The fish ( ) was used as the source for probiotics. 46 bacterial isolates were obtained from the skin, gills, guts and intestines of the guppy, (collected from a government model fish farm in Kottayam, India). Of the above isolated strains, four isolates were selected based on their inhibitory spectrum against five indicator strains, 1739, 3906, 2495, 1271 and 1424 (standard cultures collected from Microbial Type Culture Collection (MTCC) Chandigarh, India). Among the resulting isolates, two were gram-positive cocci, namely MBTU-PB2 and MBTU-PB3 and belong to the genus . The other two were gram-negative rods, namely MBTU-PB1 and MBTU-PB4, of the genera and , respectively. The basic probiotic characteristics of these isolates such as the production of bacteriocin like inhibitory substances (BLIS), antibiotic sensitivities and growth profiles were also determined. The above four isolated strains exhibited different antagonisms than the five indicator strains. During incubation, the antibacterial activity gradually increased in the inhibition zone and was influenced by the lag period ( ) and doubling time. The lag periods for most of the four selected strains were shorter than those of the indicator strains and the isolates had different growth rates ( ) than the indicator strains. All four isolates produced BLIS, however, the strains had different BLIS activities against the indicator strains. Treatment of the neutralized cell free supernatants of the selected isolates with proteases eliminated or reduced the BLIS activity, suggesting a proteinaceous nature of the inhibitory compounds. Further, the optimum BLIS activity was observed at neutral pH after 18 h of incubation. The antibiotic sensitivity assay revealed that the isolates were susceptible to routinely used antibiotics, whereas the plasmid profiles showed that the plasmids had no role in the antagonistic properties of the four isolated strains. The results showed that the isolates could be a promising source for biocontrol agents in aquacultures.