Stuart C. Clarke

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.

Diarrhoeagenic Escherichia coli—an emerging problem?

Diarrhea remains one of the main sources of morbidity and morbidity in todays world and a large proportion is caused by diarrheagenic Escherichia coli. They are a particular problem in developed countries although travelers diarrhea and hemorrhagic colitis are also a problem in developed countries. There are seven classes of diarrheagenic E. coli, namely enteropathogenic E. coli (EPEC), enterohaemorrhagic E. coli (EHEC), enteroinvasive E. coli (EIEC), enterotoxigenic E. coli (ETEC), enteroaggregative E. coli (EAggEC), diarrhea-associated hemolytic E. coli (DHEC) and cytolethal distending toxin (CDT)- producing E. coli. Many of their virulence determinants have been determined and some classes of diarrheagenic E. coli produce toxins. The virulence factors of some diarrhogenic E. coli have yet to be full determined and in the meantime they remain a large and emerging problem without the availability of effective vaccines.

Interlaboratory Comparison of PCR-Based Identification and Genogrouping of Neisseria meningitidis

ABSTRACT Twenty clinical samples (18 cerebrospinal fluid samples and 2 articular fluid samples) were sent to 11 meningococcus reference centers located in 11 different countries. Ten of these laboratories are participating in the EU-MenNet program (a European Union-funded program) and are members of the European Monitoring Group on Meningococci. The remaining laboratory was located in Burkina Faso. Neisseria meningitidis was sought by detecting several meningococcus-specific genes (crgA, ctrA, 16S rRNA, and porA). The PCR-based nonculture method for the detection of N. meningitidis gave similar results between participants with a mean sensitivity and specificity of 89.7 and 92.7%, respectively. Most of the laboratories also performed genogrouping assays (siaD and mynB/sacC). The performance of genogrouping was more variable between laboratories, with a mean sensitivity of 72.7%. Genogroup B gave the best correlation between participants, as all laboratories routinely perform this PCR. The results for genogroups A and W135 were less similar between the eight participating laboratories that performed these PCRs.

Identification of Invasive Serotype 1 Pneumococcal Isolates That Express Nonhemolytic Pneumolysin

ABSTRACT Recently, there has been an increase in invasive pneumococcal disease (IPD) caused by serotype 1 Streptococcus pneumoniae throughout Europe. Serotype 1 IPD is associated with bacteremia and pneumonia in Europe and North America, especially in neonates, and is ranked among the top five most prevalent pneumococcal serotypes in at least 10 countries. The currently licensed pediatric pneumococcal vaccine does not afford protection to this serotype. Upon screening of 252 clinical isolates of S. pneumoniae, we discovered mutations in the pneumolysin gene of two out of the four serotype 1 strains present in the study group. Analysis of an additional 28 serotype 1 isolates from patients with IPD from various Scottish Health Boards, revealed that >50% had mutations in their pneumolysin genes. This resulted in the expression of nonhemolytic forms of pneumolysin. All of the strains producing nonhemolytic pneumolysin were sequence type 306 (ST306), whereas those producing “wild-type” pneumolysin were ST227. The mutations were in a region of pneumolysin involved in pore formation. These mutations can be made in vitro to give the nonhemolytic phenotype. Pneumolysin is generally conserved throughout all serotypes of S. pneumoniae and is essential for full invasive disease; however, it appears that serotype 1 ST306 does not require hemolytically active pneumolysin to cause IPD.

Genetic Analysis of Diverse Disease-Causing Pneumococci Indicates High Levels of Diversity within Serotypes and Capsule Switching

ABSTRACT We have used multilocus sequence typing (MLST) and serotyping to build a phylogenetic framework for pneumococcal disease isolates in Scotland that provides a snapshot of the relationships between capsular type and genotype. The results show that while the MLST type correlates with the serotype, isolates within a serotype can belong to a number of individual clonal complexes or sequence types (STs). We also show that isolates of the same ST can express different capsular polysaccharides, i.e., display capsular switching, and that this phenomenon is observed both for capsular types commonly isolated from patients with invasive disease and for serogroups less commonly isolated from patients with invasive disease but which may commonly be carried asymptomatically in the human nasopharynx.

Invasive pneumococcal disease in Scotland, 1999-2001: use of record linkage to explore associations between patients and disease in relation to future vaccination policy.

A record linkage study was done to provide comprehensive data on the epidemiologic characteristics of invasive pneumococcal disease (IPD) in Scotland. The overall incidence of IPD was 11 cases/10(5) persons and 21 cases/10(5) persons <1 year of age, 51 cases/10(5) persons 1 year of age, 45 cases/10(5) elderly persons (age > or =65 years), 176-483 cases/10(5) persons with chronic medical conditions, and 562-2031 cases/10(5) persons with severe immunosuppression. The case-fatality rate was 11% among elderly persons and ranged from 3% to 13% among persons with underlying medical conditions. The most common pneumococcal serogroups associated with IPD were 14, 9, 6, 19, 23, 8, and 4. Serogroups included in the 23-valent polysaccharide vaccine caused the majority of cases of IPD. The proportion of IPD due to the 7-, 9-, and 11-valent conjugate vaccine serogroups was lower among older people and persons with underlying medical conditions.

Multilocus sequence typing

Numerous computer-based statistical packages have been developed in recent years and it has become easier to analyze nucleotide sequence data and gather subsequent information that would not normally be available. Multilocus sequence typing (MLST) is used for characterizing isolates of bacterial and fungal species and uses nucleotide sequences of internal fragments of housekeeping genes. This method is finding a place in clinical microbiology and public health by providing data for epidemiological surveillance and development of vaccine policy. It adds greatly to our knowledge of the genetic variation that can occur within a species and has therefore been used for studies of population biology. Analysis requires the detailed interpretation of nucleotide sequence data obtained from housekeeping and nonhousekeeping genes. This is due to the amount of data generated from nucleotide sequencing and the information generated from an array of analytical tools improves our understanding of bacterial pathogens. This can benefit public health interventions and the development of enhanced therapies and vaccines. This review concentrates on the analytical tools used in MLST and their use in the clinical microbiology and public health fields.

Comparison of Commercial DNA Extraction Kits for Extraction of Bacterial Genomic DNA from Whole-Blood Samples

ABSTRACT The demand for molecular diagnostic tests in medical microbiology has highlighted the need for efficient methods of DNA extraction. In addition, it is preferable for these methods to be automated. An example of such a requirement is for the confirmation of meningococcal disease where rapid, sensitive, and specific procedures are required for public health management purposes. Previous studies have shown that whole blood is the preferred method for the isolation of bacterial DNA in meningococcal disease, and in this study, we compare five commercially available kits for the extraction of bacterial genomic DNA from whole-blood samples. These include kits in a 96-well binding plate, 96-well filter plate, and metallic bead formats. The method for all five kits is described, and the sensitivity, specificity, ease of automation, and overall efficiency are determined.

Presence of Nonhemolytic Pneumolysin in Serotypes of Streptococcus pneumoniae Associated with Disease Outbreaks

Pneumolysin is an important virulence factor of the human pathogen Streptococcus pneumoniae. Sequence analysis of the ply gene from 121 clinical isolates of S. pneumoniae uncovered a number of alleles. Twenty-two strains were chosen for further analysis, and 14 protein alleles were discovered. Five of these had been reported previously, and the remaining 9 were novel. Cell lysates were used to determine the specific hemolytic activities of the pneumolysin proteins. Six strains showed no hemolytic activity, and the remaining 16 were hemolytic, to varying degrees. We report that the nonhemolytic allele reported previously in serotype 1, sequence type (ST) 306 isolates is also present in a number of pneumococcal isolates of serotype 8 that belong to the ST53 lineage. Serotype 1 and 8 pneumococci are known to be associated with outbreaks of invasive disease. The nonhemolytic pneumolysin allele is therefore associated with the dominant clones of outbreak-associated serotypes of S. pneumoniae.

Declining serotype coverage of new pneumococcal conjugate vaccines relating to the carriage of Streptococcus pneumoniae in young children

BACKGROUND Asymptomatic carriage of the opportunistic pathogen Streptococcus pneumoniae is known to precede the development of invasive disease. Young children are one of the major reservoirs for pneumococci and worldwide over 700,000 children under two years old die due to invasive pneumococcal disease each year. Heptavalent conjugate vaccine (PCV-7) was introduced into the UK childhood immunisation schedule in September 2006. Our objective was to assess the emergence of colonising serotypes in young children in the three years following PCV-7 implementation. METHODS Time-series prevalence survey set in the paediatric outpatients department of a large UK teaching hospital. Participants were children aged four years and under attending the outpatients department during PCV-7 introduction (October 2006-February 2007) and in the same months of the two subsequent years. The main outcome measure was prevalence of pneumococcal carriage by serotype. RESULTS The rate of pneumococcal nasopharyngeal carriage remained stable during the three year period. We observed a significant 69% (95% CI, -40% to -118%, p<0.0001) decrease in carriage of PCV-7 serotypes during PCV-7 implementation and a concomitant increase in the proportion of non PCV-7 serotypes. The most prevalent emerging non-vaccine serotypes were 6C, 11A, 19A and 22F. By March 2009, PCV-13 was predicted to cover only 33.3% (95% CI, 24.2-42.5%) of strains carried in the study population. CONCLUSIONS Although the overall pneumococcal carriage rate remained stable between 2006 and 2009, we observed a significant decrease in the serotype coverage of PCV-7 and PCV-13. PCV-7 was highly successful in reducing carriage of vaccine serotypes. However, the increase in the proportion of non-vaccine serotypes found both in our study and causing invasive disease currently in the UK, underlines the importance of continued surveillance of carriage and disease.