Members of the genus Streptomyces are rich source of medically important natural products. Industrial bioprocesses with Streptomyces species often face difficulties caused by high viscosity of the cultures, thus increasing energy demands of the bio-processes. The viscosity of the culture is influenced by strain morphology. In recent years, several proteins that effect mycelial growth and morphology in liquid cultures (including CslA, SsgA) were identified. In this work, we aimed to engineer strains with specific morphological phenotypes of reduced viscosity in liquid culture. Using bioinformatics approaches we identified ssgA and cslA gene homologues in S. clavuligerus ATTC 27064. We engineered mutant strains of S. clavuligerus with deleted genes ssgA and cslA by applying the CRISPR/Cas system. We also constructed strains where ssgA and cslA genes were over-expressed by in-trans chromosome integration. The phenotypes of the genetically modified strains were evaluated in liquid medium on the laboratory and pilot plant scale. Morphological differences between the genetically modified strains and the wild-type strain were determined by image analysis of hyphal length and level of branching (for SsgA) or by analyzing the size and structure of pellets (for CslA). Our results show that the strain with ssgA over-expressing had shorter hyphae with more branches. The viscosity and energy demand in the pilot scale fermentors with the culture over-expressing ssgA was much lower compared to the wild type strain culture. The strain with the cslA deletion formed smaller pellets with longer hyphae on the periphery of the pellets and a smaller “dead” core in the pellet center. Both strains with changed morphologyzapia therefore show potential for improvement of the bioprocess for clavulanic acid production.
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