Loss-of-Function Mutations in the Dpp and Opp Permeases Render Erwinia amylovora Resistant to Kasugamycin and Blasticidin S

Abstract

Extensive use of the antibiotic streptomycin to control fire blight disease of apples and pears, caused by the enterobacterial plant pathogen Erwinia amylovora, leads to the development of streptomycin-resistant strains in the United States and elsewhere. Kasugamycin (Ksg) has been permitted to be used as an alternative or replacement to control this serious bacterial disease. In this study, we investigated the role of two major peptide ATP-binding cassette transporter systems in E. amylovora, the dipeptide permease (Dpp) and oligopeptide permease (Opp), in conferring sensitivity to Ksg and blasticidin S (BcS). Minimum inhibitory concentration and spot dilution assays showed that the dpp deletion mutants exhibited slightly enhanced resistance to Ksg in rich medium, whereas the opp mutant exhibited slightly enhanced resistance to Ksg in minimal medium and BcS in rich medium. Deletion of both dpp and opp conferred a higher level of resistance to Ksg in both rich and minimal media, whereas deletion of opp alone was sufficient to confer high level of resistance to BcS in minimal medium. In addition, bioinformatic analysis combined with reverse transcription-quantitative polymerase chain reaction showed that the Rcs phosphorelay system negatively regulates opp expression and the rcsB mutant was more sensitive to both Ksg and BcS in minimal medium as compared with the wild type. An electrophoresis motility shift assay further confirmed the direct binding of the RcsA/RcsB proteins to the promoter region of the opp operon. However, neither the Dpp nor the Opp permeases contributed to disease progress on immature pears, hypersensitive response on tobacco leaves, or exopolysaccharide amylovoran production. These results suggested that Ksg and BcS employ the Dpp and Opp permeases to enter E. amylovora cells and the Dpp and Opp permeases act synergistically for illicit transport of antibiotics.

Prospective of Essential Oils of the Genus Mentha as Biopesticides: A Review

“…Additionally, there are several strains of bacterial pathogens which have developed resistance against commercial bactericides due to the change in climate. Plant bacteriologists reported that plant pathogenic bacteria readily acquire resistance to copper bactericides and streptomycin ( Stall and Thayer, 1962 ; Marco and Stall, 1983 ; Cuppels and Elmhirst, 1999 ). Therefore, researchers looked for the alternatives that would resolve the resistance problems, be less harmful and would be effective to control the bacterial pathogens.…”

Homologues of the RNA binding protein RsmA in Pseudomonas syringae pv. tomato DC3000 exhibit distinct binding affinities with non‐coding small RNAs and have distinct roles in virulence

“…It was compromised in motility and had reduced exopolysaccharide (EPS) amylovoran and expression of type III secretion system (T3SS) genes. In addition, CsrA in E. amylovora may indirectly affected uptakes of antibiotics through the Rcs system (Ancona et al , ; Ge et al , ; Lee et al , ). In Pectobacterium carotovorum , overexpression of rsmA inhibited motility, biofilm formation, EPS and secondary metabolite, antibiotics and pigment productions (Mukherjee et al , ).…”

The stringent response regulator (p) ppGpp mediates virulence gene expression and survival in Erwinia amylovora

“…Kim and colleagues suggested that peptide transporters provide peptides containing essential amino acids for both survival and infection in Salmonella [53]. Previous study showed that both Opp and Dpp are hijacked for importing antibiotics, but are dispensable for virulence in E. amylovora [54]. Taken together, these results suggested that (p) ppGpp positively regulates peptide uptake systems in WT, but negatively regulates genes involved in amino acid uptake systems and amino acid biosynthesis in the HMM environment.…”

“…On the other hand, (p) ppGpp regulates gene expression indirectly by sigma factor competition [2]. High concentration of (p) ppGpp inhibits sigma factor σ 70 , which allows more free RNAPs interact with alternative sigma factors, including σ 54 , to activate genes in response to stresses [3,8]. Moreover, (p) ppGpp also influences gene expression other than through RNAP [9] by directly down-regulating stable RNA (rRNA and tRNA) and genes related to transcription and translation, while directly upregulating amino acid biosynthesis genes [1,3,8,10,11].…”