Clonal analysis of meningococci during a 26 year period prior to the introduction of meningococcal serogroup C vaccines

Meningococcal disease remains a public health burden in the UK and elsewhere. Invasive Neisseria meningitidis, isolated in Scotland between 1972 and 1998, were characterised retrospectively to examine the serogroup and clonal structure of the circulating population. 2607 isolates causing invasive disease were available for serogroup and MLST analysis whilst 2517 were available for multilocus sequence typing (MLST) analysis only. Serogroup distribution changed from year to year but serogroups B and C were dominant throughout. Serogroup B was dominant throughout the 1970s and early 1980s until serogroup C became dominant during the mid-1980s. The increase in serogroup C was not associated with one particular sequence type (ST) but was associated with a number of STs, including ST-8, ST-11, ST-206 and ST-334. This is in contrast to the increase in serogroup C disease seen in the 1990s that was due to expansion of the ST-11 clonal complex. While there was considerable diversity among the isolates (309 different STs among the 2607 isolates), a large proportion of isolates (59.9%) were associated with only 10 STs. These data highlight meningococcal diversity over time and the need for ongoing surveillance during the introduction of new meningococcal vaccines

The molecular characterisation of Neisseria meningitidis in Scotland before, during and after the initial introduction of the meningococcal serogroup C conjugate (MenC) vaccines

The aim of this project was to develop and introduce an MLST system for all meningococci causing invasive disease in Scotland starting in 1999, and subsequently throughout 2000, 2001 and 2002. MLST was introduced because strains within the electrophoretic type 37 (ET-37) complex, particularly those of ST11, are often indistinguishable by traditional methods. Nucleotide sequencing was performed on seven housekeeping genes and one outer membrane protein gene, porA. Data was analysed using databases and software available through the MLST website (www.mlst.net) and the porA websites (neisseria.org/nm/typing/porA/ and http://www.show.scot.nhs.uk/smprl/). The introduction of MLST in Scotland was used as a routine method for the characterization of Neisseria meningitidis isolates in 1999. Coincidentally this is just prior to the introduction of the MenC vaccines in November 1999. All N. meningitidis isolates from invasive disease were analysed. This has given valuable data highlighting the effect the MenC has on Serogroup C disease and its decline over the length of the vaccination campaign. At the same time, the overall numbers of cases were monitored including the effect on serogroup B between 1999 and 2002. This has been linked with historical data such as the typical cyclical pattern seen with serogroups B and C over the past decade. The incidences of ET-37 strains in serogroup C disease were assessed along with ET-37 strains of other serogroups. The number of different ST's isolated were monitored from the introduction of the MenC vaccine and this has been linked with their corresponding serogroups and subtypes. A decrease in serogroup C disease was observed, however this has been accompanied by an increase in serogroup B disease. The occurrence of capsule switch has been assessed and applied to the whole data set for subsequent conclusion. This thesis clearly shows that MLST is extremely important for the surveillance of meningococcal disease over a period of years and, and that this study, has been effective not only in monitoring the impact of the MenC vaccines, but also providing a detailed genotypic representation of strains now commonly associated with disease. In addition to enhanced sequence based characterisation, development of a fluorescent -based PCR protocol using N. meningitidis specific probes has been developed to markedly improved sensitivity and specificity of meningococcal DNA detection. Using an automated platform employing Taqman chemistry, a detection system employs fluorescence-based chemistry that eliminates post PCR processing and has provided accurate real-time quantitative PCR. This development has identified previously undetectable quantities of meningococcal DNA, which have been subsequently used in a modified MLST system to produce genotypic results, which would not normally be created

Exponential growth, high prevalence of SARS-CoV-2, and vaccine effectiveness associated with the Delta variant

SARS-CoV-2 infections were rising during early summer 2021 in many countries associated with the Delta variant. We assessed RT-PCR swab-positivity in the REal-time Assessment of Community Transmission-1 (REACT-1) study in England. We observed sustained exponential growth with average doubling time (June-July 2021) of 25 days driven by complete replacement of Alpha variant by Delta, and by high prevalence at younger less-vaccinated ages. Unvaccinated people were three times more likely than double-vaccinated people to test positive. However, after adjusting for age and other variables, vaccine effectiveness for double-vaccinated people was estimated at between ~50% and ~60% during this period in England. Increased social mixing in the presence of Delta had the potential to generate sustained growth in infections, even at high levels of vaccination

Increased Genetic Diversity of Neisseria meningitidis Isolates after the Introduction of Meningococcal Serogroup C Polysaccharide Conjugate Vaccines

During the 1990s, the incidence of meningococcal disease was high in the United Kingdom. This was due primarily to an increase in serogroup C disease, particularly that within the ET-37/ST-11 genetic lineage. Serogroup C meningococcal polysaccharide conjugate vaccines were introduced in the United Kingdom in 1999, but the sequence types of meningococci causing disease since that time have not yet been reported. We have used serogrouping and multilocus sequence typing to characterize meningococci from patients with invasive disease over a 4-year period and show that there is a significant increase in genetic diversity but no genetic evidence of capsule switching

Molecular methods for the detection and characterization of Neisseria meningitidis

Neisseria meningitidis remains a common global cause of morbidity and mortality. The laboratory confirmation of meningococcal disease is, therefore, very important for individual patient management and for public health management. Through surveillance schemes, it provides long-term epidemiologic data that can be used to inform vaccine policy. Traditional methods, such as latex agglutination and the enzyme-linked immunosorbent assay, are still used, but molecular methods are now also established. In this review, molecular methods for the laboratory confirmation and characterization of meningococci are described. PCR is an invaluable tool in modern biology and can be used to predict the group, type and subtype of meningococci. It is now also used in a fluorescence-based format for increased sensitivity and specificity. The method also provides the amplified DNA for other techniques, such as multilocus sequence typing. Other methods for the discrimination of meningococci have also played and continue to play an important part in epidemiology. For example, pulsed-field gel electrophoresis is highly discriminatory, whilst multilocus enzyme electrophoresis provided the basis for the description of global meningococcal clones and formed the foundation for multilocus sequence typing. Other less commonly used methods, such as matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and pyrosequencing, may increasingly find their way into microbiology reference laboratories. Nevertheless, nucleotide sequencing and laboratory automation have aided the introduction of many methods and provide data that are digitally based and, therefore, highly accurate and portabl

Automation of a fluorescence-based multiplex PCR for the laboratory confirmation of common bacterial pathogens

A fluorescence-based multiplex PCR was automated for the simultaneous detection of Neisseria meningitidis, Streptococcus pneumoniae and Haemophilus influenzae in clinical samples from patients with suspected meningitis. Sensitivity of one to two genome copies per 100 microl sample and specificity of 100% for each organism were shown. Automation of DNA extraction, liquid handling, PCR and analysis are achieved on a single platform, which enables a high throughput and rapid turnaround of clinical samples that, in turn, leads to faster diagnosis. This is ultimately beneficial to the treatment of the patient and for public health management.</p

Nucleotide sequence-based typing of meningococci directly from clinical samples

The unpredictable characteristics of meningococcal disease (MD) make outbreaks complicated to monitor and consequently lead to high levels of public anxiety. Traditional molecular techniques have been utilized in order to understand better the epidemiology of MD, but some have disadvantages such as being highly specialized and labour-intensive, with low reproducibility. Some of these problems have been overcome by using multilocus sequence typing (MLST). This technique exploits the unambiguous nature and electronic portability of nucleotide sequencing data for the characterization of micro-organisms. The need for enhanced surveillance of MD after the introduction of serogroup C conjugate vaccines means that it is important to gain typing information from the infecting organism in the absence of a culture isolate. Here, the application of MLST for the laboratory confirmation and characterization of Neisseria meningitidis directly from clinical samples is described. This involved using a newly designed set of primers that were complementary to nucleotide sequences external to the existing MLST primers already in use for culture-based MLST of meningococci. This combination has produced a highly sensitive procedure to allow the efficient genotypic characterization of meningococci directly from clinical samples.</p

RESEARCH Neisseria meningitidis Sequence Type and Risk for Death, Iceland

Invasive meningococcal infections are hyperendemic in Iceland, a relatively isolated country in the mid-Atlantic. We performed a nationwide study on all viable meningococcal strains (N = 362) from 1977 to 2004. We analyzed the association of patient’s age and sex, meningococcal serogroups, and sequence types (STs) with outcomes. Overall, 59 different STs were identified, 19 of which were unique to Iceland. The most common STs were 32 (24.6%), 11 (19.9%), and 10 (10.2%). The unique ST-3492 ranked fourth (7.7%). The most common serogroups were B (56.4%), C (39.8%), and A (2.2%). Age (p&lt;0.001) and infection with a unique ST (p = 0.011) were independently associated with increased death rates, whereas isolation of meningococci from cerebrospinal fluid only was associated with lower death rates (p = 0.046). This study shows evolutionary trends of meningococcal isolates in a relatively isolated community and highlights an association between unique STs and poor outcome. Invasive infections caused by Neisseria meningitidis (meningococci) cause high rates of illness and death worldwide (1–3). Meningococci have frequently caused epidemics in Iceland, a relatively isolated community in the mid-Atlantic (4,5). To more fully understand the phylogeny of meningococcal strains, various typing methods have been used, including serogroup and serotype classifications. Epidemiologic studies have used more discriminating methods, such as multilocus enzyme electrophoresis, based on electrophoretic variation of several chromosomally encoded cytoplasmic “housekeeping ” enzymes (6). More recently, sequence-based molecular methods have bee

Association of STs with predominant clonal complexes from Scotland 1972–98.

<p>Association of STs with predominant clonal complexes from Scotland 1972–98.</p