Isolation of polyhydroxyalkanoates-producing bacteria using a combination of phenotypic and genotypic approach.

AIMS: To develop an efficient approach using a combination of phenotypic and genotypic methods for isolation of environmental bacteria that produce mid-chain-length polyhydroxyalkanoates (mcl-PHAs). METHODS AND
RESULTS: A viable-colony staining method using Nile red was used to screen for PHA-producing bacteria followed by a polymerase chain reaction (PCR) screen using primers to amplify the partial nucleic acid sequence of the phaC1 synthase gene for confirmation. Microbes containing lipophilic storage compounds isolated from environmental samples could readily be detected by the colony staining method. They were further examined by Sudan Black staining to highlight the inclusions inside the cells. These isolates were subsequently subjected to PCR analysis. As a result, more than a hundred strains were identified as PHA-positive isolates from this screening approach.
CONCLUSIONS: These results conclusively demonstrate that environmental bacterial strains able to accumulate the PHAs could readily be obtained by this screening method. SIGNIFICANCE AND IMPACT OF THE STUDY: We propose a polyphasic approach using a combination of phenotypic and genotypic screening method to rapidly screen and identify bacteria able to produce significant amounts of mcl-PHAs from environment. This approach can be adopted as a rapid screen for micro-organisms able to accumulate PHAs to be used for potential manufacture and other industrial applications.