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A Pseudomonas sp PS-102 recovered from Muttukkadu brackish water lagoon, situated south of Chennai, showed significant activity against a number of shrimp pathogenic vibrios. Out of the 112 isolates of bacterial pathogens comprising Vibrio harveyi, V. vulnificus, V. parahaemolyticus, V. alginolyticus, V. fluvialis, and Aeromonas spp, 73% were inhibited in vitro by the cell-free culture supernatant of Pseudomonas sp PS-102 isolate. The organism produced yellowish fluorescent pigment on King's B medium, hydrolysed starch and protein, and produced 36.4% siderophore units by CAS assay and 32 μM of catechol siderophores as estimated by Arnow's assay. The PS-102 isolate showed wide ranging environmental tolerance with, temperatures from 25 to 40°C, pH from 6 to 8, salinity from 0 to 36 ppt, while the antagonistic activity peaked in cultures grown at 30°C, pH 8.0 and at 5 ppt saline conditions. The antagonistic activity of the culture supernatant was evident even at 30% v / v dilution against V. harveyi. The preliminary studies on the nature of the antibacterial action indicated that the antagonistic principle as heat stable and resistant to proteolytic, lipolytic and amylolytic enzymes. Pseudomonas sp PS 102 was found to be safe to shrimp when PL-9 stage were challenged at 10 7 CFU ml − 1 and by intramuscular injection into of ∼5 g sub-adults shrimp at 10 5 to 10 8 CFU. Further, its safety in a mammalian system, tested by its pathogenicity to mice, was also determined and its LD 50 to BALB/c mice was found to be 10 9 CFU. The results of this study indicated that the organism Pseudomonas sp PS 102 could be employed as a potential probiont in shrimp and prawn aquaculture systems for management and control of bacterial infections.

A continuous cell line (SISK) from kidney of sea bass, Lates calcarifer, has been established and characterized. The cell line was maintained in Leibovitz' L-15 supplemented with 15% fetal bovine serum. This cell line has been subcultured more than 100 times over a period of 2 years. The SISK cell line consists of predominantly of epithelial-like cells. These cells showed strong positive for epithelial markers such as cytokeratin 19 and pancytokeratin. The cells were able to grow at temperature between 25 and 32°C with optimum temperature of 28°C. The growth rate of sea bass kidney cells increased as the FBS proportion increased from 2% to 20% at 28°C with optimum growth at the concentrations of 15% or 20% FBS. The distribution of chromosome number was 30 to 56 with a modal peak at 48 chromosomes. Polymerase chain reaction products were obtained from SISK cells and tissues of sea bass with primer sets of microsatellite markers of sea bass. Five fish viruses were tested on this cell line to determine its susceptibility to these viruses and this was found to be susceptible to MABV NC1 and nodavirus, and the infection was confirmed by RT-PCR and CPE. This suggests that the SISK cell line has good potential for the isolation of various fish viruses. This cell line has been shown to be susceptible to bacterial extracellular products. The SISK cell line is the India's first marine fish cell line.

Aeromonas spp. are ubiquitous aquatic organisms, associated with multitude of diseases in several species of animals, including fishes and humans. In the present study, water samples from two ornamental fish culture systems were analyzed for the presence of Aeromonas. Nutrient agar was used for Aeromonas isolation, and colonies (60 No) were identified through biochemical characterization. Seven clusters could be generated based on phenotypic characters, analyzed by the programme NTSYSpc, Version 2.02i, and identified as: Aeromonas caviae (33.3%), A. jandaei (38.3%) and A. veronii biovar sobria (28.3%). The strains isolated produced highly active hydrolytic enzymes, haemolytic activity and slime formation in varying proportions. The isolates were also tested for the enterotoxin genes (act, alt and ast), haemolytic toxins (hlyA and aerA), involved in type 3 secretion system (TTSS: ascV, aexT, aopP, aopO, ascF-ascG, and aopH), and glycerophospholipid-cholesterol acyltransferase (gcat). All isolates were found to be associated with at least one virulent gene. Moreover, they were resistant to frequently used antibiotics for human infections. The study demonstrates the pathogenic potential of Aeromonas, associated with ornamental fish culture systems suggesting the emerging threat to public health.

The cichlid oscar Astronotus ocellatus has worldwide commercial value in the pet fish industry because of its early maturation, relatively high fecundity, ability to identify its caretaker and also to alter colouration amongst conspecifics. Pathogenic strains of Aeromonas veronii resistant to multiple antibiotics were isolated from A. ocellatus individuals showing signs of infectious abdominal dropsy. The moribund fish showed haemorrhage in all internal organs, and pure cultures could be obtained from the abdominal fluid. The isolates recovered were biochemically identified as A. veronii biovar sobria and genetically confirmed as A. veronii based on 16S rRNA gene sequence analysis (GenBank accession no. FJ573179). The RAPD profile using 3 primers (OPA-3, OPA-4 and OPD-20) generated similar banding patterns for all isolates. They displayed cytotoxic and haemolytic activity and produced several exoenzymes which were responsible for the pathogenic potential of the isolates. In the representative isolate MCCB 137, virulence genes such as enterotoxin act, haemolytic toxin aerA, type 3 secretion genes such as aexT, ascV and ascF-ascG, and gcat (glycerophospholipidcholesterol acyltransferase) could be amplified. MCCB 137 exhibited a 50% lethal dose (LD 50 ) of 10 5.071 colony-forming units ml -1 in goldfish and could be subsequently recovered from lesions as well as from the internal organs. This is the first description of a virulent A. veronii from oscar. KEY WORDS: Aeromonas veronii · Oscar · Astronotus ocellatus · Virulence · Ornamental fishesResale or republication not permitted without written consent of the publisher

This is the first report of an isolate of V. harveyi which utilizes glycerol as the sole carbon source for PHB production with high biomass yield. This isolate could be of use as candidate species for commercial PHB production using glycerol as the feed stock or as source of genes for recombinant PHB production or for synthetic biology.

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 gl⁻¹), 20% FBS, N-phenylthiourea (0.2 mM), 0.06 μg ml⁻¹ chloramphenicol, 100 μg ml⁻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-2'-deoxyuridine (BrdU) labeling assay revealed its incorporation in 22 ± 7% of cells at 24h. Susceptibility of the cells to WSSV was confirmed by immunofluorescence assay using a monoclonal antibody against 28 kDa envelope protein of WSSV. A convenient method for determining virus titer as MTT(50)/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 IC(50). 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.

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