Disruption of rsmA gene of Pectobacterium carotovorum subsp. carotovorum LY34 and effect on pathogenicity

Abstract

The rsmA gene was cloned from soft-rot bacterium Pectobacterium carotovorum subsp. carotovorum LY34 (Pcc LY34), and its role in pathogenicity was investigated by marker exchange mutagenesis. From a cosmid library of Pcc LY34 genomic DNA, a positive clone carrying the rsmA gene was selected, and the gene was cloned by polymerase chain reaction (PCR) amplification. The gene is 186 bp in size and encodes a protein of 62 amino acids with a predicted molecular mass of 6,839 Da. The calculated pI of the RsmA is 8.16. The phylogenetic tree showed that the RsmA of Pcc LY34 appeared genetically identical to the CsrA of Pectobacterium atrosepticum SCRI1043 (100% identity) and similar to the CsrA of Yersinia pestis KIM10+ (98.3%). The gene was disrupted by the Kmr gene, and the cells became mutated (i.e., RsmA− mutant). The pathogenicity test revealed that the disease rating of the RsmA− mutant only differed slightly from that of the wild type on a slice of potato tuber and a Chinese cabbage stalk. These results suggest that RsmA is not an essential factor for the pathogenicity of Pcc LY34 and that the rsmA gene of Pcc LY34 is not completely derepressed in the RsmA− mutant for virulence-related genes, contrary to the results of Erwinia carotovora subsp. carotovora RsmA− mutant, which proved hypervirulent for celery petioles. These results showed that the microenvironmental conditions of the host and/or strain of pathogen are important for the coordination of virulence gene expression.