Primary hemocyte culture of Penaeus monodon as an in vitro model for white spot syndrome virus titration, viral and immune related gene expression and cytotoxicity assays

Immortal cell lines have not yet been reported from Penaeus monodon, which delimits the prospects of investigating the associated viral pathogens especially white spot syndrome virus (WSSV). In this context, a method of developing primary hemocyte culture from this crustacean has been standardized by employing modified double strength Leibovitz-15 (L-15) growth medium supplemented with 2% glucose, MEM vitamins (1 ), tryptose phosphate broth (2.95 g l 1), 20% FBS, N-phenylthiourea (0.2 mM), 0.06 lgml 1 chloramphenicol, 100 lgml 1 streptomycin and 100 IU ml 1 penicillin and hemolymph drawn from shrimp grown under a bio-secured recirculating aquaculture system (RAS). In this medium the hemocytes remained viable up to 8 days. 5-Bromo-20-deoxyuridine (BrdU) labeling assay revealed its incorporation in 22 ± 7% of cells at 24 h. Susceptibility of the cells to WSSV was confirmed by immunofluoresence assay using a monoclonal antibody against 28 kDa envelope protein of WSSV. A convenient method for determining virus titer as MTT50/ml was standardized employing the primary hemocyte culture. Expression of viral genes and cellular immune genes were also investigated. The cell culture could be demonstrated for determining toxicity of a management chemical (benzalkonium chloride) by determining its IC50. The primary hemocyte culture could serve as a model for WSSV titration and viral and cellular immune related gene expression and also for investigations on cytotoxicity of aquaculture drugs and chemicals

Structural, Spectral, Biological and Electrochemical Studies Of Some 3d Transition Metal Complexes of O-N-S and N-N-S Donor Ligands

The primary aim of these investigations was to probe the spectroscopic, electrochemical, biological and single crystal X-ray diffraction studies of some selected transition metal complexes of 4N-monosubstituted thiosemicarbazones. Transition metal complexes with thiosemicarbazones exhibit a wide range of stereochemistries and possess potential biological activity. Metal complexes of thiosemicarbazones are proved to have improved pharmacological and therapeutic effects. The studies are conducted to bring about a fair understanding of the structure activity relationship and to develop certain effective and economical metal-based antimicrobial agents. Study showed that the thiosemicarbazones have antibacterial, antiviral and antiproliferative properties and hence used against tuberculosis, leprosy, psoriasis, rheumatism, trypanosomiasis and coccidiosis. Certain thiosemicarbazones showed a selective inhibition of HSV and HIV infections. The insolubility of most thiosemicarbazones in water causes difficulty in the oral administration in clinical practice. Transition metal complexes are found to have more activity than uncombined thiosemicarbazones. They exhibit a variety of denticity and can be varied by proper substitution. The stereochemistry assumed by the thiosemicarbazones during the coordination with transition metal ions depends on the factors such as preparative conditions and availability of additional bonding site in the ligand moiety and charge of the ligand. The resulting complexes exhibited a wide range of stereochemistries and have biomimic activity and potential application as sensors.

Structural, Spectral, Biological and Electrochemical Studies Of Some 3d Transition Metal Complexes of O-N-S and N-N-S Donor Ligands

The primary aim of these investigations was to probe the spectroscopic, electrochemical, biological and single crystal X-ray diffraction studies of some selected transition metal complexes of 4N-monosubstituted thiosemicarbazones. Transition metal complexes with thiosemicarbazones exhibit a wide range of stereochemistries and possess potential biological activity. Metal complexes of thiosemicarbazones are proved to have improved pharmacological and therapeutic effects. The studies are conducted to bring about a fair understanding of the structure activity relationship and to develop certain effective and economical metal-based antimicrobial agents. Study showed that the thiosemicarbazones have antibacterial, antiviral and antiproliferative properties and hence used against tuberculosis, leprosy, psoriasis, rheumatism, trypanosomiasis and coccidiosis. Certain thiosemicarbazones showed a selective inhibition of HSV and HIV infections. The insolubility of most thiosemicarbazones in water causes difficulty in the oral administration in clinical practice. Transition metal complexes are found to have more activity than uncombined thiosemicarbazones. They exhibit a variety of denticity and can be varied by proper substitution. The stereochemistry assumed by the thiosemicarbazones during the coordination with transition metal ions depends on the factors such as preparative conditions and availability of additional bonding site in the ligand moiety and charge of the ligand. The resulting complexes exhibited a wide range of stereochemistries and have biomimic activity and potential application as sensors

Biochemical and molecular investigations on Salmonella serovars from seafood

The present investigation was envisaged to determine the prevalence and identify the different Salmonella serovar in seafood from Cochin area. Though, the distribution of Salmonella serovars in different seafood samples of Cochin has been well documented, the present attempt was made to identify the different Salmonella serovars and determine its prevalence in various seafoods. First pan of this investigation involved the isolation and identification of Salmonella strains with the help of different conventional culture methods. The identified isolates were used for the further investigation i.e. serotyping, this provides the information about the prevalent serovars in seafood. The prevalent Salmonella strains have been further characterized based on the utilization of different sugars and amino acids, to identify the different biovar of a serovar.A major research gap was observed in molecular characterization of Salmonella in seafood. Though, previous investigations reported the large number of Salmonella serovars from food sources in India, yet, very few work has been reported regarding genetic characterization of Salmonella serovars associated with food. Second part of this thesis deals with different molecular fingerprint profiles of the Salmonella serovars from seafood. Various molecular typing methods such as plasmid profiling, characterization of virulence genes, PFGE, PCR- ribotyping, and ERIC—PCR have been used for the genetic characterization of Salmonella serovars.The conventional culture methods are mainly used for the identification of Salmonella in seafood and most of the investigations from India and abroad showed the usage of culture method for detection of Salmonella in seafood. Hence, development of indigenous, rapid molecular method is most desirable for screening of Salmonella in large number of seafood samples at a shorter time period. Final part of this study attempted to develop alternative, rapid molecular detection method for the detection of Salmonella in seafood. Rapid eight—hour PCR assay has been developed for detection of Salmonella in seafood. The performance of three different methods viz., culture, ELISA and PCR assays were evaluated for detection of Salmonella in seafood and the results were statistically analyzed. Presence of Salmonella cells in food and enviromnental has been reported low in number, hence, more sensitive method for enumeration of Salmonella in food sample need to be developed. A quantitative realtime PCR has been developed for detection of Salmonella in seafood. This method would be useful for quantitative detection of Salmonella in seafood.

Detection of sandal spike phytoplasma using immunological and molecular techniques

Spike disease in sandal is generally diagnosed by the manifestation of external symptoms. Attempts have been made to detect the diseased plants by determining the length/breadth ratio of leaves (lyengar, 1961) and histochemical tests using Mann's stain (Parthasarathi et al., 1966), Dienes' stain (Ananthapadmanabha et a/., 1973) aniline blue and Hoechst 33258 (Ghosh et a/., 1985, Rangaswamy, 1995). But most of these techniques are insensitive, indirect detection methods leading to misinterpretation of results. Moreover, to identify disease resistant sandal trees, highly sensitive techniques are needed to detect the presence of the pathogen. In sandal forests, several host plants of sandal like Zizyphus oenop/ea (Fig. 1.3) also exhibit the yellows type disease symptoms. Immunological and molecular assays have to be developed to confirm the presence of sandal spike phytoplasma in such hosts. The major objectives of the present work includes:In situ detection of sandal spike phytoplasma by epifluorescence microscopy and scanning electron microscopy.,Purification of sandal spike phytoplasma and production of polyclonal antibodies.,Amino acid and total protein estimation of sandal spike phytoplasma.,Immunological detection of sandal spike phytoplasma., Molecular detection of sandal spike phytoplasma.,Screening for phytoplasma in host plants of spike disease affected sandal using immunological and molecular techniques.

Growth response of phytoplankton exposed to industrial effluents in river Periyar

The thesis entitled Growth Response of Phytoplankton Exposed to Industrial Effluents in River Periyar. The present investigation has been conducted in two phases: field observation and algal assays. The monthly distribution of hydrographic features is represented graphically. The sampling year has been divided into three seasons: monsoon (June to September), postmonsoon (October to January) and premonsoon (February to May). The data were analysed using Student's t-test to find whether there was any significant difference between surface and bottom samples. The spatial variation of the variables was assessed by Page's L (trend) test (Ray Meddis, 1975). The standard procedure for algal toxicity test (Ward and Parrish, 1982) was followed throughout the study. Statistical analysis (Page's L (trend) test) showed that there was no significant difference in Secchi disc transparency between the stations. The field observations as well as the laboratory assays confirm that the rate of discharge in river Periyar during premonsoon is insufficient to effect dilution of wastewater received in the industrial zone.