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ThesisItem Open Access ACUTE TOXICITY AND BEHAVIORAL CHANGES OF THE BRACKISHWATER FISH (ETROPLUS SURATENSIS) EXPOSED TO ORGANOPHOSPHORUS PESTICIDE – MALATHION(KARNATAKA VETERINARY, ANIMAL AND FISHERIES SCIENCES UNIVERSITY, BIDAR, 2022) PAUNIKAR SUCHITA SURESH; LAKSHMIPATHIMalathion is one of the most commonly used pesticides in agriculture. This study was aimed to investigate the acute toxicity of malathion as an aquatic pollutant on the behaviour and haematological indices in pearl spot ( Etroplus suratensis). A static renewal bioassay was conducted to determine the lethal toxicity (LC50) of Malathion pesticide to Etroplus suratensis. The lethal toxicity of Malathion to fingerlings of Pearl spot exposed for 96 h was found to be 1.40 ppm. Study was conducted in accumulation phase. In accumulation phase the effect of sub lethal concentrations of Malathion (1/10th of LC50 i.e. 0.14 ppm and 1/5th of LC50 i.e. 0.28 ppm) was studied on physiology, haematology of Malathion for a period of 28 days. During accumulation phase physiological parameters such as oxygen consumption rate, ammonia excretion rate, food consumption rate, O:N ratio decreased significantly at both sub lethal concentrations of Malathion as compared to control group the behavioural changes to control group in a natural manner, i.e., they were active for feeding and alert to slightest of the disturbance with their well synchronized movements. While in toxic media, Pearl spot exhibited disrupted shoaling behaviour, localization to the bottom of the test chamber and independency ( spreading out) in swimming. In haematology study RBC, Hb, PCV decreased and WBC, MCHC, MCV & MCH increased significantly (p < 0.05) at both sub lethal concentrations of malathion as compared to control group.ThesisItem Open Access MOLECULAR BASED IDENTIFICATION OF VIRUSES BELONGING TO FAMILY IRIDOVIRIDAE AND ALLOHERPESVIRIDAE FROM CULTURED FISH(KARNATAKA VETERINARY, ANIMAL AND FISHERIES SCIENCES UNIVERSITY, BIDAR, 2022) SHIVAKUMAR; T. SURESHDiseases are of great concern in fish production and its expansion. Similar to other animals, fish can also suffer from different diseases. Disease is a main agent affecting fish mortality and a major constraint in aquaculture. The global economic loss caused by diseases is about 6.5 billion USD (2017). The habitat of the fish makes it more susceptible to the pathogens. Emerging and re-emerging viral diseases like ISKNV, RSIV, KHV, VNN, SDDV, TRBIV and LCDV are known to cause huge economical loss in wild and cultured. Although prevalence of more than 125 different viruses have been reported in fish culture around the globe including many Asian countries. In the present study, several ornamental and cultured fishes were screened for the viruses of family Iridoviridae and Alloherpesviridae. Moribund and clinically unhealthy samples were collected from different location of Karnataka and were checked for viral infection by using specific primers through conventional PCR method. A total of 130 samples were collected, among them 1 Gold gourami was found to be positive for the presence of ISKNV and 4 Asian Sea bass were positive for RSIV. Further confirmation was done by sequence analysis of positive samples, where sequenced ISKNV strain was more similar (99.74 - 99.88 %) to the Malaysia and Australian ISKNV strains and RSIV strains were more similar (98.77-99.65 %) to Indian and Japanese RSIV strains. Phylogenetic trees of ISKNV and RSIV revealed that our ISKNV strain and RSIV strains formed a clade with the reference ISKNV and RSIV strains. These results suggested, that the occurrence of iridoviral infection is more than the viruses of family Alloherpesviridae in ornamental and cultured fishes in Karnataka.ThesisItem Open Access PHENOTYPIC AND MOLECULAR BASED IDENTIFICATION OF MAJOR PARASITIC DISEASES IN CULTURED FISHES(KARNATAKA VETERINARY, ANIMAL AND FISHERIES SCIENCES UNIVERSITY, BIDAR, 2022) NIROSHA,; S.K.GIRISHAquaculture is the rapid growing food producing sector in India. Disease is one of the major limiting factors responsible for huge economic loss in aquaculture. Aquaculture is prone to various diseases from infectious agents like Virus, Bacteria and Parasites. The information available on the parasites is scant in India hence this study was undertaken by aiming for parasitic disease in aquaculture environments. A total 120 samples were screened for 7 major groups of parasites such as, Protozoans, Myxozoans, Microsporidians, Trematodes, Nematodes, Cestodes and Crustacean parasites. The identification was done on the basis of morphological level by using brightfield microscope and molecular level by following standard molecular diagnostic techniques. Out of 120 samples, 10 fishes were found to be infected with Lerneae, 4 fishes with Argulus, 4 fishes with Ichthyophthirius multifiliis, 3 fishes with Dactylogyrus, 1 fish with Camallanus and 1 fish with Tetrahymena. The prevalence of infection was found to be 8.33%, 3.33%. 3.33%, 2.55%, 0.83% and 0.83% in Lernaea, Argulus, Ichthyophthirius multifiliis, Dactylogyrus, Tetrahymena and Camallanus respectively. Identification of the parasites to their species level by using the microscopic method was highly difficult due to its high degree of morphological intraspecific diversity and interspecific similarities and there is a high risk of misidentification as different species. Therefore, in this study, molecular technique i.e polymerase chain reaction was employed to identify the parasites to their species level. The phylogenetic analysis revealed that Lernaea showed maximum homogeneity with L. cyprinaceae from Russia, Argulus with A. japonicus from Japan, Dactylogyrus with D. inexpectatus from USA, I. multifiliis with isolates from Canada, China and Philippines, Tetrahymena with T. pigmentosa from USA and Camallanus with C. cotti from China.ThesisItem Open Access FISH IMMUNE GENE EXPRESSION AGAINST RED SEABREAM IRIDOVIRUS (RSIV) INFECTION(KARNATAKA VETERINARY, ANIMAL AND FISHERIES SCIENCES UNIVERSITY, BIDAR, 2022) NIDHI, D. C.,; T .SURESHDiseases are one of the major constraints in aquaculture and are also the limiting factor for economic and socio-economic development. Emerging viral diseases like Red sea bream iridovirus (RSIV) are known to cause huge economical loss in wild and cultured fish. The virus mainly targets the spleen and kidney of the host and wears out the immune system, making host susceptible for secondary infections. In the present study SISS cell lines was used for virus propagation. Significant cytopathic effect (CPE) like multiple vacuolation, rounding of cells was observed in SISS-infected cells. Virus concentration in culture supernatant collected from infected cell lines was tested by virus titration and yielded viral titres of 106 TCID50/ml. These results indicate the suitability of the SISS cell lines for RSIV propagation. Further, quantitive real-time PCR (qRT-PCR) was carried out to analyze the host defense response in Asian sea bass (Lates calcarifer) challenged with RSIV. Major immune related genes encoding pattern recognition receptors like TLR-3, pro inflammatory cytokines (IFNγ1, TNFα, Casp-1 and IL-8), anti-inflammatory gene (IL-10) and antiviral proteins (MxA, viperin and perforin) were analyzed. There was an up regulation of all immune genes analyzed against RSIV infection at different time intervals in head kidney and spleen. The results revealed varying expression profiles and clear transcriptional activation of these immune related genes in kidney and spleen, which will contribute to better understand the pathogenesis and host defensive system in RSIV invasion.ThesisItem Open Access COMPARATIVE EVALUATION OF PHYTOREMEDIATION OF SELECTED AQUATIC WEEDS(KARNATAKA VETERINARY, ANIMAL AND FISHERIES SCIENCES UNIVERSITY, BIDAR., 2022) NARENDRA KUMAR MAURYA; shivakumar mThe pollution of surface and subsurface water is a severe unhealthy as a result of increased urbanisation and rapid population rise. Heavy metals such as copper (Cu), nickel (Ni), cadmium (Cd), chromium (Cr), lead (Pb), and argon (Ar) are found in waste water, causing toxicity in humans and the environment. Phytoremediation is the process of reducing pollutant concentrations in polluted soils, water, or air by using plants that can hold, eliminate metals, pesticides, solvents, explosives, crude oil and its derivatives, and other contaminants from the media in which they are present. A trial of 28 days was conducted on aquatic macrophytes such as Eichhornia crassipes, Azolla pinnata, Pistia stratiotes, and Lemna minor that offer significant potential for heavy metal clean up, such as Cadmium (Cd) and Lead (Pb). The experimental setup was done with 2, 4, 10 and 15 mg L-1 CdCl2 and Pb (NO3)2, which were separately harvested at 0, 7, 14, 21 and 28 days and the Cd and Pb concentration in the roots and shoot tissues were estimated by atomic absorption spectroscopy (AAS). The Cd metal accumulation in E. crassipes and P. stratiotes treated in 15 mg L-1 metal solution observed to be highest in roots and shoots at 21st day and 28th day as 52.02 mg kg-1 and 34.61 mg kg-1; 53.82 mg kg-1 and 46.83 mg kg-1 respectively. The Cd accumulation in A. pinnata and L. minor observed to be highest in 15 mg L-1 treated metal solution as 27.26 mg kg-1 and 18.72 mg kg-1 respectively. The Pb metal accumulation in E. crassipes and P. stratiotes treated in 15 mg L-1 metal solution observed to be highest in roots and shoots at 21st day and 28th day as 26.18 mg kg-1 and 34.61 mg kg-1; 73.02 mg kg-1 and 51.44 mg kg-1 respectively. The Pb accumulation in A. pinnata and L. minor observed to be highest in 15 mg L-1 treated metal solution as 28.19 mg kg-1 (28th day) and 18.40 mg kg-1 (14th day). The bioaccumulation factor of E. crassipes, A. pinnata, P. stratiotes, and L. minor of Cd and Pb metal were 78.95, 84.09, 178.53, 45.79 and 89.91, 77.36, 365.10, 242.67 respectively. The translocation factor of E. crassipes and P. stratiotes of Cd and Pb metal were 1.15 and 1.15; 1.04 and 0.94 respectively.ThesisItem Open Access STUDIES ON BIOACCUMULATION OF HEAVY METALS IN SELECTED CRABS OFF MANGALURU COAST(KARNATAKA VETERINARY, ANIMAL AND FISHERIES SCIENCES UNIVERSITY, BIDAR – 585 401, 2022) NANDAN Y.A.,; RAMCHANDRA NAIK A.T.Heavy metal contamination is hazardous and a severe problem in the marine environment because it is long-lasting and difficult to oxidise, degrade, remove or convert to less dangerous components by biological or chemical processes. The inshore waters off Mangaluru receive a substantial amount of pollution from nearby industries and companies, including wastes from oil refineries, fertilisers, pesticides and other contaminants. Decapods have been extensively researched as bioindicators of metal concentrations. The crab fishery is primarily supported by Portunus pelagicus and P. sanguinolentus, both of which have high export demand and are important component of the country's small-scale coastal fisheries. Commercially important crab populations are found in coastal areas with high levels of heavy metals. Considering the polluted state of marine ecosystem, an attempt has been made to know the concentrations of five heavy metals concentration in water, plankton, sediment and different organs of P. pelagicus and P. sanguinolentus. The order of heavy metal concentration in different organs of P. pelagicus was Cu>Zn>Ni>Pb>Cd in gills; Zn>Cu> Ni>Pb>Cd in muscles and Zn>Cu>Cd>Ni>Pb in hepatopancreas. However, in P. sanguinolentus, the heavy metal concentration was in the order of Cu>Zn>Ni>Cd>Pb in gills; Zn>Cu>Ni>Pb>Cd in muscles and Zn> Cu > Cd > Ni>Pb in hepatopancreas. Whereas abundance of heavy metal concentrations in the waters of study area was Ni>Zn>Pb>Cu>Cd. The abundance of heavy metal concentrations in the plankton of study area were in the sequence of Cu>Zn>Ni>Pb>Cd. While abundance of heavy metal concentrations in the sediment of study area was in the sequence of Zn>Cu>Ni>Pb>Cd.ThesisItem Open Access BIOACCUMULATION OF HEAVY METALS IN NAGAVARA LAKE, BENGALURU(KARNATAKA VETERINARY, ANIMAL AND FISHERIES SCIENCES UNIVERSITY, BIDAR, 2023) MANOJ M.,; SHIVAKUMAR MThe contamination of heavy metals in the aquatic environments is hazardous due to their toxicity, low solubility, persistence and biological accumulation. Lakes are important ecosystems of great value to humans and are prone to pollution from native and non-native sources also the various anthropogenic sources that interrupt the functioning of aquatic ecosystem. Fishes have been extensively examined as bioindicators of metal accumulation. Fish is an important source of food for humans, because of the pharmaceutical benefits of omega 3 polyunsaturated fatty acids, which is available in fish oil. But wide ranges of contaminants, including heavy metals are continuously introduced into the aquatic environments and fish from polluted waters seriously threaten human health due to the bioaccumulation of heavy metals in muscle and other tissues. On the whole pollution state of freshwater ecosystem, an experiment has been carried out to know the accumulation of four heavy metals concentration in water, sediment and different organs of Oreochromis mossambicus collected from Nagavara lake. Among the four metals lead (Pb) and cadmium (Cd) were not detected in any of the tissue samples of the fish. The abundance of heavy metal concentrations in the waters of study area was Zn>Cu> Pb>Cd. Whereas the abundance of heavy metal concentrations in the sediment of study area was in the sequence of Cu>Zn>Pb>Cd. The order of heavy metal accumulation in different organs of O. mossambicus was Zn>Cu>Pb-Cd in gill tissue, Zn>Cu>Pb-Cd in the muscle tissue, Zn>Cu>Pb-Cd in kidney and Zn>Cu>Pb-Cd in the liver. Key words: Nagavara lake, Bioaccumulation, Heavy metalsThesisItem Open Access PROTECTIVE EFFICACY OF FORMALIN-INACTIVATED VACCINE AGAINST RED SEABREAM IRIDOVIRUS (RSIV) IN ASIAN SEABASS (LATES CALCARIFER)(KARNATAKA VETERINARY, ANIMAL AND FISHERIES SCIENCES UNIVERSITY, BIDAR, 2022) MAHESH, A. S.,; SURESH TRed seabream iridovirus (RSIV) was recently reported in India from cage cultures of Asian seabass causing high mortality and economic losses. Currently, no antiviral therapy or vaccines are available for the control of this pathogen. Hence, the present study was aimed to develop formalin inactivated vaccine to investigate the vaccine efficacy in Asian seabass. The virus used in the study was initially isolated from diseased Asian seabass and stored at -80oC. Confirmation of the RSIV was done by extracting the virus DNA from spleen and kidney tissues of infected fish. The results of the PCR assay showed a 568bp targeting DNA polymerase gene of the RSIV. Further, RSIV was propagated using cell lines under in-vitro conditions using the virus extracted from the spleen and kidney tissues of infected fish was inoculated onto SISK cell lines. Cytopathic effect (CPE) was observed in RSIV infected cells from 2nd post infection, which included rounding and enlargement of cells followed by cell death on the 7th day. The harvested virus stock was quantified by endpoint dilution assay and found to contain 106.3 TCID50/ml. Inactivation of the virus was performed using 0.1% of formalin for 10 days in 4°C and confirmed by negative CPE in SISK cells after 2-3 blind passages and then formalin inactivated virus (FIV) was stored in 4°C until use. Further, efficacy of FIV was carried out against RSIV in Asian seabass. Vaccinated fish did not exhibit typical clinical symptoms of RSIV infection and mortality is less, therefore the FIV was safer to use. After 21dpv fish were challenged with virulent RSIV-LC strain and after 14dpc relative percentage survival (RPS) was calculated. Asian seabass immunized with FIV showed high level of protection with RPS value of 86.6%. Therefore, further investigations for assess its efficacy against RSIV challenge in field trail and commercialization are require.ThesisItem Open Access BIOACCUMULATION AND DEPURATION OF PESTICIDE ACEPHATE BASED ON HISTOLOGICAL CHANGES IN COMMON CARP (Cyprinus carpio) (LINNAEUS, 1758)(KARNATAKA VETERINARY, ANIMAL AND FISHERIES SCIENCES UNIVERSITY, BIDAR, 2022) BHOOLESHWARI,; SHIVAKUMAR M.The extensive use of pesticides has led to adverse effects to human health as well as other non-target species and the environment. Pesticides can thus act both indirectly to decrease the food supply, and directly by increasing concentration in the food chain itself on the ultimate predator. Only around 0.1 percent of pesticides are believed to reach the intended organisms, with the rest polluting the environment and causing environmental harm. They can be transferred through food chain, to fish and ultimately reach to humans. Acephate, an organic compound pesticide and its metabolite methamidophos are common organophosphorus insecticide used for crop protection. High uses of acephate and methamidophos have induced health issues and environmental pollution. Fishes are often at the front line of manifesting toxicological effects when exposed to pesticides in aquatic ecosystems. The present study was carried out to investigate the bioaccumulation and depuration of pesticide acephate based on histological changes of Cyprinus carpio. Acute and Chronic toxicity of acephate were analysed by histological changes and physiological changes during exposure. The lethal toxicity of Acephate to fingerlings of Cyprinus carpio exposed for 96h was found to be 850.41ppm. Study was conducted in accumulation phase (sublethal toxicity studies) for the 28days and after completion of accumulation phase fishes were exposed to depuration phase (exposed in clear water) for the 28days. The effect of sublethal concentrations of Acephate ( 1/5th of LC50 i.e.170.082 ppm and 1/10th of LC50 i.e. 85.041 ppm ) was studied on physiological, histological of acephate for a period of 28 days. In accumulation phase physiological parameters such as oxygen consumption rate, ammonia-N excretion rate, food consumption rate, Oxygen:Nitrogen ratio had 98 decreased significantly and at the end of the depuration phase it has slightly increased at both sublethal concentrations of acephate as compared to control group. Damages in vital organs of the fish tissue i.e. gills, liver and kidney resulted that the use of acephate is toxic for the fishes even in the lower concentrations. After 28 days of accumulation, the fishes are exposed to freshwater which showed slowly recover in tissues structure. By this, it can be concluded that fishes can remove accumulated toxic content from body if we provide cleaner environment. Hence, the present investigation recommends the utility of physiological and histological responses as diagnostic tools to assess toxic effects in fishes. All the results were statistically significant at P<0.05. KEY WORDS:Acephate, Lethal toxicity, Cyprinus carpio, Bioaccumulation, Depuration, Physiology and Histology .
