Peter Muir

Evolutionary Dynamics and Temporal/Geographical Correlates of Recombination in the Human Enterovirus Echovirus Types 9, 11, and 30

ABSTRACT The relationship between virus evolution and recombination in species B human enteroviruses was investigated through large-scale genetic analysis of echovirus type 9 (E9) and E11 isolates (n = 85 and 116) from 16 European, African, and Asian countries between 1995 and 2008. Cluster 1 E9 isolates and genotype D5 and A E11 isolates showed evidence of frequent recombination between the VP1 and 3Dpol regions, the latter falling into 23 (E9) and 43 (E11) clades interspersed phylogenetically with 46 3Dpol clades of E30 and with those of other species B serotypes. Remarkably, only 2 of the 112 3Dpol clades were shared by more than one serotype (E11 and E30), demonstrating an extremely large and genetically heterogeneous recombination pool of species B nonstructural-region variants. The likelihood of recombination increased with geographical separation and time, and both were correlated with VP1 divergence, whose substitution rates allowed recombination half-lives of 1.3, 9.8, and 3.1 years, respectively, for E9, E11, and E30 to be calculated. These marked differences in recombination dynamics matched epidemiological patterns of periodic epidemic cycles of 2 to 3 (E9) and 5 to 6 (E30) years and the longer-term endemic pattern of E11 infections. Phylotemporal analysis using a Bayesian Markov chain Monte Carlo method, which placed recombination events within the evolutionary reconstruction of VP1, showed a close relationship with VP1 lineage expansion, with defined recombination events that correlated with their epidemiological periodicity. Whether recombination events contribute directly to changes in transmissibility that drive epidemic behavior or occur stochastically during periodic population bottlenecks is an unresolved issue vital to future understanding of enterovirus molecular epidemiology and pathogenesis.

Identification of hepatitis C virus seroconversion resulting from nosocomial transmission on a haemodialysis unit: Implications for infection control and laboratory screening

Hepatitis C virus (HCV) seroconversion was detected by routine screening in a haemodialysis patient, Patient 1. Serological investigations were undertaken over the following 3 months to determine if further transmission to other patients on the unit had occurred. No additional cases were identified. Twenty‐two haemodialysis patients known to have HCV infection were investigated using molecular epidemiological methods to determine if transmission between patients had occurred. HCV viraemia was demonstrated by polymerase chain reaction in 19 of 22 patients (86%). Genotyping showed that eight patients were infected with genotype 1, three with genotype 3 and eight, including Patient 1, with genotype 2. Phylogenetic analysis of viral sequences from the eight patients with genotype 2 revealed three, including Patient 1,with a novel subtype of HCV type 2, and revealed close similarity between viral sequences from patient 1 and those from one other patient, suggesting transmission. This was consistent with haemodialysis histories. Among other patients with genotype 2, there were two with subtype 2a and three others with three separate novel subtypes, as yet undesignated. With the exception of patient 1, all patients infected with novel subtypes were of Afro‐Caribbean origin. The HCV prevalence among patients on the haemodialysis unit was high (14%), which may reflect the ethnicity of our haemodialysis population. This case emphasises the risk of nosocomial transmission and the importance of infection control procedures on haemodialysis units, and highlights the usefulness of molecular epidemiological techniques for the investigation of outbreaks of HCV infection. J. Med. Virol. 59:135–140, 1999.

Multicenter Proficiency Testing of Nucleic Acid Amplification Methods for the Detection of Enteroviruses

ABSTRACT A multicenter study of molecular detection of enteroviruses was conducted using a proficiency panel. Of 70 data sets, 46 (66%) reported correct results for samples containing at least 1 50% infective dose per ml and for negative samples. Variation in performance between laboratories demonstrates the need for ongoing quality control.

2015 UK National Guideline on the management of non-gonococcal urethritis

We present the updated British Association for Sexual Health and HIV guideline for the management of non-gonococcal urethritis in men. This document includes a review of the current literature on its aetiology, diagnosis and management. In particular it highlights the emerging evidence that azithromycin 1u2009g may result in the development of antimicrobial resistance in Mycoplasma genitalium and that neither azithromycin 1u2009g nor doxycycline 100u2009mg twice daily for seven days achieves a cure rate of >90% for this micro-organism. Evidence-based diagnostic and management strategies for men presenting with symptoms suggestive of urethritis, those confirmed to have non-gonococcal urethritis and those with persistent symptoms following first-line treatment are detailed.

Relationships Between Rhinitis Symptoms, Respiratory Viral Infections and Nasopharyngeal Colonization With Streptococcus pneumoniae, Haemophilus influenzae and Staphylococcus aureus in Children Attending Daycare

Background: Nasal bacterial colonization is often dubbed “asymptomatic.” We hypothesized that rhinitis, common in preschool children, is associated with bacterial colonization and that respiratory viruses, which cause rhinitis, interact with bacteria in ways which promote transmission. Methods: Five hundred eighty-five children (4.2–73.6 months) attending daycare had clinical information, a rhinitis score and nasal swabs collected in February 2009. Swabs in soya tryptone glucose glycerine broth were cultured for Streptococcus pneumoniae (Sp), Haemophilus influenzae (Hi) and Staphylococcus aureus and analyzed by real-time polymerase chain reaction for respiratory viruses, both semiquantitatively. Results: Rhinitis symptoms, carriage of Sp and Hi and viral infection fell, whereas S. aureus carriage rates rose with age. Significant, age-independent associations between rhinitis symptoms and detection of Hi (P < 0.033) and Hi colonization density (P < 0.027) were observed. Of the 42% with detected viruses, most (78%) had picornavirus infection. There was a significant age-independent association between viral infection (and viral load, picornavirus infection and picornaviral load) and detection of Sp (P = 0.020, 0.035, 0.005, 0.014) and between viral infection and viral load and Sp colonization density (P = 0.024, 0.028). Conclusions: Hi may promote its own transmission by inducing or amplifying rhinitis in children. There is a close quantitative relationship between respiratory viral infection, including picornavirus infection and Sp colonization. These findings have implications for understanding disease pathogenesis and formulating prevention strategies using vaccines.

Development and Assay of RNA Transcripts of Enterovirus Species A to D, Rhinovirus Species A to C, and Human Parechovirus: Assessment of Assay Sensitivity and Specificity of Real-Time Screening and Typing Methods

ABSTRACT Nucleic acid amplification methods such as the PCR have had a major impact on the diagnosis of viral infections, often achieving greater sensitivities and shorter turnaround times than conventional assays and an ability to detect viruses refractory to conventional isolation methods. Their effectiveness is, however, significantly influenced by assay target sequence variability due to natural diversity and rapid sequence changes in viruses that prevent effective binding of primers and probes. This was investigated for a diverse range of enteroviruses (EVs; species A to D), human rhinoviruses (HRVs; species A to C), and human parechovirus (HPeV) in a multicenter assay evaluation using a series of full-length prequantified RNA transcripts. RNA concentrations were quantified by absorption (NanoDrop) and fluorescence methods (RiboGreen) prior to dilution in buffer supplemented with RNase inhibitors and carrier RNA. RNA transcripts were extremely stable, showing minimal degradation after prolonged storage at temperatures between ambient and −20°C and after multiple freeze-thaw cycles. Transcript dilutions distributed to six referral laboratories were screened by real-time reverse transcriptase PCR assays using different primers and probes. All of the laboratories reported high assay sensitivities for EV and HPeV transcripts approaching single copies and similar amplification kinetics for all four EV species. HRV detection sensitivities were more variable, often with substantially impaired detection of HRV species C. This could be accounted for in part by the placement of primers and probes to genetically variable target regions. Transcripts developed in this study provide reagents for the ongoing development of effective diagnostics that accommodate increasing knowledge of genetic heterogeneity of diagnostic targets.

The TARGET cohort study protocol: a prospective primary care cohort study to derive and validate a clinical prediction rule to improve the targeting of antibiotics in children with respiratory tract illnesses

BackgroundChildren with respiratory tract infections are the single most frequent patient group to make use of primary care health care resources. The use of antibiotics remains highly prevalent in young children, but can lead to antimicrobial resistance as well as reinforcing the idea that parents should re-consult for similar symptoms. One of the main drivers of indiscriminate antimicrobial use is the lack of evidence for, and therefore uncertainty regarding, which children are at risk of poor outcome. This paper describes the protocol for the TARGET cohort study, which aims to derive and validate a clinical prediction rule to identify children presenting to primary care with respiratory tract infections who are at risk of hospitalisation.Methods/designThe TARGET cohort study is a large, multicentre prospective observational study aiming to recruit 8,300 children aged ≥3 months and <16 years presenting to primary care with a cough and respiratory tract infection symptoms from 4 study centres (Bristol, London, Oxford and Southampton). Following informed consent, symptoms, signs and demographics will be measured. In around a quarter of children from the Bristol centre, a single sweep, dual bacterial-viral throat swab will be taken and parents asked to complete a symptom diary until the child is completely well or for 28 days, whichever is sooner. A review of medical notes including clinical history, re-consultation and hospitalisations will be undertaken. Multivariable logistic regression will be used to identify the independent clinical predictors of hospitalisation as well as the prognostic significance of upper respiratory tract microbes. The clinical prediction rule will be internally validated using various methods including bootstrapping.DiscussionThe clinical prediction rule for hospitalisation has the potential to help identify a small group of children for hospitalisation and a much larger group where hospitalisation is very unlikely and antibiotic prescribing would be less warranted. This study will also be the largest natural history study to date of children presenting to primary care with acute cough and respiratory tract infections, and will provide important information on symptom duration, re-consultations and the microbiology of the upper respiratory tract.

Serotype-Specific Detection of Coxsackievirus A16 in Clinical Specimens by Reverse Transcription-Nested PCR

ABSTRACT We describe the development of a coxsackievirus A16 (CVA16) serotype-specific PCR which correctly differentiated between CVA16 and other enterovirus serotypes of both laboratory isolates and clinical specimens. The assay will be useful for monitoring CVA16 outbreaks and studying the disease association, epidemiology, and evolution of this common enterovirus serotype.

Development and internal validation of a clinical rule to improve antibiotic use in children presenting to primary care with acute respiratory tract infection and cough: a prognostic cohort study

Summary Background Antimicrobial resistance is a serious threat to public health, with most antibiotics prescribed in primary care. General practitioners (GPs) report defensive antibiotic prescribing to mitigate perceived risk of future hospital admission in children with respiratory tract infections. We developed a clinical rule aimed to reduce clinical uncertainty by stratifying risk of future hospital admission. Methods 8394 children aged between 3 months and 16 years presenting with acute cough (for ≤28 days) and respiratory tract infection were recruited to a prognostic cohort study from 247 general practitioner practices in England. Exposure variables included demographic characteristics, parent-reported symptoms, and physical examination signs. The outcome was hospital admission for respiratory tract infection within 30 days, collected using a structured, blinded review of medical records. Findings 8394 (100%) children were included in the analysis, with 78 (0·9%, 95% CI 0·7%–1·2%) admitted to hospital: 15 (19%) were admitted on the day of recruitment (day 1), 33 (42%) on days 2–7; and 30 (39%) on days 8–30. Seven characteristics were independently associated (p<0·01) with hospital admission: age <2 years, current asthma, illness duration of 3 days or less, parent-reported moderate or severe vomiting in the previous 24 h, parent-reported severe fever in the previous 24 h or a body temperature of 37·8°C or more at presentation, clinician-reported intercostal or subcostal recession, and clinician-reported wheeze on auscultation. The area under the receiver operating characteristic (AUROC) curve for the coefficient-based clinical rule was 0·82 (95% CI 0·77–0·87, bootstrap validated 0·81). Assigning one point per characteristic, a points-based clinical rule consisting of short illness, temperature, age, recession, wheeze, asthma, and vomiting (mnemonic STARWAVe; AUROC 0·81, 0·76–0·85) distinguished three hospital admission risk strata: very low (0·3%, 0·2–0·4%) with 1 point or less, normal (1·5%, 1·0–1·9%) with 2 or 3 points, and high (11·8%, 7·3–16·2%) with 4 points or more. Interpretation Clinical characteristics can distinguish children at very low, normal, and high risk of future hospital admission for respiratory tract infection and could be used to reduce antibiotic prescriptions in primary care for children at very low risk. Funding National Institute for Health Research (NIHR).

Clinical presentation and microbiological diagnosis in paediatric respiratory tract infection: a systematic review

BACKGROUNDnAntibiotic prescribing decisions for respiratory tract infection (RTI) in primary care could be improved if clinicians could target bacterial infections. However, there are currently no evidence-based diagnostic rules to identify microbial aetiology in children presenting with acute RTIs.nnnAIMnTo analyse evidence of associations between clinical symptoms or signs and detection of microbes from the upper respiratory tract (URT) of children with acute cough.nnnDESIGN AND SETTINGnSystematic review and meta-analysis.nnnMETHODnA literature search identified articles reporting relationships between individual symptoms and/or signs, and microbes detected from URT samples. Associations between pathogens and symptoms or signs were summarised, and meta-analysis conducted where possible.nnnRESULTSnThere were 9984 articles identified, of which 28 met inclusion criteria. Studies identified 30 symptoms and 41 signs for 23 microbes, yielding 1704 potential associations, of which only 226 (13%) have presently been investigated. Of these, relevant statistical analyses were presented for 175 associations, of which 25% were significant. Meta-analysis demonstrated significant relationships between respiratory syncytial virus (RSV) detection and chest retractions (pooled odds ratio [OR] 1.9, 95% confidence interval [CI] = 1.6 to 2.3), wheeze (pooled OR 1.7, 95% CI = 1.5 to 2.0), and crepitations/crackles (pooled OR 1.7, 95% CI = 1.3 to 2.2).nnnCONCLUSIONSnThere was an absence of evidence for URT pathogens other than RSV. The meta-analysis identified clinical signs associated with RSV detection, suggesting clinical presentation may offer some, albeit poor, diagnostic value. Further research is urgently needed to establish the value of symptoms and signs in determining microbiological aetiology and improve targeting of antibiotics in primary care.