Colonization of dermal arterioles by Neisseria meningitidis provides a safe haven from neutrophils.

The human pathogen Neisseria meningitidis can cause meningitis and fatal systemic disease. The bacteria colonize blood vessels and rapidly cause vascular damage, despite a neutrophil-rich inflammatory infiltrate. Here, we use a humanized mouse model to show that vascular colonization leads to the recruitment of neutrophils, which partially reduce bacterial burden and vascular damage. This partial effect is due to the ability of bacteria to colonize capillaries, venules and arterioles, as observed in human samples. In venules, potent neutrophil recruitment allows efficient bacterial phagocytosis. In contrast, in infected capillaries and arterioles, adhesion molecules such as E-Selectin are not expressed on the endothelium, and intravascular neutrophil recruitment is minimal. Our results indicate that the colonization of capillaries and arterioles by N. meningitidis creates an intravascular niche that precludes the action of neutrophils, resulting in immune escape and progression of the infection.

Type IV pilus retraction enables sustained bacteremia and plays a key role in the outcome of meningococcal sepsis in a humanized mouse model.

Neisseria meningitidis (the meningococcus) remains a major cause of bacterial meningitis and fatal sepsis. This commensal bacterium of the human nasopharynx can cause invasive diseases when it leaves its niche and reaches the bloodstream. Blood-borne meningococci have the ability to adhere to human endothelial cells and rapidly colonize microvessels. This crucial step enables dissemination into tissues and promotes deregulated inflammation and coagulation, leading to extensive necrotic purpura in the most severe cases. Adhesion to blood vessels relies on type IV pili (TFP). These long filamentous structures are highly dynamic as they can rapidly elongate and retract by the antagonistic action of two ATPases, PilF and PilT. However, the consequences of TFP dynamics on the pathophysiology and the outcome of meningococcal sepsis in vivo have been poorly studied. Here, we show that human graft microvessels are replicative niches for meningococci, that seed the bloodstream and promote sustained bacteremia and lethality in a humanized mouse model. Intriguingly, although pilus-retraction deficient N. meningitidis strain (ΔpilT) efficiently colonizes human graft tissue, this mutant did not promote sustained bacteremia nor induce mouse lethality. This effect was not due to a decreased inflammatory response, nor defects in bacterial clearance by the innate immune system. Rather, TFP-retraction was necessary to promote the release of TFP-dependent contacts between bacteria and, in turn, the detachment from colonized microvessels. The resulting sustained bacteremia was directly correlated with lethality. Altogether, these results demonstrate that pilus retraction plays a key role in the occurrence and outcome of meningococcal sepsis by supporting sustained bacteremia. These findings open new perspectives on the role of circulating bacteria in the pathological alterations leading to lethal sepsis.

Normocellular Community-Acquired Bacterial Meningitis in Adults: A Nationwide Population-Based Case Series.

STUDY OBJECTIVE: This study sought to describe the clinical presentation of normocellular community-acquired bacterial meningitis in adults. METHODS: Using the prospective, nationwide, population-based database of the Danish Study Group of Infections of the Brain, the study identified all adults with normocellular community-acquired bacterial meningitis who were treated at departments of infectious diseases in Denmark from 2015 through 2018. Normocellular community-acquired bacterial meningitis was defined as a cerebrospinal fluid leukocyte count of up to 10×106/L combined with detection of bacteria in the cerebrospinal fluid. Outcome was categorized according to the Glasgow Outcome Scale at discharge. RESULTS: Normocellular cerebrospinal fluid was observed in 12 of 696 (2%) patients with community-acquired bacterial meningitis. The median age was 70 years (range 17 to 92 years), and 8 of 12 (67%) patients were male. All patients had symptoms suggestive of community-acquired bacterial meningitis and pathogens identified by culture (Streptococcus pneumoniae, n=10; Staphylococcus aureus, n=1) or polymerase chain reaction (Neisseria meningitidis; n=1) of the cerebrospinal fluid. Bacteremia was found in 9 of 12 (75%) patients, and 1 of 12 (8%) presented with septic shock. None of the patients had serious underlying immunocompromising conditions. The median times from admission to lumbar puncture and meningitis treatment were 2.5 hours (interquartile range 1.1 to 3.9 hours) and 2.6 hours (interquartile range 0.9 to 22.8 hours). In 3 of 11 (27%) patients, empiric treatment for community-acquired bacterial meningitis was interrupted by a normal cerebrospinal fluid cell count. The overall case-fatality rate was 3 of 12 (25%); meningitis treatment was interrupted in 1 of these patients, and 8 of 12 (67%) had a Glasgow Outcome Scale score of 1 to 4 at discharge. CONCLUSION: Normocellular community-acquired bacterial meningitis is not very common, but it is important to consider and may be associated with a pneumococcal cause.

Genomic analysis of the meningococcal ST-4821 complex-Western clade, potential sexual transmission and predicted antibiotic susceptibility and vaccine coverage.

INTRODUCTION: The ST-4821 complex (cc4821) is a leading cause of serogroup C and serogroup B invasive meningococcal disease in China where diverse strains in two phylogenetic groups (groups 1 and 2) have acquired fluoroquinolone resistance. cc4821 was recently prevalent among carriage isolates in men who have sex with men in New York City (USA). Genome-level population studies have thus far been limited to Chinese isolates. The aim of the present study was to build upon these with an extended panel of international cc4821 isolates. METHODS: Genomes of isolates from Asia (1972 to 2017), Europe (2011 to 2018), North America (2007), and South America (2014) were sequenced or obtained from the PubMLST Neisseria database. Core genome comparisons were performed in PubMLST. RESULTS: Four lineages were identified. Western isolates formed a distinct, mainly serogroup B sublineage with alleles associated with fluoroquinolone susceptibility (MIC <0.03 mg/L) and reduced penicillin susceptibility (MIC 0.094 to 1 mg/L). A third of these were from anogenital sites in men who have sex with men and had unique denitrification gene alleles. Generally 4CMenB vaccine strain coverage was reliant on strain-specific NHBA peptides. DISCUSSION: The previously identified cc4821 group 2 was resolved into three separate lineages. Clustering of western isolates was surprising given the overall diversity of cc4821. Possible association of this cluster with the anogenital niche is worthy of monitoring given concerns surrounding antibiotic resistance and potential subcapsular vaccine escape.

Complement interactions with the pathogenic Neisseriae: clinical features, deficiency states, and evasion mechanisms.

Neisseria gonorrhoeae causes the sexually transmitted infection gonorrhea, while Neisseria meningitidis is an important cause of bacterial meningitis and sepsis. Complement is a central arm of innate immune defenses and plays an important role in combating Neisserial infections. Persons with congenital and acquired defects in complement are at a significantly higher risk for invasive Neisserial infections such as invasive meningococcal disease and disseminated gonococcal infection compared to the general population. Of note, Neisseria gonorrhoeae and Neisseria meningitidis can only infect humans, which in part may be related to their ability to evade only human complement. This review summarizes the epidemiologic and clinical aspects of Neisserial infections in persons with defects in the complement system. Mechanisms used by these pathogens to subvert killing by complement and preclinical studies showing how these complement evasion strategies may be used to counteract the global threat of meningococcal and gonococcal infections are discussed.

Clinical and economic burden of invasive meningococcal disease: Evidence from a large German claims database.

BACKGROUND: Limited data is available to describe clinical characteristics, long-term outcomes, healthcare resource use and the attributable costs of invasive meningococcal disease (IMD) in Germany. We aimed to examine demographic and clinical characteristics as well as healthcare resource use and related costs. METHODS: We conducted a retrospective cohort study based on the InGef database in patients with IMD between 2009 and 2015. Cases were identified based on hospital main discharge diagnoses of IMD. Demographics, clinical characteristics, 30-day and 1-year mortality as well as IMD-related complications and sequelae in IMD cases were examined. In addition, short and long-term costs and healthcare resource use in IMD cases were analyzed and compared to an age- and sex-matched control group without IMD. RESULTS: The study population comprised 164 IMD cases between 2009 and 2015. The mean length of the IMD-related hospitalization was 13 days and 38% of all cases presented with meningitis only, 35% with sepsis only, 16% with both and 11% with other IMD. The 30-day and one-year mortality were 4.3% and 5.5%, respectively. Approximately 13% of IMD cases had documented IMD-related complications at hospital discharge and 24% suffered from sequelae during follow-up. The IMD-related hospitalization was associated with mean costs of € 9,620 (standard deviation: € 22,197). The difference of mean costs between IMD cases and matched non-IMD controls were € 267 in the first month and € 1,161 from one month to one year after discharged from IMD-related hospitalization. During the later follow-up period, the mean overall costs and costs associated with individual healthcare sectors were also higher for IMD cases without reaching statistical significance. CONCLUSIONS: IMD resulted in severe complications and sequelae and was associated with extensive costs and increased healthcare resource use in Germany, especially in the first year after IMD diagnosis and due the IMD-related hospitalization.

Chimeric hepatitis B virus core particles displaying Neisserial surface protein A confer protection against virulent Neisseria meningitidis serogroup B in BALB/c mice.

PURPOSE: The primary goal of the present study was to explore and evaluate the highly conserved Neisserial surface protein A (NspA) molecule, fused with truncated HBV virus-like particles (VLPs), as a candidate vaccine against the virulent Neisseria meningitidis serogroup B (NMB). METHODS: NspA was inserted into the major immunodominant region of the truncated hepatitis B virus core protein (HBc; amino acids 1-144). The chimeric protein, HBc-N144-NspA, was expressed from a prokaryotic vector and generated HBc-like particles, as determined by transmission electron microscopy. Further, the chimeric protein and control proteins were used to immunize mice and the resulting immune responses evaluated by flow cytometry, enzyme-linked immunosorbent assay, and analysis of serum bactericidal activity (SBA) titer. RESULTS: Evaluation of the immunogenicity of the recombinant HBc-N144-NspA protein showed that it elicited the production of high levels of NspA-specific total IgG. The SBA titer of HBc-N144-NspA/F reached 1:16 2 weeks after the last immunization in BALB/c mice, when human serum complement was included in the vaccine. Immunization of HBc-N144-NspA, even without adjuvant, induced high levels of IL-4 and a high IgG1 to IgG2a ratio, confirming induction of an intense Th2 immune response. Levels of IL-17A increased rapidly in mice after the first immunization with HBc-N144-NspA, indicating the potential for this vaccine to induce a mucosal immune response. Meanwhile, the immunization of HBc-N144-NspA without adjuvant induced only mild inflammatory infiltration into the mouse muscle tissue. CONCLUSION: This study demonstrates that modification using HBc renders NspA a candidate vaccine, which can trigger protective immunity against NMB.

An Overview of Neisseria meningitidis.

Neisseria meningitidis (the meningococcus) is a member of the normal nasopharyngeal microbiome in healthy individuals, but can cause septicemia and meningitis in susceptible individuals. In this chapter we provide an overview of the disease caused by N. meningitidis and the schemes used to type the meningococcus. We also review the adhesions, virulence factors, and phase variable genes that enable it to successfully colonize the human host. Finally, we outline the history and current status of meningococcal vaccines and highlight the importance of continued molecular investigation of the epidemiology and the structural analysis of the antigens of this pathogen to aid future vaccine development.

Cellulitis: An unusual manifestation of Neisseria meningitidis infection.

INTRODUCTION: Neisseria meningitidis infections often cause severe meningitis as well as bacteraemia. However, cellulitis in meningococcal diseases have rarely been described. Here, we report a case of right lower limb cellulitis caused by N. meningitidis. CASE REPORT: A 69-year-old woman presented with fever and lower limb swelling. She had diabetes mellitus, hypertension, dyslipidaemia and a history of surgical resection of vulvar carcinoma. N. meningitidis was isolated from her blood culture. DISCUSSION: This report provides additional evidence in support of N. meningitidis as a cause of cellulitis.

AMEND study protocol: a case-control study to assess the long-term impact of invasive meningococcal disease in Australian adolescents and young adults.

INTRODUCTION: Invasive meningococcal disease (IMD) primarily causes disease in young children and adolescents and can cause long-term disability. Many countries are considering implementation of meningococcal B and/or meningococcal ACWY vaccines to control meningococcal disease. Estimating the cost-effectiveness of meningococcal vaccine programme is hampered due to a lack of good quality costing and burden of disease data. This study aims to address this evidence gap by assessing the clinical, physical, neurocognitive, economic and societal impact of IMD on adolescents and young adults. METHODS AND ANALYSIS: A case-control study of 64 participants with confirmed IMD (15-24 years 11 months at time of disease) and 64 control participants (17-34 years 11 months) will be conducted in Australia from 2016 to 2020. All participants will undergo a neurocognitive assessment, full medical examination, pure tone audiometry assessment and complete quality of life and behavioural questionnaires. Meningococcal cases will be assessed 2-10 years posthospitalisation and a subset of cases will be interviewed to explore in depth their experiences of IMD and its impact on their life. Primary outcome measures include general intellectual functioning from the Wechsler Adult Intelligence Scale and overall quality of life from the Health Utilities Index. Secondary outcome measures include academic achievement, executive functioning, behaviour, hearing, psychological and physical functioning. Outcome measures will be compared between cases and controls using independent t-tests or ORs, or if any significant confounders are identified, adjusted analyses (analysis of covariance or adjusted ORs) will be conducted. Thematic analysis will be used to analyse transcribed interviews and a costing model will be used to project lifetime costs. ETHICS AND DISSEMINATION: The Adolescent MENingococcal Disease (AMEND) study has been approved by the Human Research Ethics Committee of the Women's and Children's Health Network (HREC/14/WCHN/024). The results will be disseminated via peer-reviewed publications, conference presentations, study participants, and meningococcal and meningitis foundations. TRIAL REGISTRATION NUMBER: NCT03798574.

Epidemiology, clinical presentation, risk factors, intensive care admission and outcomes of invasive meningococcal disease in England, 2010-2015.

The epidemiology of invasive meningococcal disease (IMD) is constantly changing as new strains are introduced into a population and older strains are removed through vaccination, population immunity or natural trends. Consequently, the clinical disease associated with circulating strains may also change over time. In England, IMD incidence has declined from 1.8/100,000 in 2010/2011 to 1.1/100,000 in 2013/2014, with a small increase in 2014/2015 to 1.3/100,000. Between 01 January 2011 and 30 June 2015, MenB was responsible for 73.0% (n = 2489) of 3411 laboratory-confirmed IMD cases, followed by MenW (n = 371, 10.9%), MenY (n = 373, 10.9%) and MenC (n = 129, 3.8%); other capsular groups were rare (n = 49, 1.4%). Detailed questionnaires were completed for all 3411 laboratory-confirmed cases. Clinical presentation varied by capsular group and age. Atypical presentations were uncommon (244/3411; 7.2%), increasing from 1.2% (41/3411) in children to 3.5% (120/3411) in older adults. Known IMD risk factors were rare (18/3411; 0.5%) and included complement deficiency (n = 11), asplenia (n = 6) or both (n = 1). Nearly a third of cases required intensive care (1069/3411; 31.3%), with rates highest in adults. The 28-day CFR was 6.9% (n = 237), with the lowest rates in 0-14 year-olds (85/1885, 4.5%) and highest among 85+ year-olds (30/94, 31.9%). These observations provide a useful baseline for the current burden of IMD in a European country with enhanced national surveillance.

Lactobacillus paracasei feeding improves the control of secondary experimental meningococcal infection in flu-infected mice.

BACKGROUND: The use of probiotics to improve anti-microbial defence, such as for influenza infections, is increasingly recommended. However, no data are available on the effect of probiotics on flu-associated secondary bacterial infections. There is strong evidence of a spatiotemporal association between influenza virus infection and invasive Neisseria meningitidis. We thus investigated the effect of feeding mice Lactobacillus paracasei CNCM I-1518 in a mouse model of sequential influenza-meningococcal infection. METHODS: We intranasally infected BALB/c mice with a strain of influenza A virus (IAV) H3N2 that was first adapted to mice. Seven days later, a secondary bacterial infection was induced by intranasal administration of bioluminescent N. meningitidis. During the experiment, mice orally received either L. paracasei CNCM I-1518 or PBS as a control. The effect of L. paracasei administration on secondary bacterial infection by N. meningitidis was evaluated. RESULTS: Oral consumption of L. paracasei CNCM I-1518 reduced the weight loss of infected mice and lowered the bioluminescent signal of infecting meningococci. This improvement was associated with higher recruitment of inflammatory myeloid cells, such as interstitial monocytes and dendritic cells, to the lungs. CONCLUSIONS: Our data highlight the role of the gut-lung axis. L. paracasei CNCM I-1518 may boost the defence against IAV infection and secondary bacterial infection, which should be further studied and validated in clinical trials.

Fatal meningococcal septicemia without meningeal signs, contribution of the peripheral smear in diagnosis: Report of a case.

Acute meningococcemia is characterized by extensive purpurae consisting of both petechiae and ecchymoses. This condition can be rapidly fatal without treatment due to shock and severe consumptive coagulopathy. We report a case of fatal meningococcal septicemia in a military recruit who presented with fever and associated rapidly progressive purpuric rash (purpura fulminans) without any meningeal signs. Evaluation revealed evidence of disseminated intravascular coagulopathy and multiorgan failure. Diplococci were demonstrated in peripheral blood neutrophils and monocytes. On autopsy, extensive hemorrhages were found in both adrenals, lungs, liver, skin, and kidneys with secondary hemophagocytic lymphohistiocytosis in bone marrow. This report highlights useful information obtained from examination of peripheral blood smear in purpura fulminans.

Fully 3D printed integrated reactor array for point-of-care molecular diagnostics.

Molecular diagnostics that involve nucleic acid amplification tests (NAATs) are crucial for prevention and treatment of infectious diseases. In this study, we developed a simple, inexpensive, disposable, fully 3D printed microfluidic reactor array that is capable of carrying out extraction, concentration and isothermal amplification of nucleic acids in variety of body fluids. The method allows rapid molecular diagnostic tests for infectious diseases at point of care. A simple leak-proof polymerization strategy was developed to integrate flow-through nucleic acid isolation membranes into microfluidic devices, yielding a multifunctional diagnostic platform. Static coating technology was adopted to improve the biocompatibility of our 3D printed device. We demonstrated the suitability of our device for both end-point colorimetric qualitative detection and real-time fluorescence quantitative detection. We applied our diagnostic device to detection of Plasmodium falciparum in plasma samples and Neisseria meningitides in cerebrospinal fluid (CSF) samples by loop-mediated, isothermal amplification (LAMP) within 50 min. The detection limits were 100 fg for P. falciparum and 50 colony-forming unit (CFU) for N. meningitidis per reaction, which are comparable to that of benchtop instruments. This rapid and inexpensive 3D printed device has great potential for point-of-care molecular diagnosis of infectious disease in resource-limited settings.

[Acute meningococcal disease in children and adolescents].

Meningococcal disease is a rapidly progressing infection, which continues to cause deaths among children and adolescents. In this review, clinical signs and initial treatment of acute childhood meningococcal disease is described. Operational flow charts have been developed for assessment of non-blanching rash and initial treatment of meningococcal disease.

The sweet side of the pathogenic Neisseria: the role of glycan interactions in colonisation and disease.

Glycomics is a rapidly growing field that focuses on the structure and function of carbohydrates (glycans) in biological systems. Glycan interactions play a major role in infectious disease, at all stages of colonisation and disease progression. Neisseria meningitidis, the cause of meningococcal sepsis and meningitis, and Neisseria gonorrhoeae, which causes the sexually transmitted infection gonorrhoea, are responsible for significant morbidity and mortality worldwide. Neisseria meningitidis displays a range of surface glycosylations including capsule polysaccharide, lipooligosaccharide and O-linked glycoproteins. While N. gonorrhoeae does not have a capsule, it does express both lipooligosaccharide and O-linked glycoproteins. Neisseria gonorrhoeae also has the ability to scavenge host sialic acids, while several N. meningitidis serogroups can synthesise sialic acid. Surface expressed sialic acid is key in serum resistance and survival in the host. On the host side, the pathogenic Neisseria protein adhesins such as Opc and NHBA bind to host glycans for adherence and colonisation of host cells. Essentially, from both the bacterial and host perspective, glycan interactions are fundamental in colonisation and disease of pathogenic Neisseria. The key aspects of glycobiology of the pathogenic Neisseria are reviewed herein.

Natural resistance to Meningococcal Disease related to CFH loci: Meta-analysis of genome-wide association studies.

Meningococcal disease (MD) remains an important infectious cause of life threatening infection in both industrialized and resource poor countries. Genetic factors influence both occurrence and severity of presentation, but the genes responsible are largely unknown. We performed a genome-wide association study (GWAS) examining 5,440,063 SNPs in 422 Spanish MD patients and 910 controls. We then performed a meta-analysis of the Spanish GWAS with GWAS data from the United Kingdom (combined cohorts: 897 cases and 5,613 controls; 4,898,259 SNPs). The meta-analysis identified strong evidence of association (P-value ≤ 5 × 10-8) in 20 variants located at the CFH gene. SNP rs193053835 showed the most significant protective effect (Odds Ratio (OR) = 0.62, 95% confidence interval (C.I.) = 0.52-0.73; P-value = 9.62 × 10-9). Five other variants had been previously reported to be associated with susceptibility to MD, including the missense SNP rs1065489 (OR = 0.64, 95% C.I.) = 0.55-0.76, P-value = 3.25 × 10-8). Theoretical predictions point to a functional effect of rs1065489, which may be directly responsible for protection against MD. Our study confirms the association of CFH with susceptibility to MD and strengthens the importance of this link in understanding pathogenesis of the disease.

Risk of Invasive Meningococcal Disease in Men Who Have Sex with Men: Lessons Learned from an Outbreak in Germany, 2012-2013.

BACKGROUND: We undertook investigations in response to an invasive meningococcal disease (IMD) outbreak in men who have sex with men (MSM) in Berlin 2012-2013 to better understand meningococcal transmission and IMD risk in MSM. METHODS: We retrospectively searched for further IMD cases in MSM in Germany through local health departments and undertook exploratory interviews. We performed antigen sequence typing, characterized fHbp and aniA genes of strains with the outbreak finetype and reviewed epidemiologically or spatiotemporally linked cases from 2002-2014. RESULTS: Among the 148 IMD-cases notified from 01.01.2012-30.09.2013 in 18-59 year-old men we identified 13 MSM in 6 federal states: 11 serogroup C (MenC, all finetype C:P1.5-1,10-8:F3-6), 2 MenB. Interviews with 7 MSM revealed frequent meeting of multiple partners online or via mobile apps and illicit drug use as potential risk factors. MenC incidence was 13-fold higher in MSM than non-MSM. MenC isolates from 9/11 MSM had a novel fHbp allele 766. All C:P1.5-1,10-8:F3-6 strains from MSM versus 16/23 from non-MSM had intact aniA genes (p = 0.04). Although definitive evidence for transmission among MSM in epidemiological or spatiotemporal clusters in 2002-2014 was lacking, clusters were more frequent in men aged 20-49 years. Molecular analysis of C:P1.5-1,10-8:F3-6 strains revealed cases with intact aniA since 2007, mainly associated with fHbp361, fHbp766 and fHbp813, all involving one or more MSM. CONCLUSIONS: MenC incidence was elevated in MSM during the study period. Multiple casual sexual contacts and illicit drug use were common in affected MSM. In all strains from MSM we detected an intact aniA gene coding for a nitrite reductase, which permits survival in microanaerobic environments and could play a role in meningococcal transmission in MSM through urogenital colonization. Furthermore, meningococcal transmission among MSM may be sustained over large areas and thus require modified spatiotemporal scanning algorithms for timely detection and control.

Experimental Evidence of Bacterial Colonization of Human Coronary Microvasculature and Myocardial Tissue during Meningococcemia.

Meningococcal septic shock is associated with profound vasoplegia, early and severe myocardial dysfunction, and extended skin necrosis responsible for a specific clinical entity designated purpura fulminans (PF). PF represents 90% of fatal meningococcal infections. One characteristic of meningococcal PF is the myocardial dysfunction that occurs in the early phase of sepsis. Furthermore, hemodynamic studies have shown that the prognosis of meningococcal sepsis is directly related to the degree of impairment of cardiac contractility during the initial phase of the disease. To gain insight into a potential interaction of Neisseria meningitidis with the myocardial microvasculature, we modified a previously described humanized mouse model by grafting human myocardial tissue to SCID mice. We then infected the grafted mice with N. meningitides Using the humanized SCID mouse model, we demonstrated that N. meningitidis targets the human myocardial tissue vasculature, leading to the formation of blood thrombi, infectious vasculitis, and vascular leakage. These results suggest a novel mechanism of myocardial injury in the course of severe N. meningitidis sepsis that is likely to participate in primary myocardial dysfunction.