Genetic variants linked to the phenotypic outcome of invasive disease and carriage of Neisseria meningitidis.

Neisseria meningitidis can be a human commensal in the upper respiratory tract but is also capable of causing invasive diseases such as meningococcal meningitis and septicaemia. No specific genetic markers have been detected to distinguish carriage from disease isolates. The aim here was to find genetic traits that could be linked to phenotypic outcomes associated with carriage versus invasive N. meningitidis disease through a bacterial genome-wide association study (GWAS). In this study, invasive N. meningitidis isolates collected in Sweden (n=103) and carriage isolates collected at Örebro University, Sweden (n=213) 2018-2019 were analysed. The GWAS analysis, treeWAS, was applied to single-nucleotide polymorphisms (SNPs), genes and k-mers. One gene and one non-synonymous SNP were associated with invasive disease and seven genes and one non-synonymous SNP were associated with carriage isolates. The gene associated with invasive disease encodes a phage transposase (NEIS1048), and the associated invasive SNP glmU S373C encodes the enzyme N-acetylglucosamine 1-phosphate (GlcNAC 1-P) uridyltransferase. Of the genes associated with carriage isolates, a gene variant of porB encoding PorB class 3, the genes pilE/pilS and tspB have known functions. The SNP associated with carriage was fkbp D33N, encoding a FK506-binding protein (FKBP). K-mers from PilS, tbpB and tspB were found to be associated with carriage, while k-mers from mtrD and tbpA were associated with invasiveness. In the genes fkbp, glmU, PilC and pilE, k-mers were found that were associated with both carriage and invasive isolates, indicating that specific variations within these genes could play a role in invasiveness. The data presented here highlight genetic traits that are significantly associated with invasive or carriage N. meningitidis across the species population. These traits could prove essential to our understanding of the pathogenicity of N. meningitidis and could help to identify future vaccine targets.

Neisseria meningitidis carriage in Swedish teenagers associated with the serogroup W outbreak at the World Scout Jamboree, Japan 2015.

The aims of the study were to estimate the carrier state of Neisseria meningitidis in Swedish teenagers and its association with an outbreak at the World Scout Jamboree in 2015 as well as to compare sensitivity of throat versus nasopharyngeal swab for optimal detection of carriage. In total, 1 705 samples (cultures n = 32, throat swabs n = 715, nasopharyngeal swabs n = 958) from 1 020 Jamboree participants were collected and sent to the National Reference Laboratory for Neisseria meningitidis for culture and molecular analysis. The overall positivity for N. meningitidis was 8% (83/1 020), whereas 2% (n = 22) belonged to a known sero/genogroup while the majority (n = 61) were non-groupable. Throat sample is clearly the sampling method of choice, in 56 individuals where both throat and nasopharynx samples were taken, N. meningitidis was detected in both throat and nasopharynx in eight individuals, in 46 individuals N. meningitidis was only detected in the throat and in two individuals only in the nasopharynx. Carriage studies are important to provide knowledge of the current epidemiology and association between carrier isolates and disease-causing isolates in a given population. Therefore, planning for a carriage study in Sweden is in progress.

Whole-Genome Sequencing of Emerging Invasive Neisseria meningitidis Serogroup W in Sweden.

Invasive disease caused by Neisseria meningitidis serogroup W (MenW) has historically had a low incidence in Sweden, with an average incidence of 0.03 case/100,000 population from 1995 to 2014. In recent years, a significant increase in the incidence of MenW has been noted in Sweden, to an average incidence of 0.15 case/100,000 population in 2015 to 2016. In 2017 (1 January to 30 June), 33% of invasive meningococcal disease cases (7/21 cases) were caused by MenW. In the present study, all invasive MenW isolates from Sweden collected in 1995 to June 2017 (n = 86) were subjected to whole-genome sequencing to determine the population structure and to compare isolates from Sweden with historical and international cases. The increase of MenW in Sweden was determined to be due to isolates belonging to the South American sublineage of MenW clonal complex 11, namely, the novel U.K. 2013 lineage. This lineage was introduced in Sweden in 2013 and has since been the dominant lineage of MenW.

Evolutionary epidemiology of Neisseria meningitidis strains in Belarus compared to other European countries.

INTRODUCTION: Meningococcal infections are major causes of death in children globally. In Belarus, the incidence of cases and fatality rate of meningococcal infections are low and comparable to the levels in other European countries. AIM: In the present study, the molecular and epidemiological traits of Neisseria meningitidis strains circulating in Belarus were characterized and compared to isolates from other European countries. MATERIALS AND METHODS: Twenty N. meningitidis strains isolated from patients (n = 13) and healthy contacts (n = 7) during 2006­2012 in Belarus were selected for multilocus sequence typing (MLST), genosubtyping and FetA typing. TheSTs of the Belarusian strains were phylogenetically compared to the STs of 110 selected strains from 22 other European countries. RESULTS: Overall, eleven different genosubtypes were observed, there were seven variants of variable region of the fet Agene detected. The majority of the STs (95%) found in Belarus were novel and allthose were submitted to the Neisseria MLST database for assignment. Several newly discovered alleles of fumC (allele 451) and gdh (allele 560 and 621) appeared to be descendants of alleles which are widespread in Europe, and single aroE alleles (602 and 603) occurred as a result of separate evolution. CONCLUSIONS: N. meningitidis strains circulating in Belarus are heterogeneous and include sequence types, possibly, locally evolved in Belarus as well as representatives of widespread European hyperinvasive clonal complexes.

Multicenter study for defining the breakpoint for rifampin resistance in Neisseria meningitidis by rpoB sequencing.

Identification of clinical isolates of Neisseria meningitidis that are resistant to rifampin is important to avoid prophylaxis failure in contacts of patients, but it is hindered by the absence of a breakpoint for resistance, despite many efforts toward standardization. We examined a large number (n = 392) of clinical meningococcal isolates, spanning 25 years (1984 to 2009), that were collected in 11 European countries, Argentina, and the Central African Republic. The collection comprises all clinical isolates with MICs of > or = 0.25 mg/liter (n = 161) received by the national reference laboratories for meningococci in the participating countries. Representative isolates displaying rifampin MICs of < 0.25 mg/liter were also examined (n = 231). Typing of isolates was performed, and a 660-bp DNA fragment of the rpoB gene was sequenced. Sequences differing by at least one nucleotide were defined as unique rpoB alleles. The geometric mean of the MICs was calculated for isolates displaying the same allele. The clinical isolates displaying rifampin MICs of > 1 mg/liter possessed rpoB alleles with nonsynonymous mutations at four critical amino acid residues, D542, H552, S548, and S557, that were absent in the alleles found in all isolates with MICs of < or = 1 mg/liter. Rifampin-susceptible isolates could be defined as those with MICs of < or = 1 mg/liter. The rpoB allele sequence and isolate data have been incorporated into the PubMLST Neisseria database (http://pubmlst.org/neisseria/). The rifampin-resistant isolates belonged to diverse genetic lineages and were associated with lower levels of bacteremia and inflammatory cytokines in mice. This biological cost may explain the lack of clonal expansion of these isolates.

Antibiotic susceptibility of invasive Neisseria meningitidis isolates from 1995 to 2008 in Sweden--the meningococcal population remains susceptible.

The susceptibility to 7 antibiotics was determined for all Swedish invasive Neisseria meningitidis isolates from 1995 to 2008 (N = 717). In general, these remain highly susceptible to the antibiotics recommended for use. Accordingly, penicillin G remains effective for the treatment of invasive meningococcal disease and ciprofloxacin appropriate for prophylaxis.

Antibiotic susceptibility and characteristics of Neisseria meningitidis isolates from the African meningitis belt, 2000 to 2006: phenotypic and genotypic perspectives.

Up-to-date information regarding the antibiotic susceptibility of Neisseria meningitidis strains from African countries is highly limited. Our aim was to comprehensively describe the antibiotic susceptibilities of a selection of N. meningitidis isolates recovered between 2000 and 2006 from 18 African countries, mainly those within the meningitis belt. Susceptibilities to 11 antibiotics were determined using Etest for 137 N. meningitidis isolates (stringently selected from 693 available isolates). The isolates were also characterized by serogrouping, multilocus sequence typing, genosubtyping, and penA allele identification. All N. meningitidis isolates were susceptible to ceftriaxone, chloramphenicol, and ciprofloxacin. No isolate produced beta-lactamase. Only three isolates (2%) displayed reduced susceptibility to penicillin G. The two isolates with the highest penicillin G MICs were the only isolates showing reduced susceptibility to ampicillin and cefuroxime. One of these isolates was also resistant to penicillin V. One percent of isolates displayed reduced susceptibility to rifampin, while 52% of the isolates were resistant to tetracycline, 74% were resistant to erythromycin, and 94% were resistant to sulfadiazine. The MICs of rifampin and tetracycline seemed to be associated with the serogroup of the isolates. In total, 18 sequence types (STs), 10 genosubtypes, and 8 different penA alleles were identified; the most common were ST-7, P1.20,9,35-1, and penA4, respectively. A high level of correlation was found between ST, genosubtype, and penA allele. In conclusion, N. meningitidis isolates from the African meningitis belt remain highly susceptible to the antibiotics used. Regarding beta-lactam antibiotics, rare isolates showed a reduced susceptibility to penicillins, but the expanded-spectrum cephalosporins are not affected at present.

Prevalence and sequence variations of the genes encoding the five antigens included in the novel 5CVMB vaccine covering group B meningococcal disease.

During the recent years, projects are in progress for designing broad-range non-capsular-based meningococcal vaccines, covering also serogroup B isolates. We have examined three genes encoding antigens (NadA, GNA1030 and GNA2091) included in a novel vaccine, i.e. the 5 Component Vaccine against Meningococcus B (5CVMB), in terms of gene prevalence and sequence variations. These data were combined with the results from a similar study, examining the two additional antigens included in the 5CVMB (fHbp and GNA2132). nadA and fHbp v. 1 were present in 38% (n=36), respectively 71% (n=67) of the isolates, whereas gna2132, gna1030 and gna2091 were present in all the Neisseria meningitidis isolates tested (n=95). The level of amino acid conservation was relatively high in GNA1030 (93%), GNA2091 (92%), and within the main variants of NadA and fHbp. GNA2132 (54% of the amino acids conserved) appeared to be the most diversified antigen. Consequently, the theoretical coverage of the 5CVMB antigens and the feasibility to use these in a broad-range meningococcal vaccine is appealing.