingly used in studies related to population genetics and conservation biology of endangered species. DNA isolation for such studies requires a source of biological material that is easy to collect, non-bulky and reliable. Further, the sampling strategies based on non-invasive procedures are desirable, especially for the endangered fish species. In view of above, a rapid DNA extraction method from fish scales has been developed with the use of a modified lysis buffer that require about 2 hr duration. This methodology is non-invasive, less expensive and reproducible with high efficiency of DNA recovery. The DNA extracted by this technique, have been found suitable for performing restriction enzyme digestion and PCR amplification. Therefore, the present DNA extraction procedure can be used as an alternative technique in population genetic studies pertaining to endangered fish species. The technique was also found equally effective for DNA isolation from fre...

The present study was undertaken to evaluate the lethal toxicity and stress of commercial formulations of carbosulfan (Aatank) insecticide, glyphosate (Roundup) and atrazine (Rasayanzine) herbicides toward freshwater air-breathing fish Channa punctatus (Bloch). The 96 h LC 50 values, determined in a semi-static system by probit analysis as 0.268, 32.540 and 42.380 mg/l for carbosulfan, glyphosate and atrazine, respectively, indicated that the fish were more sensitive to carbosulfan than the other two herbicides. There were large variations in the safe levels estimated by different methods for the pesticides. In addition to dose and dose-time dependent increase in mortality rate, stress signs in the form of behavioral changes were observed in response to the test pesticides.

Fish cell lines provide an important in vitro system for investigations in physiology, virology, toxicology, carcinogenesis and transgenesis. The initiation of development of fish cell lines was done at the level of primary cell culture in India and no permanent cell line was established until 2006. The initial efforts made for development of primary cell cultures from gill tissue of mrigal (Cirrhinus mrigala) and rohu (Labeo rohita) and further attempts using variety of other tissues, such as heart and caudal fin of Indian major carps, gave the momentum to fish cell culture work in India. The daunting task of establishing a National Repository of Fish Cell Lines (NRFC) at National Beaureu of Fish Genetic Resources, Lucknow has been possible with the financial support of Department of Biotechnology, Ministry of Science & Technology, Govt. of India. This article throws light on different aspects of NRFC, such as the details of fish cell lines available with NRFC, process of establishme...

Genetic structure of four populations of Metapenaeus affinis from Maharashtra, Orissa, Kerala and Tamil Nadu in India was studied using RAPD markers. Five selective primers provided distinct and consistent RAPD profiles in all the four populations. The bands in the range 225-1,900 bp were scored for consistent results. The RAPD profiles generated by all the five primers revealed varying degrees of polymorphism, ranging from 25.00% (primer E-03) to 65.00% (primer E-06). Nei's (Nei M, Natl Acad Sci Proc USA 70:3321-3323, 1973) genetic diversity (h) among the four populations varied from 0.2565 ± 0.2146 (Orissa population) to 0.3576 ± 0.1897 (Maharashtra population).

The Indian freshwater air-breathing teleost fish Channa punctatus (Bloch) was exposed to acute concentrations of the organochlorine pesticide endosulfan. In flow-through bioassays the 24, 48, 72, and 96 h LC(50) values were estimated as 19.67, 12.95, 10.15, and 7.75 ppb, respectively. DNA damage (single-strand breaks) was also studied in gill and kidney tissues at single-cell levels in the specimens, exposed to different acute doses of endosulfan, by applying single-cell gel electrophoresis or comet assay. Dose-dependent responses were observed in DNA damage in both tissues. A comparison of DNA damage in both tissues at different doses indicated that the gill cells were more sensitive to the pesticide exposure than the kidney cells. This study explored the utility of the comet assay for in vivo laboratory studies using fish for screening the genotoxic potential of various agents.