PCR-based Screening Approach: A Rapid Method to Detect the Biosynthetic Potential of Antimicrobials in Actinobacterial Strains.

This study aimed to investigate the PCR-based screening strategy for the prediction of the antimicrobial biosynthetic potential of the selected strains originated from an extreme environment (Cholistan Desert, Pakistan). The biosynthetic potential was determined by using both molecular and culture-dependent screening approaches. The four biosynthetic genes clusters, including the , , cyp P450 hydroxylase (), and glycopeptide genes, were investigated in the selected strains by PCR amplification, sequencing, and by subsequent bioinformatics approaches. Among the 40 selected strains, 33 strains possessed the gene, 17 strains carried the gene, four strains were found to have the gene, and none of the strain carried gene. The strains including NR-1, NR-10, NR-14, and NR-15 were investigated for antifungal activity against , , and sp. The extracts were analyzed for chemical profiling (TLC and HPLC-UV), and a unique pattern of secondary metabolites was observed. The selected strains exhibited pronounced antifungal activity against the fungal test strains with the zone of inhibition up to 17, 18, and 19 mm, respectively. The study depicts that gene-based screening can be successfully applied to identify potentially bioactive strains by usin a single screening process. This PCR-based approach is rapid and can be used for sorting out and selecting the potential candidate among actinobacterial culture collections. Such a preselection or strain prioritization consequently decreases the time and efforts required for selecting the potential bioactive strain, which then can be subjected to the detailed chemical analysis. This study aimed to investigate the PCR-based screening strategy for the prediction of the antimicrobial biosynthetic potential of the selected strains originated from an extreme environment (Cholistan Desert, Pakistan). The biosynthetic potential was determined by using both molecular and culture-dependent screening approaches. The four biosynthetic genes clusters, including the , , cyp P450 hydroxylase (), and glycopeptide genes, were investigated in the selected strains by PCR amplification, sequencing, and by subsequent bioinformatics approaches. Among the 40 selected strains, 33 strains possessed the gene, 17 strains carried the gene, four strains were found to have the gene, and none of the strain carried gene. The strains including NR-1, NR-10, NR-14, and NR-15 were investigated for antifungal activity against , , and sp. The extracts were analyzed for chemical profiling (TLC and HPLC-UV), and a unique pattern of secondary metabolites was observed. The selected strains exhibited pronounced antifungal activity against the fungal test strains with the zone of inhibition up to 17, 18, and 19 mm, respectively. The study depicts that gene-based screening can be successfully applied to identify potentially bioactive strains by usin a single screening process. This PCR-based approach is rapid and can be used for sorting out and selecting the potential candidate among actinobacterial culture collections. Such a preselection or strain prioritization consequently decreases the time and efforts required for selecting the potential bioactive strain, which then can be subjected to the detailed chemical analysis.