Genomic surveillance of Neisseria meningitidis serogroup W in Portugal from 2003 to 2019.

In recent years, a change in the epidemiology of meningococcal disease caused by Neisseria meningitidis serogroup W (MenW) has been observed worldwide, with the emergence of new sublineages associated with a higher rate of fatal cases. The present study intends to describe the epidemiology of invasive meningococcal disease (IMD) due to MenW in Portugal between 2003 and 2019, and to genetically characterize population structure. Despite MenW has a low incidence in Portugal, having almost disappeared from 2008 to 2015, since 2016, the number of MenW cases has been steadily increasing at a rate of ~ twofold per year, with more than 80% of the characterized isolates belonging to clonal complex 11 (cc11). Core-genome phylogeny of 25 Portuguese (PT) MenW isolates showed a strain clustering mainly either with the Original UK or the UK 2013 sublineages. Our study also reported for the first time the presence of distinct prophages with a notable overrepresentation of an ~ 32-35-kb PS_1-like prophage found in MenW cc11 genomes. The presence of the PS_1-like prophage in almost all 4723 cc11 genomes selected from Neisseria PubMLST database regardless of the capsular group they belong to suggests an ancestral acquisition of this mobile element prior to capsular switching events. Overall, by mimicking the scenario observed worldwide, this study reinforces the importance of a close monitoring of MenW disease, especially from cc11, in order to promptly adapt the vaccination plan for IMD control in Portugal. Moreover, future studies are needed to understand the putative contribution of prophages to fitness and virulence of PT MenW strains.

Development and Immunogenicity of a Brazilian Glycoconjugate vaccine against Meningococcal W in a Pilot Scale.

Recent changes in the epidemiology of meningococcal have been reported and meningococcal group W (MenW) has become the third most prevalent group isolated in Brazil in the last 10 years. In this study we have developed a conjugate vaccine for MenW using a modified reductive amination conjugation method through a covalent linkage between periodate-oxidized MenW non-O-acetylated polysaccharide and hydrazide-activated monomeric tetanus toxoid. Process control of bulks was done by physicochemical analysis including polysaccharide and protein quantification, high performance liquid chromatography - size exclusion chromatography, capillary electrophoresis, and hydrogen nuclear magnetic resonance. Conjugate bulks were best produced with concentration of polysaccharide twice as high as protein, at room temperature, and pH approximately 6.0. A scaled-up bulk (100 mg scale) was formulated and inoculated intramuscularly in mice in a dose-response study (0.1, 0.5, 1.0 and 10.0 µg of polysaccharide/dose). The immunogenicity of conjugate bulks was determined by serum bactericidal assay and ELISA assays of serum from immunized mice. ELISA and SBA titers revealed high titers of IgG and demonstrated the functionality of the antibodies produced in all doses studied 15 days after the third dose. However, significant differences were observed among them by ELISA. In conclusion, this study established the best conditions to produce MenW conjugate bulks and showed the efficacy of the obtained conjugate bulk in induce a good immune response in mice. Further experiments will need to be done to scale up the conjugation reaction and then allow the use of this conjugate in clinical trials.

Neisseria meningitidis Serogroup Z Meningitis in a Child With Complement C8 Deficiency and Potential Cross Protection of the MenB-4C Vaccine.

Complement deficient patients are susceptible to rare meningococcal serogroups. A 6-year-old girl presented with serogroup Z meningitis. This led to identification of a C8 deficiency. The MenB-4C vaccine induced cross-reactive antibodies to serogroup Z and increased in vitro opsonophagocytic killing and may thus protect complement deficient patients.

Carbohydrate based meningococcal vaccines: past and present overview.

Neisseria meningitidis is a major cause of bacterial meningitidis worldwide. Children less than five years and adolescents are particularly affected. Nearly all invasive strains are surrounded by a polysaccharide capsule, based on which, 12 N. meningitidis serogroups are differentiated. Six of them, A, B, C, W, X, and Y, cause the vast majority of infections in humans. Mono- and multi-valent carbohydrate-based vaccines against meningococcal infections have been licensed or are currently in clinical development. In this mini-review, an overview of the past and present approaches for producing meningococcal glycoconjugate vaccines is provided.

Genome-wide methylome analysis of two strains belonging to the hypervirulent Neisseria meningitidis serogroup W ST-11 clonal complex.

A rising incidence of meningococcal serogroup W disease has been evident in many countries worldwide. Serogroup W isolates belonging to the sequence type (ST)-11 clonal complex have been associated with atypical symptoms and increased case fatality rates. The continued expansion of this clonal complex in the later part of the 2010s has been largely due to a shift from the so-called original UK strain to the 2013 strain. Here we used single-molecule real-time (SMRT) sequencing to determine the methylomes of the two major serogroup W strains belonging to ST-11 clonal complex. Five methylated motifs were identified in this study, and three of the motifs, namely 5'-GATC-3', 5'-GAAGG-3', 5'-GCGCGC-3', were found in all 13 isolates investigated. The results showed no strain-specific motifs or difference in active restriction modification systems between the two strains. Two phase variable methylases were identified and the enrichment or depletion of the methylation motifs generated by these methylases varied between the two strains. Results from this work give further insight into the low diversity of methylomes in highly related strains and encourage further research to decipher the role of regions with under- or overrepresented methylation motifs.

Peritonitis primària per N. meningitidis serogrup B / Peritonitis primaria por N. Meningitidis serogrupo B / Primary peritonitis by serogroup B N. Meningitidis

INTRODUCCIÓ: Neisseria meningitidis és un dels principals causants de sèpsia I meningitis adquirida a la comunitat en pacients pediàtrics. Se'n descriuen presentacions atípiques, com la peritonitis primària. Presentem aquest cas per la seva excepcionalitat: una patologia infreqüent en edat pediàtrica causada per un germen poc habitual. CAS CLÍNIC: Es presenta un lactant de 4 mesos que acut al servei d'Urgències amb els seus pares per febre I lesions cutànies disseminades de 24 hores d'evolució, amb vòmits I diarrea. Davant la sospita de sèpsia, s'inicien maniobres de reanimació inicial, amb càrrega de volum I antibioteràpia, prèvia extracció d'hemocultiu. Tot I la millora clínica inicial, persisteix la palpació dolorosa a l'hipogastri. Es valora juntament amb Cirurgia I es fa una tomografia computada abdominal que mostra hipercaptació de nanses d'intestí prim I líquid intraabdominal compatible amb procés inflamatori peritoneal. Es fa una laparoscòpia exploradora I s'obté abundant líquid purulent; es revisen íntegrament budell I colon sense observar lesions. Es manté a dieta absoluta durant 48 hores després de la cirurgia, amb progressió alimentària posterior sense incidències. S'obtenen resultats microbiològics: hemocultiu positiu per N. meningitidis B I reacció en cadena de la polimerasa N. meningitidis positiva en sang I en líquids cefalorraquidi I peritoneal. COMENTARIS: La peritonitis primària en pacients pediàtrics sans és una entitat de baixa incidència. Entre els gèrmens descrits en la literatura és molt poc freqüent trobar N. meningitidis. És necessari pensar en aquesta entitat davant d'un pacient amb malaltia invasiva per N. meningitidis I exploració abdominal patològica. La base del tractament són la cirurgia I l'antibioteràpia endovenosa

INTRODUCCIÓN: Neisseria meningitidis es una de las principales causas de sepsis y meningitis adquirida en la comunidad en pacientes pediátricos. Se describen presentaciones atípicas entre las cuales consta de forma excepcional la peritonitis primaria. Este caso ha sido seleccionado por su excepcionalidad, pues se trata de una patología poco frecuente en edad pediátrica y causada por un germen poco habitual. CASO CLÍNICO: Se presenta a un lactante de 4 meses que acude al servicio de Urgencias con sus padres por fiebre y lesiones cutáneas diseminadas de 24 horas de evolución, junto con vómitos y diarrea. Ante la sospecha de sepsis se realizan maniobras de reanimación inicial con carga de volumen y antibioterapia, con previa toma de hemocultivo. A pesar de la mejoría clínica inicial, persiste palpación dolorosa en hipogastrio. Se valora junto con Cirugía y se realiza tomografía computarizada abdominal que muestra hipercaptación de asas de intestino delgado, asociado a líquido intra-abdominal compatible con proceso inflamatorio peritoneal. Se realiza laparoscopia exploradora y se obtiene abundante líquido purulento y esfacelos; se revisan de forma íntegra intestino y colon sin observar lesiones. Se mantiene a dieta absoluta durante 48 horas tras la cirugía, con progresión alimentaria posterior y sin incidencias. Se obtienen resultados microbiológicos: hemocultivo positivo para N. meningitidis B sensible y reacción en cadena de la polimerasa N. meningitidis B positiva en sangre y en líquidos cefalorraquídeo y peritoneal. COMENTARIOS: La peritonitis primaria en pacientes pediátricos sanos es una entidad de baja incidencia. Entre los gérmenes descritos en la literatura es muy poco frecuente encontrar N. meningitidis. Es necesario pensar en esta entidad ante un paciente con infección invasiva por N. meningitidis con exploración abdominal patológica. La base de su tratamiento son la cirugía y la antibioterapia endovenosa

INTRODUCTION: Neisseria meningitidis is one of the main causes of community acquired sepsis and meningitis in children. Atypical presentations have been described, one of them is primary peritonitis. CASE REPORT: We present a 4-month-old infant admitted to the emergency department with fever and disseminated petechia of 24 hours of evolution along with vomiting and diarrhea. With the diagnosis of sepsis, resuscitation maneuvers were started with intravenous fluids and antibiotics after a blood culture was obtained. In spite of the initial clinical improvement, painful palpation persisted in hypogastrium. The patient was assessed together with surgery and an abdominal computed tomography was performed, which showed a small intestine loop enlargement, associated with intraabdominal fluid that suggested peritonitis. Exploratory laparoscopy was performed, obtaining abundant purulent fluid and necrotic and inflammatory tissue. The entire bowel and colon were inspected and no lesions were found. The patient remained fasting for 48 hours after surgery. Enteral nutrition was started afterwards with no complications. N. menigitidis B was isolated in blood culture and polymerase chain reaction positive for N. meningitidis B was positive in blood and in cerebrospinal and peritoneal fluids. COMMENTS: Primary peritonitis in healthy pediatric patients is rare. N. meningitidis is very rare causative pathogen. Primary peritonitis should be considered when having a patient with invasive infection by N. meningitidis with abnormal abdominal examination. Treatment includes surgery and intravenous antibiotics

Surveillance of invasive meningococcal disease in the south of Brazil: considerations of immunization programme.

Invasive meningococcal disease (IMD) is a major cause of meningitis and septicaemia worldwide. The switches in serogroup predominance contribute to the unpredictable nature of the disease with significant health impacts. The aim of this study was to determine the epidemiological profile of IMD in Rio Grande do Sul, Santa Catarina and Paraná, three states in the south of Brazil. All meningitis cases confirmed by clinical and/or laboratory criteria notified to the national information system for notifiable diseases between 2015 and 2019 were analysed. Proportions of serogroup and incidence by age were calculated. A total of 17 894 cases of IMD were reported during this period. Of these, 9029 cases (50 %) were due to serogroup C. Furthermore, serogroup W was responsible for almost half of the cases among children younger than 5 years old during 2017 and 2018, with an overall incidence of 33.3 cases per 100 000 infants. Despite the reduction in serogroup C after the introduction of meningococcal C conjugate vaccine into a childhood immunization programme in Brazil, it remains a significant healthcare issue in the south of the country. Changes in disease epidemiology were observed and serogroup W was the most common among children below 5 years of age in 2017 and 2018. Although future cost-effectiveness studies are necessary, our results could have future implications for meningococcal vaccination programmes.

Antibiotic susceptibility and molecular analysis of invasive Neisseria meningitidis recovered in the Republic of Ireland, 1996 to 2016.

This study examined the antimicrobial susceptibility of invasive meningococcal disease (IMD)-associated Neisseria meningitidis recovered in the Republic of Ireland between 1996 and 2016. In total, 1359 isolates representing over one-third of all laboratory-confirmed cases of IMD diagnosed each epidemiological year (EY; July 1-June 30) were analysed. All isolates were susceptible to ciprofloxacin, rifampicin and cefotaxime and 74% and 87% were susceptible to sulphonamide and penicillin, respectively. The proportion of isolates exhibiting reduced susceptibility to penicillin increased significantly during the study with no evidence of major clonal expansion or horizontal spread of a specific penA allele. Greater diversity observed among recently recovered meningococci and specifically among isolates exhibiting reduced penicillin susceptibility contributed to the overall increase in penA allele diversity throughout. The emergence and dissemination of strains with phenotypic and genotypic reduced susceptibility to penicillin increase the need for continued surveillance of antimicrobial susceptibility of meningococci in the Republic of Ireland especially in view of the recommendation of penicillin G as empiric treatment of choice for pre-hospital management.

Emergence of a novel urogenital-tropic Neisseria meningitidis.

PURPOSE OF REVIEW: Neisseria meningitidis (Nm) is primarily associated with asymptomatic nasopharyngeal carriage and invasive meningococcal disease (sepsis and meningitis), but like N. gonorrhoea (Ng), Nm can colonize urogenital and rectal mucosal surfaces and cause disease. First noted in 2015, but with origins in 2011, male urethritis clusters caused by a novel Nm clade were reported in the USA (the US_NmUC). This review describes research developments that characterize this urogenital-tropic Nm. RECENT FINDINGS: The US_NmUC evolved from encapsulated Nm serogroup C strains. Loss of capsule expression, lipooligosaccharide (LOS) sialylation, genetic acquisition of gonococcal alleles (including the gonococcal anaerobic growth aniA/norB cassette), antimicrobial peptide heteroresistance and high surface expression of a unique factor-H-binding protein, can contribute to the urethra-tropic phenotype. Loss-of-function mutations in mtrC are overrepresented in clade isolates. Similar to Ng, repeat US_NmUC urethritis episodes can occur. The US_NmUC is now circulating in the UK and Southeast Asia. Genomic sequencing has defined the clade and rapid diagnostic tests are being developed for surveillance. SUMMARY: The US_NmUC emerged as a cause of urethritis due to acquisition of gonococcal genetic determinants and phenotypic traits that facilitate urogenital tract infection. The epidemiology and pathogenesis of this urogenital-tropic pathogen continues to be defined.

Emergence of ciprofloxacin-resistant Neisseria meningitidis B from asymptomatic carriers during an outbreak in Peru, 2017.

Asymptomatic carriers are a likely source of transmission of Neisseria meningitidis to close contacts who are placed at a higher risk for invasive meningococcal disease (IMD). Although N. meningitidis ciprofloxacin-resistance is rare, there have been an increase in the reports of resistant isolates mainly in patients diagnosed with IMD, and little is known about the N. meningitidis ciprofloxacin-resistance in the carrier populations. We performed a pharyngeal carriage study during a 2017 military setting outbreak in Peru, caused by a ciprofloxacin-resistant N. meningitidis B. The isolates analysed came from two hospitalized cases and six asymptomatic carriers. Whole-genome sequence-based analysis was performed and showed that strains carrying the Thr91Ile mutation, in the gene encoding for subunit A of DNA gyrase (gyrA), were responsible for the fluoroquinolone resistance (MICs ≥0.256 µg ml-1) and were closely related to highly virulent strains from France, Norway and the UK. Phylogenetic analysis of the gyrA gene revealed that likely these Peruvian isolates acquired resistance through horizontal gene transfer from Neisseria lactamica. Our study provides evidence for the emergence and propagation of ciprofloxacin-resistant N. meningitidis B from asymptomatic carriers, and recommends the introduction of serogroup B vaccines for high-risk populations.

Molecular insights into meningococcal carriage isolates from Burkina Faso 7 years after introduction of a serogroup A meningococcal conjugate vaccine.

In 2010, Burkina Faso completed the first nationwide mass-vaccination campaign of a meningococcal A conjugate vaccine, drastically reducing the incidence of disease caused by serogroup A meningococci. Since then, other strains, such as those belonging to serogroups W, X and C, have continued to cause outbreaks within the region. A carriage study was conducted in 2016 and 2017 in the country to characterize the meningococcal strains circulating among healthy individuals following the mass-vaccination campaign. Four cross-sectional carriage evaluation rounds were conducted in two districts of Burkina Faso, Kaya and Ouahigouya. Oropharyngeal swabs were collected for the detection of Neisseria meningitidis by culture. Confirmed N. meningitidis isolates underwent whole-genome sequencing for molecular characterization. Among 13 758 participants, 1035 (7.5 %) N. meningitidis isolates were recovered. Most isolates (934/1035; 90.2 %) were non-groupable and primarily belonged to clonal complex (CC) 192 (822/934; 88 %). Groupable isolates (101/1035; 9.8 %) primarily belonged to CCs associated with recent outbreaks in the region, such as CC11 (serogroup W) and CC10217 (serogroup C); carried serogroup A isolates were not detected. Phylogenetic analysis revealed several CC11 strains circulating within the country, several of which were closely related to invasive isolates. Three sequence types (STs) were identified among eleven CC10217 carriage isolates, two of which have caused recent outbreaks in the region (ST-10217 and ST-12446). Our results show the importance of carriage studies to track the outbreak-associated strains circulating within the population in order to inform future vaccination strategies and molecular surveillance programmes.

Epidemiology and antibiotic resistance profile of bacterial meningitis in Morocco from 2015 to 2018.

Over a 4-year study period from 2015 to 2018, altogether 183 isolates of bacterial meningitis were collected from 12 hospitals covering the entire Moroccan territory. Neisseria meningitidis represented 58.5%, Streptococcus pneumoniae 35.5%, and Haemophilus influenzae type b 6%. H. influenzae type b mainly affected 5-year-olds and unvaccinated adults. N. meningitidis serogroup B represented 90.7% followed by serogroup W135 with 6.5%. Decreased susceptibility to penicillin G (DSPG) for all isolates accounted for 15.7%, with 11.6% being resistant to penicillin G (PG) and 4.1% decreased susceptibility. Cumulative results of all strains showed 2.7% decreased susceptibility to amoxicillin and 3.3% resistant, 2.2% of isolates were resistant to third-generation cephalosporin and 2.2% were decreased susceptible, 5.5% were resistant to chloramphenicol and 2.7% were resistant to rifampin. The frequency of DSPG observed in our study is more common in S. pneumoniae than in N. meningitidis (P < 0.05). These isolates have been found to be highly susceptible to antibiotics used for treatment and prophylaxis chemotherapy and the observed resistance remains rare. The impact of introduction of conjugate vaccines against H. influenzae type b and S. pneumoniae (PCVs) is an advantage in reducing meningitis cases due to these two species.

Diversification in immunogenicity genes caused by selective pressures in invasive meningococci.

We studied population genomics of 486 Neisseria meningitidis isolates causing meningitis in the Netherlands during the period 1979-2003 and 2006-2013 using whole-genome sequencing to evaluate the impact of a hyperendemic period of serogroup B invasive disease. The majority of serogroup B isolates belonged to ST-41/44 (41 %) and ST-32 complex (16 %). Comparing the time periods, before and after the decline of serogroup B invasive disease, there was a decrease of ST-41/44 complex sequences (P=0.002). We observed the expansion of a sub-lineage within ST-41/44 complex sequences being associated with isolation from the 1979-2003 time period (P=0.014). Isolates belonging to this sub-lineage expansion within ST-41/44 complex were marked by four antigen allele variants. Presence of these allele variants was associated with isolation from the 1979-2003 time period after correction for multiple testing (Wald test, P=0.0043 for FetA 1-5; P=0.0035 for FHbp 14; P=0.012 for PorA 7-2.4 and P=0.0031 for NHBA two peptide allele). These sequences were associated with 4CMenB vaccine coverage (Fisher's exact test, P<0.001). Outside of the sub-lineage expansion, isolates with markedly lower levels of predicted vaccine coverage clustered in phylogenetic groups showing a trend towards isolation in the 2006-2013 time period (P=0.08). In conclusion, we show the emergence and decline of a sub-lineage expansion within ST-41/44 complex isolates concurrent with a hyperendemic period in meningococcal meningitis. The expansion was marked by specific antigen peptide allele combinations. We observed preliminary evidence for decreasing 4CMenB vaccine coverage in the post-hyperendemic period.

Epidemiology of invasive meningococcal disease in Cyprus 2004 to 2018.

BackgroundDespite progress in the management of invasive meningococcal disease (IMD) it causes significant mortality and sequelae.AimThis study aims to describe the epidemiology and clinical characteristics of IMD in Cyprus and discuss the current immunisation programmes.MethodsThis is a retrospective study of all cases of IMD notified to the Ministry of Health between 2004 and 2018. Demographic, epidemiological, clinical and microbiological data were collected when a new case was notified. Risk factors associated with mortality were investigated using univariable logistic regression.Results54 cases of IMD were recorded, an overall incidence of 0.4 cases per 100,000 population. The incidence rate was highest among infants (7.2/100,000) and adolescents (1.4/100,000). Case fatality rate was 10.4%. Serogroup B accounted for 24 of 40 cases caused by known serogroup. Serogroups W and Y comprised nine cases and were responsible for most fatal cases. Serogroup C was the cause in only four cases. There was an increase in the odds of death with increasing age, while the presence of meningitis in the clinical picture was found to be associated with lower odds of death.ConclusionDespite the low incidence of IMD in Cyprus, it remains an important cause of morbidity and mortality. Serogroup B is the most frequent serogroup, while incidence of serogroups W and Y is rising. Monitoring new cases and yearly evaluation of the immunisation programmes by the National Immunization Technical Advisory Group (NITAG) is essential for successful control of the disease.

Neisseria meningitidis aislada de muestras de hombres que tienen sexo con hombres / Neisseria meningitidis isolated from patients in men who have sex with men

Resumen En los períodos 2000-2004 y 2014-2015 se investigó la presencia de Neisseria meningitidis en 1.143 y 544 hombres que tienen sexo con hombres respectivamente, atendidos en el marco de un programa de enfermedades de transmisión sexual. Se determinó la prevalencia de este agente, su distribución en serogrupos y su sensibilidad a los antimicrobianos. Se obtuvieron hisopados faríngeos, rectales y uretrales, que se sembraron en medio selectivo Thayer Martin modificado. La identificación se realizó mediante pruebas bioquímicas convencionales y por espectrometría de masas (MALDI-TOF). En el segundo período estudiado, sobre 85 aislamientos procedentes de faringes se investigaron los serogrupos B, C, W e Y mediante PCR. Se determinó la CIM de penicilina, ceftriaxona, rifampicina, azitromicina y ciprofloxacina en 66 aislamientos obtenidos en el primer período y en 102 logrados en el segundo. La prevalencia de N. meningitidis fue del 17,8% en el primer período y del 28,1% en el segundo; este microorganismo se aisló más frecuentemente de fauces. Los serogrupos hallados fueron B (31,5%), Y (7,6%) y W (3,3%), con un 9,8% de aislamientos no capsulados; los restantes corresponderían a otros serogrupos. El 34,8% y el 63,7% de los aislados estudiados correspondientes al primer y segundo período, respectivamente, tuvieron sensibilidad intermedia a la penicilina, y un 11,8% de los evaluados en el segundo período fueron resistentes a dicho antibiótico. Todos los aislados estudiados fueron sensibles a ceftriaxona y a ciprofloxacina (excepto 3, con CIM entre 0,25 y 0,5(g/ml), el 3% fueron resistentes a rifampicina y el 2% fueron no sensibles a azitromicina. La portación de N. meningitidis en hombres que tienen sexo con hombres fue elevada y hubo un alto porcentaje de cepas no sensibles a penicilina. El serogrupo B fue prevalente.

Abstract During the periods 2000-2004 and 2014-2015, Neisseria meningitidis was investigated in men who have sex with men, 1143 and 544 respectively, who consulted in the sexually-transmitted disease program. Prevalence, serogroup distribution and susceptibility to antibiotics were determined. Pharyngeal, rectal and urethral swabs were cultivated on selective Thayer-Martin modified medium. The identification was performed by biochemical tests and mass spectrometry by MALDI-TOF. Serogroups B, C, W and Y were investigated by PCR in 85 isolates recovered from the pharynx belonging to the second period. MICs of penicillin, ceftriaxone, rifampicin, azithromycin and ciprofloxacin were determined for 66 and 102 isolates from periods 1 and 2 respectively, according to CLSI. The prevalence of N. meningitidis was 17.8% and 28.1%, in periods 1 and 2 respectively; the isolates were mainly recovered from the pharynx. The distribution of serogroups was B 31.5%; Y 7.6%; W 3.3% and 9.8% non-capsulated and the rest would belong to other serogroups. Isolates classified as intermediate to penicillin were 34.8% and 63.7% (first and second periods, respectively); moreover, 11.8% of the isolates from the second period were resistant. All isolates were susceptible to ceftriaxone, to ciprofloxacin (except 3 isolates with MIC values between 0.25 and 0.5(g/ml), 3% were resistant to rifampicin and 2% were not susceptible to azithromicin. The prevalence of N. meningitidis carriage in men who have sex with men was high with a high rate of penicillin non-susceptible isolates. B was the prevalent serogroup.

A diagnostic challenge in clinical laboratory: Misidentification of Neisseria subflava as Neisseria meningitidis by MALDI-TOF MS.

MALDI-TOF MS provides fast, easy to perform and cost-effective diagnosis in clinical microbiology laboratories, however in some cases results of MALDI-TOF MS should be confirmed with additional tests. This confirmation is especially important for causes of life-threatening infections like Neisseria meningitidis. In our laboratory, three isolates were identified as N. meningitidis by Bruker MALDI Biotyper (BD, USA) between April 2018 and March 2019 from clinical specimens of blood, sputum, and urine. 16S rRNA sequencing was performed for further investigation. Two of the isolates were identified as Neisseria subflava and only one was confirmed as N. meningitidis by sequencing. These results show that MALDI-TOF MS is not always reliable in the diagnosis of N. meningitidis and clinical microbiologists should confirm these results with additional tests. Also, clinical correlations should be determined. Accurate identification of this microorganism is very important because of the necessity of prophylactic antimicrobial usage and biosafety precautions. Enlarged databases of Neisseria species are needed to overcome this problem.

Evolutionary analysis of gyrA gene from Neisseria meningitidis bacterial strains of clonal complex 4821 collected in China between 1978 and 2016.

BACKGROUND: Neisseria meningitidis (N.meningitidis) bacteria belonging to clonal complex 4821 (CC4821) have been mainly reported in China and have been characterized by a high resistance rate to ciprofloxacin (CIP). The aim of this study was to assess the evolution of the DNA gyrase A (gyrA) gene from N.meningitidis CC4821 strains collected in China between 1978 and 2016. The complete sequence of gyrA gene from 77 strains are reported in this study and analyzed in the context of publicly available sequences from N. meningitidis of other CCs as well as other Neisseria species. RESULTS: The phylogenetic analysis of CC4821 gyrA gene reveals at least 5 distinct genetic clusters. These clusters are not CC4821-specific showing that gyrA evolution is independent of CC4821 evolution. Some clusters contain sequences from other Neisseria species. Recombination within N.meningitidis strains and between Neisseria species was identified in SimPlot analysis. Finally, amino acid substitutions within GyrA protein were analyzed. Only one position, 91 (83 in E.coli gyrA gene), was linked to CIP resistance. Thirty-one additional putative resistance markers were identified, as amino acid substitutions were only found in resistant strains. CONCLUSIONS: The evolution of gyrA gene of CC4821 N.meningitidis strains is not dependent on CC4821 evolution or on CIP resistance phenotype. Only amino acid 91 is linked to CIP resistance phenotype. Finally, recombination inter- and intra-species is likely to result in the acquisition of various resistance markers, 31 of them being putatively mapped in the present study. Analyzing the evolution of gyrA gene within CC4821 strains is critical to monitor the CIP resistance phenotype and the acquisition of new resistance markers. Such studies are necessary for the control of the meningococcal disease and the development of new drugs targeting DNA gyrase.

Detection of the United States Neisseria meningitidis urethritis clade in the United Kingdom, August and December 2019 - emergence of multiple antibiotic resistance calls for vigilance.

Since 2015 in the United States (US), the US Neisseria meningitidis urethritis clade (US_NmUC) has caused a large multistate outbreak of urethritis among heterosexual males. Its 'parent' strain caused numerous outbreaks of invasive meningococcal disease among men who have sex with men in Europe and North America. We highlight the arrival and dissemination of US_NmUC in the United Kingdom and the emergence of multiple antibiotic resistance. Surveillance systems should be developed that include anogenital meningococci.