Department of Microbiology and Molecular Genetics , University of the Punjab , Quid-i-Azam Campus , Lahore , Pakistan.
Pol J Microbiol. 2020;69(2):1-11. doi: 10.33073/pjm-2020-016.
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.
本研究旨在探索基于 PCR 的筛选策略，以预测源自极端环境（巴基斯坦乔利斯坦沙漠）的选定菌株的抗菌生物合成潜力。通过分子和基于培养的筛选方法来确定生物合成潜力。通过 PCR 扩增、测序和随后的生物信息学方法，研究了选定菌株中的四个生物合成基因簇，包括、、cyp P450 羟化酶（）和糖肽基因。在 40 株选定的菌株中，有 33 株菌株具有基因，有 17 株菌株携带基因，有 4 株菌株发现具有基因，没有一株菌株携带基因。对 NR-1、NR-10、NR-14 和 NR-15 等菌株进行了抗真菌活性研究，以对抗、和 sp。对提取物进行了化学分析（TLC 和 HPLC-UV），并观察到独特的次生代谢物图谱。选定的菌株对真菌测试菌株表现出明显的抗真菌活性，抑菌圈直径分别达到 17、18 和 19mm。该研究表明，基于基因的筛选可以通过使用单一筛选过程成功地用于鉴定潜在的生物活性菌株。这种基于 PCR 的方法快速，可以用于在放线菌培养物中筛选和选择潜在的候选物。这种预选或菌株优先级排序可以减少选择潜在生物活性菌株所需的时间和精力，然后可以对其进行详细的化学分析。
