Nonpathogenic Neisseria Species Research Articles

THE DIFFERENTIAL EXTRACTION METHOD OF NEISSERIA MENINGITIDIS AND NON-PATHOGENIC SPECIES FOR PROTEIN PROFILING BY SDS-PAGE

Invasive meningococcal disease (IMD) is an acute, severe, and potentially fatal infection caused by Neisseria meningitidis and is a global public health burden. Given the high fatality rate linked to acute bacterial meningitis, it is imperative to initiate treatment and diagnosis concurrently in most cases. While a culture demonstrating the growth of N. meningitidis from blood or cerebrospinal fluid (CSF) remains the gold standard, however, its sensitivity diminishes following antibiotic administration. Therefore, in this study, differentially extracted whole cell protein (WCP), surface depleted whole cell protein (sdWCP), and cell surface protein (CSP) profiles were analysed by using SDS-PAGE to characterise and classify pathogenic N. meningitidis and non-pathogenic species. This study provides clear evidence that SDS-PAGE yields distinct protein patterns across Men A, B, C, and Y, clinical isolates 1 and 2, N. sicca, N. cinerea, and M. catarrhalis. It effectively demonstrates that SDS-PAGE protein profiling can serve as a reliable method for characterising and separating bacterial proteins at both the species and strain levels. Despite the availability of more advanced technologies, SDS-PAGE remains a valuable tool, particularly in settings where advanced equipment and knowledgeable personnel are lacking. Our approach employs various extraction techniques, has successfully defined and characterised N. meningitidis, non-pathogenic Neisseria species, and M. catarrhalis.

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

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

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.

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.

A Low Virulence Infective Tenosynovitis Triggered by a Neisseria Species.

A persistent infective tenosynovitis, caused by a non-pathogenic Neisseria species and involving the little finger and the thumb of a middle-aged woman, forming a horseshoe-type infection is presented. An extensive volar approach was utilised for excision of the granulomatous synovium and drainage of turbid fluid. Neisseria spp was isolated by tissue microbiology and targeted antibiotic therapy for 6 weeks was required to eradicate infection. The presence of low virulence microorganisms which might act as true pathogens should always be considered as a causative factor leading to persistent infections that require open surgical debridement and washout. Level of Evidence: Level V (Therapeutic).

The Human Microbiome as a Focus of Antibiotic Discovery: Neisseria mucosa Displays Activity Against Neisseria gonorrhoeae.

Neisseria gonorrhoeae infections are a serious global health problem. This organism has developed disturbing levels of antibiotic resistance, resulting in the need for new approaches to prevent and treat gonorrhea. The genus Neisseria also includes several members of the human microbiome that live in close association with an array of microbial partners in a variety of niches. We designed an undergraduate antibiotic discovery project to examine a panel of nonpathogenic Neisseria species for their ability to produce antimicrobial secondary metabolites. Five strains belonging to the N. mucosa species group displayed activity against other Neisseria in delayed antagonism assays; three of these were active against N. gonorrhoeae. The antimicrobial compound secreted by N. mucosa NRL 9300 remained active in the presence of catalase, trypsin, and HEPES buffer, and effectively inhibited a DNA uptake mutant of N. gonorrhoeae. Antimicrobial activity was also retained in an ethyl acetate extract of plate grown N. mucosa NRL 9300. These data suggest N. mucosa produces an antimicrobial secondary metabolite that is distinct from previously described antigonococcal agents. This work also serves as a demonstration project that could easily be adapted to studying other members of the human microbiome in undergraduate settings. We offer the perspective that both introductory and more advanced course-based and apprentice-style antibiotic discovery projects focused on the microbiome have the potential to enrich undergraduate curricula and we describe transferrable techniques and strategies to facilitate project design.

A role for fibroblast growth factor receptor 1 in the pathogenesis of Neisseria meningitidis

Neisseria meningitidis (the meningococcus) remains an important cause of human disease, including meningitis and sepsis. Adaptation to the host environment includes many interactions with specific cell surface receptors, resulting in intracellular signalling and cytoskeletal rearrangements that contribute to pathogenesis. Here, we assessed the interactions between meningococci and Fibroblast Growth Factor Receptor 1-IIIc (FGFR1-IIIc): a receptor specific to endothelial cells of the microvasculature, including that of the blood-brain barrier. We show that the meningococcus recruits FGFR1-IIIc onto the surface of human blood microvascular endothelial cells (HBMECs). Furthermore, we demonstrate that expression of FGFR1-IIIc is required for optimal invasion of HBMECs by meningococci. We show that the ability of N. meningitidis to interact with the ligand-binding domain of FGFR1-IIIc is shared with the other pathogenic Neisseria species, N. gonorrhoeae, but not with commensal bacteria including non-pathogenic Neisseria species.

Neisseria meningitidis has acquired sequences within the capsule locus by horizontal genetic transfer

Background: Expression of a capsule from one of serogroups A, B, C, W, X or Y is usually required for Neisseria meningitidis ( Nme) to cause invasive meningococcal disease. The capsule is encoded by the capsule locus, cps, which is proposed to have been acquired by a formerly capsule null organism by horizontal genetic transfer (HGT) from another species. Following identification of putative capsule genes in non-pathogenic Neisseria species, this hypothesis is re-examined. Methods: Whole genome sequence data from Neisseria species, including Nme genomes from a diverse range of clonal complexes and capsule genogroups, and non- Neisseria species, were obtained from PubMLST and GenBank. Sequence alignments of genes from the meningococcal cps, and predicted orthologues in other species, were analysed using Neighbor-nets, BOOTSCANing and maximum likelihood phylogenies. Results: The meningococcal cps was highly mosaic within regions B, C and D. A subset of sequences within regions B and C were phylogenetically nested within homologous sequences belonging to N. subflava, consistent with HGT event in which N. subflava was the donor. In the cps of 23/39 isolates, the two copies of region D were highly divergent, with rfbABC’ sequences being more closely related to predicted orthologues in the proposed species N. weixii (GenBank accession number CP023429.1) than the same genes in Nme isolates lacking a capsule. There was also evidence of mosaicism in the rfbABC’ sequences of the remaining 16 isolates, as well as rfbABC from many isolates. Conclusions: Data are consistent with the en bloc acquisition of cps in meningococci from N. subflava, followed by further recombination events with other Neisseria species. Nevertheless, the data cannot refute an alternative model, in which native meningococcal capsule existed prior to undergoing HGT with N. subflava and other species. Within-genus recombination events may have given rise to the diversity of meningococcal capsule serogroups.

Neisseria meningitidis has acquired sequences within the capsule locus by horizontal genetic transfer.

Background: Expression of a capsule from one of serogroups A, B, C, W, X or Y is usually required for Neisseria meningitidis ( Nme) to cause invasive meningococcal disease. The capsule is encoded by the capsule locus, cps, which is proposed to have been acquired by a formerly capsule null organism by horizontal genetic transfer (HGT) from another species. Following identification of putative capsule genes in non-pathogenic Neisseria species, this hypothesis is re-examined. Methods: Whole genome sequence data from Neisseria species, including Nme genomes from a diverse range of clonal complexes and capsule genogroups, and non- Neisseria species, were obtained from PubMLST and GenBank. Sequence alignments of genes from the meningococcal cps, and predicted orthologues in other species, were analysed using Neighbor-nets, BOOTSCANing and maximum likelihood phylogenies. Results: The meningococcal cps was highly mosaic within regions B, C and D. A subset of sequences within regions B and C were phylogenetically nested within homologous sequences belonging to N. subflava, consistent with HGT event in which N. subflava was the donor. In the cps of 23/39 isolates, the two copies of region D were highly divergent, with rfbABC' sequences being more closely related to predicted orthologues in the proposed species N. weixii(GenBank accession number CP023429.1) than the same genes in Nme isolates lacking a capsule. There was also evidence of mosaicism in the rfbABC' sequences of the remaining 16 isolates, as well as rfbABC from many isolates. Conclusions: Data are consistent with the en bloc acquisition of cps in meningococci from N. subflava, followed by further recombination events with other Neisseria species. Nevertheless, the data cannot refute an alternative model, in which native meningococcal capsule existed prior to undergoing HGT with N. subflava and other species. Within-genus recombination events may have given rise to the diversity of meningococcal capsule serogroups.

Characterization of capsule genes in non-pathogenic Neisseria species.

The genus Neisseria comprises a diverse group of commensal bacteria, which typically colonize the mucosal surfaces of humans and other animals. Neisseria meningitidis, the meningococcus, is notable for its potential to cause invasive meningococcal disease (IMD) in humans; however, IMD is comparatively rare, and meningococci normally colonize the nasopharynx asymptomatically. Possession of a polysaccharide capsule has been shown to be a prerequisite for disease in almost all IMD cases, and was previously considered unique to N. meningitidis, and potentially acquired by horizontal genetic transfer (HGT). Nevertheless, the capsule must also have some role in asymptomatic colonization and/or transmission, consistent with the existence of six non-disease-associated meningococcal capsule serogroups. In this study, full complements of putative capsule genes were identified in non-pathogenic Neisseria species, including Neisseria subflava and Neisseria elongata. These species contained genes for capsule transport and translocation homologous to those of N. meningitidis, as well as novel putative capsule synthesis genes. Phylogenetic analyses were consistent with the proposal that these genes were acquired by the meningococcus through HGT. In contrast with previous evolutionary models, however, the most parsimonious explanation of these data was that capsule transport genes had been lost in the common ancestor of the meningococcus, gonococcus, and their close relatives, and then reacquired by some meningococci. The most likely donor of the meningococcal transport genes was another Neisseria species.

Microbiological analysis of dental plaque in children with Down syndrome

Topical focus of modern dentistry is improving the prevention of dental diseases in children with Down syndrome. The goal of this work is to assess the species composition of the microflora of dental plaque in children with Down syndrome in comparison with conditionally healthy children. Materials and methods. In this work we have used the results of bacteriological research of clinical material of 10 children with Down syndrome and 15 conditionally healthy children. Swabs from cervical part of the teeth have been taken with the help of a sterile disposable tampon. Microbiological research has included a discharge of microorganisms, identification by morphological, cultural and biochemical properties of cultures. The research has been conducted on clinical strains of microorganisms discharged from dental plaque in children with Down syndrome and conditionally healthy children. The results. In the investigated material of children with Down syndrome, we have detected gram-positive microorganisms, such as Streptococcus spp with α-hemolytic activity, Bacillus spp and Candida fungi, as well as gram-negative bacteria: E. aerogenes , K. pneumoniae, P. aeruginosa, Neisseria spp ( Table 1 ). In the microflora of dental plaque, facultative anaerobic microorganisms have been discharged more often than aerobes. Conclusions. In control and main group, dental plaque microflora is more often presented by associations of bacteria and Candida fungi. The discharge rate of Streptococcus spp with α-hemolytic activity and Candida albicans is equal in both groups. Nonpathogenic Neisseria species have been presented in 9.1 % of strains in the main group, whereas in the control group there have been 35 % of strains. In children with Down syndrome we have detected gram-negative bacilli – enterobacteria with a density of microbial colonization of E. aerogenes Ig (4.66 ± 0.6) cfu/g, pseudomonads and gram-positive spore-forming bacilli – Ig (5.0 ± 0.0) cfu/g, which are representatives of the allochthonous (transitory, fickle) microflora of the oral cavity. Their long stay in the oral cavity prevents the effect of nonspecific protective factors, as well as antagonistic activity of autochthonous microflora. In case of some disorders of physiological state, representatives of allochthonous microflora can linger in the oral cavity, multiply and cause pathological processes.

Neisseria cinerea isolates can adhere to human epithelial cells by type IV pilus-independent mechanisms.

In pathogenic Neisseria species the type IV pili (Tfp) are of primary importance in host-pathogen interactions. Tfp mediate initial bacterial attachment to cell surfaces and formation of microcolonies via pilus-pilus interactions. Based on genome analysis, many non-pathogenic Neisseria species are predicted to express Tfp, but aside from studies on Neisseria elongata, relatively little is known about the formation and function of pili in these organisms. Here, we have analysed pilin expression and the role of Tfp in Neisseria cinerea. This non-pathogenic species shares a close taxonomic relationship to the pathogen Neisseria meningitidis and also colonizes the human oropharyngeal cavity. Through analysis of non-pathogenic Neisseria genomes we identified two genes with homology to pilE, which encodes the major pilin of N. meningitidis. We show which of the two genes is required for Tfp expression in N. cinerea and that Tfp in this species are required for DNA competence, similar to other Neisseria. However, in contrast to the meningococcus, deletion of the pilin gene did not impact the association of N. cinerea to human epithelial cells, demonstrating that N. cinerea isolates can adhere to human epithelial cells by Tfp-independent mechanisms.

Production of Antimicrobial Substances by Nonpathogenic Neisseria Species

The emergence of multidrug resistant Neisseria gonorrhoeae is a serious public health concern. Compounds known as bacteriocins represent one novel approach to antimicrobial development. Bacteriocin...

Laboratory identification methods of nonpathogenic Neisseria species

Laboratory identification methods of nonpathogenic Neisseria species

EFFECT OF BICLOTYMOL ON MOUTH CAVITY MUCOSAL MICROBIOTA IN CHILDREN

научный биометрический журнал, который знакомит читателя с клиническими рекомендациями и особенностями применения лекарственных и вакцинных средств у детей, предоставляет исчерпывающую информацию о воздействии лекарственных средств на плод, о проводимых в

Distribution and diversity of the haemoglobin–haptoglobin iron-acquisition systems in pathogenic and non-pathogenic Neisseria

A new generation of vaccines containing multiple protein components that aim to provide broad protection against serogroup B meningococci has been developed. One candidate, 4CMenB (4 Component MenB), has been approved by the European Medicines Agency, but is predicted to provide at most 70–80 % strain coverage; hence there is a need for second-generation vaccines that achieve higher levels of coverage. Prior knowledge of the diversity of potential protein vaccine components is a key step in vaccine design. A number of iron import systems have been targeted in meningococcal vaccine development, including the HmbR and HpuAB outer-membrane proteins, which mediate the utilization of haemoglobin or haemoglobin–haptoglobin complexes as iron sources. While the genetic diversity of HmbR has been described, little is known of the diversity of HpuAB. Using whole genome sequences deposited in a Bacterial Isolate Genome Sequence Database (BIGSDB), the prevalence and diversity of HpuAB among Neisseria were investigated. HpuAB was widely present in a range of Neisseria species whereas HmbR was mainly limited to the pathogenic species Neisseria meningitidis and Neisseria gonorrhoeae. Patterns of sequence variation in sequences from HpuAB proteins were suggestive of recombination and diversifying selection consistent with strong immune selection. HpuAB was subject to repeat-mediated phase variation in pathogenic Neisseria and the closely related non-pathogenic Neisseria species Neisseria lactamica and Neisseria polysaccharea but not in the majority of other commensal Neisseria species. These findings are consistent with HpuAB being subject to frequent genetic transfer potentially limiting the efficacy of this receptor as a vaccine candidate.

Neisseria lactamica antigens complexed with a novel cationic adjuvant

Colonization of the nasopharynx by non-pathogenic Neisseria species, including N. lactamica, has been suggested to lead to the acquisition of natural immunity against Neisseria meningitidis in young children. The aim of this study was to identify a model complex of antigens and adjuvant for immunological preparation against N. meningitidis B, based on cross reactivity with N. lactamica outer membrane vesicles (OMV) antigens and the (DDA-BF) adjuvant. Complexes of 25 µg of OMV in 0.1 mM of DDA-BF were colloidally stable, exhibiting a mean diameter and charge optimal for antigen presentation. Immunogenicity tests for these complexes were performed in mice. A single dose of OMV/DDA-BF was sufficient to induce a (DTH) response, while the same result was achieved only after two doses of OMV/alum. In addition, to achieve total IgG levels that are similar to a single immunization with OMV/DDA-BF, it was necessary to give the mice a second dose of OMV/alum. Moreover, the antibodies induced from a single immunization with OMV/DDA-BF had an intermediate avidity, but antibodies with a similar avidity were only induced by OMV/alum after two immunizations. The use of this novel cationic adjuvant for the first time with a N. lactamica OMV preparation revealed good potential for future vaccine design.

Use of a dual priming oligonucleotide system to detect multiple sexually transmitted pathogens in clinical specimens

To evaluate a new dual priming oligonucleotide (DPO)-based multiplex polymerase chain reaction (PCR) assay for detection of six sexually transmitted pathogens, including Chlamydia trachomatis, Neisseria gonorrhoeae, Mycoplasma genitalium, Mycoplasma hominis, Ureaplasma urealyticum and Trichomonas vaginalis. Using 130 clinical specimens, the results obtained by the multiplex PCR, previously established in-house PCR and COBAS Amplicor PCR assays were compared. The specimens frequently contained multiple pathogens (34/130 specimens). The multiplex PCR assay had an overall sensitivity of 96% and specificity of 100% compared to the in-house PCR assay at >20 microg ml(-1) of DNA concentrations in samples and there was no cross-reaction with nonpathogenic Neisseria species that cause the majority of false-positive results with the COBAS Amplicor PCR assay. The DPO-based multiplex PCR assay detected the six sexually transmitted pathogens in clinical specimens with a high sensitivity and specificity, although its sensitivity was dependent on the DNA content of the samples. It is the first report about the new DPO-based technique to detect multiple sexually transmitted pathogens in a single assay, which has considerable potential to diagnose the infections accurately and rapidly.

Molecular diagnosis ofNeisseria gonorrhoeaeinfection in the United States

Gonorrhea, with its Neisseria gonorrhoeae etiology, is the second most common bacterial sexually transmitted infection in the United States and is most often transmitted through asymptomatic individuals. N. gonorrhoeae is a fastidious organism and can be susceptible to killing during transport to the laboratory; hence, diagnostic methods that do not require organism viability are becoming more commonplace in the clinical setting. This review summarizes traditional and molecular-based diagnostic modalities specific to N. gonorrhoeae. Several commercially available, FDA-approved molecular methods to diagnose N. gonorrhoeae infection include nucleic acid hybridization, signal amplification, polymerase chain reaction, strand displacement amplification and transcription-mediated amplification. Molecular-based methods are rapid, reliable and effective genital specimen screening measures, especially when applied to areas of high disease prevalence. However, clinical and analytical sensitivity for some commercial systems decreases dramatically when testing urine samples. Moreover, crossreactivity to non-pathogenic Neisseria species has been documented for selected methods. Therapeutic decisions may increasingly become compromised owing to a lack of cultured N. gonorrhoeae for antimicrobial susceptibility testing. In vitro experiments suggest that transcription-mediated amplification has greater analytical sensitivity than the other molecular-based methods now available. The development of future molecular testing could address conundrums associated with confirmatory testing, medicolegal testing, testing for cure and detection of antimicrobial-resistant strains.

Absence of Mucosal Immunity in the Human Upper Respiratory Tract to the Commensal Bacteria Neisseria lactamica but Not Pathogenic Neisseria meningitidis during the Peak Age of Nasopharyngeal Carriage

The normal flora that colonizes the mucosal epithelia has evolved diverse strategies to evade, modulate, or suppress the immune system and avoid clearance. Neisseria lactamica and Neisseria meningitidis are closely related obligate inhabitants of the human upper respiratory tract. N. lactamica is a commensal but N. meningitidis is an opportunistic pathogen that occasionally causes invasive disease such as meningitis and septicemia. We demonstrate that unlike N. meningitidis, N. lactamica does not prime the development of mucosal T or B cell memory during the peak period of colonization. This cannot be explained by the induction of peripheral tolerance or regulatory CD4(+)CD25(+) T cell activity. Instead, N. lactamica mediates a B cell-dependent mitogenic proliferative response that is absent to N. meningitidis. This mitogenic response is associated with the production of T cell-independent polyclonal IgM that we propose functions by shielding colonizing N. lactamica from the adaptive immune system, maintaining immunological ignorance in the host. We conclude that, in contrast to N. meningitidis, N. lactamica maintains a commensal relationship with the host in the absence of an adaptive immune response. This may prolong the period of susceptibility to colonization by both pathogenic and nonpathogenic Neisseria species.