Development and clinical validation of multiplex TaqMan® assays for rapid diagnosis of viral gastroenteritis.

There is a need to provide rapid, sensitive, and often high throughput detection of pathogens in diagnostic virology. Viral gastroenteritis is a serious health issue often leading to hospitalization in the young, the immunocompromised and the elderly. The common causes of viral gastroenteritis include rotavirus, norovirus (genogroups I and II), astrovirus, and group F adenoviruses (serotypes 40 and 41). This article describes the work-up of two internally controlled multiplex, probe-based PCR assays and reports on the clinical validation over a 3-year period, March 2007 to February 2010. Multiplex assays were developed using a combination of TaqMan™ and minor groove binder (MGB™) hydrolysis probes. The assays were validated using a panel of 137 specimens, previously positive via a nested gel-based assay. The assays had improved sensitivity for adenovirus, rotavirus, and norovirus (97.3% vs. 86.1%, 100% vs. 87.8%, and 95.1% vs. 79.5%, respectively) and also more specific for targets adenovirus, rotavirus, and norovirus (99% vs. 95.2%, 100% vs. 93.6%, and 97.9% vs. 92.3%, respectively). For the specimens tested, both assays had equal sensitivity and specificity for astrovirus (100%). Overall the probe-based assays detected 16 more positive specimens than the nested gel-based assay. Post-introduction to the routine diagnostic service, a total of 9,846 specimens were processed with multiplex 1 and 2 (7,053 pediatric, 2,793 adult) over the 3-year study period. This clinically validated, probe-based multiplex testing algorithm allows highly sensitive and timely diagnosis of the four most prominent causes of viral gastroenteritis.

Immunovirological Correlates in Human Rabies Treated With Therapeutic Coma

A 37-year-old woman was admitted to hospital and over the next 5 days developed a progressive encephalitis. Nuchal skin biopsy, analyzed using a Rabies TaqMan(C) PCR, demonstrated rabies virus RNA. She had a history in keeping with exposure to rabies whilst in South Africa, but had not received pre- or post-exposure prophylaxis. She was treated with a therapeutic coma according to the

Association of the G4 rotavirus genotype with gastroenteritis in adults

Rotavirus is the most common etiological cause of acute viral gastroenteritis in infants and young children worldwide, yet its role in the adult population is less well understood. We have recently identified rotavirus as the causative agent of severe diarrhea in adults, specifically in two gastroenteritis outbreaks in separate care for the elderly homes. Strain typing has shown the continued presence of P[8]G1, the emergence of P[8]G9, and the reemergence of P[8]G4. A total of 26 community cases and 6 outbreak cases of rotavirus infection, positive via a molecular screening assay, were subsequently amplified using VP4 and VP7 specific primers (Con2/Con3 and 1A/1B primer sets, respectively). The age range of patients investigated was from

Prevalence of human papillomavirus in women attending cervical screening in the UK and Ireland:New data from northern Ireland and a systematic review and meta-analysis

There is substantial international variation in human papillomavirus (HPV) prevalence; this study details the first report from Northern Ireland and additionally provides a systematic review and meta-analysis pooling the prevalence of high-risk (HR-HPV) subtypes among women with normal cytology in the UK and Ireland. Between February and December 2009, routine liquid based cytology (LBC) samples were collected for HPV detection (Roche Cobas® 4800 [PCR]) among unselected women attending for cervical cytology testing. Four electronic databases, including MEDLINE, were then searched from their inception till April 2011. A random effects meta-analysis was used to calculate a pooled HR-HPV prevalence and associated 95% confidence intervals (CI). 5,712 women, mean age 39 years (±SD 11.9 years; range 20-64 years), were included in the analysis, of which 5,068 (88.7%), 417 (7.3%) and 72 (1.3%) had normal, low, and high-grade cytological findings, respectively. Crude HR-HPV prevalence was 13.2% (95% CI, 12.7-13.7) among women with normal cytology and increased with cytological grade. In meta-analysis the pooled HR-HPV prevalence among those with normal cytology was 0.12 (95% CIs, 0.10-0.14; 21 studies) with the highest prevalence in younger women. HPV 16 and HPV 18 specific estimates were 0.03 (95% CI, 0.02-0.05) and 0.01 (95% CI, 0.01-0.02), respectively. The findings of this Northern Ireland study and meta-analysis verify the prevalent nature of HPV infection among younger women. Reporting of the type-specific prevalence of HPV infection is relevant for evaluating the impact of future HPV immunization initiatives, particularly against HR-HPV types other than HPV 16 and 18. J. Med. Virol. 85:295-308, 2013. © 2012 Wiley Periodicals, Inc. Copyright © 2012 Wiley Periodicals, Inc.

HPV prevalence and type-distribution in cervical cancer and premalignant lesions of the cervix: A population-based study from Northern Ireland

Assessment of Human papillomavirus (HPV) prevalence and genotype distribution is important for monitoring the impact of prophylactic HPV vaccination. This study aimed to demonstrate the HPV genotypes predominating in pre-malignant and cervical cancers in Northern Ireland (NI) before the vaccination campaign has effect. Formalin fixed paraffin embedded tissue blocks from 2,303 women aged 16-93 years throughout NI were collated between April 2011 and February 2013. HPV DNA was amplified by PCR and HPV genotyping undertaken using the Roche^® linear array detection kit. In total, 1,241 out of 1,830 eligible samples (68.0%) tested positive for HPV, with the majority of these [1,181/1,830 (64.5%)] having high-risk (HR) HPV infection; 37.4% were positive for HPV-16 (n=684) and 5.1% for HPV-18 (n=93). HPV type-specific prevalence was 48.1%, 65.9%, 81.3%, 92.2%, and 64.3% among cervical intraepithelial neoplasias (CIN) Grades I-III, squamous cell carcinomas (SCC) and adenocarcinoma (AC) cases, respectively. Most SCC cases (81.3%) had only one HPV genotype detected and almost a third (32.0%) of all cervical pathologies were HPV negative including 51.9% of CIN I (n=283), 34.1% CIN II (n=145), 18.7% of CIN III (n=146), 7.8% of SCC (n=5), and 35.7% of AC (n=5) cases. This study provides important baseline data for monitoring the effect of HPV vaccination in NI and for comparison with other UK regions. The coverage of other HR-HPV genotypes apart from 16 and 18, including HPV-45, 31, 39, and 52, and the potential for cross protection, should be considered when considering future polyvalent vaccines.

Developing virtual patients for medical microbiology education

The landscape of medical education is changing as students embrace the accessibility and interactivity of e-learning. Virtual patients are e-learning resources that may be used to advance microbiology education. Although the development of virtual patients has been widely considered, here we aim to provide a coherent approach for clinical educators.