Stability of PorA during a meningococcal disease epidemic

Meningococci causing New Zealand's epidemic, which began in 1991, are defined as group B, serosubtype P1.4 (subtype P1.7-2,4), belonging to the ST-41/ST-44 complex, lineage III. Of the 2,358 group B isolates obtained from disease cases from 1991 through 2003, 85.7% (2,021 of 2,358) were determined to be serosubtype P1.4. Of the remaining isolates, 156 (6.6%) were not serosubtypeable (NST). Molecular analysis of the porA gene from these B:NST meningococcal isolates was used to determine the reason. Most NST isolates (156, 88.5%) expressed a PorA that was distinct from P1.7-2,4 PorA. Fifteen isolates expressed variants of P1.7-2,4 PorA, and a further three expressed P1.7-2,4 PorA without any sequence variation. These three isolates expressed P1.7-2,4 PorA at very low levels, as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis, and showed variation in the porA promoter region. Among the 15 meningococcal isolates expressing variants of P1.7-2,4 PorA, 11 different sequence variations were found. Compared with the P1.7-2,4 PorA sequence, the sequences of these variants contained deletions, insertions, or single-nucleotide substitutions in the VR2 region of the protein. Multilocus restriction typing was used to assess the clonal derivations of B:NST case isolates. Meningococcal isolates expressing distinct PorA proteins belonged mostly to clonal types that were unrelated to the epidemic strain, whereas all meningococcal isolates expressing variants of P1.7-2,4 PorA belonged to the ST-41/ST-44 complex, lineage III. These results, together with those obtained serologically, demonstrate that the P1.7-2,4 PorA protein of meningococci responsible for New Zealand's epidemic has remained relatively stable over 13 years and support the use of a strain-specific outer membrane vesicle vaccine to control the epidemic.     

Assessing the risk of laboratory-acquired meningococcal disease

Neisseria meningitidis is infrequently reported as a laboratory-acquired infection. Prompted by two cases in the United States in 2000, we assessed this risk among laboratorians. We identified cases of meningococcal disease that were possibly acquired or suspected of being acquired in a laboratory by placing an information request on e-mail discussion groups of infectious disease, microbiology, and infection control professional organizations. A probable case of laboratory-acquired meningococcal disease was defined as illness meeting the case definition for meningococcal disease in a laboratorian who had occupational exposure to an N. meningitidis isolate of the same serogroup within 14 days of illness onset. Sixteen cases of probable laboratory-acquired meningococcal disease occurring worldwide between 1985 and 2001 were identified, including six U.S. cases between 1996 and 2000. Nine cases (56%) were serogroup B; seven (44%) were serogroup C. Eight cases (50%) were fatal. All cases occurred among clinical microbiologists. In 15 cases (94%), isolate manipulation was performed without respiratory protection. We estimated that an average of three microbiologists are exposed to the 3,000 meningococcal isolates seen in U.S. laboratories yearly and calculated an attack rate of 13/100,000 microbiologists between 1996 and 2001, compared to 0.2/100,000 among U.S. adults in general. The rate and case/fatality ratio of meningococcal disease among microbiologists are higher than those in the general U.S. population. Specific risk factors for laboratory-acquired infection are likely associated with exposure to droplets or aerosols containing N. meningitidis. Prevention should focus on the implementation of class II biological safety cabinets or additional respiratory protection during manipulation of suspected meningococcal isolates.     

Human opsonins induced during meningococcal disease recognize transferrin binding protein complexes

Patient serum opsonins against transferrin binding protein A+B (TbpA+B) complexes from two Neisseria meningitidis strains (K454 and B16B6, with 85- and 68-kDa TbpB, respectively) were quantified by a functional phagocytosis and oxidative burst assay. TbpA+B complexes adsorbed to fluorescent beads were opsonized with individual acute and convalescent sera from 40 patients infected by a variety of meningococcal strains. Flow cytometric quantitation of leukocyte phagocytosis products (PP) demonstrated that disease-induced serum opsonins recognized TbpA+B, and the highest anti-TbpA+B serum opsonic activities were found between admission to hospital and 6 weeks later. The PP values obtained with TbpA+B from strain B16B6 (PP(B16B6)) were higher than those obtained with TbpA+B from strain K454 (PP(K454)), with both acute and convalescent sera (P < 0.0001), and correlated positively with higher immunoglobulin G enzyme-linked immunosorbent assay titers against TbpA+B from strain B16B6 than from strain K454 (P < 0.001). In spite of considerable variations between individuals, significant correlations were found between the PP(B16B6) and PP(K454) values, and the PP values did not depend on the variability of the TbpB proteins of the disease-causing strains. Simultaneously measured oxidative burst activity correlated closely with the PP values. We conclude that highly cross-reactive anti-TbpA+B serum opsonins are produced during meningococcal disease. The anti-TbpA+B opsonic activities were not affected by the variability of the TbpB proteins of the disease-causing strains, which further adds to the evidence for the vaccine potential of meningococcal TbpA+B complexes.     

An outbreak of group B meningococcal disease: tracing the causative strain of Neisseria meningitidis by DNA fingerprinting.

Following an outbreak of meningococcal disease in three schoolchildren in a small community in northern Norway, DNA fingerprinting, serotyping with monoclonal antibodies, serogrouping, and sulfonamide sensitivity testing were applied for characterization and tracing of the causative agent. The three case isolates were genomically indistinguishable, sulfonamide-resistant, serogroup B, serotype 15 meningococci. Throat specimens were collected from 552 healthy contacts, including all children below age 17 and their parents. Among the 36 carrier isolates (carrier rate, 6.5%) 13 showed DNA fingerprints identical, or almost identical, to the index pattern. All of these 13 isolates were sulfonamide resistant, 12 were of serotype 15, and 8 were of polysaccharide serogroup B (5 were nongroupable). These closely related isolates were almost exclusively recovered from schoolchildren of 2 of 15 small villages, one of which included the homes of two of the patients. The remaining 23 carrier isolates were nonresistant, non-type 15 meningococci of widely differing DNA restriction patterns. Our results confirm that DNA fingerprinting has potential as an efficient tool in practical meningococcal epidemiology.     

Human opsonins induced during meningococcal disease recognize outer membrane proteins PorA and PorB

Human opsonins directed against specific meningococcal outer membrane structures in sera obtained during meningococcal disease were quantified with a recently developed antigen-specific, opsonin-dependent phagocytosis and oxidative burst assay. Outer membrane vesicles (OMVs) and PorA (class 1) and PorB (class 3) proteins purified from mutants of the same strain (44/76; B:15:P1.7. 16) were adsorbed to fluorescent beads, opsonized with acute- and convalescent-phase sera from 40 patients with meningococcal disease, and exposed to human leukocytes. Flow cytometric quantitation of the resulting leukocyte phagocytosis products (PPs) demonstrated that disease-induced serum opsonins recognized meningococcal OMV components and both porins. The PPPorA and PPPorB values induced by convalescent-phase sera correlated positively with the PPOMV values. However, the PPPorB values were higher than the PPPorA values in convalescent-phase sera (medians [ranges] of 754 [17 to 1,057] and 107 [4 to 458], respectively) (P < 0.0001) and correlated positively with higher levels of immunoglobulin G against PorB than against PorA as evaluated by enzyme-linked immunosorbent assay. Extensive individual variations in the anti-OMV and antiporin serum opsonic activities between patients infected by serotypes and serosubtypes homologous and heterologous to the target antigens were observed. Simultaneously measured oxidative burst activity correlated with the opsonophagocytosis, an indication that both of these important steps in the in vitro phagocytic elimination of meningococci are initiated by opsonins directed against OMV components, including PorA and PorB. In conclusion, human patient opsonins against meningococcal OMV components and in particular PorB epitopes were identified by this new method, which might facilitate selection of opsonin-inducing meningococcal antigens for inclusion in future vaccines.     

Neisseria lactamica protects against experimental meningococcal infection

Immunological and epidemiological evidence suggests that the development of natural immunity to meningococcal disease results from colonization of the nasopharynx by commensal Neisseria spp., particularly with N. lactamica. We report here that immunization with N. lactamica killed whole cells, outer membrane vesicles, or outer membrane protein (OMP) pools and protected mice against lethal challenge by a number of diverse serogroup B and C meningococcal isolates in a model of bacteremic infection. Sera raised to N. lactamica killed whole cells, OMPs, or protein pools were found to cross-react with meningococcal isolates of a diverse range of genotypes and phenotypes. The results confirm the potential of N. lactamica to form the basis of a vaccine against meningococcal disease.     

Cross-reactivity of antibodies against PorA after vaccination with a meningococcal B outer membrane vesicle vaccine

The cross-reactivity of PorA-specific antibodies induced by a monovalent P1.7-2,4 (MonoMen) and/or a hexavalent (HexaMen) meningococcal B outer membrane vesicle vaccine (OMV) in toddlers and school children was studied by serum bactericidal assays (SBA). First, isogenic vaccine strains and PorA-identical patient isolates were compared as a target in SBA, to ensure that the vaccine strains are representative for patient isolates. Geometric mean titers (GMTs) in SBA against patient isolates with subtypes P1.5-2,10 and P1.5-1,2-2 after vaccination with HexaMen were generally lower than those against vaccine strains with the same subtype, although the percentage of vaccine responders (> or =4-fold increase in SBA after vaccination) was not affected. Using various P1.7-2,4 patient isolates, GMTs as well as the number of vaccine responders were higher than for the P1.7-2,4 vaccine strain, indicating that the use of the P1.7-2,4 vaccine strain may have underestimated the immunogenicity of this subtype in HexaMen. Secondly, the cross-reactivity of antibodies induced by MonoMen and HexaMen was studied using several patient isolates that differed from the vaccine subtypes by having minor antigenic variants of one variable region (VR), by having a completely different VR or by having a different combination of VRs. MonoMen induced P1.4-specific antibodies that were cross-reactive with P1.4 variants P1.4-1 and P1.4-3. HexaMen induced a broader cross-reactive antibody response against various patient isolates with one VR identical to a vaccine subtype or a combination of VRs included in HexaMen. Cross-reactivity, measured by a fourfold increase in SBA after vaccination, against these strains ranged from 23 to 92% depending on the subtype of the tested strain and was directed against both VR1 and VR2. The extended cross-reactivity of vaccinee sera induced by HexaMen against antigenic variants has important favorable implications for meningococcal B OMV vaccine coverage.     

Genotypic and phenotypic characterization of carriage and invasive disease isolates of Neisseria meningitidis in Finland

The relationship between carriage and the development of invasive meningococcal disease is not fully understood. We investigated the changes in meningococcal carriage in 892 military recruits in Finland during a nonepidemic period (July 2004 to January 2006) and characterized all of the oropharyngeal meningococcal isolates obtained (n = 215) by using phenotypic (serogrouping and serotyping) and genotypic (porA typing and multilocus sequence typing) methods. For comparison, 84 invasive meningococcal disease strains isolated in Finland between January 2004 and February 2006 were also analyzed. The rate of meningococcal carriage was significantly higher at the end of military service than on arrival (18% versus 2.2%; P < 0.001). Seventy-four percent of serogroupable carriage isolates belonged to serogroup B, and 24% belonged to serogroup Y. Most carriage isolates belonged to the carriage-associated ST-60 clonal complex. However, 21.5% belonged to the hyperinvasive ST-41/44 clonal complex. Isolates belonging to the ST-23 clonal complex were cultured more often from oropharyngeal samples taken during the acute phase of respiratory infection than from samples taken at health examinations at the beginning and end of military service (odds ratio [OR], 6.7; 95% confidence interval [95% CI], 2.7 to 16.4). The ST-32 clonal complex was associated with meningococcal disease (OR, 17.8; 95% CI, 3.8 to 81.2), while the ST-60 clonal complex was associated with carriage (OR, 10.7; 95% CI, 3.3 to 35.2). These findings point to the importance of meningococcal vaccination for military recruits and also to the need for an efficacious vaccine against serogroup B isolates.     

Elevated procalcitonin as a diagnostic marker in meningococcal disease

Patients with meningococcal disease who seek medical attention can create a diagnostic dilemma for clinicians due to the nonspecific nature of the disease’s presentation. This study assesses the diagnostic accuracy of procalcitonin levels in the setting of meningococcal disease. Two emergency department cohorts (A and B) were studied between 2002 and 2005, during the current epidemic of serogroup B meningococcal disease in New Zealand. Cohort A consisted of 171 patients, all with confirmed meningococcal disease (84 children, 87 adults). Cohort B consisted of a large (n=1,524) consecutively recruited population of febrile patients who presented to the emergency department, 28 of whom had confirmed meningococcal disease. Within the meningococcal disease cohort (cohort A), the geometric mean procalcitonin level was 9.9 ng/ml, with levels being higher in children than in adults (21.6 vs. 4.6 ng/ml, p=0.01). The overall sensitivity of elevated procalcitonin, using a cutoff of 2.0 ng/ml in children and 0.5 ng/ml in adults, was 0.93 (95%CI: 0.88–0.96). Despite the higher cutoff level for paediatric patients, a trend towards greater sensitivity existed in children (0.96 vs. 0.90; p=0.08). Elevated procalcitonin was correlated with whole blood meningococcal load (r=0.50) and Glasgow Meningococcal Sepsis Prognostic Score (r=0.40). Within the cohort of patients who were febrile on presentation (cohort B), the specificity of elevated procalcitonin in meningococcal disease was 0.85 (95% CI: 0.83–0.87), the positive and negative likelihood ratios were 6.1 and 0.08, respectively, and the sensitivity of elevated procalcitonin (0.93; 95% CI: 0.76–0.99) was corroborated. Measurement of procalcitonin is a useful tool in patients with nonspecific febrile illnesses when the possibility of meningococcal disease is present. The diagnostic accuracy surpasses that of current early laboratory markers, allowing results to be used to guide decisions about patient management.     

Dynamics of meningococcal long-term carriage among university students and their implications for mass vaccination

In the 1997-98 academic year, we conducted a longitudinal study of meningococcal carriage and acquisition among first-year students at Nottingham University, Nottingham, United Kingdom. We examined the dynamics of long-term meningococcal carriage with detailed characterization of the isolates. Pharyngeal swabs were obtained from 2,453 first-year students at the start of the academic year (October), later on during the autumn term, and again in March. Swabs were immediately cultured on selective media, and meningococci were identified and serologically characterized. Nongroupable strains were genetically grouped using a PCR-based assay. Pulsed-field gel electrophoresis was used to determine the link between sequential isolates. Of the carriers initially identified in October, 44.1% (98 of 222) were still positive later on in the autumn (November or December); 57.1% of these remained persistent carriers at 6 months. Of the index carriers who lost carriage during the autumn, 16% were recolonized at 6 months. Of 344 index noncarriers followed up, 22.1% acquired carriage during the autumn term and another 13.7% acquired carriage by March. Overall, 43.9% (397 of 904) of the isolates were noncapsulated (serologically nongroupable); by PCR-based genogrouping, a quarter of these belonged to the capsular groups B and C. The ratio of capsulated to noncapsulated forms for group B and C strains was 2.9 and 0.95, respectively. Sequential isolates of persistent carriers revealed that individuals may carry the same or entirely different organisms at different times. We identified three strains that clearly switched their capsular expression on and off at different times in vivo. One student developed invasive meningococcal disease after carrying the same organism for over 7 weeks. The study revealed a high rate of turnover of meningococcal carriage among students. Noncapsulated organisms are capable of switching their capsular expression on and off (both ways) in the nasopharynx, and group C strains are more likely to be noncapsulated than group B strains. Carriage of a particular meningococcal strain does not necessarily protect against colonization or invasion by a homologous or heterologous strain.     

Fatal outcome from meningococcal disease--an association with meningococcal phenotype but not with reduced susceptibility to benzylpenicillin

Penicillin has been the mainstay of treatment for meningococcal disease. Isolates of Neisseria meningitidis that are less susceptible to penicillin have been reported in several countries and in recent years have become more common. The clinical significance of this reduced susceptibility has not been investigated on a large scale. Hence, N. meningitidis isolates from culture-confirmed cases of meningococcal disease in England and Wales, between 1993 and 2000, were routinely serogrouped, serotyped and tested for susceptibility to penicillin. These data were linked to death registrations and analysed retrospectively. The changing trends in susceptibility were described and multivariate logistic regression was used to examine associations between strain characteristics and fatal outcome. The frequency of N. meningitidis isolates less susceptible to penicillin increased from < 6% in 1993 to > 18% in 2000. In particular, isolates expressing serogroup C with serotype 2b and serogroup W135 had a higher frequency of reduced penicillin susceptibility (49% and 55%, respectively). There was no evidence of an association between fatal outcome and infection with a less penicillin-susceptible isolate. Fatal outcome was associated with serogroup and serotype, with the odds of death for cases infected with C:2a and B:2a strains three-fold higher when compared with the baseline. For this large dataset the serogroup and serotype of the infecting strain influenced mortality from meningococcal disease and may be markers for hypervirulence. No association was found between reduced penicillin susceptibility and fatal outcome, but the increasing frequency of isolates less susceptible to penicillin highlights the need for continued surveillance.     

Measurement of antibodies against meningococcal capsular polysaccharides B and C in enzyme-linked immunosorbent assays: towards an improved surveillance of meningococcal disease.

In order to improve the surveillance of serogroup B and C meningococcal diseases, enzyme-linked immunosorbent assays (ELISAs) specific for anti-B immunoglobulin M (IgM) and anti-C IgM and IgG antibodies were developed. The tests were evaluated by using paired sera from 122 patients with and 101 patients without laboratory evidence of meningococcal disease. Fifty-three of 67 patients (79%) with culture-confirmed serogroup B disease had an anti-B IgM antibody response; anti-B IgM levels waned rapidly in children < or = 4 years of age. Twenty-four of 25 patients (96%) with culture-confirmed serogroup C disease had an anti-C IgM and/or IgG antibody response (IgM, 92%; IgG, 68%). In patients without evidence of meningococcal disease, 19% of children < or = 4 years of age and 69% of those > 4 years of age had intermediate anti-B IgM titers. In contrast, only 1 and 5% of these patients had intermediate titers of anti-C IgM and anti-C IgG, respectively. The ELISAs were shown to be powerful tools for discriminating between serogroup B and C diseases in 96 to 100% of culture-confirmed cases. For 90% of patients with culture-negative meningococcal disease, a serogroup-specific diagnosis could be established by examination of paired sera in the ELISAs. As serogroup B and C meningococci account for practically all cases of meningococcal disease in industrialized countries, the availability of these tests may improve surveillance and prevention.     

Cross-reacting serum opsonins in patients with meningococcal disease.

We have examined the opsonic activity of sera from patients with Neisseria meningitidis (B:15:P1.16) infections against different meningococcal strains, using flow cytometry and luminol-enhanced chemiluminescence. A marked increase in the phagocytosis of ethanol-fixed meningococcal strains of different serogroups, serotypes, and serosubtypes was demonstrated in the presence of convalescence sera compared with acute sera. Convalescence sera also caused a significant increase of leukocyte oxidative metabolism during phagocytosis, as measured by luminol-enhanced chemiluminescence. The sera contained a broad range of opsonins cross-reacting with serogroup A, B, C, W-135, and Y meningococci of different serotypes and serosubtypes, indicating that the cross-reacting opsonins recognized surface epitopes other than those determined by current serotyping schemes.     

Invasive meningococcal disease in Quebec, Canada, due to an emerging clone of ST-269 serogroup B meningococci with serotype antigen 17 and serosubtype antigen P1.19 (B:17:P1.19)

During periods of endemic meningococcal disease, serogroup B Neisseria meningitidis is responsible for a significant percentage of invasive diseases, and no particular clone or strain predominates (F. E. Ashton and D. A. Caugant, Can. J. Microbiol. 47: 293-289, 2001), However, in the winter of 2004 to 2005, a cluster of serogroup B meningococcal disease occurred in one region in the province of Québec, Canada. The N. meningitidis strain responsible for this cluster of cases was identified as sequence type ST-269 with the antigenic formula B:17:P1.19. Retrospective analysis of isolates from 2000 onwards showed that this clone first emerged in the province of Québec in 2003. The emergence of this clone of serogroup B meningococci occurred after a mass vaccination against serogroup C N. meningitidis, suggesting possible capsule replacement.     

Continuing diversification of Neisseria meningitidis W135 as a primary cause of meningococcal disease after emergence of the serogroup in 2000

The occurrence of a clonal outbreak of serogroup W135 (of the electrophoretic type 37 [ET-37] clonal complex) meningococcal disease among Hajj pilgrims in 2000 has led to enhanced surveillance of the evolution of this particular serogroup, formerly considered rare, in invasive infections. Since the first case of meningococcal disease due to a serogroup W135 strain was detected in France in 1994, all isolates were characterized phenotypically. We further used phenotypic and genotypic approaches to type the 101 serogroup W135 strains isolated from patients with invasive meningococcal diseases in France in 2001 and 2002. Overall, 55% of these isolates had Hajj strain-related phenotypes (60 and 52% in 2001 and 2002, respectively), although only 45% belonged to the ET-37 clonal complex. Moreover, pulsed-field gel electrophoresis of the ET-37 clonal complex isolates showed that only 32% of the serogroup W135 isolates were indistinguishable from the 2000 Hajj-related strain. Our results suggest the continuous emergence of new genetic lineages of serogroup W135 independently of the 2000 global outbreak.     

Antibodies to meningococcal class 1 outer membrane proteins in South African complement-deficient and complement-sufficient subjects.

Inhibition assays were used to investigate human serum antibodies to the meningococcal class 1 outer membrane proteins. We adapted the whole-cell enzyme-linked immunosorbent assay technique to determine the ability of sera to inhibit the binding of murine subtyping monoclonal antibodies. Serum samples from 33 South African subjects with a deficiency in the sixth component of complement as well as serum samples from various groups of complement-sufficient subjects were investigated. Subjects were subdivided according to whether they were (i) convalescent from Neisseria meningitidis infections, (ii) nonconvalescent, or (iii) controls. Preliminary subtyping investigations had shown that P1.2 was present on 36% of meningococcal clinical isolates from Cape Province, South Africa. Assays with the anti-P1.2 antibodies showed the presence of high antibody levels in many deficient sera and moderately elevated levels in some sera from the complement-sufficient convalescent patients. P1.2, P1.4, P1.15, and P1.16 are epitopes situated on loop 4 of the class 1 outer membrane proteins, whereas P1.7 is on loop 1. Inhibition assays showed that human sera that inhibited binding by P1.2 monoclonal antibodies tended to inhibit the other monoclonal antibodies directed to loop 4 epitopes. This suggests that the epitopes recognized by the human antibodies are not exactly the same as the epitopes recognized by the murine monoclonal antibodies and raises the possibility of the importance of other epitopes.     

Bacteriocins (meningocins) in Norwegian isolates of Neisseria meningitidis: possible role in the course of a meningococcal epidemic

In the Neisseria meningitidis strain MC58 (serogroup B; ET-5 complex) genome three putative islands of horizontally transferred DNA (IHTs) have been identified. IHT-A2 codes for eight hypothetical proteins and two disrupted open reading frames with similarity to a secretion protein (NMB0097) and an ABC transporter (NMB0098). The strains MC58 and 44/76 (shown here) are meningocin resistant/weakly sensitive. None of these strains are meningocin producers. However, NMB0097 and NMB0098 homologues with open reading frames are found in meningocin producers (N. meningitidis P241 (serogroup A; systemic isolate) and BT878 (serogroup B; carrier isolate), and also in strain FAM18 (serogroup C; ET-37 complex). Knocking out either of the two genes in the strain BT878 yielded mutants that did not secrete meningocin. A similarly disrupted tolC mutant in strain BT878 still released meningocin. Among systemic meningococcal isolates prior to and at the onset (mid-1973 to the end of 1974) of the epidemic peaking in 1975 in North Norway, 12 of 30 (40%) isolates of serogroup A were meningocin producers. However, the rate for serogroup B was 1 of 45 (2.2%). Serogroup B meningocin-resistant/weakly sensitive non-producers dominated in the region from mid-1975 and spread to the rest of the country from then on. No producers were found in selected pharyngeal isolates from healthy carriers collected in Svalbard in the early spring of 1975. Our results suggest that meningocinogeny has played a part in the change from serogroup A to serogroup B among isolates in North Norway during the first half of 1975.     

Increasing microbiological confirmation and changing epidemiology of meningococcal disease on Merseyside, England

Objectives   To determine, for the last 5 years in children on Merseyside with clinical meningococcal disease (MCD), the impact on diagnostic yield of newer bacteriologic methods; bacterial antigen detection (AD) and polymerase chain reaction (PCR).
Methods   Prospective data collection at Royal Liverpool Children's Hospital over two epochs: 1 September 1992 to 30 April 1994 (epoch A, n  = 126) and 17 November 1997 to 15 September 1998 (epoch B, n  = 85).
Results   Epoch A was compared with epoch B. Diagnosis was confirmed by detection of meningococci in 78 of 126 (61.9%) versus 64 of 85 (75.3%, P  = 0.04), but with a significantly lower rate of positive blood and cerebrospinal fluid culture in the later epoch. The proportion of cases receiving penicillin pretreatment was unchanged at 32%, but the proportion undergoing lumbar puncture decreased significantly. Median ages were higher in epoch B: 1.7 years versus 2.49 years ( P  = 0.013, Mann–Whitney). There was a significant increase in the proportion of cases due to serogroup C (14/78 (18%) versus 30/64 (46.9%), P  = 0.001).
Conclusions   Culture detection of meningococci from children with MCD has reduced, as less lumbar punctures are done. However, improved diagnosis by PCR and AD has increased microbiological confirmation overall. Serogroup C disease and the median age of cases continue to rise.     

Molecular epidemiology of an outbreak of meningococcal disease in a university community.

Over a 2-month period, five cases of serogroup C meningococcal disease occurred in Iowa City, Iowa. Two patients were unacquainted university students who had independently visited another university with endemic meningococcal disease. Isolates from these patients had DNA fingerprints identical to those of the isolates responsible for infections on the other campus. Three cases for which the patients' isolates had a different DNA fingerprint were linked to visiting a local tavern. To disrupt the outbreak, the University of Iowa offered free meningococcal vaccine to all students. This report demonstrates that outbreaks of meningococcal disease may be due to more than one circulating strain and illustrates the utility of pulsed-field gel electrophoresis in defining the molecular epidemiology of meningococcal infections.     

Genomic fingerprinting of Neisseria meningitidis associated with group C meningococcal disease in Canada.

A single electrophoretic type (ET15) of Neisseria meningitidis has been associated with an increased incidence of group C meningococcal disease in Canada. Genomic fingerprinting through pulsed-field gel electrophoresis of chromosomal DNA was used to characterize the clonal relationship among meningococcal isolates of different electrophoretic types and among isolates within ET15. The genomic fingerprints of the ET15 isolates, while similar as a group, were sufficiently distinct to confirm linkage for four pairs of strains from focal outbreaks and differed markedly from those of the other common electrophoretic types, ET5, ET9, and ET21.