Sample preparation methods for isolation of Mycobacterium spp. from cultured fish and environmental samples

Fish mycobacteriosis' is a problem to which more than 150 species offish are susceptible. In order to isolate non-pathogenic and fish pathogenic m.ycobacterial species from environm.ental samples, three different methods were evaluated. Shaking and membrane filtration methods were adopted to retrieve maxim.um acidfast strains from fish and environmental samples. Decontamination with 4% sodium hydroxide facilitated the isolation of acid-fast bacteria in the selective media, killing all the contam.inants like heterotrophs and saprophytes. Centrifugation procedure at 4000 r.p. mfor 20 minutes eliminated the contaminants from the sample and allowed only acid-fast bacilli to grow in the selective media. Centrifugation was carried out twice with distilled water. Peizer TB and LJ slopes worked well for selective isolation of fish pathogenic Mycobacteria/rom the centrifugated samples. It is believed that all the three sample preparation methods will be useful tool to study the fastidious fish pathogenic Mycobacteria from environmental samples