A mutant of Brucella suis bearing a Tn5 insertion in norD, the last gene of the operon norEFCBQD, encoding nitric oxide reductase, was unable to survive under anaerobic denitrifying conditions. The norD strain exhibited attenuated multiplication within nitric oxide-producing murine macrophages and rapid elimination in mice, hence demonstrating that norD is essential for Brucella virulence.The gram-negative intracellular pathogen Brucella sp. is the causative agent of brucellosis, which is transmissible to humans from domestic animal species that are infected with B. abortus, B. suis, or B. melitensis.Multiplication inside macrophages allows Brucella to be carried throughout the host organism and to colonize specific organs. The pathogenicity of brucellae and chronicity are based on the ability of the pathogen to adapt to the environmental conditions that it encounters in its replicative niche (18) and to avoid the killing mechanisms within macrophages (3,8,10,14).Intensive studies were performed at the genetic level to investigate the factors that are essential for the adaptation of Brucella to the intracellular conditions (5). Analysis of the intramacrophagic virulome (17) confirmed that the type IV secretion system encoded by virB (19) is, to date, the main virulence factor of B. suis. It also revealed that the B. suis replicative niche is characterized by low levels of nutrients and oxygen. The latter parameter was previously observed in the phagosomes of stimulated macrophages (13). Complete genome sequences (6,12,20) have revealed that Brucella possesses all of the genes that are necessary for a complete denitrification pathway resulting in the reduction of nitrate to nitrogen. Genes encoding the four reductases Nar (nitrate reductase), Nir (nitrite reductase), Nor (nitric oxide reductase), and Nos (nitrous oxide reductase) constitute a "denitrification island" in B. suis which shares numerous similarities with that of Sinorhizobium meliloti (21); both organisms belong to the ␣-subclass of the proteobacteria. The respiratory system could allow Brucella to survive under very low oxygen tension, using nitrogen oxides as terminal electron acceptors. Bacteria may also take advantage of denitrification to cope with nitric oxide (NO) production in the macrophages during the innate response against infection. NO production by infected macrophages is a major defense system in control of Brucella infection in mice (9, 16) and possibly, although more controversial, in human infection, as revealed by the use of human macrophages transfected with inducible NO synthase (7). Despite the low levels of NO that were released by human macrophages, the nitric oxide reductase of Neisseria meningitidis was found to confer intracellular resistance to NO and allowed its utilization, resulting in the optimal survival of this bacterium in nasopharyngeal mucosa (22). In fact, during an infection of murine macrophages producing high levels of NO, B. abortus displayed increased late survival (23). The authors of that work suggested th...

No abstract

Background. Clostridium difficile infection is often considered to result from recent acquisition of a C difficile isolate in a healthcare setting. However, C difficile spores can persist for long periods of time, suggesting a potentially large community environmental reservoir. The objectives of this study were to assess community environmental contamination of toxigenic C difficile and to assess strain distribution in environmental versus clinical isolates.Methods.From 2013 to 2015, we collected community environmental swabs from homes and public areas in Houston, Texas to assess C difficile contamination. All positive isolates were tested for C difficile toxins A and B, ribotyped, and compared with clinical C difficile isolates obtained from hospitalized patients in Houston healthcare settings.Results.A total of 2538 environmental samples were collected over the study period. These included samples obtained from homes (n = 1079), parks (n = 491), chain stores (n = 225), fast food restaurants (n = 123), other commercial stores (n = 172), and hospitals (n = 448). Overall, 418 environmental isolates grew toxigenic C difficile (16.5%; P < .001) most commonly from parks (24.6%), followed by homes (17.1%), hospitals (16.5%), commercial stores (8.1%), chain stores (7.6%), and fast food restaurants (6.5%). A similar distribution of ribotypes was observed between clinical and environmental isolates with the exception that ribotype 027 was more common in clinical isolates compared with environmental isolates (P < .001).Conclusions.We identified a high prevalence of toxigenic C difficile from community environs that were similar ribotypes to clinical isolates. These findings suggest that interventions beyond isolation of symptomatic patients should be targeted for prevention of C difficile infection.

ObjectivesRidinilazole (SMT19969) is a narrow-spectrum, non-absorbable antimicrobial with activity against Clostridium difficile undergoing clinical trials. The purpose of this study was to assess the pharmacological activity of ridinilazole and assess the effects on cell morphology.MethodsAntibiotic killing curves were performed using the epidemic C. difficile ribotype 027 strain, R20291, using supra-MIC (4× and 40×) and sub-MIC (0.125×, 0.25× and 0.5×) concentrations of ridinilazole. Following exposure, C. difficile cells were collected for cfu counts, toxin A and B production, and morphological changes using scanning electron and fluorescence microscopy. Human intestinal cells (Caco-2) were co-incubated with ridinilazole-treated C. difficile growth medium to determine the effects on host inflammatory response (IL-8).ResultsTreatment at supra-MIC concentrations (4× and 40× MIC) of ridinilazole resulted in a significant reduction in vegetative cells over 72 h (4 log difference, P < 0.01) compared with controls without inducing spore formation. These results correlated with a 75% decrease in toxin A production (P < 0.05) and a 96% decrease in toxin B production (P < 0.05). At sub-MIC levels (0.5× MIC), toxin A production was reduced by 91% (P < 0.01) and toxin B production was reduced by 100% (P < 0.001), which resulted in a 74% reduction in IL-8 release compared with controls (P < 0.05). Sub-MIC (0.5×)-treated cells formed filamentous structures ∼10-fold longer than control cells. Following fluorescence labelling, the cell septum was not forming in sub-MIC-treated cells, yet the DNA was dividing.ConclusionsRidinilazole had robust killing effects on C. difficile that significantly reduced toxin production and attenuated the inflammatory response. Ridinilazole also elicited significant cell division effects suggesting a potential mechanism of action.

No abstract