Neuroimaging in encephalitis: analysis of imaging findings and interobserver agreement.

AIM: To assess the role of imaging in the early management of encephalitis and the agreement on findings in a well-defined cohort of suspected encephalitis cases enrolled in the Prospective Aetiological Study of Encephalitis conducted by the Health Protection Agency (now incorporated into Public Health England). MATERIALS AND METHODS: Eighty-five CT examinations from 68 patients and 101 MRI examinations from 80 patients with suspected encephalitis were independently rated by three neuroradiologists blinded to patient and clinical details. The level of agreement on the interpretation of images was measured using the kappa statistic. The sensitivity, specificity, and negative and positive predictive values of CT and MRI for herpes simplex virus (HSV) encephalitis and acute disseminated encephalomyelitis (ADEM) were estimated. RESULTS: The kappa value for interobserver agreement on rating the scans as normal or abnormal was good (0.65) for CT and moderate (0.59) for MRI. Agreement for HSV encephalitis was very good for CT (0.87) and MRI (0.82), but only fair for ADEM (0.32 CT; 0.31 MRI). Similarly, the overall sensitivity of imaging for HSV encephalitis was ∼80% for both CT and MRI, whereas for ADEM it was 0% for CT and 20% for MRI. MRI specificity for HSV encephalitis between 3-10 days after symptom onset was 100%. CONCLUSION: There is a subjective component to scan interpretation that can have important implications for the clinical management of encephalitis cases. Neuroradiologists were good at diagnosing HSV encephalitis; however, agreement was worse for ADEM and other alternative aetiologies. Findings highlight the importance of a comprehensive and multidisciplinary approach to diagnosing the cause of encephalitis that takes into account individual clinical, microbiological, and radiological features of each patient.

Diagnostic strategy used to establish etiologies of encephalitis in a prospective cohort of patients in England.

The laboratory diagnostic strategy used to determine the etiology of encephalitis in 203 patients is reported. An etiological diagnosis was made by first-line laboratory testing for 111 (55%) patients. Subsequent testing, based on individual case reviews, resulted in 17 (8%) further diagnoses, of which 12 (71%) were immune-mediated and 5 (29%) were due to infection. Seventy-five cases were of unknown etiology. Sixteen (8%) of 203 samples were found to be associated with either N-methyl-d-aspartate receptor or voltage-gated potassium channel complex antibodies. The most common viral causes identified were herpes simplex virus (HSV) (19%) and varicella-zoster virus (5%), while the most important bacterial cause was Mycobacterium tuberculosis (5%). The diagnostic value of testing cerebrospinal fluid (CSF) for antibody was assessed using 139 samples from 99 patients, and antibody was detected in 46 samples from 37 patients. Samples collected at 14 to 28 days were more likely to be positive than samples taken 0 to 6 days postadmission. Three PCR-negative HSV cases were diagnosed by the presence of virus-specific antibody in the central nervous system (CNS). It was not possible to make an etiological diagnosis for one-third of the cases; these were therefore considered to be due to unknown causes. Delayed sampling did not contribute to these cases. Twenty percent of the patients with infections with an unknown etiology showed evidence of localized immune activation within the CNS, but no novel viral DNA or RNA sequences were found. We conclude that a good standard of clinical investigation and thorough first-line laboratory testing allows the diagnosis of most cases of infectious encephalitis; testing for CSF antibodies allows further cases to be diagnosed. It is important that testing for immune-mediated causes also be included in a diagnostic algorithm.

Causality in acute encephalitis: defining aetiologies.

Defining the causal relationship between a microbe and encephalitis is complex. Over 100 different infectious agents may cause encephalitis, often as one of the rarer manifestations of infection. The gold-standard techniques to detect causative infectious agents in encephalitis in life depend on the study of brain biopsy material; however, in most cases this is not possible. We present the UK perspective on aetiological case definitions for acute encephalitis and extend them to include immune-mediated causes. Expert opinion was primarily used and was supplemented by literature-based methods. Wide usage of these definitions will facilitate comparison between studies and result in a better understanding of the causes of this devastating condition. They provide a framework for regular review and updating as the knowledge base increases both clinically and through improvements in diagnostic methods. The importance of new and emerging pathogens as causes of encephalitis can be assessed against the principles laid out here.

Evaluation of a measles vaccine campaign by oral-fluid surveys in a rural Kenyan district: interpretation of antibody prevalence data using mixture models.

We evaluated the effectiveness of a measles vaccine campaign in rural Kenya, based on oral-fluid surveys and mixture-modelling analysis. Specimens were collected from 886 children aged 9 months to 14 years pre-campaign and from a comparison sample of 598 children aged 6 months post-campaign. Quantitative measles-specific antibody data were obtained by commercial kit. The estimated proportions of measles-specific antibody negative in children aged 0-4, 5-9 and 10-14 years were 51%, 42% and 27%, respectively, pre- campaign and 18%, 14% and 6%, respectively, post-campaign. We estimate a reduction in the proportion susceptible of 65-78%, with approximately 85% of the population recorded to have received vaccine. The proportion of 'weak' positive individuals rose from 35% pre-campaign to 54% post-campaign. Our results confirm the effectiveness of the campaign in reducing susceptibility to measles and demonstrate the potential of oral-fluid studies to monitor the impact of measles vaccination campaigns.

A randomised placebo-controlled trial to explore the effect of suppressive therapy with acyclovir on genital shedding of HIV-1 and herpes simplex virus type 2 among Zimbabwean sex workers.

OBJECTIVES: To determine the effect of daily acyclovir on genital shedding of HIV-1 and herpes simplex virus type 2 (HSV-2) in a randomised placebo-controlled trial among rural Zimbabwean sex workers. METHODS: 214 women were recruited and tested for HIV-1 and HSV-2 antibodies, HIV plasma viral load, CD4 lymphocyte count and genital swabs for qualitative detection of HIV-1 and HSV-2 genital shedding. Women were randomly assigned to acyclovir 400 mg twice a day for 12 weeks or matching placebo and were followed weekly to detect HIV-1 or HSV-2 genital shedding. Shedding analyses were only undertaken on 125 women co-infected with HSV-2 and HIV-1. Data were analysed using logistic regression, with random effects modelling used to account for repeated measurements on the same women. RESULTS: All women were randomly assigned to acyclovir or placebo; 125 of whom were co-infected with HIV-1 and HSV-2. 69 women were randomly assigned to acyclovir and 56 to placebo. Although twice daily acyclovir reduced rates of HSV-2 genital shedding, (adjusted odds ratio (AOR) 0.24; 95% CI 0.12 to 0.48; less than p<0.001), it had no effect on the proportion of visits at which HIV-1 shedding was detected (AOR 1.08; 95% CI 0.48 to 2.42; p = 0.9). Adherence varied between participants but even when adherence was high (as determined by pill count and extent of HSV-2 suppression) HIV-1 shedding was not reduced. CONCLUSION: Among these HIV-1 and HSV-2-seropositive women, suppressive acyclovir therapy had no effect on the rate of HIV genital shedding despite a reduction in genital HSV-2. Treatment adherence and its measurement clearly affect the interpretation of these results.

Viral gastroenteritis outbreaks in deployed British troops during 2002-7.

OBJECTIVES: The aim of this study was to see what lessons could be learnt from the suspected viral gastroenteritis outbreaks that have occurred in deployed British troops during 2002-7. METHOD: Epidemiological and laboratory data from identifiable outbreaks were reviewed, including epidemic curves and the results of PCR testing for enteropathic viruses. RESULTS: The epidemic curves of outbreaks varied predictably in accordance with the size of the population at risk and whether this population was constant or expanding. Of 11 outbreaks identified, 10 (91%) had a proven viral cause and 10 (91%) occurred in Iraq. Of 84 enteropathic viruses identified, 61 (73%) were noroviruses and these included both unknown strains and those that were common in the UK and Europe. Of the 10 viral outbreaks, 3 (30%) occurred in medical units, 5 (50%) were associated with large-scale relief in place (RiP) deployments and 5 (50%) involved >3 different viruses, which is strongly suggestive of food or water contamination. CONCLUSION: These findings can help to predict future viral gastroenteritis outbreaks and target improved prevention strategies appropriately. However, more systematic studies are now required.

The seroepidemiology of varicella zoster virus among pregnant Bangladeshi and white British women in the London Borough of Tower Hamlets, UK.

We investigated the comparative seroepidemiology of varicella zoster virus (VZV) in pregnant women of two ethnic groups, white British and Bangladeshi, living in an inner city area of London, United Kingdom. Women aged 16-45 years were recruited from antenatal clinics of the Royal London Hospital in the Borough of Tower Hamlets. Complete data were obtained from 275 white British and 765 Bangladeshi women. VZV antibody prevalence was 93.1% (95% CI 89.4-95.8) and 86.0% (95% CI 83.3-88.4) respectively. Women who were born in Bangladesh and lived there at least until the age of 15 years had the lowest odds of being immune (OR 0.37, 95% CI 0.22-0.63). This implies they will have an increased risk of varicella during pregnancy. Women arriving in the United Kingdom in adulthood should be screened routinely during pregnancy and vaccination offered postpartum if they are susceptible.

Serological and molecular epidemiology of measles virus outbreaks reported in Ethiopia during 2000-2004.

Twenty-eight outbreaks in six regions and two major cities in Ethiopia from 2000 to 2004 were investigated, with the collection of 207 venous blood and/or oral fluid samples. Measles diagnosis was confirmed by detection of measles-specific IgM and/or detection of measles virus by polymerase chain reaction (PCR). Of 176 suspected cases tested for specific measles IgM, 142 (81%) were IgM positive. Suspected cases in vaccinated children were much less likely to be laboratory confirmed than in unvaccinated children (42% vs. 83%, P < 0.0001). Of 197 samples analyzed by RT-PCR measles virus genome was detected in 84 (43%). A total of 58 wild-type measles viruses were characterized by nucleic acid sequence analysis of the nucleoprotein (N) and hemagglutinin (H) genes. Two recognized genotypes (D4 and B3) were identified. Each outbreak comprised only a single genotype and outbreaks of each genotype tended to occur in distinct geographical locations. B3 was first observed in 2002, and has now been the cause of three documented outbreaks near to the border of Sudan. D4 genotype was previously observed in an outbreak in 1999 and occurs in more diverse locations throughout the country. These data yield insights into geographical and age-related sources of continued transmission. Refinement of measles control measures might include targeting older age groups (5-14 years) and strengthening routine immunization particularly where importation of cases is a concern.

Evaluation of a commercial rubella IgM assay for use on oral fluid samples for diagnosis and surveillance of congenital rubella syndrome and postnatal rubella.

BACKGROUND: Clinical diagnosis (surveillance) of rubella is unreliable and laboratory confirmation is essential. Detection of virus specific IgM in serum is the most commonly used method. However, the use of serum necessitates the drawing of blood, either through venipuncture or finger/heel prick, which can be difficult in young babies. Oral fluid samples have proved useful as an alternative, less invasive sample for virus specific IgM detection however until recently no commercial rubella IgM tests were available, restricting the usefulness of this approach. OBJECTIVES: To evaluate the performance of the Microimmune Rubella IgM capture EIA using oral fluid samples from outbreaks as well as in cases of suspected congenital rubella syndrome (CRS). STUDY DESIGN: Paired serum and oral fluids were collected from cases during a rubella outbreak in three provinces in Turkey. Matched serum and oral fluid samples were collected from children with suspected CRS in an active surveillance programme at the Aravind Eye Hospital in South India. Serum samples were collected as part of the measles surveillance programme in Ethiopia. RESULTS: On serum samples the sensitivity and specificity of the Microimmune Rubella IgM capture EIA compared to Behring Enzygnost rubella IgM test was 96.9% (62/64; 95% CI 94.2-100%) and 100% (53/53; 95% CI 93.2-100%). On oral fluids compared to matched Behring results on serum the sensitivity was 95.5% (42/44; 95% CI 84.5-99.4%). The sensitivity and specificity of Microimmune Rubella IgM capture EIA on oral fluids from suspected CRS cases compared to serum results using Behring Enzygnost IgM assay was 100% (95% CI 84.5-100%) and 100% (95% CI 95.8-100.0%) respectively. CONCLUSION: Microimmune Rubella IgM capture EIA has adequate performance for diagnosis and surveillance of rubella in outbreak using either serum or oral fluid specimens.

Measles viral load may reflect SSPE disease progression.

Subacute sclerosing panencephalitis (SSPE) is a rare, slowly progressive neurological disorder caused by the persistent infection with measles virus (MV). Despite much research into SSPE, its pathology remains obscure. We examined autopsy tissues of eight SSPE patients by real time quantitative PCR, immunohistochemistry and immunoblotting to determine viral load. MV N, M and H gene RNA could be detected in the central nervous system (CNS) of all patients and in two non-CNS tissues of one patient. The viral burden between patients differed up to four-fold by quantitative PCR and corresponded with detection of MV protein. The level of both viral RNA and antigen in the brain may correlate with disease progression.

Institutional risk factors for outbreaks of nosocomial gastroenteritis: survival analysis of a cohort of hospital units in South-west England, 2002-2003.

Nosocomial outbreaks of gastroenteritis are a major burden on hospital inpatient services, costing an estimated pound115 million annually to the English National Health Service. We actively followed-up 171 inpatient units from four major acute hospitals and 11 community hospitals in South-west England for one year. Outbreaks of gastroenteritis were ascertained through an active surveillance network using standard clinical definitions. Survival analysis Cox regression models using an outbreak of gastroenteritis as the endpoint were fitted to identify institutional and operational attributes related to increased outbreak rates at the level of the care unit. Greater number of beds in unit [hazard ratio (HR) 1.22 (per 10 additional beds), 95% confidence intervals (CI) 0.96-1.55] was associated with increased hazard, as were geriatric (HR 2.6, 95%CI 1.6-4.3) and general medical (HR 1.7, 95%CI 1.1-2.6) care units. The average length of stay on a unit was inversely associated with outbreak incidence [HR=0.89 (per additional week of stay), 95%CI 0.80-0.99]. Larger care units and those with higher throughput have increased rates of gastroenteritis outbreaks. These results should guide infection control policy and support the design of hospitals with smaller care units.

Detection of multiple enteric virus strains within a foodborne outbreak of gastroenteritis: an indication of the source of contamination.

An outbreak of acute gastroenteritis of suspected viral aetiology occurred in April 2003 in the British Royal Fleet Auxiliary ship (RFA) Argus deployed in the Northern Arabian Gulf. There were 37 cases amongst a crew of 400 personnel. Of 13 samples examined from cases amongst the crew, six enteric viruses were detected by reverse transcriptase polymerase chain reaction (RT-PCR). Five different viruses were identified including, three norovirus genotypes, a sapovirus and a rotavirus. No multiple infections were detected. A common food source was implicated in the outbreak and epidemiological analysis showed a statistically significant association with salad as the source of the outbreak, with a relative risk of 3.41 (95% confidence interval of 1.7-6.81) of eating salad on a particular date prior to the onset of symptoms. Faecal contamination of the salad at source was the most probable explanation for the diversity of viruses detected and characterized.

A chimeric bovine enteric calicivirus: evidence for genomic recombination in genogroup III of the Norovirus genus of the Caliciviridae.

The Norovirus genus of the Caliciviridae encompasses viruses that cause outbreaks of gastroenteritis in human and viruses that have been associated with diarrhea in cattle. The two bovine noroviruses, Bo/Newbury2/76/UK and Bo/Jena/80/DE, represent two distinct genetic clusters in the newly described genogroup III. In the present study, Jena-like polymerase sequences were identified for the first time in the UK, but one of these, Bo/Thirsk10/00/UK, was a chimeric virus. Bo/Thirsk10/00/UK had a Jena-like polymerase gene but Newbury2-like capsid and ORF3 genes by comparison of their genome organization, nucleotide, and amino acid identities and phylogenetic analyses. The present study is one of few studies to clearly demonstrate the existence of chimeric genomes in the Norovirus genus and the first, to our knowledge, to identify a chimeric genome in genogroup III. It provides additional support that genomic recombination is part of the natural evolution of noroviruses and is relevant to the diagnosis and immunological control of norovirus diarrhea outbreaks.

Noroviruses associated with acute gastroenteritis in a children's day care facility in Rio de Janeiro, Brazil.

Noroviruses (Norwalk-like viruses) are an important cause of gastroenteritis worldwide. They are the most common cause of outbreaks of gastroenteritis in the adult population and occur in nursing homes for the elderly, geriatric wards, medical wards, and in hotel and restaurant settings. Food-borne outbreaks have also occurred following consumption of contaminated oysters. This study describes the application of a reverse transcription-polymerase chain reaction (RT-PCR) assay using random primers (PdN6) and specific Ni and E3 primers, directed at a small region of the RNA-dependent RNA polymerase-coding region of the norovirus genome, and DNA sequencing for the detection and preliminary characterisation of noroviruses in outbreaks of gastroenteritis in children in Brazil. The outbreak samples were collected from children <5 years of age at the Bertha Lutz children's day care facility at Oswaldo Cruz Foundation (Fiocruz), Rio de Janeiro, that occurred between 1996 and 1998, where no pathogen had been identified. At the Bertha Lutz day care center facility, only Fiocruz's employee children are provided for, and they come from different social, economic and cultural backgrounds. Three distinct genogroup II strains were detected in three outbreaks in 1997/98 and were most closely related to genotypes GII-3 (Mexico virus) and GII-4 (Grimsby virus), both of which have been detected in paediatric and adult outbreaks of gastroenteritis worldwide.

Rubella.

Maternal rubella is now rare in many developed countries that have rubella vaccination programmes. However, in many developing countries congenital rubella syndrome (CRS) remains a major cause of developmental anomalies, particularly blindness and deafness. WHO have provided recommendations for prevention of CRS, and, encouragingly, the number of countries introducing rubella vaccination programmes has risen. However, declining uptake rates due to concerns about the measles-mumps-rubella vaccine in the UK, and increasing numbers of cases in some European countries coupled with poor uptake rates might jeopardise this progress. Surveillance of postnatally and congenitally acquired infection is an essential component of CRS prevention since rubella is difficult to diagnose on clinical grounds alone. Laboratory differentiation of rubella from other rash-causing infections, such as measles, parvovirus B19, human herpesvirus 6, and enteroviruses in developed countries, and various endemic arboviruses is essential. Reverse transcriptase PCR and sequencing for diagnosis and molecular epidemiological investigation and detection of rubella-specific IgG and IgM salivary antibody responses in oral fluid are now available.

A population-based seroprevalence study of hepatitis A virus using oral fluid in England and Wales.

Population-based seroprevalence studies provide important data on susceptible groups and the potential for future outbreaks. However, the invasive nature of serum collection has limited studies. This paper describes the first postal population-based survey using noninvasive oral fluid technology to collect antibody prevalence data in conjunction with extensive risk factor data to assess the distribution of immunity to common viral infections in England and Wales. These results pertain to hepatitis A virus (HAV). Approximately 5,500 oral fluid samples were collected between August 2001 and May 2002, as well as individual risk factor data through a questionnaire, from persons aged less than 45 years randomly sampled from general practices countrywide. Samples were tested for immunoglobulin G-specific antibody marking a past infection or immunity to HAV using an antibody-capture enzyme-linked immunosorbent assay. The age-specific HAV seroprevalences indicated a low incidence of infection (overall seroprevalence of 18.9% (95% confidence interval: 17.0, 20.9) and of 9.2% (95% confidence interval: 7.1, 11.3) after the exclusion of vaccinees). Vaccination proved the most important determinant of seropositivity. Ethnic minority groups were underrepresented, and adjustment increased the overall prevalence to 20.1% and to 12.1% in unvaccinated individuals. The availability of comprehensive risk factor data allowed the description of two risk profiles related to natural infection and vaccination.

Noroviruses associated with acute gastroenteritis in a children's day care facility in Rio de Janeiro, Brazil

Noroviruses (Norwalk-like viruses) are an important cause of gastroenteritis worldwide. They are the most common cause of outbreaks of gastroenteritis in the adult population and occur in nursing homes for the elderly, geriatric wards, medical wards, and in hotel and restaurant settings. Food-borne outbreaks have also occurred following consumption of contaminated oysters. This study describes the application of a reverse transcription-polymerase chain reaction (RT-PCR) assay using random primers (PdN6) and specific Ni and E3 primers, directed at a small region of the RNA-dependent RNA polymerase-coding region of the norovirus genome, and DNA sequencing for the detection and preliminary characterisation of noroviruses in outbreaks of gastroenteritis in children in Brazil. The outbreak samples were collected from children <5 years of age at the Bertha Lutz children's day care facility at Oswaldo Cruz Foundation (Fiocruz), Rio de Janeiro, that occurred between 1996 and 1998, where no pathogen had been identified. At the Bertha Lutz day care center facility, only Fiocruz's employee children are provided for, and they come from different social, economic and cultural backgrounds. Three distinct genogroup II strains were detected in three outbreaks in 1997/98 and were most closely related to genotypes GII-3 (Mexico virus) and GII-4 (Grimsby virus), both of which have been detected in paediatric and adult outbreaks of gastroenteritis worldwide.

A novel method of oral fluid collection to monitor immunity to common viral infections.

Serological surveys among representative population samples have proved rare given their reliance on invasive sample collection. We therefore completed the first population-based postal survey of immunity in England and Wales using new oral fluid technology. This paper examines the feasibility of this new methodological approach. Nearly 5500 oral fluid samples were collected, with individual demographic and social data via a questionnaire, from persons under 45 years of age recruited through general practices. Instructions were accurately followed with only 1% of samples returned without risk-factor data. The overall response rate was 40%. Response was independently associated with age, sex and location. Response was highest in children aged 5-14 years, adult females and in rural locations. This approach allowed the successful collection of comprehensive individual risk data, but response rates in adults must be improved if oral fluid surveys are to routinely complement serological surveillance.

Validation of a modified commercial assay for the detection of rubella-specific IgG in oral fluid for use in population studies.

Matching serum and oral fluid (saliva) samples were collected from 369 subjects in Tunisia in 2002, from a city in the north and a rural district in the south. Rubella-specific IgG was detected in sera by commercial ELISA (Dade Behring) and in matching oral fluids by two methods, a previously described IgG-capture ELISA (GACELISA) [J. Clin. Microbiol. 37 (1999) 391] and the Dade Behring ELISA with the assay protocol modified for use with oral fluids. Total IgG concentration of oral fluids was also measured. Rubella-specific IgG was detected in 289 (78.3%) sera overall. Differences in the age distribution of the study population in the north and south led to a higher prevalence being found in the north (86.2%) than in the south (71.8%). This difference was reflected in the oral fluid rubella-specific IgG results. With GACELISA, rubella-specific IgG was detected in 79.4% of oral fluids from the north and 69.7% from the south and with the modified Dade Behring assay, in 81.4% of oral fluids from the north and in 64.9% from the south. The sensitivity and specificity of GACELISA in comparison to results from the matching sera were 92.4 and 93.2%, respectively. The sensitivity and specificity of the modified Dade Behring ELISA were 89.8 and 92.0%, respectively. Total IgG concentration in oral fluid showed a weak correlation (r=0.19) with the modified Dade Behring results and with samples where total IgG was >7.5 mg/l, the sensitivity and specificity were 94.4 and 90.0%, respectively. Twenty-nine oral fluids, which gave false negative rubella-specific IgG results with the modified Dade Behring ELISA, had a lower mean total IgG concentration than 256 oral fluids which gave concordant positive results (7.0mg/l versus 15.8 mg/l, P<0.001). The study validated the modified Dade Behring ELISA, providing an alternative to the GACELISA for assessing levels of rubella immunity for population studies using oral fluid samples.

Primary human herpesvirus-6 and -7 infections, often coinciding, misdiagnosed as measles in children from a tropical region of Brazil.

We investigated primary human herpesvirus-6 and -7 (HHV-6, HHV-7) infections as a cause of rashes incorrectly diagnosed as measles in Brazilian children. Sera from 124 patients, aged 4 months to 17 years, from the states of Rio de Janeiro and Espirito Santo, in whom measles, rubella and parvovirus B19 infections had been excluded, were studied using indirect immunofluorescence antibody avidity tests; 38 (31%) had evidence of primary HHV-6 and/or HHV-7 infections. Twenty four children had primary HHV-6 infection, either recent or coincident with the rash, and similarly 31 had primary HHV-7 infection. Remarkably, almost half (17) of primary infections were dual HHV-6 and HHV-7 infections with the majority, 12 (71%), in children less than 1 year old. HHV-7 infection occurred earlier than previously reported, perhaps due to socioeconomic and tropical conditions in this region of Brazil, and thus coincided with the HHV-6 infections. This study also highlights the difficulties of diagnosing a rash illness on clinical grounds alone.