The effect of the hybrid histidine kinase gene inactivation on cell morphology in Azospirillum baldaniorum and Azospirillum brasilense

Objective. The aim of the work is to test the hypothesis about the effect of the chromosome-located HSHK-RR gene encoding a hybrid multisensory histidine kinase on the morphology of bacteria of the genus Azospirillum. Material and methods. The strains A. baldaniorum Sp245, Lafˉ mutant Sp245.1062 (Sp245-HSHKDRR-Km), A. brasilense Sp7 and Sp7.10030 (Sp7-HSHKDRR-Km) were used in the work. Bacterial morphology and motility were characterized by electron microscopy. The relative level of gene transcription was determined by real-time PCR. Results. The hybrid multisensory histidine kinase (HSHK-RR) genes in Sp245 (AZOBR_150176) and Sp7 (AMK58_10030) strains are 95% homologous. Replacement of the internal fragment of the HSHK-RR gene with the kanamycin resistance gene resulted resulted in changes in the morphology and motility of cells in semi-liquid agar in the Sp245-HSHKDRR-Km and Sp7-HSHKDRR-Km mutants. The mutant cells were shorter than the cells of the strains Sp245 and Sp7 and moved more slowly in semi-liquid agar, forming colonies of a significantly smaller diameter. Both mutants retained the synthesis of a functioning polar flagellum (Fla). However, in the case of Sp7-HSHKDRR-Km, up to 45% of the cells synthesized rare or single lateral flagella (Laf) on agar media, whereas in Sp245-HSHKDRR-Km, no more than 15% of such cells were present. Conclusion. It was established that Laf biogenesis is under multistage control, including a two-component signal transduction system involving HSHK-RR encoded by chromosomal genes in Sp245 and Sp7, respectively, strains AZOBR_150176 and AMK58_10030. Phenotype analysis of mutants for the AZOBR_150176 and AMK58_10030 genes showed that the studied HSHK-RR modulates the motility of Sp245 and Sp7 under mechanical stress, participating in the transcription control of the flagellin laf1 genes and components of the export system of lateral flagella assembly.