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International audience; Meningococci spread via respiratory droplets, whereas the closely related gonococci are transmitted sexually. Several outbreaks of invasive meningococcal disease have been reported in Europe and the United States among men who have sex with men (MSM). We recently identified an outbreak of serogroup C meningococcal disease among MSM in Germany and France. In this study, genomic and proteomic techniques were used to analyze the outbreak isolates. In addition, genetically identical urethritis isolates were recovered from France and Germany and included in the analysis. Genome sequencing revealed that the isolates from the outbreak among MSM and from urethritis cases belonged to a clade within clonal complex 11. Proteome analysis showed they expressed nitrite reductase, enabling anaerobic growth as previously described for gonococci. Invasive isolates from MSM, but not urethritis isolates, further expressed functional human factor H binding protein associated with enhanced survival in a newly developed transgenic mouse model expressing human factor H, a complement regulatory protein. In conclusion, our data suggest that urethritis and outbreak isolates followed a joint adaptation route including adaption to the urogenital tract.

Respiratory infectious diseases are the third cause of worldwide death. The nasopharynx is the portal of entry and the ecological niche of many microorganisms, of which some are pathogenic to humans, such as Neisseria meningitidis and Moraxella catarrhalis. These microbes possess several surface structures that interact with the actors of the innate immune system. In our attempt to understand the past evolution of these bacteria and their adaption to the nasopharynx, we first studied differences in cell wall structure, one of the strongest immune-modulators. We were able to show that a modification of peptidoglycan (PG) composition (increased proportion of pentapeptides) and a cell shape change from rod to cocci had been selected for along the past evolution of N. meningitidis. Using genomic comparison across species, we correlated the emergence of the new cell shape (cocci) with the deletion, from the genome of N. meningitidis ancestor, of only one gene: yacF. Moreover, the reconstruction of this genetic deletion in a bacterium harboring the ancestral version of the locus together with the analysis of the PG structure, suggest that this gene is coordinating the transition from cell elongation to cell division. Accompanying the loss of yacF, the elongation machinery was also lost by several of the descendants leading to the change in the PG structure observed in N. meningitidis. Finally, the same evolution was observed for the ancestor of M. catarrhalis. This suggests a strong selection of these genetic events during the colonization of the nasopharynx. This selection may have been forced by the requirement of evolving permissive interaction with the immune system, the need to reduce the cellular surface exposed to immune attacks without reducing the intracellular storage capacity, or the necessity to better compete for adhesion to target cells., Author Summary The nasopharynx hosts an important microbial community that comprises some well-known pathogens such as Neisseria meningitidis and Moraxella catarrhalis. In some circumstances, it also represents the portal of entry of systemic infections such as septicemia and meningitis, or infections of the respiratory system, middle ear, eye, central nervous system and joints of humans, caused by N. meningitidis and M. catarrhalis, respectively. In this article, we demonstrated that both bacteria underwent a similar cell shape evolution that resulted in a transition from a bacillus to a coccus. This was consequently accompanied by a change, similar for both bacteria, in the structure of the PG, the major bacterial cell shape determinant and also a strongly recognized molecule by the immune system. In our efforts in understanding the evolutionary events that led to the cell shape transition in N. meningitidis, we identified two genetic deletion events required for the shape transition, i.e. of yacF (zapD) and the cell elongation machinery. Furthermore, we delineated the importance of YacF (ZapD) in the coordination of the cell elongation and division. Finally, we suggest that this transition was selected to reduce the cell surface sensible to immune attacks and to redistribute surface appendages, such as pili, to acquire new properties of cell adhesion or movement necessary for the proper colonization of the nasopharynx.

International audience; Peptidoglycan O-acetylation is a modification found in many bacteria. In Gram-positive pathogens, it contributes to virulence by conferring resistance to host lysozyme. However, in Gram-negative pathogens, its contribution to physiology and virulence is unknown. We examined the contribution of patA, patB and ape1 to peptidoglycan O-acetylation in the major human pathogen Neisseria meningitidis (Nm). Using genetic expression of all possible combinations of the three genes in Escherichia coli and Nm, we confirmed that PatA and PatB were required for PG O-acetylation, while ApeI removed the O-acetyl group. ApeI was active on all O-acetylated muropeptides produced by PatA and PatB during heterologous expression in E. coli and was also active on several PG structures in vitro. Interestingly, in Nm, ApeI was found to preferentially de-O-acetylate muropeptides with tripeptide stems (GM3), suggesting that its activity is highly regulated. Accordingly, de-O-acetylation of GM3 regulated glycan chain elongation and cell size. Additionally, the virulence of Nm lacking ApeI was drastically reduced suggesting that regulation of glycan chain length by O-acetylation contributes to bacterial fitness in the host. Altogether, our results suggest that ApeI represents an attractive target for new drug development.

International audience; Neisseria meningitidis is a human pathogenic bacterium responsible for life threatening and rapidly evolving invasive infections. Several bacterial virulence factors may play primordial roles during host-bacteria interactions. The meningococcal factor H binding protein, fHbp, interacts with the complement negative regulator, factor H (fH), to enhance meningococcal survival. fHbp is a major component in recombinant vaccines against meningococci that are under development. In 2010, we detected variations in fhbp gene during an outbreak provoked by serogroup C isolates belonging to the clonal complex, ST-11. We therefore explored 680 meningococcal isolates (88% of all invasive isolates in 2009 and 2010) by DNA sequencing of fhbp gene. The level of fHbp at the bacterial surface was determined by ELISA and flow cytometry using anti-fHbp antibodies. We also analyzed the interaction of fHbp with human fH as well as the deposition of C3b complement component. We observed important sequence diversity of fHbp in particular within regions known to interact with fH. The distribution of fhbp alleles differed among meningococcal serogroups and clonal complexes. This diversity affected directly binding of fH to fHbp and seemed to influence the deposition of the complement C3b component on the bacterial surface. However, bacterial killing by anti-fHbp antibodies was still achieved and required a minimum level of fHbp at the bacterial surface regardless the binding to fH or sequence diversity. These data have impacts on our understanding of the role of fHbp in meningococcal pathogenesis. They also provide data on the diversity of fhbp before the introduction of vaccines targeting fHbp and stress the need to include characterization of fHbp in typing schemes of meningococcal isolates.