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The present study investigates the distribution and dynamics of gonadotropin-releasing hormone I (GnRH I) and bradykinin in the air-breathing catfish, Heteropneustes fossilis, in relation to the reproductive cycle. Changes in bradykinin, bradykinin B2-receptor, and ovarian GnRH I regulation were demonstrated during the reproductive cycle. The localization of GnRH I, bradykinin, and their respective receptors in the ovaries was investigated by immunohistochemistry, while their levels were quantified by slot/western blot followed by densitometry. GnRH I and its receptor were mainly localized in the cytoplasm of oocytes during the early previtellogenic phase. However, as the follicles grew larger, immunoreactivity was observed in the granulosa and theca cells of the late previtellogenic follicles. The ovaries showed significantly higher expression of GnRH I protein and its receptor during the early to mid-previtellogenic phase, suggesting their involvement in follicular development. Bradykinin and bradykinin B2-receptor showed a distribution pattern similar to that of GnRH I and its receptor. This study further suggested the possibility that bradykinin regulates GnRH I synthesis in the ovary. Thus, we show that the catfish ovary has a GnRH-bradykinin system and plays a role in follicular development and oocyte maturation in H. fossilis.

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This investigation was done to determine how much zinc (Zn) the stinging catfish, Heteropneustes fossilis, needs in its diet. Five isonitrogenous (34.5% protein) and isolipidic (6.0% lipid) diets were prepared to contain graded levels of Zn (0, 10, 20, 30, and 40 mg kg-1), supplied as zinc sulfate (ZnSO4·7H2O), and referred to as Zn0, Zn10, Zn20, Zn30, and Zn40, respectively. A total of 600 fish (initial body weight: 1.41 ± 0.02 g) were stocked in 15 glass aquaria (40 fish/aquarium), each with 180 L water capacity. For ten weeks, each diet was hand fed to three groups of fish twice daily until they appeared satisfied. The highest weight gain and specific growth rate, and lowest feed conversion ratio were recorded in fish fed with a 30 mg Zn kg-1 diet. Zn contents in bone and muscle linearly increased up to 30 mg kg-1 Zn and then remained stable, while iron (Fe) and copper (Cu) contents in bone and muscle had an inverse pattern with the inclusion level of dietary Zn. Increasing dietary Zn levels up to 30 mg kg-1 was found to improve values of hematological parameters such as red blood cell (RBC), white blood cell (WBC), haemoglobin (Hb), and haematocrit (HCT). These values, however, decreased when the dietary Zn level was further increased. The serum alkaline phosphatase level was the highest in fish fed a diet containing 30 mg kg-1 of Zn. Regression analyses based on weight gain, specific growth rate, and bone and muscle Zn concentrations indicated that the optimum dietary Zn requirement for stinging catfish was in a range of 27.4-36.5 mg kg-1.

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A 12 week feeding trial was conducted to evaluate the effects of dietary zinc levels on Heteropneustes fossilis. Triplicate groups of fish were fed isoproteic (CP; 400 g/kg) and isocaloric (GE; 17.89 kJ/g) diets increasing levels of zinc (0, 5, 10, 15, 20, 25, 30 mg/kg) achieved by supplementing zinc sulphate heptahydrate to basal diet. Analysed concentrations of zinc in diets were 10.68, 15.83, 21.34, 26.74, 30.61, 34.91 and 41.34 mg/kg. Growth indices increased linearly (P<0.05) up to 26.74 mg/kg Zn. The protein and ash content of whole body also improved significantly up to 26.74 mg/kg Zn. Whole body fat content showed inverse pattern. Haematological parameters also showed an improving trend with the increase in dietary zinc up to 26.74 mg/kg and then levelled off. Activities of antioxidant enzymes were improved with the increase in dietary zinc level up to 26.74 mg/kg followed by no significant change (P>0.05). Serum lysozyme activity also exhibited the similar pattern. Immune response in terms of the activities of lysozyme, alkaline phosphatase and myeloperoxidase was also improved with the increase in dietary zinc levels up to 26.74 mg/kg. Dietary zinc levels affected significantly the whole body as well as vertebrae mineralization. Broken-line regression analysis of weight gain, vertebrae zinc activity, serum superoxide dismutase and protease activity against increasing amounts of dietary zinc revealed that the inclusion of zinc in diet in the range of 26.82-29.84 mg/kg is optimum for growth, haematological indices, antioxidant status, immune response and tissue mineralization in fingerling H. fossilis. The information obtained from present study would be helpful in formulating the zinc-balanced commercial feeds to improve the growth and health status of this important fish, thus contributing to aquaculture production and strengthening the food security.

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Biofloc technology aims to maximize fish farming productivity by effectively breaking down ammonia and nitrite, promoting healthy flocculation, and enhancing the growth and immunity of cultured animals. However, a major limitation in this field is the suitable starter microbial culture and narrow number of fish species that have been tested with the biofloc system. Here, we investigated various microbial inoculum containing beneficial microbes with probiotics, immunostimulatory and flocs development and bioremediation properties would lead to the development of ideal biofloc development. Three treatment groups with different microbial combinations, viz., group 1 [Bacillus subtilis (AN1) + Pseudomonas putida (PB3) + Saccharomyces cerevisiae (ATCC-2601)], group 2 [B. subtilis (AN2) + P. fluorescens (PC3) + S. cerevisiae (ATCC-2601)] and group 3 [B. subtilis (AN3) + P. aeruginosa (PA2) + S. cerevisiae (ATCC-2601)] were used and compared with the positive control (pond water without microbial inoculums) and negative control (clear water: without microbial inoculums and carbon sources) on biofloc development and its characteristic features to improve the water quality and growth of fish. We demonstrated that microbial inoculums, especially group 2, significantly improve the water quality and microbiota of flocs and gut of the test animal, Heteropneustes fossilis. The study further demonstrates that biofloc system supplemented with microbial inoculums positively regulates gut histomorphology and growth performance, as evidenced by improved villous morphology, amylase, protease and lipase activity, weight gain, FCR, T3, T4 and IGF1 levels. The inoculums induced an antioxidative response marked by significantly higher values of catalase (CAT) and superoxide dismutase (SOD) activity. Furthermore, the supplementation of microbial inoculums enhances both specific and non-specific immune responses and significantly elevated levels of immune genes (transferrin, interleukin-1ß and C3), and IgM was recorded. This study provides a proof-of-concept approach for assessing microbial inoculums on fish species that can be further utilized to develop biofloc technology for use in sustainable aquaculture.

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The current study was performed to explore potential toxic effect of nickel oxide nanoparticles (NiO NPs) on muscle tissue of catfish, Heteropneustes fossilis. Fishes were exposed to different concentrations of NiO NPs (12 mg/L, 24 mg/L, 36 mg/L and 48 mg/L) for a period of 14 days. Results revealed that NiO NPs caused significant increase in Ni accumulation, metallothionein content, lipid peroxidation and activity of different antioxidant enzymes (catalase, glutathione s transferase and glutathione reductase) while decrease in activity of superoxide dismutase (p < 0.05). Data also reported induction of Na+/K+ ATPase activity initially and then its decrease in concentration dependent manner. Fourier transform infrared spectroscopy revealed shift and changes in spectra of muscle of NiO NPs treated fishes. Fluctuations in activity of aspartate amino transferase, alanine amino transferase and alkaline phosphatase were also noticed. Nutritional contents like protein, lipid, and moisture significantly reduced while glucose and ash percent increased.

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In terms of hatchery-based seed production, one of the most important aquaculture species in Bangladesh is the stinging catfish (Heteropneustes fossilis). Scientific and evidence-based embryonic and larval development research on this fish species in the context of climate change is limited. This experimental study was conducted via induced breeding of stinging catfish using a conventional hatchery system, rearing the larvae in hapas placed in ponds. A series of microscopic observations using a trinocular digital microscope and an analysis of the relationship between larval growth and climate-driven water quality parameters such as temperature, pH, dissolved oxygen, total dissolved solids, alkalinity, and ammonia were performed. During embryonic development, the first cleavage was observed between 30 and 35 min of post-fertilization. Embryonic development (ranging from the 2-cell to the pre-hatching stage) took 21:00 h. Hatching occurred at 22:30 to 23:00 h after fertilization, with an average larvae length of 2.78 ± 0.04 mm. In the post-hatching stage, four pairs of tiny barbels appeared at 36:00 h, and the larvae started feeding exogenously after 72:00 h. These larvae fully absorbed their yolk sacs on the 6th day and attained an average length of 6.44 ± 0.06 mm. Aerial respiration of the larvae was investigated through naked-eye observation on the 10th day of hatching. The average length of the larvae was 32.00 ± 2.0 mm at the end of the 30-day post-hatching observation period. Bivariate correlation analysis showed significant correlations between key climatic variables and water quality parameters under hapa-based larval-rearing conditions. According to canonical correlation analysis, the first canonical function revealed the highest significant correlation between the two sets of variables (r1 = 0.791). The response variable weight of larvae (6.607) was linked to two explanatory variables: pH (0.321) and dissolved oxygen (0.265). For the second canonical correlation function, a positive correlation (0.431) was observed between the two sets of variables. Larval weight (-18.304) was observed to be linked to climatic variables, including air temperature (-0.316) and surface pressure (0.338). Results of this study reveal the subtle correlation between larval growth and water quality driven by climatic variables.

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The stinging catfish Heteropneustes fossilis is a champion survivor under hypertonic stress and is suggested to be a profitable candidate for culture in slightly saline water in coastal regions. Fish gills are an essential site of osmoregulation and other physiological processes. To investigate the stress responses and mechanisms of salinity tolerance in stinging catfish, we sampled gills tissues from control and hypertonicity (100 mM NaCl solution) treated adult catfish and assessed for transcriptomic profiling by high throughput sequencing. The raw data generated was filtered and assembled for de novo transcriptome assembly. The final contig assembly produced a total of 1,71,478 unigene transcripts with an average transcript length of 898 bp and a GC content of 45%. A total of 22,231 transcripts matched with Chordata with BLAST search and were functionally annotated, out of which 21,814 were best-hit transcripts aligned with the UniProt database. Comparative transcriptomic analysis revealed that a total of 1951 genes were differentially expressed in the gills of NaCl-treated fish compared to the control. Functional and enrichment analysis of the Differentially expressed genes demonstrated that several GO pathway terms were significantly over-represented, such as 'catalytic activity', 'hydrolase activity' in molecular function category, 'membrane', 'integral component of membrane' in cellular component category and 'metabolic process', 'regulation of transcription' in biological process category. The functional analysis study of DEGs demonstrated that tolerance to hypertonic stress by stinging catfish is associated with a few pathways related to stress response, immune response, biosynthesis, metabolism, molecular transport, cytoskeleton remodeling, apoptosis, cell signaling, transcriptional regulation, etc. The present study provides a novel insight into the molecular responses of the air-breathing stinging catfish against salinity stress, which could elucidate the underlying mechanisms of adaptation of this stenohaline species under various environmental constraints.

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A thorough investigation of the water permeability of H. fossilis aquaporin 1 (hfAQP1) in a hypertonic environment can provide a useful insight into the understanding of the underlying molecular mechanism of its high tolerance to salinity. Here, we constructed a 3 D homology model of hfAQP1 by taking Bos taurus AQP1, AQP0, and human AQP2 as templates using I-TASSER. The model obtained has similar structural organizations with mammalian AQP1s in all aspects. We investigated the water permeability of the modeled hfAQP1 in a 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) membrane under neutral and 100 mM hypersalinity by subjecting each system to a 100 ns molecular dynamics simulation. Our results show that hypersalinity hinders water permeation across the membrane through the hfAQP1 channel. A change in the intermolecular distance between key residues of the ar/R selectivity filter along with charge redistribution resulted in the accommodation of only 2-6 water molecules inside the channel at once under hypersaline conditions. We investigated the mRNA expression pattern of hfaqp1 in osmoregulatory organs of H. fossilis in response to 100 mM hypertonicity by using qPCR analysis. The transcript was downregulated in kidney and GI tract, but upregulated in the Gills. Thus, the catfish survive in a hypertonic environment by reducing the transport of water in its cellular systems and downregulating the expression of the hfaqp1 gene. The results observed in our study can shed more light on the functionality of AQP1 in catfishes under salinity stress and aid in future researches on solving more gating mechanisms involved in its regulation.Communicated by Ramaswamy H. Sarma.

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Aquaporins (AQPs) are a superfamily of transmembrane channel proteins that are responsible for the transport of water and some other molecules to and from the cell, mainly for osmoregulation under anisotonicity. We investigated here the expression patterns of different AQP isoforms and also during exposure to hypertonicity (300 mOsmol/L) for 48 h in juvenile stages of air-breathing stinging catfish (Heteropneustes fossilis). A total of 8 mRNA transcripts for different isoforms of AQPs and their translated proteins could be detected in the anterior and posterior regions of S1, S2, and S3 stages of juveniles of stinging catfish at variable levels. In general, more expression of mRNAs for different aqp genes was seen in the S2 and S3 juveniles than in the S1 juveniles. Most interestingly, exposure to hypertonicity of S2 juveniles for a period of 48 h led to increased expression of most of the aqp genes both at transcriptional and translational levels, except for aqp3 in the anterior and posterior regions and aqp1 in the anterior region, showing maximum expression at later stages of hypertonic exposure. Thus, it is evident that AQPs play crucial roles in maintaining the water and ionic balances under anisotonic conditions even at the early developmental stages of stinging catfish as a biochemical adaptational strategy to survive and grow in anisotonic environment.

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The toxic effects of Zinc oxide nanoparticles (nZnO) on Branchiura sowerbyi and Heteropneustes fossilis, was assessed in a 96-hour acute exposure regime using behavioral (including loss-of balance and clumping tendencies) and physiological (mucus secretion and oxygen consumption) endpoints. While the relationship between behavioral, physiological biomarkers, and exposure concentrations was assessed using correlation analysis, nZnO toxicity was further predicted using the General Unified Threshold model for Survival (GUTS). The time-dependent lethal limits for acute nZnO toxicity (LC50) on B. sowerbyi were estimated to be 0.668, 0.588, 0.448, and 0.400 mg/l, respectively, at 24, 48, 72, and 96 h whereas for H. fossilis the LC50 values are 0.954, 0.905, 0.874 and 0.838 mg/l. Threshold effect values i.e., LOEC (Lowest Observed Effect Concentration), NOEC (No Observed Effect Concentration), and MATC (Maximum Acceptable Toxicant Concentration) threshold effect values at 96 h were higher for fish compared to the oligochaete. For B. sowerbyi, the GUTS-SD (stochastic death) model is a better predictor of nanoparticle exposure effects compared to the GUTS-IT (individual tolerance) model, however in the case of H. fossilis, the reverse pattern was observed. Oxygen consumption rate was negatively correlated to mortality under acute exposure duration. The strong negative correlation between mortality and oxygen consumption strongly suggests a metabolic-toxicity pathway for nZnO exposure effects. The higher toxicity threshold values i.e., LOEC, NOEC, and MATC for fish compared to the oligochaete invertebrate indicates greater risks for invertebrates compared to vertebrates, with resultant implications for local habitat trophic relationships.

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A new genus, Mystocestus, is proposed to accommodate a new species, Mystocestus anindoi n. g., n. sp. from Mystus vittatus (Bloch) (type host) in West Bengal and Mystus cavasius (Hamilton) (Siluriformes: Bagridae) in Maharashtra, India. The new genus is most similar to Lucknowia Gupta, 1961 in the shape of the body, which is elongate, slightly tapering towards the anterior end, and scolex, which is digitiform, but differs in the shape of the ovary, which is H-shaped (vs. inverted A-shaped in Lucknowia), the absence of a seminal receptacle (present in Lucknowia) and exclusively cortical vitelline follicles (vs. some follicles in the medulla in the latter genus). Molecular data support the erection of the new genus and place it close to Bovienia Fuhrmann, 1931, species of which can be easily distinguished by exclusively lateral vitelline follicles (lateral and median in the new genus), the presence of a seminal receptacle (absent in Mystocestus) and scolex shape (digitiform, with blunt or slightly concave anterior edge in the new genus vs. small, unspecialized or spatulate in Bovienia). The convoluted taxonomy of tapeworms placed in Mystoides Mathur, 1992 is critically reviewed to clean up taxonomic chaos in Indo-Malayan caryophyllideans. Mystoides was erected in an unpublished PhD thesis and thus its generic name becomes unavailable and also, its type species is conspecific with Lucknowia fossilisi Gupta, 1961 from the stinging catfish, Heteropneustes fossilis (Bloch). Other species of this genus are also conspecific with L. fossilisi. In addition, specific names of most of these taxa are unavailable because they were described in unpublished theses or conference abstracts. Based on recent revisions of Indo-Malayan caryophyllideans, the following nine genera with 15 species are considered valid (numbers of species of individual genera are provided in parentheses): Bovienia (3), Djombangia (1), Lucknowia (2), Lytocestus (2), Mystocestus (1), Pseudocaryophyllaeus (2) (all family Lytocestidae); Adenoscolex (1), Lobulovarium (2), Paracaryophyllaeus (1) (all family Caryophyllaeidae).

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The skin mucosa of fish serves as a primary barrier against pathogens. In lesion sites in diseased fish, the mucosal barrier is expected to be compromised, with a substantial presence of potential pathogens. An understanding of the skin microbiome and its functional repertoire would provide important insights into host-microbe interactions, which has important implications for prophylactic measures in aquaculture. This study revealed the skin microbiomes and their functional annotations from healthy and diseased stinging catfish (Heteropneustes fossilis) based on 16S rRNA metagenomics. The OTUs consisted of four major phyla, Proteobacteria, Bacteroidota, Actinobacteriota and Firmicutes. Among members of the predominant phyla, Proteobacteria were rich in healthy fishes, but Bacteroidota and Firmicutes were significantly differentiated in healthy and diseased fish. The diversified microbiome was high in the skin of healthy fishes and did not significantly differ from that of the diseased groups. At the genus level, Pseudomonas showed the highest abundance in healthy fish but was nearly absent in diseased fish, whereas Flavobacterium showed the highest abundance in diseased fish. Linear discriminant analysis identified two phyla (Bacteroidota, Firmicutes) and two genera (Flavobacterium, Allorhizobium) that were consistently identified in diseased fishes. Functional prediction analysis specified that the genes related to physiological functions such as metabolism, immune and digestive systems and environmental adaptations could be highly expressed in diseased fishes. The present study indicates that the compositions, richness and functions of the bacterial community could influence the health status of cultured stinging catfish. Aquaculture-associated pathogenic bacteria may be identified, and preventive measures can be taken for the surveillance of fish health.

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The stinging catfish, Heteropneustes fossilis (Bloch, 1974) is a commercially important fish species in Asia. This is an important food fish as is enriched with high amounts of protein, iron and calcium. The current research demonstrates the population structure, size at sexual maturity, spawning- and peak-spawning season and fecundity of H. fossilis in an important wetland ecosystem - Gajner Beel in northwestern Bangladesh with an aim of its sustainable conservation through induced breeding and aquaculture practices. A total of 426 stinging catfish captured from the Gajner Beel through monthly sampling from January to December 2019 used in the study. Total length (TL), standard length (SL) and body weight (BW) of individual fishes were measured. The size ranges were with 6.70-24.10 cm TL, 1.37-83.94 g BW. Gonads were removed carefully through ventral dissection and weighted. Lm was 14.02, 13.5, 13.0 and 15.0 cm based on maximum length (Lmax ), TL vs. GSI (%), TL vs. SL and logistic model, respectively. Monthly variations of GSI and maturation stages were confirmed in April to August as spawning season and June as peak spawning month. Fulton's condition factor (KF ) was found to be with significant relations with GSI values. Fecundity was 1,730 to 23,870 and significantly correlated with both TL and BW. Temperature has been increasing 0.029 °C/year with the falling of rainfall at 2.96 mm/year in the study area. Environmental factors -Temperature, rainfall, dissolved oxygen and pH were found to be significantly correlated with GSI. We found the optimal range of temperature (29-31 °C), rainfall (350-380 mm), dissolved oxygen (5.0-6.0 mg/l) and pH (7.1-7.5) for spawning of H. fossilis. The paper recommended the policy guidelines to pave the ways of the aquaculture, conservation and management of H. fossilis in the changing eco-climatic events through specific management measures.

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In the present study, Water Deprived Condition (WPC, up to 18 h) induced hypoxia and altered oxidative stress (OS) physiology along with responses of respiratory chain enzyme in Heteropneustes fossilis are described . The body O2 saturation level in the fish was declined with respect to air exposure. Higher levels of lipid peroxidation and protein carbonylation were recorded in the tissue of fish exposed to 6 h of WPC stress. The regulation of the mitochondrial complex and antioxidant enzymes, small antioxidant molecules indicated that the fish can moderately survive up to 6 h of air exposure. Probably with the onset of metabolic depression, it can critically resist the dehydration stress up to 18 h. Although the activities of glutathione peroxidase and reductase were elevated, activities of antioxidant enzymes such as superoxide dismutase and catalase were insufficient to combat WPC induced ROS and OS generated under hypoxia. The small antioxidant molecules played a key role in elimination of ROS. The elevated complex II activity was probably responsible for resisting the complex I, II and IV mediated electron leakage events in mitochondria of the fish under WPC. The total H2O2 removing capacity was less under WPC while the total units of all calculated antioxidants were alleviated signifying an interesting mechanism of WPC induced OS in the fish.

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Reproduction plays an important role in fish population efficiency and its resiliency to fishing and environment changes. The present study described the comprehensive information on reproductive feature of stinging catfish, Heteropneustes fossilis (Bloch 1794), including size at sexual maturity, spawning season, and fecundity using 622 female individuals sampling by the use of gill net, cast net, and square lift net from January to December 2019 in the Ganges River. We calculated the influences of various environmental parameters which include temperature, dissolved oxygen, pH, and rainfall on the reproductive feature of H. fossilis in the Ganges River. For every specimen, total length (TL), standard length (SL), and body weight (BW) were estimated by measuring board and electronic weighing scale. With ventral dissection of fishes, female gonads were cautiously removed and measured to 0.01 g precision. The gonadosomatic index (GSI), modified gonadosomatic index (MGSI), and Dobriyal index (DI) were used to assess the size at sexual maturity (Lm) and spawning season. According to the results of these indices, Lm was obtained 15.5 cm in TL. Also, TL50 was determined through logistic function as 15.5 cm in TL. Moreover, the highest GSI, MGSI, and DI values indicated the spawning season as of March-August, with peak in May-June. Total fecundity (FT) varied from 2059 to 59,984 with a mean of 25,028 ± 15,048. Temperature and rainfall was statistically correlated with GSI. In addition, long climatic data series analysis denoted that yearly mean atmospheric temperature is rising in 0.028 °C/year and yearly mean rainfall is declining in 2.98 mm/year which may suggest a potential shift of the spawning period of the species in the future if this trend persists. The results of our study might be more useful in imposing particular management and conservation for H. fossilis in the Ganges River and the surroundings.

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During the study fishes of the species Channa striatus, Heteropneustes fossilis, Anabas testudineus, Channa punctata were collected and examined for ecto as well as endoparasites. However, a new species of parasitic nematode of the genus Camallanus Railliet and Henry (Rev Parasitol 39:117-129, 1915) is described only from intestinal region of Siluriformes fish Heteropneustes fossilis Bloch (1794). Buccal capsule of the new species is slightly longer than width marked internally by 9-10 beaded longitudinal ridges in lateral view, 1-2 dorsal and ventral ridges incomplete that differs from other related species. It also differs on the basis of body measurements, maximum width/length ratio of buccal capsule, number and length of spicule, tridents nature, number and arrangement of caudal papillae in the posterior end. Due to differences in many aspects from the related congeners, the present species is proposed as new to the science. The present study deals with the morphological and morphometric description of Camallanus sonaiensis sp.n. from Assam, India.

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Heteropneustes fossilis is an air-breathing teleost inhabiting environments with very poor O2 conditions, and so it has evolved to cope with hypoxia. In the gills and respiratory air-sac, the sites for O2 sensing and the response to hypoxia rely on the expression of acetylcholine (Ach) acting via its nicotinic receptor (nAChR). This study examined the expression patterns of neuronal markers and some compounds in the NECs of the gills and respiratory air sac having an immunomodulatory function in mammalian lungs. Mucous cells, epithelial cells and neuroepithelial cells (NECs) were immunopositive to a variety of both neuronal markers (VAChT, nAChR, GABA-B-R1 receptor, GAD679) and the antimicrobial peptide piscidin, an evolutionary conserved humoral component of the mucosal immune system in fish. We speculate that Ach release via nAChR from mucous cells may be modulated by GABA production in the NECs and it is required for the induction of mucus production in both normoxic and hypoxic conditions. The presence of piscidin in mucous cells may act in synergy with the autocrine/paracrine signals of Ach and GABA binding to GABA B R1B receptor that may play a local immunomodulatory function in the mucous epithelia of the gills and the respiratory air sac. The potential role of the NECs in the immunobiological behaviour of the gill/air-sac is at moment a matter of speculation. The extent to which the NECs as such may participate is elusive at this stage and waits investigation.

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The information on the circadian characteristics of catfish in shoals is scanty. We examined the circadian locomotor activity rhythm of catfish Heteropneustes fossilis either singly housed (SS1) or in shoals of four (SS4) and six (SS6) under different light regimens. We carried out the study sequentially under LD1 (12:12), constant darkness (DD), LD2, continuous light (LL), LD3, and DL (12:12). Each condition was for at least 10-12 consecutive days. We analyzed the time-series data by employing Cosinor rhythmometry to detect circadian rhythm characteristics in locomotor activity at a fixed time window with a τ = 24 h. Results indicated that singly housed or shoals exhibited statistically significant (p < .001) circadian rhythm in locomotor activity under LD conditions with a higher activity level during the dark phase. Further, we observed free-running rhythms in locomotor activity under DD and LL, irrespective of the shoal sizes. In phase inverse DL, both singly housed and shoals demonstrated higher activity in the dark phase. The two-way ANOVA results revealed a significant effect of the factor 'light regimen' on amplitude and acrophase; the factor 'shoal size' produced a statistically significant impact on the mesor only. Both shoals showed significantly higher mesor than singly housed fish. The circadian amplitude declined under constant conditions of DD and LL. The locomotor activity rhythm exhibited a free-running pattern with a τFR greater than 24 h under both DD and LL conditions. We conclude that light is a more prominent factor for the entrainment of circadian activity in catfish H. fossilis. However, the extent of social aggregation (shoal size) has little or no effects on the characteristics of circadian locomotor activity rhythm in H. fossilis.

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Endotoxin, the outer cell wall membrane lipopolysaccharide component of the Gram-negative bacteria is a factor responsible for a number of complications/disorders and plays important role in the associated with pathophysiological complications and pathogenesis of many diseases in animals. Unlike higher animals which are extremely sensitive to endotoxin, fish are found to be resistant to endotoxic shock and earlier studies though limited have demonstrated the patho-physiological, immuno-endocrinological and immuno-neurological effects of LPS/endotoxin in aquatic animals including fish. Herein in the present investigation, the effect of pure endotoxin on immuno-haematological parameters of stinging catfish, Heteropneustes fossilis ranging from 50-60 g was studied by intraperitoneally injecting 0.1, 0.05 and 0.01 mg endotoxin per fish. H. fossilis yearlings were found to resist the endotoxin concentration up to 0.1 mg without any mortality. While, no change in immune parameters was recorded in stinging catfish injected with low dose of endotoxin (0.01 mg), most of the immune parameters were found to be significantly elevated in catfish injected with 0.05 mg endotoxin. Different serum and immune parameters like protein, globulin, lysozyme, respiratory burst activity, myeloperoxidase activity, natural agglutination titre were found to be significantly high (p < 0.01) at a dose of 0.05 mg endotoxin per fish. On the contrary, most of these parameters were decreased at high dose i.e., 0.1 mg endotoxin per fish, thereby indicating the immuno-suppressive effect of the endotoxin. The findings of the modulation of innate immunity also corroborated with the results of Aeromonas hydrophila pathogen challenge study with highest percent of mortality in group injected with 0.1 mg endotoxin per fish and least percentage in group injected with 0.05 mg endotoxin per fish.

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Seven isonitrogenous (400 g/kg crude protein) and isocaloric (17.89 kJ/g gross energy) purified diets (casein-gelatin based) with different concentrations of calcium (0, 1, 2, 3, 4, 5 and 6 g/kg) supplemented with calcium lactate at the levels of 0, 5.4, 10.8, 16.3, 21.8, 27.27 and 32.73 g/kg were prepared and fed for 12 weeks to triplicate groups of Heteropneustes fossilis (7.46 ± 0.03 g) to determine the optimum dietary calcium requirement. Analysed values of calcium in the diets were 2.41, 3.82, 4.56, 5.99, 6.71, 7.40 and 8.19 g/kg, respectively. Absolute weight gain, specific growth rate, protein retention efficiency, protein gain and feed conversion ratio of fish fed diets with increasing levels of dietary calcium improved up to 5.99 g/kg and then levelled off. Whole-body protein, moisture and ash contents improved up to 5.99 g/kg dietary calcium and stabilized thereafter. However, whole-body fat exhibited reverse pattern and decreased with incremental levels of dietary calcium up to 5.99 g/kg. Whole-body and vertebrae mineralization was also significantly affected (p < .05) by the increasing dietary calcium levels. Alkaline phosphatase activity improved significantly (p < .05) up to 6.71 g/kg, and no change was recorded beyond this level. Serum calcium and phosphorus concentrations were not influenced (p > .05) by dietary calcium levels. The Ca-P ratio remained static in the whole body, vertebrae and serum. Broken-line regression analysis of data obtained on growth, mineralization and serum ALP activity against increasing levels of dietary calcium reflected the optimum calcium requirement between 5.77-6.81 g/kg diet.

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