Commensal Neisseria Species Research Articles

Carriage and antimicrobial susceptibility of commensal Neisseria species from the human oropharynx

Commensal Neisseria (Nc) mainly occupy the oropharynx of humans and animals. These organisms do not typically cause disease; however, they can act as a reservoir for antimicrobial resistance genes that can be acquired by pathogenic Neisseria species. This study characterised the carriage and antimicrobial susceptibility profiles of Nc from the oropharynx of 50 participants. Carriage prevalence of Nc species was 86% with 66% of participants colonised with more than one isolate. Isolates were identified by MALDI-ToF and the most common species was N. subflava (61.4%). Minimum inhibitory concentrations (MICs) to penicillin, ceftriaxone, ciprofloxacin, azithromycin, tetracycline, and gentamicin were determined by agar dilution and E-test was used for cefixime. Using Ng CLSI/EUCAST guidelines, Nc resistance rates were above the WHO threshold of 5% resistance in circulating strains for changing the first line treatment empirical antimicrobial: 5% (CLSI) and 13 (EUCAST) for ceftriaxone and 29.3% for azithromycin. Whole genome sequencing of 30 Nc isolates was performed, which identified AMR genes to macrolides and tetracycline. Core gene MLST clustered Nc into three main groups. Gonococcal DNA uptake sequences were identified in two Nc clusters. This suggests that Nc have the potential AMR gene pool and transfer sequences that can result in resistance transfer to pathogenic Neisseria within the nasopharyngeal niche.

Genetic Manipulation of Neisseria gonorrhoeae and Commensal Neisseria Species.

The sexually transmitted pathogen, Neisseria gonorrhoeae, undergoes natural transformation at high frequency. This property has led to the rapid dissemination of antibiotic resistance markers and the panmictic structure of the gonococcal population. However, high-frequency transformation also makes N. gonorrhoeae one of the easiest bacterial species to manipulate genetically in the laboratory. Techniques have been developed that result in transformation frequencies >50%, allowing the identification of mutants by screening and without selection. Constructs have been created to take advantage of this high-frequency transformation, facilitating genetic mutation, complementation, and heterologous gene expression. Similar methods have been developed for N. meningitidis and nonpathogenic Neisseria including N. mucosa and N. musculi. Techniques are described for genetic manipulation of N. gonorrhoeae and commensal Neisseria species, as well as for growth of these fastidious organisms. © 2024 The Author(s). Current Protocols published by Wiley Periodicals LLC. Basic Protocol 1: Spot transformation of Neisseria gonorrhoeae on agar plates Basic Protocol 2: Spot transformation of commensal Neisseria on agar plates Basic Protocol 3: Transformation of Neisseria gonorrhoeae in liquid culture Basic Protocol 4: Electroporation of Neisseria gonorrhoeae Basic Protocol 5: Creation of unmarked mutations using a positive and negative selection cassette Basic Protocol 6: In vitro mutagenesis of Neisseria gonorrhoeae chromosomal DNA using EZ-Tn5 Basic Protocol 7: Chemical mutagenesis Basic Protocol 8: Complementation on the Neisseria gonorrhoeae chromosome Alternate Protocol 1: Complementation with replicating plasmids Alternate Protocol 2: Complementation on the Neisseria musculi or Neisseria mucosa chromosome Basic Protocol 9: Preparation of chromosomal DNA from Neisseria gonorrhoeae grown on solid medium Alternate Protocol 3: Preparation of chromosomal DNA from Neisseria gonorrhoeae grown in broth Support Protocol: Preparing PCR templates from Neisseria gonorrhoeae colonies.

Cyclization increases bactericidal activity of arginine-rich cationic cell-penetrating peptide for Neisseria gonorrhoeae.

We previously reported that a linear cationic 12-amino acid cell-penetrating peptide (CPP) was bactericidal for Neisseria gonorrhoeae. In this study, our objectives were to determine the effect of cyclization of the linear CPP on its antibacterial activity for N. gonorrhoeae and cytotoxicity for human cells. We compared the bactericidal effect of 4-hour treatment with the linear CPP to that of CPPs cyclized by a thioether or a disulfide bond on human challenge and multi-drug resistant (MDR) strains of N. gonorrhoeae grown in cell culture media with 10% fetal bovine serum (FBS). The effect of lipooligosaccharide (LOS) sialylation on bactericidal activity was analyzed. We determined the ability of the CPPs to treat human cells infected in vitro with N. gonorrhoeae, to reduce the inflammatory response of human monocytic cells to gonococci, to kill strains of three commensal Neisseria species, and to inhibit gonococcal biofilms. The cyclized CPPs killed 100% of gonococci from all strains at 100 µM and >90% at 20 µM and were more potent than the linear form. The thioether-linked but not the disulfide-linked CPP was less cytotoxic for human cervical cells compared to the linear CPP. LOS sialylation had minimal effect on bactericidal activity. In treating infected human cells, the thioether-linked CPP at 20 µM killed >60% of extra- and intracellular bacteria and reduced TNF-α expression by THP-1 cells. The potency of the CPPs for the pathogenic and the commensal Neisseria was similar. The thioether-linked CPP partially eradicated gonococcal biofilms. Future studies will focus on determining efficacy in the female mouse model of gonorrhea.IMPORTANCENeisseria gonorrhoeae remains a major cause of sexually transmitted infections with 82 million cases worldwide in 2020, and 710,151 confirmed cases in the US in 2021, up 25% from 2017. N. gonorrhoeae can infect multiple tissues including the urethra, cervix, rectum, pharynx, and conjunctiva. The most serious sequelae are suffered by infected women as gonococci ascend to the upper reproductive tract and cause pelvic inflammatory disease, chronic pelvic pain, and infertility in 10%-20% of women. Control of gonococcal infection is widely recognized as increasingly challenging due to the lack of any vaccine. N. gonorrhoeae has quickly developed resistance to all but one class of antibiotics and the emergence of multidrug-resistant strains could result in untreatable infections. As such, gonorrhea is classified by the Center for Disease Control (CDC) as an urgent public health threat. The research presented herein on new therapeutics for gonorrhea has identified a cyclic cell-penetrating peptide (CPP) as a potent molecule targeting N. gonorrhoeae.

45 years of tetracycline post exposure prophylaxis for STIs and the risk of tetracycline resistance: a systematic review and meta-analysis

There is considerable interest in the use of doxycycline post exposure prophylaxis (PEP) to reduce the incidence of bacterial sexually transmitted infections (STIs). An important concern is that this could select for tetracycline resistance in these STIs and other species. We searched PubMed and Google Scholar, (1948–2023) for randomized controlled trials comparing tetracycline PEP with non-tetracycline controls. The primary outcome was antimicrobial resistance (AMR) to tetracyclines in all bacterial species with available data. Our search yielded 140 studies, of which three met the inclusion criteria. Tetracycline PEP was associated with an increasedprevalence of tetracycline resistance in Neisseria gonorrhoeae, but this effect was not statistically significant (Pooled OR 2.3, 95% CI 0.9-3.4). PEP had a marked effect on the N. gonorrhoeae tetracycline MIC distribution in the one study where this was assessed. Prophylactic efficacy was 100% at low MICs and 0% at high MICs. In the one study where this was assessed, PEP resulted in a significant increase in tetracycline resistance in commensal Neisseria species compared to the control group (OR 2.9, 95% CI 1.5-5.5) but no significant effect on the prevalence of tetracycline resistance in Staphylococcus aureus. The available evidence suggests that PEP with tetracyclines could be associated with selecting tetracycline resistance in N. gonorrhoeae and commensal Neisseria species.

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.

Characterizing the diversity and commensal origins of penA mosaicism in the genus Neisseria.

Mosaic penA alleles formed through horizontal gene transfer (HGT) have been instrumental to the rising incidence of ceftriaxone-resistant gonococcal infections. Although interspecies HGT of regions of the penA gene between Neisseria gonorrhoeae and commensal Neisseria species has been described, knowledge concerning which species are the most common contributors to mosaic penA alleles is limited, with most studies examining only a small number of alleles. Here, we investigated the origins of recombinant penA alleles through in silico analyses that incorporated 1700 penA alleles from 35 513 Neisseria isolates, comprising 15 different Neisseria species. We identified Neisseria subflava and Neisseria cinerea as the most common source of recombinant sequences in N. gonorrhoeae penA. This contrasted with Neisseria meningitidis penA, for which the primary source of recombinant DNA was other meningococci, followed by Neisseria lactamica. Additionally, we described the distribution of polymorphisms implicated in antimicrobial resistance in penA, and found that these are present across the genus. These results provide insight into resistance-related changes in the penA gene across human-associated Neisseria species, illustrating the importance of genomic surveillance of not only the pathogenic Neisseria, but also of the oral niche-associated commensals from which these pathogens are sourcing key genetic variation.

Gonococcal resistance to zoliflodacin could emerge via transformation from commensal Neisseria species. An in-vitro transformation study

One of the most promising new treatments for gonorrhoea currently in phase 3 clinical trials is zoliflodacin. Studies have found very little resistance to zoliflodacin in currently circulating N. gonorrhoeae strains, and in-vitro experiments demonstrated that it is difficult to induce resistance. However, zoliflodacin resistance may emerge in commensal Neisseria spp., which could then be transferred to N. gonorrhoeae via transformation. In this study, we investigated this commensal-resistance-pathway hypothesis for zoliflodacin. To induce zoliflodacin resistance, ten wild-type susceptible isolates belonging to 5 Neisseria species were serially passaged for up to 48 h on gonococcal agar plates containing increasing zoliflodacin concentrations. Within 7 to 10 days, all strains except N. lactamica, exhibited MICs of ≥ 4 µg/mL, resulting in MIC increase ranging from 8- to 64-fold. The last passaged strains and their baseline were sequenced. We detected mutations previously reported to cause zoliflodacin resistance in GyrB (D429N and S467N), novel mutations in the quinolone resistance determining region (QRDR) (M464R and T472P) and mutations outside the QRDR at amino acid positions 28 and 29 associated with low level resistance (MIC 2 µg/mL). Genomic DNA from the laboratory evolved zoliflodacin-resistant strains was transformed into the respective baseline wild-type strain, resulting in MICs of ≥ 8 µg/mL in most cases. WGS of transformants with decreased zoliflodacin susceptibility revealed presence of the same zoliflodacin resistance determinants as observed in the donor strains. Two inter-species transformation experiments were conducted to investigate whether zoliflodacin resistance determinants of commensal Neisseria spp. could be acquired by N. gonorrhoeae. N. gonorrhoeae strain WHO P was exposed to (i) pooled genomic DNA from the two resistant N. mucosa strains and (ii) a gyrB amplicon of the resistant N. subflava strain 45/1_8. Transformants of both experiments exhibited an MIC of 2 µg/mL and whole genome analysis revealed uptake of the mutations detected in the donor strains. This is the first in-vitro study to report that zoliflodacin resistance can be induced in commensal Neisseria spp. and subsequently transformed into N. gonorrhoeae.

Antimicrobial susceptibility of commensal Neisseria spp. in parents and their children in Belgium: a cross-sectional survey.

commensal Neisseria species are part of the oropharyngeal microbiome and play an important role in nitrate reduction and protecting against colonization by pathogenic bacteria. They do, however, also serve as a reservoir of antimicrobial resistance. Little is known about the prevalence of these species in the general population, how this varies by age and how antimicrobial susceptibility varies between species. we assessed the prevalence and antimicrobial susceptibility of commensal Neisseria species in the parents (n=38) and children (n=50) of 35 families in Belgium. various commensal Neisseria (n=5) could be isolated from the participants. Most abundant were N. subflava and N. mucosa. Neisseria subflava was detected in 77 of 88 (87.5%) individuals and N. mucosa in 64 of 88 (72.7%). Neisseria mucosa was more prevalent in children [41/50 (82%)] than parents [23/38 (60.5%); P<.05], while N. bacilliformis was more prevalent in parents [7/36 (19.4%)] than children [2/50 (4%); P<.05]. Neisseria bacilliformis had high ceftriaxone minimum inhibitory concentrations (MICs; median MIC 0.5mg/l; IQR 0.38-0.75). The ceftriaxone MICs of all Neisseria isolates were higher in the parents than in the children. This could be explained by a higher prevalence of N. bacilliformis in the parents. the N. bacilliformis isolates had uniformly high ceftriaxone MICs which warrant further investigation.

Non-pathogenic Neisseria species of the oropharynx as a reservoir of antimicrobial resistance: a cross-sectional study.

Commensal Neisseria species of the oropharynx represent a significant reservoir of antimicrobial resistance determinants that can be transferred to Neisseria gonorrhoeae. This aspect is particularly crucial in 'men having sex with men' (MSM), a key population in which pharyngeal co-colonization by N. gonorrhoeae and non-pathogenic Neisseria species is frequent and associated with the emergence of antimicrobial resistance. Here, we explored the antimicrobial susceptibility of a large panel of non-pathogenic Neisseria species isolated from the oropharynx of two populations: a group of MSM attending a 'sexually transmitted infection' clinic in Bologna (Italy) (n=108) and a group of males representing a 'general population' (n=119). We collected 246 strains, mainly belonging to N. subflava (60%) and N. flavescens (28%) species. Their antimicrobial susceptibility was evaluated assessing the minimum inhibitory concentrations (MICs) for azithromycin, ciprofloxacin, cefotaxime, and ceftriaxone using E-test strips. Overall, commensal Neisseria spp. showed high rates of resistance to azithromycin (90%; median MICs: 4.0 mg/L), and ciprofloxacin (58%; median MICs: 0.12 mg/L), whereas resistance to cephalosporins was far less common (<15%). Neisseria strains from MSM were found to have significantly higher MICs for azithromycin (p=0.0001) and ciprofloxacin (p<0.0001) compared to those from the general population. However, there was no significant difference in cephalosporin MICs between the two groups. The surveillance of the antimicrobial resistance of non-pathogenic Neisseria spp. could be instrumental in predicting the risk of the spread of multi-drug resistant gonorrhea. This information could be an early predictor of an excessive use of antimicrobials, paving the way to innovative screening and prevention policies.

Phylogenomic analysis of the understudied Neisseriaceae species reveals a poly- and paraphyletic Kingella genus.

Taxonomic classification and phylogenetic analysis of the Neisseriaceae family have focused on the pathogens Neisseria meningitidis and Neisseria gonorrhoeae. Less is known about the relationships of commensal Neisseria species and other Neisseriaceae genera, raising the possibility that the phylogeny of this family may not agree with current taxonomy. In this study, we used available nucleotide sequences and a phylogenetic approach to assess the Kingella genus and its relatives. We found that this genus is both paraphyletic and polyphyletic. Kingella potus is more closely related to Neisseria bacilliformis than to other Kingella species. The Alysiella and Simonsiella genera form a distinct clade within the Kingella genus that is closely related to the pathogens K. kingae and K. negevensis. We find a phylogenetic relationship between Conchiformibius, Alysiella, Simonsiella, and Kingella, which we name the CASK clade. Finally, we define the gene sets that differentiate each genus of the CASK clade from one another and from the rest of the Neisseriaceae family. IMPORTANCE Understanding the evolutionary relationships between the species in the Neisseriaceae family has been a persistent challenge in bacterial systematics due to high recombination rates in these species. Previous studies of this family have focused on Neisseria meningitidis and N. gonorrhoeae. However, previously understudied Neisseriaceae species are gaining new attention, with Kingella kingae now recognized as a common human pathogen and with Alysiella and Simonsiella being unique in the bacterial world as multicellular organisms. A better understanding of the genomic evolution of the Neisseriaceae can lead to the identification of specific genes and traits that underlie the remarkable diversity of this family.

Epigenetic effects of ceftriaxone-resistant Neisseria gonorrhoeae FC428 mosaic-like sequences found in PenA sequences unique to Neisseria subflava and related species.

The aim of this study was to explore the origin of the PenA mosaic amino acid sequence in the ceftriaxone-resistant Neisseria gonorrhoeae FC428 clone. The penA sequences of 27 Neisseria subflava pharyngeal isolates were determined by the Sanger method and penA sequences of 52 isolates from nine Neisseria species were obtained from the NCBI database. Comparative analysis of each PenA sequence was performed by multiple sequence alignment using ClustalW. In vitro resistance acquisition experiments were conducted to investigate the possibility of selection pressure by cefixime-induced amino acid substitution mutations in PenA. All N. subflava strains, including two with low susceptibility to expanded-spectrum cephalosporins (ESCs), possessed the majority of the PenA FC428 sequence. Furthermore, a number of strains, but not all, of closely related species of N. subflava showed similar results. PenA FC428 sequences were also found in some strains of distantly related species. No new mutations in the penA sequence were observed in colonies with increased MIC in in vitro resistance acquisition experiments. This study provides strong evidence that the FC428 PenA mosaic sequence originated from N. subflava and related species among oral commensal Neisseria species. The results of in vitro resistance acquisition experiments also suggested that one of the PenA FC428-like sequence gene polymorphisms resulted in the expression of ESC resistance. Furthermore, many of the PenA FC428 mosaic sequences were thought to be involved in the so-called epistasis effect that regulates the expression of resistance, without directly contributing to the resistance level itself.

Identification of ceftriaxone-resistant Neisseria gonorrhoeae FC428 clone and isolates harboring a novel mosaic penA gene in Chengdu in 2019–2020

BackgroundAntimicrobial resistance in gonorrhea has become a growing global public health burden. Neisseria gonorrhoeae isolates with resistance to ceftriaxone, the last remaining first-line option, represent an emerging threat of untreatable gonorrhea.MethodsA total of ten ceftriaxone-resistant N. gonorrhoeae FC428 isolates and two isolates harboring a novel mosaic penA-232.001 allele from 160 gonococcal isolates in Chengdu in 2019–2020 was described in the present study. Multilocus sequence typing (MLST) and N. gonorrhoeae sequence typing for antimicrobial resistance (NG-STAR) were performed to characterize the isolates. Whole genome sequencing and maximum-likelihood method were performed to infer how the genetic phylogenetic tree of these isolates looks like. Recombination analysis was performed using the RDP4 software. This study was registered in the Chinese Clinical Trial Registry (ChiCTR2100048771, registration date: 20210716).ResultsThe genetic phylogeny showed that the ten FC428 isolates sporadically clustered into different phylogenetic clades, suggesting different introductions and local transmission of FC428. Two isolates showed close genetic relatedness to ceftriaxone-resistant clone A8806, which was only reported from Australia in 2013. Homologous recombination events were detected in penA between Neisseria gonorrhoeae and commensal Neisseria species (N. perflava and N. polysaccharea), providing evidence of commensal Neisseria species might serve as reservoirs of ceftriaxone resistance-mediating penA sequences in clinical gonococcal strains.ConclusionsOur results demonstrate further dissemination of FC428 in China and resurgence risks of sporadic ceftriaxone-resistant A8806 to become the next clone to spread.

Commensal Neisseria species share immune suppressive mechanisms with Neisseria gonorrhoeae.

Neisseria gonorrhoeae is a highly adapted human sexually transmitted pathogen that can cause symptomatic infections associated with localized inflammation as well as asymptomatic and subclinical infections, particularly in females. Gonococcal infection in humans does not generate an effective immune response in most cases, which contributes to both transmission of the pathogen and reinfection after treatment. Neisseria gonorrhoeae is known to evade and suppress human immune responses through a variety of mechanisms. Commensal Neisseria species that are closely related to N. gonorrhoeae, such as N. cinerea, N. lactamica, N. elongata, and N. mucosa, rarely cause disease and instead asymptomatically colonize mucosal sites for prolonged periods of time without evoking clearing immunologic responses. We have shown previously that N. gonorrhoeae inhibits the capacity of antigen-pulsed dendritic cells to induce CD4+ T cell proliferation in vitro. Much of the suppressive effects of N. gonorrhoeae on dendritic cells can be recapitulated either by outer-membrane vesicles released from the bacteria or by purified PorB, the most abundant outer-membrane protein in Neisseria gonorrhoeae. We show here that three commensal Neisseria species, N. cinerea, N. lactamica and N. mucosa, show a comparable capacity to suppress dendritic cell-induced T cell proliferation in vitro through mechanisms similar to those demonstrated previously for N. gonorrhoeae, including inhibition by purified PorB. Our findings suggest that some immune-evasive properties of pathogenic N. gonorrhoeae are shared with commensal Neisseria species and may contribute to the ability of both pathogens and commensals to cause prolonged mucosal colonization in humans.

The oropharynx of men using HIV pre-exposure prophylaxis is enriched with antibiotic resistance genes: A cross-sectional observational metagenomic study

Phenotypic studies have found high levels of antimicrobial resistance to cephalosporins, macrolides and fluoroquinolones in commensal Neisseria species in the oropharynx of men who have sex with men (MSM) using HIV pre-exposure prophylaxis (PrEP). These species include Neisseria subflava and Neisseria mucosa. This may represent a risk to pathogens like Neisseria gonorrhoeae which tend to take up antibiotic resistance genes (ARGs) from other bacteria. We aimed to explore to what extent the oropharyngeal resistome of MSM using PrEP differed from the general population. We collected oropharyngeal swabs from 32 individuals of the general population and from 64 MSM using PrEP. Thirty-two MSM had consumed antibiotics in the previous six months, whereas none of the other participants had. Samples underwent shotgun metagenomic sequencing. Sequencing reads were mapped against MEGARes 2.0 to estimate ARG abundance. ARG abundance was compared between groups by zero-inflated negative binomial regression. ARG abundance was significantly lower in the general population than in MSM (ratio 0.41, 95% CI 0.26-0.65). More specifically, this was the case for fluoroquinolones (0.33, 95% CI 0.15-0.69), macrolides (0.37, 95% CI 0.25-0.56), tetracyclines (0.41, 95% CI 0.25-0.69), and multidrug efflux pumps (0.11, 95% CI 0.03-0.33), but not for beta-lactams (1.38, 95% CI 0.73-2.61). There were no significant differences in ARG abundance between MSM who had used antibiotics and those that had not. The resistome of MSM using PrEP is enriched with ARGs, independent of recent antibiotic use. Stewardship campaigns should aim to reduce antibiotic consumption in populations at high risk for STIs.

Colonization Rate and Associated Factors of Non-Pathogenic Neisseria Species, and Moraxella catarrhalis Among Healthy School Children in Gondar, Northwest Ethiopia.

Although commensal Neisseria species inhabiting mucosal surfaces in the upper respiratory tract (URT) are rarely associated with infections, their presence in the area has been linked to the development of immunity against N. meningitidis and the source of antibiotic resistance determinants in pathogenic species. M. catarrhalis in the oropharynx of children is also a predisposing factor for otitis media. As a result, determining the oropharyngeal carriage rate of these commensal species and associated factors among healthy schoolchildren is substantial. This community-based cross-sectional study was conducted in Gondar, Northwest Ethiopia, from January to April 2019. A multi-stage and simple random sampling technique were used to select schools and participants, respectively. A total of 524 oropharyngeal swabs were collected using cotton swabs. Modified Thayer-Martin media was used for primary bacterial isolation, and battery of biochemical tests was performed to identify species. For frequencies, descriptive statistics were computed and the logistic regression model was used to see the relationship between dependent and independent variables. A total of 524 healthy schoolchildren with a mean age of 12.2 ± 2.74 years participated in this study. The overall oropharyngeal carriage rate was 21.8% (114/524). Of these, N. meningitidis, N. lactamica, N. sicca, and M. catarrhalis were identified in 53 (46.5%), 14 (12.3%), 11 (9.6%), and 36 (31.6%) children, respectively. The culture positivity rate was higher at a younger age, which was 8.1%, 11.3%, and 14.9% in ages between 15-18, 11-14, and 7-10, respectively. The oropharyngeal carriage was significantly associated with the number of students per class (>40). There is a higher proportion of carriers of commensal N. lactamica and M. catarrhalis in Gondar town schoolchildren. The oropharyngeal carriage rate was associated with a crowded classroom. The characterization of non-pathogenic Neisseria species and M. catarrhalis in the study area can support the diagnosis of patients suspected of having N. meningitis infections.

Commensal Neisseria species share immune suppressive mechanisms with Neisseria gonorrhoeae

Commensal Neisseria species share immune suppressive mechanisms with Neisseria gonorrhoeae

Genomic sequence of the non-pathogen Neisseria sp. strain MA1-1 with antibiotic resistance and virulence factors isolated from a head and neck cancer patient.

Recent research has claimed virulence factors or antimicrobial resistance in commensal or non-pathogenic Neisseria spp. This study aimed to isolate and analyze commensal microorganisms related to the genus Neisseria from the oral cavity of a patient with head and neck cancer. We successfully isolated strain MA1-1 and identified its functional gene contents. Although strain MA1-1 was related to Neisseria flava based on 16S rRNA gene sequence similarity, genomic relatedness analysis revealed that strain MA1-1 was closely related to Neisseria mucosa, reported as a commensal Neisseria species. The strain MA1-1 genome harbored genes for microaerobic respiration and the complete core metabolic pathway with few transporters for nutrients. A number of genes have been associated with virulence factors and resistance to various antibiotics. In addition, the comparative genomic analysis showed that most genes identified in the strain MA1-1 were shared with other Neisseria spp. including two well-known pathogens, Neisseria gonorrhoeae and Neisseria meningitidis. This indicates that the gene content of intra-members of the genus Neisseria has been evolutionarily conserved and is stable, with no gene recombination with other microbes in the host. Finally, this study provides more fundamental interpretations for the complete gene sequence of commensal Neisseria spp. and will contribute to advancing public health knowledge.

Neisseria mucosa Does Not Inhibit the Growth of Neisseria gonorrhoeae

Antibiotic-sparing treatments are required to prevent the further emergence of antimicrobial resistance in Neisseria gonorrhoeae. Commensal Neisseria species have previously been found to inhibit the growth of pathogenic Neisseria species. For example, a previous study found that three out of five historical isolates of Neisseria mucosa could inhibit the growth of N. gonorrhoeae. In this study, we used agar overlay assays to assess if 24 circulating and historical isolates of Neisseria mucosa could inhibit the growth of 28 circulating and historical isolates of N. gonorrhoeae. Although pitting around each colony of N. mucosa created an optical illusion of decreased growth of N. gonorrhoeae, we found no evidence of inhibition (n = 24). In contrast, positive controls of Streptococcus pneumoniae and Escherichia coli demonstrated a strong inhibitory effect against the growth of N. gonorrhoeae.

Evidence of Horizontal Gene Transfer of 50S Ribosomal Genes rplB, rplD, and rplY in Neisseria gonorrhoeae.

Horizontal gene transfer (HGT) in the penA and multidrug efflux pump genes has been shown to play a key role in the genesis of antimicrobial resistance in Neisseria gonorrhoeae. In this study, we evaluated if there was evidence of HGT in the genes coding for the ribosomal proteins in the Neisseria genus. We did this in a collection of 11,659 isolates of Neisseria, including N. gonorrhoeae and commensal Neisseria species (N. cinerea, N. elongata, N. flavescens, N. mucosa, N. polysaccharea, and N. subflava). Comparative genomic analyses identified HGT events in three genes: rplB, rplD, and rplY coding for ribosomal proteins L2, L4 and L25, respectively. Recombination events were predicted in N. gonorrhoeae and N. cinerea, N. subflava, and N. lactamica were identified as likely progenitors. In total, 2,337, 2,355, and 1,127 isolates possessed L2, L4, and L25 HGT events. Strong associations were found between HGT in L2/L4 and the C2597T 23S rRNA mutation that confers reduced susceptibility to macrolides. Whilst previous studies have found evidence of HGT of entire genes coding for ribosomal proteins in other bacterial species, this is the first study to find evidence of HGT-mediated chimerization of ribosomal proteins.

Antimicrobial Resistance Profiles of Human Commensal Neisseria Species.

Pathogenic Neisseria gonorrhoeae causes the sexually transmitted infection gonorrhea. N. gonorrhoeae has evolved high levels of antimicrobial resistance (AR) leading to therapeutic failures even in dual-therapy treatment with azithromycin and ceftriaxone. AR mechanisms can be acquired by genetic transfer from closely related species, such as naturally competent commensal Neisseria species. At present, little is known about the antimicrobial resistance profiles of commensal Neisseria. Here, we characterized the phenotypic resistance profile of four commensal Neisseria species (N. lactamica, N. cinerea, N. mucosa, and N. elongata) against 10 commonly used antibiotics, and compared their profiles to 4 N. gonorrhoeae strains, using disk diffusion and minimal inhibitory concentration assays. Overall, we observed that 3 of the 4 commensals were more resistant to several antibiotics than pathogenic N. gonorrhoeae strains. Next, we compared publicly available protein sequences of known AR genes, including penicillin-binding-protein 2 (PBP2) from commensals and N. gonorrhoeae strains. We found mutations in PBP2 known to confer resistance in N. gonorrhoeae also present in commensal Neisseria sequences. Our results suggest that commensal Neisseria have unexplored antibiotic resistance gene pools that may be exchanged with pathogenic N. gonorrhoeae, possibly impairing drug development and clinical treatment.