Search

Abstract Citrus greening, also known as Huanglongbing (HLB), is a devastating citrus plant disease caused predominantly by Liberibacter asiaticus. While nearly all Liberibacter species remain uncultured, here we used the culturable L. crescens BT-1 as a model to examine physiological changes in response to the variable osmotic conditions and nutrient availability encountered within the citrus host. Similarly, physiological responses to changes in growth temperature and dimethyl sulfoxide concentrations were also examined, due to their use in many of the currently employed therapies to control the spread of HLB. Sublethal heat stress was found to induce the expression of genes related to tryptophan biosynthesis, while repressing the expression of ribosomal proteins. Osmotic stress induces expression of transcriptional regulators involved in expression of extracellular structures, while repressing the biosynthesis of fatty acids and aromatic amino acids. The effects of osmotic stress were further evaluated by quantifying biofilm formation of L. crescens in presence of increasing sucrose concentrations at different stages of biofilm formation, where sucrose-induced osmotic stress delayed initial cell attachment while enhancing long-term biofilm viability. Our findings revealed that exposure to osmotic stress is a significant contributing factor to the long term survival of L. crescens and, possibly, to the pathogenicity of other Liberibacter species.

Abstract Background The aggressive spread of Liberibacter asiaticus, a bacterium closely associated with citrus greening, has given rise to an acute crisis in the citrus industry, making it imperative to expand the scientific knowledge base regarding L. asiaticus. Despite several endeavors to culture L. asiaticus, this bacterium has yet to be maintained in axenic culture, rendering identification and analysis of potential treatment targets challenging. Accordingly, a thorough understanding of biological mechanisms involved in the citrus host-microbe relationship is critical as a means of directing the search for future treatment targets. In this study, we evaluate the biochemical characteristics of CLIBASIA_01175, renamed LdtP (L,D-transpeptidase). Surrogate strains were used to evaluate its potential biological significance in gram-negative bacteria. A strain of E. coli carrying quintuple knock-outs of all genes encoding L,D-transpeptidases was utilized to demonstrate the activity of L. asiaticus LdtP. Results This complementation study demonstrated the periplasmic localization of mature LdtP and provided evidence for the biological role of LdtP in peptidoglycan modification. Further investigation highlighted the role of LdtP as a periplasmic esterase involved in modification of the lipid A moiety of the lipopolysaccharide. This work described, for the first time, an enzyme of the L,D-transpeptidase family with moonlighting enzyme activity directed to the modification of the bacterial cell wall and LPS. Conclusions Taken together, the data indicates that LdtP is a novel protein involved in an alternative pathway for modification of the bacterial cell, potentially affording L. asiaticus a means to survive within the host.