Estimates of the reproductive numbers and demographic reconstruction of outbreak associated with C:P1.5-1,10-8:F3-6:ST-11(cc11) Neisseria meningitidis strains.

BACKGROUND: Neisseria meningitidis can cause sporadic cases and outbreaks of invasive disease, including meningitis and sepsis. The meningococcal serogroup C (MenC) is the second most common serogroup in Italy after MenB. In this study we have estimated the reproductive numbers and the demographic reconstruction on the genomes of invasive N. meningitidis C:P1.5-1,10-8:F3-6:ST-11(cc11) strains isolated in Italy in 2012 - 2017, a period that includes the outbreak in Tuscany. METHODS: The genomes of N. meningitidis were sequenced using the Illumina MiSeq platform, through the whole genome sequencing (WGS) method and were analyzed by the core genome MLST (cgMLST) approach, using the BIGSdb Genome Comparator tool implemented on the PubMLST website. A Bayesian method was applied to study population dynamics across the entire N. meningitidis dataset. The basic reproduction number R0, which indicates the average number of secondary cases generated by a single primary case, was calculated using a Bayesian method, on the dataset and on the two subsets. The effective reproduction number R(t), defined as the average number of secondary cases per infectious case in a population, made up of susceptible and non-susceptible hosts was studied on the Tuscany dataset, with a Bayesian method. RESULTS: An increase in the effective number of the N. meningitidis infections was observed between 2013 and 2016. The estimated R0 parameter was 1.31 (95% HPD: 1.03 - 1.64), 1.22 (95% HPD: 0.90 - 1.64) and 1.4 (95% HPD: 0.91 - 1.9) for the entire dataset, first and second subset, respectively. The BDSKY estimated an initial R(t) of about 2.0 (95% HPD: 0.04 - 5.0), which showed a growing trend at the end of 2014, reaching an average value of 3.22 in 2015, and then declining below 1 from the year 2016. CONCLUSION: Monitoring the effective reproduction number can help to inform future vaccination strategies. The increase in the reproductive number for the Tuscany dataset, was consistent with the amplification event that led to the Tuscany outbreak. Subsequently, the intervention that led to the decline of the cases was followed, suggesting a high effectiveness of the vaccination campaign.

Development and pre-clinical evaluation of a synthetic oligosaccharide-protein conjugate vaccine against Neisseria meningitidis serogroup C.

Significant improvement has been made in the development of vaccines against Neisseria meningitidis infections since the introduction of polysaccharide-protein conjugate vaccines. Conventional bacterial capsular polysaccharide (PS) based conjugate vaccines require unique and expensive manufacturing facilities, complex production processes and extensive quality testing. Synthetic oligosaccharide (OS) based approach is one of the novel technologies that is being developed to simplify production of conjugate vaccines. OSs can be chemically synthesized to a desired length long enough to represent the antigenic epitopes which often present as a homogenous mixture. We prepared OSs corresponding to tetramer and octamer of N. meningitidis serogroup C (MenC) PS by organic synthesis. The MenC tetramer and octamer were further conjugated with tetanus toxoid to produce respective monovalent conjugates having the desired physico-chemical characteristics. The conjugates were evaluated in a mouse model for immunogenicity and compared with a licensed PS conjugate vaccine. Synthetic conjugates could induce anti-MenC PS IgG as well as serum bactericidal titers at levels comparable to those elicited by the licensed vaccine. The increase in length of synthetic oligomers from tetramer to octamer did not appear to increase immunogenicity. The results establish the pre-clinical proof of concept for a synthetic MenC oligosaccharide conjugate vaccine candidate.

cgMLST characterisation of invasive Neisseria meningitidis serogroup C and W strains associated with increasing disease incidence in the Republic of Ireland.

INTRODUCTION AND AIMS: Since 2013 MenC and MenW disease incidence and associated mortality rates have increased in the Republic of Ireland. From 2002/2003 to 2012/2013, the average annual MenC incidence was 0.08/100,000, which increased to 0.34/100,000 during 2013/2014 to 2017/18, peaking in 2016/17 (0.72/100,000) with an associated case fatality rate (CFR) of 14.7%. MenW disease incidence has increased each year from 0.02/100,000 in 2013/2014, to 0.29/100,000 in 2017/18, with an associated CFR of 28.6%. We aimed to characterise and relate recent MenC isolates to the previously prevalent MenC:cc11 ET-15 clones, and also characterise and relate recent MenW isolates to the novel 'Hajj' clones. METHODS: Using WGS we characterised invasive (n = 74, 1997/98 to 2016/17) and carried (n = 16, 2016/17) MenC isolates, and invasive (n = 18, 2010/11 to 2016/17) and carried (n = 15, 2016/17) MenW isolates. Genomes were assembled using VelvethOptimiser and stored on the PubMLST Neisseria Bacterial Isolate Genome Sequence Database. Isolates were compared using the cgMLST approach. RESULTS: Most MenC and MenW isolates identified were cc11. A single MenC:cc11 sub-lineage contained the majority (68%, n = 19/28) of recent MenC:cc11 disease isolates and all carried MenC:cc11 isolates, which were interspersed and distinct from the historically significant ET-15 clones. MenW:cc11 study isolates clustered among international examples of both the original UK 2009 MenW:cc11, and novel 2013 MenW:cc11clones. CONCLUSIONS: We have shown that the majority of recent MenC disease incidence was caused by strain types distinct from the MenC:cc11 ET-15 clone of the late 1990s, which still circulate but have caused only sporadic disease in recent years. We have identified that the same aggressive MenW clone now established in several other European countries, is endemic in the RoI and responsible for the recent MenW incidence increases. This data informed the National immunisation Advisory Committee, who are currently deliberating a vaccine policy change to protect teenagers.

Virulence Traits of a Serogroup C Meningococcus and Isogenic cssA Mutant, Defective in Surface-Exposed Sialic Acid, in a Murine Model of Meningitis.

In serogroup C Neisseria meningitidis, the cssA (siaA) gene codes for an UDP-N-acetylglucosamine 2-epimerase that catalyzes the conversion of UDP-N-acetyl-α-d-glucosamine into N-acetyl-d-mannosamine and UDP in the first step in sialic acid biosynthesis. This enzyme is required for the biosynthesis of the (α2→9)-linked polysialic acid capsule and for lipooligosaccharide (LOS) sialylation. In this study, we have used a reference serogroup C meningococcal strain and an isogenic cssA knockout mutant to investigate the pathogenetic role of surface-exposed sialic acids in a model of meningitis based on intracisternal inoculation of BALB/c mice. Results confirmed the key role of surface-exposed sialic acids in meningococcal pathogenesis. The 50% lethal dose (LD50) of the wild-type strain 93/4286 was about four orders of magnitude lower than that of the cssA mutant. Compared to the wild-type strain, the ability of this mutant to replicate in brain and spread systemically was severely impaired. Evaluation of brain damage evidenced a significant reduction in cerebral hemorrhages in mice infected with the mutant in comparison with the levels in those challenged with the wild-type strain. Histological analysis showed the typical features of bacterial meningitis, including inflammatory cells in the subarachnoid, perivascular, and ventricular spaces especially in animals infected with the wild type. Noticeably, 80% of mice infected with the wild-type strain presented with massive bacterial localization and accompanying inflammatory infiltrate in the corpus callosum, indicating high tropism of meningococci exposing sialic acids toward this brain structure and a specific involvement of the corpus callosum in the mouse model of meningococcal meningitis.

Tracking a serial killer: Integrating phylogenetic relationships, epidemiology, and geography for two invasive meningococcal disease outbreaks.

BACKGROUND: While overall rates of meningococcal disease have been declining in the United States for the past several decades, New York City (NYC) has experienced two serogroup C meningococcal disease outbreaks in 2005-2006 and in 2010-2013. The outbreaks were centered within drug use and sexual networks, were difficult to control, and required vaccine campaigns. METHODS: Whole Genome Sequencing (WGS) was used to analyze preserved meningococcal isolates collected before and during the two outbreaks. We integrated and analyzed epidemiologic, geographic, and genomic data to better understand transmission networks among patients. Betweenness centrality was used as a metric to understand the most important geographic nodes in the transmission networks. Comparative genomics was used to identify genes associated with the outbreaks. RESULTS: Neisseria meningitidis serogroup C (ST11/ET-37) was responsible for both outbreaks with each outbreak having distinct phylogenetic clusters. WGS did identify some misclassifications of isolates that were more distant from the outbreak strains, as well as those that should have been included based on high genomic similarity. Genomes for the second outbreak were more similar than the first and no polymorphism was found to either be unique or specific to either outbreak lineage. Betweenness centrality as applied to transmission networks based on phylogenetic analysis demonstrated that the outbreaks were transmitted within focal communities in NYC with few transmission events to other locations. CONCLUSIONS: Neisseria meningitidis is an ever changing pathogen and comparative genomic analyses can help elucidate how it spreads geographically to facilitate targeted interventions to interrupt transmission.

Meningococcus serogroup C clonal complex ST-10217 outbreak in Zamfara State, Northern Nigeria.

After the successful roll out of MenAfriVac, Nigeria has experienced sequential meningitis outbreaks attributed to meningococcus serogroup C (NmC). Zamfara State in North-western Nigeria recently was at the epicentre of the largest NmC outbreak in the 21st Century with 7,140 suspected meningitis cases and 553 deaths reported between December 2016 and May 2017. The overall attack rate was 155 per 100,000 population and children 5-14 years accounted for 47% (3,369/7,140) of suspected cases. The case fatality rate (CFR) among children 5-9 years was 10%, double that reported among adults ≥ 30 years (5%). NmC and pneumococcus accounted for 94% (172/184) and 5% (9/184) of the laboratory-confirmed cases, respectively. The sequenced NmC belonged to the ST-10217 clonal complex (CC). All serotyped pneumococci were PCV10 serotypes. The emergence of NmC ST-10217 CC outbreaks threatens the public health gains made by MenAfriVac, which calls for an urgent strategic action against meningitis outbreaks.

Neisseria meningitidis serogroup C sepsis and septic arthritis in an HIV-positive man.

A patient with well-controlled HIV-1 infection presented with fever and rigors, a widespread maculopapular rash, and severe generalised arthralgia. Sepsis of unknown aetiology was diagnosed, and treatment with broad-spectrum antimicrobials commenced. Following initial clinical improvement, a right knee septic arthritis developed. Microscopy and culture of the joint aspirate were negative for organisms but 16S rDNA PCR identified Neisseria meningitidis DNA, subsequently verified as capsular genogroup C, thus confirming a diagnosis of disseminated meningococcal sepsis with secondary septic arthritis.

Evolutionary Events Associated with an Outbreak of Meningococcal Disease in Men Who Have Sex with Men.

Meningococci spread via respiratory droplets, whereas the closely related gonococci are transmitted sexually. Several outbreaks of invasive meningococcal disease have been reported in Europe and the United States among men who have sex with men (MSM). We recently identified an outbreak of serogroup C meningococcal disease among MSM in Germany and France. In this study, genomic and proteomic techniques were used to analyze the outbreak isolates. In addition, genetically identical urethritis isolates were recovered from France and Germany and included in the analysis. Genome sequencing revealed that the isolates from the outbreak among MSM and from urethritis cases belonged to a clade within clonal complex 11. Proteome analysis showed they expressed nitrite reductase, enabling anaerobic growth as previously described for gonococci. Invasive isolates from MSM, but not urethritis isolates, further expressed functional human factor H binding protein associated with enhanced survival in a newly developed transgenic mouse model expressing human factor H, a complement regulatory protein. In conclusion, our data suggest that urethritis and outbreak isolates followed a joint adaptation route including adaption to the urogenital tract.

Genome-based study of a spatio-temporal cluster of invasive meningococcal disease due to Neisseria meningitidis serogroup C, clonal complex 11.

OBJECTIVES: To describe a spatio-temporal cluster of invasive meningococcal disease (IMD) due to serogroup C meningococci, occurred in a restricted area of Tuscany between January and October 2015, and the results of whole genome sequencing (WGS). METHODS: Surveillance activities and public health measures were implemented in the Region. Bacterial isolates from IMD cases were characterized by the National Reference Laboratory of the Istituto Superiore di Sanità (ISS), and WGS was performed on available strains. The kSNP software was used to identify core genome SNPs. RESULTS: Overall, 28 IMD cases due to meningococcus C were identified up to 31st October, 2015. Of them, 26 were due to meningococcus C:P1.5-1,10-8: F3-6:ST-11 (cc11) and 2 to C:P1.5-1,10-8: F3-6:ST-2780 (cc11). WGS of 13 meningococci isolated during the outbreak occurred in Tuscany in 2015 showed higher similarity when compared with those of 47 C: P1.5-1,10-8: F3-6:ST-11 (cc11) invasive strains from sporadic cases previously detected in Italy. CONCLUSIONS: A highly aggressive meningococcal C strain was involved in the cluster of severe IMD occurred in Tuscany, a Region with high vaccine coverage among children. Whether this was due to low herd immunity related to the short duration of vaccine protection needs further investigation.

Lipooligosaccharide Structures of Invasive and Carrier Isolates of Neisseria meningitidis Are Correlated with Pathogenicity and Carriage.

The degree of phosphorylation and phosphoethanolaminylation of lipid A on neisserial lipooligosaccharide (LOS), a major cell-surface antigen, can be correlated with inflammatory potential and the ability to induce immune tolerance in vitro. On the oligosaccharide of the LOS, the presence of phosphoethanolamine and sialic acid substituents can be correlated with in vitro serum resistance. In this study, we analyzed the structure of the LOS from 40 invasive isolates and 25 isolates from carriers of Neisseria meningitidis without disease. Invasive strains were classified as groups 1-3 that caused meningitis, septicemia without meningitis, and septicemia with meningitis, respectively. Intact LOS was analyzed by high resolution matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Prominent peaks for lipid A fragment ions with three phosphates and one phosphoethanolamine were detected in all LOS analyzed. LOS from groups 2 and 3 had less abundant ions for highly phosphorylated lipid A forms and induced less TNF-α in THP-1 monocytic cells compared with LOS from group 1. Lipid A from all invasive strains was hexaacylated, whereas lipid A of 6/25 carrier strains was pentaacylated. There were fewer O-acetyl groups and more phosphoethanolamine and sialic acid substitutions on the oligosaccharide from invasive compared with carrier isolates. Bioinformatic and genomic analysis of LOS biosynthetic genes indicated significant skewing to specific alleles, dependent on the disease outcome. Our results suggest that variable LOS structures have multifaceted effects on homeostatic innate immune responses that have critical impact on the pathophysiology of meningococcal infections.

Community-Based Outbreak of Neisseria meningitidis Serogroup C Infection in Men who Have Sex with Men, New York City, New York, USA, 2010-2013.

In September 2012, the New York City Department of Health and Mental Hygiene identified an outbreak of Neisseria meningitidis serogroup C invasive meningococcal disease among men who have sex with men (MSM). Twenty-two case-patients and 7 deaths were identified during August 2010-February 2013. During this period, 7 cases in non-MSM were diagnosed. The slow-moving outbreak was linked to the use of websites and mobile phone applications that connect men with male sexual partners, which complicated the epidemiologic investigation and prevention efforts. We describe the outbreak and steps taken to interrupt transmission, including an innovative and wide-ranging outreach campaign that involved direct, internet-based, and media-based communications; free vaccination events; and engagement of community and government partners. We conclude by discussing the challenges of managing an outbreak affecting a discrete community of MSM and the benefits of using social networking technology to reach this at-risk population.

Replacement of Neisseria meningitidis C cc11/ET-15 variant by a cc103 hypervirulent clone, Brazil 2005-2011.

Outbreaks caused by serogroup C meningococci in the northeast region of Brazil from 2005 to 2011 were associated to the emergence of variant ET-15 of cc11, which has been replaced by cc103 from 2006 to date. The increase of cc103 should be closely monitored to prevent the spread of this clone to neighbouring regions.

[Paediatric meningococcal meningitis in France: ACTIV/GPIP network results]. / Méningites à méningocoques de l'enfant en France: résultats de l'observatoire ACTIV/GPIP.

BACKGROUND: The GPIP/ACTIV (Groupe de Pathologie Infectieuse Pédiatrique and Association Clinique et Thérapeutique Infantile du Val de Marne) set up an active surveillance network to analyze the epidemiological, clinical and biological features of meningococcal meningitis. METHODS: French pediatric wards working with 166 microbiology laboratories enrolled all children (0-18 years old) with bacterial meningitis. Risk factors, signs and symptoms, vaccination status, cerebrospinal fluid analysis, treatments and case fatality rate were recorded. RESULTS: Since 2001, 1661 meningococcal meningitis were reported among 3769 (44.1%) bacterial meningitis. Mean age was 4.4- year- old (± 4.8, median 2.5) and 2/3 cases occurred in children under 5- year- old (68.8%). Serogroup B (61.3%) is preponderant following by serogroup C (27.0%). 27.5% of children had received an antibiotic treatment 24 hours before lumbar puncture. A shock is reported in 31.0% of cases. No cases of meningococcal meningitis C has been reported in children vaccinated with a conjugate vaccine. Two children vaccinated with MenBvac(®) vaccine had a meningitis B14:P1.7,16. Global case fatality rate was 6.5% but was higher (9.2%) for serogroup C than for serogroup B (5.9%) (p=0.02). CONCLUSION: This is among the largest series of microbiologically documented meningococcal meningitis to date (1661 cases). In France, meningococcal is responsible for approximately 50 % of meningitis. Effective meningococcal serogroup B vaccine and serogroup C vaccination recommendation could control the burden of meningococcal meningitis.

Absence of Neisseria meningitidis serogroup C-specific antibodies during the first year of life in the Netherlands: an age group at risk?

In The Netherlands, a single meningococcal serogroup C conjugate (MenCC) vaccination is administered to children at the age of 14 months. Here, we report the levels of MenC polysaccharide-specific antibodies in children at birth and at 3, 11, and 12 months of age and the presence of functional antibodies at 11 months of age, before infants receive their MenCC immunization. We observed a rapid decline in polysaccharide-specific antibodies after birth and no induction of naturally elicited polysaccharide-specific antibodies. Furthermore, at 11 months of age, no bactericidal antibodies are observed. These data indicate that these infants may be at risk in the period prior to MenCC immunization, if Neisseria meningitidis serogroup C starts to (re)circulate.

Impact of meningococcal serogroup C conjugate vaccines on carriage and herd immunity.

BACKGROUND: In 1999, meningococcal serogroup C conjugate (MCC) vaccines were introduced in the United Kingdom for those under 19 years of age. The impact of this intervention on asymptomatic carriage of meningococci was investigated to establish whether serogroup replacement or protection by herd immunity occurred. METHODS: Multicenter surveys of carriage were conducted during vaccine introduction and on 2 successive years, resulting in a total of 48,309 samples, from which 8599 meningococci were isolated and characterized by genotyping and phenotyping. RESULTS: A reduction in serogroup C carriage (rate ratio, 0.19) was observed that lasted at least 2 years with no evidence of serogroup replacement. Vaccine efficacy against carriage was 75%, and vaccination had a disproportionate impact on the carriage of sequence type (ST)-11 complex serogroup C meningococci that (rate ratio, 0.06); these meningococci also exhibited high rates of capsule expression. CONCLUSIONS: The impact of vaccination with MCC vaccine on the prevalence of carriage of group C meningococci was consistent with herd immunity. The high impact on the carriage of ST-11 complex serogroup C could be attributed to high levels of capsule expression. High vaccine efficacy against disease in young children, who were not protected long-term by the schedule initially used, is attributed to the high vaccine efficacy against carriage in older age groups.

Characterization of ST-4821 complex, a unique Neisseria meningitidis clone.

Ten outbreaks of a new serogroup C meningococcal disease emerged during 2003-2005 in China. The multilocus sequence typing results indicated that unique sequence type 4821 clone meningococci were responsible for these outbreaks. Herein, we determined the entire genomic DNA sequence of serogroup C isolate 053442, which belongs to ST-4821. Comparison of 053442 gene contents with other meningococcal genomes shows that they have similar characteristics, including thousands of repetitive elements and simple sequence repeats, numerous phase-variable genes, and similar virulence-related factors. However, many strain-specific regions were found in each genome. We also present the results of a genomic comparison of 28 ST-4821 complex isolates that were isolated from different serogroups using comparative genomic hybridization analysis. Genome comparison between the newly emerged hyperinvasive isolates belonging to different serogroups will further our understanding of their respective pathogenetic mechanisms.

Consecutive use of two multiplex PCR-based assays for simultaneous identification and determination of capsular status of nine common Neisseria meningitidis serogroups associated with invasive disease.

We developed two Neisseria meningitidis multiplex PCR assays to be used consecutively that allow determination of the serogroup and capsular status of serogroup A, B, C, 29E, W135, X, and Y cnl-3/cnl-1-like-containing N. meningitidis isolates by direct analysis of the amplicon size. These assays offer a rapid and simple method of serogrouping N. meningitidis.

Purulent pericarditis caused by Neisseria meningitidis serogroup C and confirmed through polymerase chain reaction.

We here report the case of a previously healthy 20-y-old male with disseminated meningococcal disease and purulent pericarditis. Polymerase chain reaction was used to identify Neisseria meningitidis serogroup C as the causative agent in the pericardial fluid.

Identification of a new Neisseria meningitidis serogroup C clone from Anhui province, China.

BACKGROUND: Outbreaks of a new serogroup C meningococcal disease emerged during 2003-04 (five outbreaks with 43 cases) and in 2004-05 (five outbreaks with 29 cases), all in Anhui province, China. We describe the molecular epidemiology and features of the causative bacterial strains. METHODS: We used pulsed-field gel electrophoresis (PFGE) and multi-locus sequence typing (MLST) to analyse the strains. FINDINGS: Of 34 strains of Neisseria meningitidis cultured during 2003-04 from Anhui province, 31 were group C meningococci, 28 of which were associated with three of five outbreaks; one from a patient and 27 from close contacts of eight patients. Of 30 strains isolated from Anhui province during 2004-05, 17 were identified as serogroup C meningococci, ten of which were associated with four of five outbreaks. In a nationwide survey, 542 strains were isolated during 2004-05; 58 were serogroup C meningococci interspersed among 11 other provinces where no serogroup C outbreak occurred. Of the 106 serogroup C strains analysed, 89 had identical PFGE patterns, designated AH1. Of 28 strains selected for MLST analyses, 25 were sequence type 4821 (ST-4821), which did not belong to any of the previously reported sequence types that can form a new hypervirulent lineage. INTERPRETATION: ST-4821 seems to be unique and caused the serogroup C meningitis outbreaks during the two seasons from 2003 to 2005 in Anhui province. The emergence of this sequence type has epidemiological importance that should be monitored for future spread in China and the rest of the world.