Immunological and molecular diagnostic techniques in fish health: present and future prospectus.

Fish health management is critical to aquaculture and fisheries as it directly affects sustainability and productivity. Fish disease diagnosis has taken a massive stride because of advances in immunological and molecular diagnostic tools which provide a sensitive, quick, and accurate means of identifying diseases. This review presents an overview of the main molecular and immunological diagnostic methods for determining the health of fish. The immunological techniques help to diagnose different fish diseases by detecting specific antigens and antibodies. The application of immunological techniques to vaccine development is also examined in this review. The genetic identification of pathogens is made possible by molecular diagnostic techniques that enable the precise identification of bacterial, viral, and parasitic organisms in addition to evaluating host reactions and genetic variation associated with resistance to disease. The combination of molecular and immunological methods has resulted in the creation of novel techniques for thorough evaluation of fish health. These developments improve treatment measures, pathogen identification and provide new information about the variables affecting fish health, such as genetic predispositions and environmental stresses. In the framework of sustainable fish farming and fisheries management, this paper focuses on the importance of these diagnostic techniques that play a crucial role in protecting fish populations and the aquatic habitats. This review also examines the present and potential future directions in immunological and molecular diagnostic techniques in fish health.

Quality assessment of springs for drinking water in the Himalaya of South Kashmir, India.

The present study describes the water quality scenario of some freshwater springs of South Kashmir during the two-year period (2013-2015) because of rising pollution risks endangering water resources globally. The accessibility to quality drinking water has become a challenge and is receiving renewed attention. A total of 96 samples from twelve springs were collected and analyzed for major drinking water quality parameters. Piper trilinear and Durov diagram depicted dominance of Ca-Mg-HCO3 hydrochemical facies and simple dissolution and mixing process. Water quality was falling in very good to excellent class and well within the desirable limits of WHO thereby indicating huge potential for meeting rising drinking water demand. The principal component analysis (PCA) revealed the generation of three components (PC1, PC2, and PC3) with higher eigenvalues of 3 or more (3-6) explaining 40, 21, and 17% of the overall variance in water quality data sets, respectively. The components obtained from PCA indicate that the parameters responsible for variations are mainly related to discharge, temperature, and dissolved oxygen (natural), nutrients (agriculture), and cation and anions (lithology). The results suggest that the hydrochemistry of springs is jointly controlled by lithology and anthropogenic inputs.

Podophyllum hexandrum ameliorates endosulfan-induced genotoxicity and mutagenicity in freshwater cyprinid fish crucian carp.

CONTEXT: Medicinal plants continue to act as a repository for novel drug leads with novel mechanisms of action. Podophyllum hexandrum Royale (Berberideceae) treats diverse conditions in folk medicine. OBJECTIVE: The antimutagenic potential of P. hexandrum was evaluated against endosulfan-induced clastogenicity in a piscine model by cytogenetic endpoints. MATERIALS AND METHODS: Podophyllum hexandrum rhizomes were subjected to successive solvent extraction. Fish were exposed to hexane, chloroform, ethyl acetate, methanol and aqueous extracts (15 mg/L each) of plant and endosulfan (0.05 mg/L) alone followed by their combination for antimutagenicity estimates. Chromosomal aberrations (CA) were made from kidney cells and micronuclei (MN) slides from peripheral blood erythrocytes at 48, 72 and 96 h. Antioxidant activity was analyzed by the DPPH assay. Phytochemical analyses were carried out using chromatographic and spectroscopic techniques. RESULTS: Endosulfan induced significant (p < .05) MN, authenticated by scanning electron microscopy, and CA in a time-dependent manner. However, methanol and ethyl acetate extracts revealed ameliorating effects. The column eluted methanolic fraction-2 (ME-F2) showed highest reduction profile of 83 and 84% in CA and MN, followed in its extent (73 and 72%) by ethyl acetate fraction-4 (EE-F4). ME-F2 and EE-F4 showed three and six major peaks when analyzed by GC-MS. To explore possible mechanism of action, ME-F2 showed potent antioxidant potential and strong correlation (R2 = .900) with antimutagenic activity, whereas EE-F4 seemed to act through a different mechanism. DISCUSSION AND CONCLUSION: This study confirms the antimutagenic potential of the subject plant with the identification of some novel compounds, justifying their use in folk medicine, and their corresponding benefit to mankind.

Distribution of Helminth Parasites in Intestines and Their Seasonal Rate of Infestation in Three Freshwater Fishes of Kashmir.

The present study was undertaken to determine the incidence of helminth parasites in fishes with special reference to water quality parameters in Dal Lake and River Jhelum and correlate the observations. Water, fish, and parasite samples were collected during different seasons from various sites and processed. Three fish species, namely, Schizothorax niger Heckel 1838, Schizothorax esocinus Heckel 1838, and Schizothorax curvifrons Heckel 1838, were recovered from these water bodies. The physicochemical parameters temperature, dissolved oxygen, pH, and free carbon dioxide showed variation vis-à-vis the season and location of the stations in water bodies. Acanthocephalan parasite Pomphorhynchus kashmirensis Kaw 1941 (27.47%) and two intestinal cestodes Bothriocephalus acheilognathi Yamaguti 1934 (30.63%) and Adenoscolex oreini Fotedar 1958 (32.43%) were recovered from all the three species of Schizothorax. All the three parasites showed higher prevalence during summer and the least prevalence during winter. Parasitic infections were prevalent more in male fishes compared to females. The presence of the parasites had reduced the condition coefficient of the infected fishes in both water bodies. The study also showed that some of the physicochemical features showed a significant positive correlation with the prevalence.

Length variation and sequence divergence in mitochondrial control region of Schizothoracine (Teleostei: Cyperinidae) species.

Schizothoracine fish commonly called snow trouts inhibit the entire network of snow and spring fed cool waters of Kashmir, India. Over 10 species reported earlier, only five species have been found, these include Schizothorax niger, Schizothorax esocinus, Schizothorax plagiostomus, Schizothorax curvifrons and Schizothorax labiatus. The relationship between these species is contradicting. To understand the evolutionary relation of these species, we examined the sequence information of mitochondrial D-loop of 25 individuals representing five species. Sequence alignment showed D-loop region highly variable and length variation was observed in di-nucleotide (TA)n microsatellite between and within species. Interestingly, all these species have (TA)n microsatellite not associated with longer tandem repeats at the 3' end of the mitochondrial control region and do not show heteroplasmy. Our analysis also indicates the presence of four conserved sequence blocks (CSB), CSB-D, CSB-1, CSB-II and CSB-III, four (Termination Associated Sequence) TAS motifs and 15bp pyrimidine block within the mitochondrial control region, that are highly conserved within genus Schizothorax when compared with other species. The phylogenetic analysis carried by Maximum likelihood (ML), Neighbor Joining (NJ) and Bayesian inference (BI) generated almost identical results. The resultant BI tree showed a close genetic relationship of all the five species and supports two distinct grouping of S. esocinus species. Besides the species relation, the presence of length variation in tandem repeats is attributed to differences in predicting the stability of secondary structures. The role of CSBs and TASs, reported so far as main regulatory signals, would explain the conservation of these elements in evolution.

Assessment of endosulfan induced genotoxicity and mutagenicity manifested by oxidative stress pathways in freshwater cyprinid fish crucian carp (Carassius carassius L.).

Over the past few decades, endosulfan, one of the polychlorinated pesticides still in use, has received considerable attention of a number of international regulations and restriction action plans worldwide. This study aimed to evaluate the cytogenetic effects of endosulfan using robust genotoxicity assays, along with the oxidative stress pathways in order to understand biochemical mechanism, in Carassius carassius L. The LC50-96 h (95% confidence limits) value of endosulfan was 0.070 (0.046-0.093) ppm; and on its basis three test concentrations (sub-lethal I: 0.052, II: 0.035 and III: 0.017 ppm) were selected for 35 d in vivo exposure. The mean concentration of endosulfan in aquaria was always constant, when analyzed by dispersive liquid-liquid micro extraction (DLLME) followed by GC-MS. Autopsy was done on days 1, 2, 3, 4, 7, 14, 21, 28 and 35 of endosulfan exposure; the micronucleus formation (MN), authenticated by scanning electron microscopy, and chromosomal aberrations (CA), were induced significantly (p<0.05) in all the treated groups, including positive control cyclophosphamide (4 ppm), when compared to negative control. Similarly lipid peroxidation (LPO) was induced significantly with the maximal at higher concentration (SL-I) on 4th day (722.45%; p<0.01). Antioxidant biomarkers like glutathione reduced, superoxide dismutase and catalase also fluctuated significantly (p<0.01) in all treatment groups. Collective findings demonstrated that genotoxic effects were invariably accompanied and correlated with increased oxidative stress and disturbance of antioxidant enzymes; and the MN and CA assays are useful tools in determining potential genotoxicity of aquatic xenobiotics and might be appropriate as a part of monitoring program.

Mitochondrial DNA variability to explore the relationship complexity of Schizothoracine (Teleostei: Cyprinidae).

Despite numerous studies on the taxonomy of a highly complex group of schizothoracine (snow trouts), with over five recognized species from Kashmir, India (Schizothorax niger, Schizothorax esocinus, Schizothorax plagiostomus, Schizothorax curvifrons and Schizothorax labiatus) based on traditional morphological data, the relationships between these species is poorly understood and the taxonomic validity is still under debate. To resolve the evolutionary relationships among these species, we sequenced mitochondrial fragments, including 16Sr RNA, Cytb and the D-loop. Separate analyses of 16S and Cytb showed intermixing of the species and 16S was found more conserved than Cytb. The D-loop was found highly variable and showed length variation between and within species. Length variation was observed in di-nucleotide (TA)n microsatellite repeats with a variable number of repeat units (n = 7-14) that did not show heteroplasmy. Central conserved sequence blocks (CSBs) in D-loop sequences were found comparable to other vertebrate species. All phylogenetic reconstructions recovered the focal taxa as a monophyletic clade within the schizothoracines. Analyses with combined mitochondrial data sets showed close genetic relationships of all the five species. In addition to a close relationship between S. niger and S. curvifrons, two distinct groupings of S. ecoscinus and S. plagiostomus were supported by all the analyses. This study gives an insight into molecular phylogeny of the species and improves our understanding of historical and taxonomic relationships derived from morphological and ecological studies.