Occurrence of functional glutamine/glutamate-acid resistance systems in Yersinia ruckeri

Background and Aims Acid resistance (AR) systems protect neutroalophilic bacteria to pH < 2.5. The most potent AR systems, extensively characterized in Escherichia coli, depend either on glutamate (Glu; AR2) or on glutamine (Gln; AR2_Q). AR2_Q rely on the glutaminase GlsA1 and GadC, an antiporter importing L-glutamine and exporting either glutamate or GABA. The latter is produced when L-glutamate is decarboxylated by glutamate decarboxylase (GadA/GadB), Notably, the genes coding for the homologues of GadB, GadC and GlsA1 are present in Yersinia ruckeri, a fish pathogen causing enteric red mouth disease and affecting species of commercial importance. Thus, AR may contribute to Y. ruckeri pathogenicity by protecting it from acid stress. Methods Y. ruckeri was grown at 25°C, pH 5.0, and spent medium assayed for GABA content using GABase. AR was assayed by diluting stationary phase culture into minimal medium at pH 2.2 with/without Gln or Glu. After 2 hours survivors were detected by plating on LB agar. Y. ruckeri gadBC and glsA1 were cloned into plasmid pBBR and the recombinant plasmids used to transform E. coli gadABC and glsA deletion mutants. Results AR assays in the presence of Gln or Glu provided evidence that both AR systems are operative in Y. ruckeri. GABA export was also detected. AR on E. coli acid-sensitive mutant strains was re-established by complementation in trans with the relevant Y. ruckeri genes. Conclusions For the first time evidence is provided that Y. ruckeri possesses fully functional AR2 and AR2_Q systems. This knowledge will shed light on the pathogenic process of the disease.