Alkaline protease from a non-toxigenic mangrove isolate of Vibrio sp. V26 with potential application in animal cell culture

Vibrio sp. V26 isolated from mangrove sediment showed 98 % similarity to 16S rRNA gene of Vibrio cholerae, V. mimicus, V. albensis and uncultured clones of Vibrio. Phenotypically also it resembled both V. cholerae and V. mimicus.Serogrouping, virulence associated gene profiling, hydrophobicity, and adherence pattern clearly pointed towards the non—toxigenic nature of Vibrio sp. V26. Purification and characterization of the enzyme revealed that it was moderately thermoactive, nonhemagglutinating alkaline metalloprotease with a molecular mass of 32 kDa. The application of alkaline protease from Vibrio sp. V26 (APV26) in sub culturing cell lines (HEp-2, HeLa and RTG-2) and dissociation of animal tissue (chick embryo) for primary cell culture were investigated. The time required for dissociation of cells as well as the viable cell yield obtained by while administeringAPV26 and trypsin were compared. Investigations revealed that the alkaline protease of Vibrio sp. V26 has the potential to be used in animal cell culture for subculturing cell lines and dissociation of animal tissue for the development of primary cell cultures, which has not been reported earlier among metalloproteases of Vibrios.

Development of Cell Culture Systems from Selected Species of Fish and Prawns

The present work deals with the development of primary cell culture and diploid cell lines from two fishes, such as Poecilia reticulata and Clarias gariepinus. The greatest difficulty experienced was the avoidance of bacterial and fungi contamination. Three types of cell cultures are commonly developed, primary cell culture, diploid cell lines and heteroploid cell lines. Primary cell culture obtained from the animal tissues that have been cultivated in vitro for the first time. They are characterized by the same chromosome number as parent tissue, cultivated in vitro for the first time, have wide range of virus susceptibility, usually not malignant, six chromatin retarded and do not grow as suspension cultures. Diploid cell lines arise from a primary cell culture at the time of subculturing. Diploid cell lines commercially used in virology are W1-38 (human embryonic lung), W1-26 (human embryonic lung) and HEX (Human embryonic kidney). Heteroploid cell lines have been subcultivated with less than 75% of the cells in the population having a diploid chromosome constitution. Tissue cultures have been extensively used in biomedical research. The main applications are in three areas, Karyological studies, Identification and study of hereditary metabolic disorders and Somatic cell genetics. Other applications are in virology and host-parasite relationships. In this study an attempt was made to preserve the ovarian tissue at low temperature in the presence of cryoprotectants so that the tissue can be retrieved at any time and a cell culture could be developed.

Cell Culture Systems from Penaeus monodon: Development and Application

National Centre for Aquatic Animal Health, Cochin University of Science and Technology

“Development of lymphoid cell culture system from Penaeus monodon and molecular approaches for its transformation

Unveiling the molecular and regulatory mechanisms that prevent in vitro transformation in shrimp remains elusive in the development of continuous cell lines, with an arduous history of over 25 years (Jayesh et al., 2012). Despite presenting challenges to researchers in developing a cell line, the billion dollar aquaculture industry is under viral threat. In addition, the regulatory mechanisms that prevent in vitro transformation and carcinoma in shrimps might provide new leads for the development of anti-ageing and anti-cancer interventions in human (Vogt, 2011) and in higher vertebrates. This highlights the importance of developing shrimp cell lines, to bring out effective prophylactics against shrimp viruses and for understanding the mechanism that induce cancer and ageing in human.. Advances in molecular biology and various gene transfer technologies for immortalization of cells have resulted in the development of hundreds of cell lines from insects and mammals, but yet not a single cell line has been developed from shrimp and other marine invertebrates. With this backdrop, the research described in this thesis attempted to develop molecular tools for induced in vitro transformation in lymphoid cells from Penaeus monodon and for the development of continuous cell lines using conventional and novel technologies to address the problems at cellular and molecular level.

Reproductive physiology of indian species of the genus perna (family mytilidae)

This thesis deals with the reproductive physiology of the two species of Indian sea mussels, namely the brown mussel perna indica and the green mussel p viridis .The major aspects of the study include ecophysiology of reproduction linking up the animal reproductive cycle of the animals with the ecological conditions of the natural mussel beds, biochemical and histochemical changes associated with reproduction and neurosecretory cycle in synchrony with the reproductive cycle. Some basic studies on gamete morphology and certain aspects of gamete physiology have been taken up. The experimental work deals with the influence of different feeding levels on gamatogenesis and maturation and the effect of ganglia ablation on spawning. The material for the investigation on perna indica were collected from the natural mussel beds at Vizhinjam near Trivandrum and on p.viridic from Elathur, near Calicut. The period of observation extended from October 1981 to December 1982.

Studies on the physiology of moulting in the penaeid prawn, penaeius indicius h. Milne edwards

Penaeid prawns form the most economically significant group in the marine and brackishwater fishery resources of India. This particular group contributes about 62% of the total prawn landings of the country. At present prawns have assumed an important place especially as a commodity supporting an export trade of sizable magnitude. Considerable interest has been shown in the last decade to increase prawn production through various culture practices, mainly due to the high demand for good quality prawns for export coupled with the stagnant and even depleting nature of marine catches. Available informations suggest that among the 15 species of shrimps and prawns occurring in Indian waters, which are deemed suitable for aquaculture, the Indian white prawn §.indicus is identified as one of the most important commercial species. Considering the increasing importance as an accepted species for prawn culture, £.indicus was selected for the present study. In the life history of prawns, moulting is an important event, which enables the animal to achieve growth. This dynamic physiological event continues through out the life span ofthe prawn, linking almost all biological activities with this process. Hence, a good knowledge pertaining to the physiology of moulting is imperative to understand the growth process. This knowledge will be of great use in the scientific prawn farming, so as to achieve high prawn production.