UDP-N-acetylglucosamine 1-carboxyvinyl-transferase from Acinetobacter calcoaceticus

Abstract We have analyzed the sequence downstream of rpoN from Zcinetobacter calcoaceticus and identified an open reading frame encoding a protein with high similarity to UDP- N -acetylgucosamine 1-carboxyvinyl-transferase (MurZ). Multicopy plasmids encoding this enzyme conferred phosphomycin resistance to A. calcoaceticus . The polar effect of a rpoN mutation on the phosphomycin resistance level suggests that murZ is, in part, cotranscribed with rpoN . These observations confirm that A. calcoaceticus represents the first exceptin from a conserved genetic context of rpoN observed in several other Gram-negative bacteria.     

Novel persistence genes in Pseudomonas aeruginosa identified by high-throughput screening

Salmonella enterica polymyxin B (PM) resistance is modulated mainly by substitutions of the acyl chains and the phosphate groups on the lipid A moiety of lipopolysaccharide. These modifications are mediated by genes under the control of the PmrA/PmrB and PhoP/PhoQ two-component regulatory systems. In this study, a deletion in the gene encoding the alternative σ⁵⁴ factor, rpoN , was shown to increase PM resistance without affecting protamine sensitivity. The results presented here showed that the increased polymyxin resistance observed in the Δ rpoN mutant occurs through a PmrA/PhoP-independent pathway. Downregulation of one or more genes belonging to the RpoN regulon may provide an additional mechanism of defence against membrane-permeabilizing antimicrobial peptides that helps the pathogen to survive in different environments.     

Characterization of an rpoN mutant of Mesorhizobium ciceri

AIMS: To study the genetic basis of C(4)-dicarboxylate transport (Dct) in relation to symbiotic nitrogen fixation in Mesorhizobium ciceri. METHODS AND RESULTS: A Tn5-induced mutant strain (TL16) of M. ciceri, unable to grow on C(4)-dicarboxylates, was isolated from the wild-type strain TAL 620. The mutant lacked activities of the enzymes, which use C(4)-dicarboxylates as substrate. The sequencing of the 3.2kb EcoRI fragment, which was the site of Tn5 insertion, revealed three complete and two partial open reading frames. In the mutant, Tn5 interrupted the rpoN gene, of which only one copy was there. Complementation and biochemical studies suggest that the M. ciceri rpoN activity is required for C(4)-Dct, maturation of bacteroids and symbiotic nitrogen fixation. The fine structure of the ineffective nodules produced by TL16 on Cicer arietinum L changed in comparison with those produced by the wild type. CONCLUSIONS: The mutant strain TL16 suffered a disruption in the rpoN gene. Only one copy of rpoN gene is present in M. ciceri. The mutation abolishes Dct activity. It additionally abolishes the symbiotic nitrogen fixation activity of the bacteroids in the nodules. SIGNIFICANCE AND IMPACT OF THE STUDY: This first document in M. ciceri shows that a functional rpoN gene is essential for the transport of dicarboxylic acids and symbiotic nitrogen fixation.     

LuxO controls extracellular protease, haemolytic activities and siderophore production in fish pathogen Vibrio alginolyticus

Aims: To characterize the luxO gene in fish pathogen Vibrio alginolyticus MVP01 and investigate its roles in regulation of extracellular products (ECP) and siderophore production. Methods and Results: The luxO gene was cloned from V. alginolyticus MVP01. Genetic analysis revealed that it encoded a protein with high similarity to other LuxO homologues. The luxO in-frame deletion mutant and rpoN null mutant were constructed with suicide plasmids. We demonstrated that sole deletion in LuxO increased the secretion of extracellular protease and haemolytic products, but decreased siderophore production for V. alginolyticus MVP01. Mutants with null rpoN displayed significantly enhanced protease level and siderophore production while notable reduction in haemolytic activities of ECP. Conclusions: Vibrio alginolyticus harbours functional luxO gene that regulates the secretion of extracellular protease and haemolytic materials as well as siderophore production in either σ⁵⁴ dependent or independent manners. Significance and Impact of the Study: The current study demonstrated that V. alginolyticus MVP01 produces extracellular protease and haemolytic activity material as well as siderophore, which may be characteristics of the virulence of the strain. Revelations that secretion of these products is under the regulation of LuxO and σ⁵⁴ as well as the potential quorum sensing systems in V. alginolyticus MVP01 will expedite the understanding of vibriosis pathogenesis.