Peptidoglycan fragment release and NOD activation by commensal Neisseria species from humans and other animals.

Neisseria gonorrhoeae, a human restricted pathogen, releases inflammatory peptidoglycan (PG) fragments that contribute to the pathophysiology of pelvic inflammatory disease. The genus Neisseria is also home to multiple species of human- or animal-associated Neisseria that form part of the normal microbiota. Here we characterized PG release from the human-associated nonpathogenic species Neisseria lactamica and Neisseria mucosa and animal-associated Neisseria from macaques and wild mice. An N. mucosa strain and an N. lactamica strain were found to release limited amounts of the proinflammatory monomeric PG fragments. However, a single amino acid difference in the PG fragment permease AmpG resulted in increased PG fragment release in a second N. lactamica strain examined. Neisseria isolated from macaques also showed substantial release of PG monomers. The mouse colonizer Neisseria musculi exhibited PG fragment release similar to that seen in N. gonorrhoeae with PG monomers being the predominant fragments released. All the human-associated species were able to stimulate NOD1 and NOD2 responses. N. musculi was a poor inducer of mouse NOD1, but ldcA mutation increased this response. The ability to genetically manipulate N. musculi and examine effects of different PG fragments or differing amounts of PG fragments during mouse colonization will lead to a better understanding of the roles of PG in Neisseria infections. Overall, we found that only some nonpathogenic Neisseria have diminished release of proinflammatory PG fragments, and there are differences even within a species as to types and amounts of PG fragments released.

Mutation of mltG increases peptidoglycan fragment release, cell size, and antibiotic susceptibility in Neisseria gonorrhoeae.

IMPORTANCE: Neisseria gonorrhoeae is unusual in that the bacteria release larger amounts of cell wall material as they grow as compared to related bacteria, and the released cell wall fragments induce inflammation that leads to tissue damage in infected people. The study of MltG revealed the importance of this enzyme for controlling cell wall growth, cell wall fragment production, and bacterial cell size and suggests a role for MltG in a cell wall synthesis and degradation complex. The increased antibiotic sensitivities of mltG mutants suggest that an antimicrobial drug inhibiting MltG would be useful in combination therapy to restore the sensitivity of the bacteria to cell wall targeting antibiotics to which the bacteria are currently resistant.

Mutation of mltG increases peptidoglycan fragment release, cell size, and antibiotic susceptibility in Neisseria gonorrhoeae.

Infection with the Gram-negative species Neisseria gonorrhoeae leads to inflammation that is responsible for the disease symptoms of gonococcal urethritis, cervicitis, and pelvic inflammatory disease. During growth these bacteria release significant amounts of peptidoglycan (PG) fragments which elicit inflammatory responses in the human host. To better understand the mechanisms involved in PG synthesis and breakdown in N. gonorrhoeae, we characterized the effects of mutation of mltG. MltG has been identified in other bacterial species as a terminase that stops PG strand growth by cleaving the growing glycan. Mutation of mltG in N. gonorrhoeae did not affect bacterial growth rate but resulted in increased PG turnover, more cells of large size, decreased autolysis under non-growth conditions, and increased sensitivity to antibiotics that affect PG crosslinking. An mltG mutant released greatly increased amounts of PG monomers, PG dimers, and larger oligomers. In the mltG background, mutation of either ltgA or ltgD, encoding the lytic transglycosylases responsible for PG monomer liberation, resulted in wild-type levels of PG monomer release. Bacterial two-hybrid assays identified positive interactions of MltG with synthetic penicillin-binding proteins PBP1 and PBP2 and the PG-degrading endopeptidase PBP4 (PbpG). These data are consistent with MltG acting as a terminase in N. gonorrhoeae and suggest that absence of MltG activity results in excessive PG growth and extra PG in the sacculus that must be degraded by lytic transglycosylases including LtgA and LtgD. Furthermore, absence of MltG causes a cell wall defect that is manifested as large cell size and antibiotic sensitivity.

Cervical cerclage for singleton pregnant patients on vaginal progesterone with progressive cervical shortening.

BACKGROUND: Premature cervical ripening plays a significant role in spontaneous preterm birth. Vaginal progesterone is the recommended treatment in singleton pregnancy with incidental short cervix. There is lack of evidence on whether it is beneficial to reinforce the cervix with cerclage when the cervical length becomes progressively shortened <10 mm while on vaginal progesterone. OBJECTIVE: Our aims are to determine whether cerclage with vaginal progesterone will: (1) reduce the overall spontaneous preterm birth rate, (2) prolong pregnancy latency, and (3) improve neonatal outcomes compared to vaginal progesterone alone. STUDY DESIGN: This was a retrospective cohort study at the University of Illinois at Chicago of all women with singleton pregnancy on vaginal progesterone for incidental short cervix, cervical length <20 mm. Only those with progressive cervical length shortening <10 mm who delivered at the University of Illinois at Chicago from January 2013 through December 2016 were included. The decision to perform cerclage was based on individual physician preference. Demographic data; information on serial cervical length status; medical, obstetric, and social history; cerclage vs no cerclage; and neonatal outcomes were compared. RESULTS: A total of 310 women with incidental short cervix on vaginal progesterone were identified, and of these, 75 had progressive shortening cervical length <10 mm and met inclusion criteria. Among the women with extremely shortened cervical length <10 mm, 36 women (48%) had cervical cerclage plus vaginal progesterone, and 39 women (52%) continued on vaginal progesterone alone. The baseline characteristics, mean cervical length (5.06 vs 5.52 mm), and mean gestational age at diagnosis of extreme short cervix (21.5 vs 21.3 weeks) were similar between women who received cerclage vs those who did not, respectively. The mean gestational age at delivery was significantly greater for those with cerclage (34 weeks and 3 days vs 27 weeks and 2 days; P < .001). The rate of spontaneous preterm birth at <37, 35, 32, 28, and 24 weeks were significantly lower in the cerclage group: 44.1% vs 84.2%, 38.2% vs 81.6%, 23.5% vs 78.9%, 14.7% vs 63.2%, and 11.8% vs 39.5%, respectively. The rate of spontaneous preterm birth <37 weeks remained significant after controlling for confounders (relative risk, 0.11; 95% confidence interval, 0.03-0.41; P < .001). The average pregnancy latency was 14 weeks in the cerclage combined with vaginal progesterone group compared to vaginal progesterone alone group. Neonatal intensive care unit admission and development of respiratory distress syndrome were significantly lower in the cerclage group compared to vaginal progesterone alone group: 13 (36.1%) vs 23 (65.7%) (relative risk, 0.55; 95% confidence interval, 0.34-0.90; P = .018) and 8 (22.2%) vs 17 (43.6%) (relative risk, 0.59; 95% confidence interval, 0.29-0.90; P = .027), respectively. Neonates of women with cerclage were also significantly less likely to develop necrotizing enterocolitis or experience neonatal death. CONCLUSION: Our study showed that cerclage plus vaginal progesterone in women with extremely shortened cervix significantly decreased overall spontaneous preterm birth rates, prolonged pregnancy latency by 2-fold, and decreased the overall neonatal morbidity and mortality.