Mortalities due to pathogenic bacteria are a major problem in aquaculture, especially in larval rearing systems. Use of antibiotics to overcome this problem is not an option any more due to the increasing antibiotic resistance among pathogens. The present study aims to understand the diversity of bacteria with antagonistic properties in the tropical estuarine habitats of Cochin, located along the southwest coast of India, and to use them as an alternative to antibiotics in aquaculture. Among the 4,870 isolates screened, approximately 1 % showed significant antibacterial activity against six common aquaculture pathogens belonging to the genera Aeromonas and Vibrio. The antagonistic bacteria were identified as Bacillus (81 %) and Pseudomonas (19 %) using biochemical and 16S rRNA gene sequence homology. The isolates showing stable and higher levels of antibacterial activity were subjected to enzymatic expression profile, antibiotic resistance pattern and abiotic stress tolerance assays. As a result, five Pseudomonas spp. and four Bacillus spp., were identified as promising antagonistic isolates that could be exploited as probionts or microbial products (MP's), to control bacterial diseases in aquaculture rearing systems.

Documenting bacteria present in healthy individuals forms the first step in understanding the effects of microbial manipulation in aquaculture systems. Among the commensal microflora, gut microbiota has attracted extensive attention owing to their role in host metabolism and health maintenance. Basic knowledge on normal gut microbes within a particular host species is thus essential to determine how successfully these microbes can be manipulated and engineered for sustainable aquaculture systems. In spite of the good aquaculture potential of Mangrove red snapper, Lutjanus argentimaculatus, the information on microbial communities associated with the gut of this fish, and their contribution towards digestive efficiency and disease resistance is scarce. Therefore, an attempt was made to elucidate the abundance and diversity of cultivable gut microbes of wild caught L. argentimaculatus along with their digestive exoenzyme profiles and prohibitory effect against fish pathogens. Results on abundance showed similar gut bacterial loads as that of other marine fish imposing the less contribution of microflora to the volume of gut materials in fish. Eleven distinct bacterial species including two proposed novel vibrios were identified. An incidental observation of Morganella morganii throughout samples is an alarming signal, emphasizing the need for immediate de-gutting to avoid histamine intoxication. Abundance of digestive enzyme producers and excellent enzymatic potential of some isolates suggested the contribution of digestive enzymes may supplement to the symbiosis between gut flora and host and the information is of interest to aquaculture nutritionists/commercial industries. Interestingly, some isolates demonstrated estimable co-aggregation with aquatic pathogens, indicating their involvement in disease resistance and the results correlated well with gut microbial diversity. These findings highlight the significant role of gut microbes towards nutritional physiology and disease resistance of this aquaculture candidate in natural ecosystem. The culturable microbiota profiles of wild fish generated in the study can be applied for measuring the quality of husbandry routines in aquaculture facility of this marine fish. Overall, the present study fetches insights on the gut microbiome of healthy L. argentimaculatus which forms a platform for follow-up studies. The study may also help in the development of "functional" fish feeds for L. argentimaculatus. The investigation also demonstrated some potential digestive enzyme-producing isolates having probiotic applications in commercial aquaculture. Documenting the bacteria present in healthy individuals is the first step

Pseudomonas aeruginosa is a ubiquitous Gram-negative bacterium having a versatile metabolic potential and great ecological and clinical significance. The geographical distribution of P. aeruginosahas revealed the existence of an unbiased genetic arrangement in terrestrial isolates. In contrast, there are very few reports about P. aeruginosa strains from marine environments. The present work was aimed at studying the distribution of P. aeruginosa in coastal waters along the Indian Peninsula and understanding the environmental influence on genotypic, metabolic and phenotypic characteristics by comparing marine and clinical isolates. Of the 785 marine isolates obtained on selective media, only 32 (~4.1%) were identified as P. aeruginosa, based on their fatty acid methyl ester profiles. A low Euclidian distance value (< 2.5) obtained from chemotaxonomic analysis suggested that all the environmental (coastal and marine) isolates originated from a single species. While UPGMA analyses of AP-PCR and phenotypic profiles separated the environmental and clinical isolates, fatty acid biotyping showed overlapping between most clinical and environmental isolates. Our study revealed the genetic diversity among different environmental isolates of P. aeruginosa. While biogeographical separation was not evident based solely on phenotypic and metabolic typing, genomic and metatranscriptomic studies are more likely to show differences between these isolates. Thus, newer and more insightful methods are required to understand the ecological distribution of this complex group of bacteria.

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Antimicrobial compounds from the natural source have gained greater relevance because of their wide spectrum of possible applications, especially in the aquaculture industry where pathogenic threat and antibacterial resistance are serious concerns. In this regard, Pseudomonas aeruginosa MBTDCMFRI Ps04 (P. aeruginosa Ps04) strain isolated from the tropical estuarine habitats of Cochin was evaluated for its antibacterial potential against major aquaculture pathogens. The physiological conditions for the maximum production of the active metabolite were also optimized. An activity-guided approach was employed further to isolate and characterize the secondary metabolite responsible for the inhibitory potential. It was found that the cell free supernatant (CFS) of P. aeruginosa Ps04 exhibited strong antibacterial activity against major aquaculture pathogens belonging to the genus Vibrio and Aeromonas and retained its potential even at 30% (v/v) dilution. The highest antibacterial activity was detected from 3rd day culture, grown in glycerol alanine media (1% each) as carbon and nitrogen source, respectively, at 30°C, pH 7.0 and at a salinity of 20 parts per thousand (ppt). The activity of the antagonistic principle was found to be stable against variations in pH (pH 2-pH 12), temperature (up to 120°C) and enzymatic treatments. Bioassay-guided purification followed by spectroscopic characterization of active fractions of P. aeruginosa Ps04 revealed that the compound 4-Hydroxy-11-methylpentacyclo [11.8.0.0 2,3 .0 11, 12 .0 16,17 ]henicosa-1,3,5,8(9),17-penten-14-one is responsible for its major antibacterial activity. The results of this study indicated that P. aeruginosa Ps04 has beneficial antibacterial properties which could be used in developing novel antimicrobial therapeutics against a variety of aquaculture pathogens.

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