OPTIMIZED NUTRIENT-CHLORAMPHENICOL AGAR MEDIUM FOR SELECTIVE ISOLATION OF PSEUDOMONAS FLUORESCENS

Commercially available medium for selective isolation of Pseudomonas are relatively expensive. Incorporation of chloramphenicol into nutrient agarhas been used in the selective isolation of Pseudomonasfluorescens. The aim of this study was to apply optimization in modification of nutrient-chloramphenicol agar medium (NCAM) so as to optimize it for selective isolation of Pseudomonas species. Water sample determined to contain fluorescent greenish-pigment producing bacteria (FGPB) was used for the optimization study. Medium component selected for the optimization process were pH, NaCl concentration (NC), and chloramphenicol concentration (CC), with value ranges of 7.0 – 8.2, 0.5 – 1.0 %, and 40 – 60 μg/ml, respectively. Fifteen media variations were derived from combination of these ranges using Box-Behnken design matrix, and were used in culturing the water sample for isolation of FGPB. Relationship between obtained proportions of bacterial populations that are FGPB and chosen media components were calculated using polynomial equation for three factors design. Prediction profiles derived from the equation showed that the highest proportion of bacterial populations that are FGPB was achievable at pH, NC, and CC combination values of 7.0, 0.2 %, and 60 μg/ml, respectively. NCAM with this optimized combination increased the efficiency of selectively isolating FGPB from the water sample from 0.008 to 0.011 %. Physicochemical/biochemical testing showed that the FGPBisolates were Pseudomonas fluorescens. It is concluded that NCAM with pH 7.0, NaCl concentration of 0.2 %, and chloramphenicol concentration of 60 μg/ml can be used for selective isolation of Pseudomonas fluorescens with optimum efficiency.