1,721,095 research outputs found
Control of pneumococcal disease in the United Kingdom – the start of a new era
No full textIn 2000, a multi-valent pneumococcal conjugate vaccine, known as Prevnar, was licensed for use ininfants and young children in the USA. The subsequent introduction of the vaccine into thechildhood immunization schedule in that country led to a significant decrease in pneumococcaldisease. The vaccine is effective against invasive and non-invasive pneumococcal infection, can beused in young children as well as adults and, like all conjugate vaccines, provides long-lastingimmunity. Moreover, it reduces the incidence of antibiotic resistance because a number ofresistant serotypes are targeted by the vaccine. Prevnar, also known as Prevenar, has since beenlicensed in numerous countries, including the UK.On 8 February 2006, the Departments of Health inEngland, Scotland and Wales announced the inclusion of Prevenar in the childhood immunizationschedule. This announcement has important implications for pneumococcal infection, diseasesurveillance and immunization policy in the UK
Pyrosequencing: nucleotide sequencing technology with bacterial genotyping applications
No full textPyrosequencing is a relatively new method for real-time nucleotide sequencing. It has rapidly found applications in DNA sequencing, genotyping, single nucleotide polymorphism analysis, allele quantification and whole-genome sequencing within the areas of microbiology, clinical genetics and pharmacogenetics. It is fast becoming a real alternative to the traditional Sanger sequencing method although, at present, read lengths are normally limited to approximately 70 nucleotides. The pyrosequencing method involves four main stages: first, target DNA is amplified using PCR; second, double-stranded DNA is converted to single-stranded DNA templates; third, oligonucleotide primers are hybridized to a complementary sequence of interest; and, finally, the pyrosequencing reaction itself, in which a reaction mixture of enzymes and substrates catalyses the synthesis of complementary nucleotides. Data are shown as a collection of signal peaks in a pyrogram. Pyrosequencing is increasingly used for bacterial detection, identification and typing, and, recently, a commercial system became available for the identification of bacterial isolates. Pyrosequencing can also be partially or fully automated, thus enabling the high-throughput analysis of samples. Wider use of pyrosequencing may occur in the future if longer nucleotide reads are made possible, which will enable its expansion into larger nucleotide sequencing such as multilocus sequence typing and whole-genome sequencing.</p
Detection of Neisseria meningitidis, Streptococcus pneumoniae, and Haemophilus influenzae in blood and cerebrospinal fluid using fluorescence-based PCR
No full textThe polymerase chain reaction (PCR) is a fundamental part of modern molecular biology. Fluorescence-based PCR methods also are now available, which enable rapid, specific, and sensitive assays for the amplification and analysis of deoxyribonucleic acid (DNA). These methods are performed in closed-tube format, thereby reducing the risk of contamination between stages. In addition, post-PCR processing, such as clean-up steps and gel electrophoresis, are eliminated as the results are read via an integrated fluorimeter. An example of this methodology is fluorescence-based PCR using dual-labeled probes, termed dual-labeled end-point fluorescence PCR. This method uses oligonucleotide probes that are dual-labeled with a reporter dye and quencher dye. The method has the advantage that DNA extraction, liquid handling, PCR, and analysis also can be fully automated. In this chapter, the simultaneous detection of Neisseria meningitidis, Streptococcus pneumoniae, and Haemophilus influenzae from clinical samples is described
Diarrhoeagenic Escherichia coli—an emerging problem?
No full textDiarrhea remains one of the main sources of morbidity and morbidity in today's world and a large proportion is caused by diarrheagenic Escherichia coli. They are a particular problem in developed countries although traveler's 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.</p
Nucleotide sequence‐based typing of bacteria and the impact of automation
No full textDNA-based typing methods are increasingly important for the characterisation of bacteria. They are used to monitor the epidemiology of pathogens with public health significance and also to help understand the evolution and population biology of bacteria. However, these methods require accuracy and reproducibility and are often of a high-throughput nature. Laboratory automation is therefore the key to the successful implementation of such methods. This review describes the impact of automation on DNA-based typing methods, particularly multi-locus sequence typing (MLST), and the method components that can be automated.</p
The costs associated with the public health management of a cluster of meningococcal infection in England
No full textThe human and public health costs of meningococcal disease are substantial, with 201 cases reported in England between April and June 2013. It is estimated that serogroup B accounts for 85–90% of all cases of meningococcal disease. A vaccine against serogroup B meningococcal disease was approved by the European Medicines Agency in November 2012. In March 2014 the UK Joint Committee on Vaccination and Immunisation recommended that the vaccine be offered to babies at 2, 4 and 12 months of age, provided it can be procured at a cost effective price.Using a real cluster of two cases of meningococcal infection in the UK, we estimate that the cost of managing two cases was 17 times more than the cost of managing a single case (£5584.39 versus £317.72, respectively). We recommend that vaccine cost effectiveness models should take a full account of the costs involved in managing cases and clusters of meningococcal disease
Pyrosequencing: sequence typing at the speed of light
No full textNucleotide sequencing is an established method for gaining information relating to partial gene, whole gene, or whole genome sequence. Here we describe some of the background leading to the advent of modern nucleotide sequencing and how it has led to the development of Pyrosequencing, a relatively new method for real-time nucleotide sequencing. In particular, we describe how this method can be used for typing bacterial pathogens.</p
Genotypic characterization of Neisseria meningitidis using Pyrosequencing
No full textPyrosequencing involves the synthesis of single-stranded deoxyribonucleic acid leading to rapid and accurate analysis of nucleotide sequences. This article describes the development of typing assays for the characterization of Neisseria meningitidis using Pyrosequencing. This involved developing methods for the nucleotide sequence analysis of important variable regions contained on a major capsule gene and on outer membrane protein genes that are used for grouping, typing, and subtyping meningococci. To achieve this, primers were designed for amplification of three genes, siaD, porB, and porA from the four main serogroups B, C, Y, and W135. To facilitate throughput and reproducibility, the method was also automated. Data from 717 isolates have shown that Pyrosequencing can be used for the single nucleotide polymorphism and sequence-analysis characterization of meningococci.</p
A novel method for preparing single-stranded DNA for pyrosequencing
No full textPyrosequencing technology is a powerful genotyping tool that requires the generation of single stranded DNA. Currently, two simple, solid-phase-based methods are available for this, but they require special equipment, they are not automated, and they are relatively expensive because of the need for biotinylated polymerase chain reaction primers. In this article, an enzymatic liquid-phase method for the generation of high-quality, single-stranded DNA, and its novel use for Pyrosequencing are described. The method has also been fully automated.</p
Increased genetic diversity of Neisseria meningitidis isolates after the introduction of meningococcal serogroup C polysaccharide conjugate vaccines
No full textDuring 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.</p
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