Abstract
Recent research has shown that pathogen virulence can be altered by exposure to antibiotics, even when the growth rate is unaffected. Investigating this phenomenon provides new insights into understanding the virulence mechanisms of bacterial pathogens. This study investigates the phenotypic and transcriptomic responses of the rice pathogenic bacterium Acidovorax avenae subsp. avenae (Aaa) strain RS-1 to ß-lactam antibiotics especially Ampicillin (Amp). Our results indicate that exposure to Amp does not influence bacterial growth and biofilm formation, but alters the virulence, colonization capacity, composition of extracellular polymeric substances and secretion of Type VI secretion system (T6SS) effector Hcp. This attenuation in virulence is linked to unique or differential expression of known virulence-associated genes based on genome-wide transcriptomic analysis. The reliability of expression data generated by RNA-Seq was verified with quantitative real-time PCR of 21 selected T6SS genes, where significant down-regulation in expression of hcp gene, corresponding to the reduction in secretion of Hcp, was observed under exposure to Amp. Hcp is highlighted as a potential target for Amp, with similar changes observed in virulence-associated phenotypes between exposure to Amp and mutation of hcp gene. In addition, Hcp secretion is reduced in knockout mutants of 4 differentially expressed T6SS genes.
Highlights
Acidovorax avenae subsp. avenae (Aaa) strain RS-1 has been reported to have strong virulence to rice plants[1,2,3,4,5]
Growth of strain RS-1 was differentially affected by the three different ß-lactam antibiotics
The virulence of strain RS-1 was attenuated under exposure to ß-lactam antibiotics especially Amp, which caused a 190.36%, 81.74% and 129.91% increase in emergence, root length and plant height, respectively, compared to the pathogen control (Fig. 2; Table S2)
Summary
Acidovorax avenae subsp. avenae (Aaa) strain RS-1 has been reported to have strong virulence to rice plants[1,2,3,4,5]. A preliminary study revealed the naturally occurring ß-lactam-resistance of strain RS-1, which makes it necessary to examine the pathogenic consequence of β -lactam antibiotics exposure in this rice bacterial pathogen. A number of studies have been performed to explore the underlying causes for the changes in virulence of human and animal bacterial pathogens when exposure to antibiotics[17,18,19]. Little information was available about genome-wide analysis of gene expression under exposure to antibiotics, which will provide a comprehensive insight into molecular mechanisms of virulence in bacterial pathogen. Phenotypic changes and the genome-wide transcriptomic response of strain RS-1 were examined following exposure to ß-lactam antibiotics. Secreted levels of the T6SS effector, Hcp, were measured by western blot analysis and ELISA, and its interaction with other T6SS components was investigated by using both a bacterial two-hybrid assay and a GST pull-down assay
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