Abstract
Normally, cultivation of most Streptomyces species for the purpose of mutant-screening is an arduous process, needing a large amount of material. Moreover, the quantitative analysis of corresponding metabolites is done using HPLC, which is both expensive and time-consuming, and hence not ideal for screening of large numbers of isolates. A novel two-step high-throughput screening strategy has been designed to efficiently identify high-titer virginiamycin-producing mutants of Streptomyces virginiae. The high-throughput screening strategy described here can be used to cultivate a large number of mutant S. virginiae strains in 24-well deep-well microtiter plates and rapidly evaluate a large number of strains with varying virginiamycin titers. This is done based on the cylinder-plate bioassay method using Micrococcus luteus CMCC (B) 28001 as an indicator without compromising the accuracy. After one round of microwave-mediated mutagenesis, S. virginiae strain number 74 identified from nearly 1244 isolates, was shown to have the highest titer. The titer of virginiamycin was approximately 61.26 ± 2.53 mg/L, which is nearly three times compared to the value for the parent strain. Thus, this approach holds promise in facilitating the screening of high-titer virginiamycin producing strains.