Detection of IgG antibody to Epstein-Barr virus viral capsid antigen in saliva by antibody capture radioimmunoassay.

A 'G' antibody capture radioimmunoassay (GACRIA) to detect IgG to Epstein-Barr virus (EBV) viral capsid antigen (VCA) in saliva is described. The monoclonal antibody to EBV VCA used in the GACRIA bound non-specifically when testing saliva samples having a total IgG content of less than 2.0 mg/l, so giving false positive results. This problem was overcome by including 0.5% EBV-negative human serum in the monoclonal antibody diluent. The performance of the assay was then evaluated by comparing the GACRIA test on serum and saliva samples to indirect immunofluorescence assay (IFA) results on sera using a panel of paired serum/saliva samples. Compared to the corresponding serum IFA the saliva GACRIA had a sensitivity and specificity of 93.5 and 100%, respectively. Although less sensitive than IFA on serum samples, the saliva GACRIA is sufficiently sensitive to be used for epidemiological screening and will enable testing for anti-EBV VCA to be carried out easily and on a wide scale.

Detection of measles specific IgG in oral fluid using an FITC/anti-FITC IgG capture enzyme linked immunosorbent assay (GACELISA).

An IgG antibody capture enzyme linked immunosorbent assay (GACELISA) for the detection of measles specific IgG in oral fluid was developed using an FITC/anti-FITC amplification system. The GACELISA was evaluated by testing paired oral fluid and serum samples from 787 subjects in an epidemiological study of measles in rural Ethiopia. Oral fluids were tested by GACELISA and corresponding serum samples by a sensitive indirect ELISA for measles IgG (Behring Enzygnost). By comparison with the serum measles IgG assay, the oral fluid GACELISA had a sensitivity of 97.4% (95% confidence intervals: 95.9, 98.2) and a specificity of 90.0% (81.9, 94.3), with no significant differences observed by age group. Total IgG concentrations were measured on a subset of 160 oral fluids by an in-house ELISA. This showed that false negative GACELISA results tended to occur in samples with low concentrations of total IgG, although the trend was not statistically significant. It is concluded that the overall performance of the GACELISA was satisfactory, showing close agreement to the serum ELISA, and has potential to serve as an easily transferable tool for large scale epidemiological studies as required for the World Health Organisation's programme for the global control of measles.

The burden of infection with cytomegalovirus in England and Wales: how many women are infected in pregnancy?

A serological survey was used to investigate the epidemiology of cytomegalovirus (CMV) infection in England and Wales. A total of 5237 sera representing the complete age range were used reflecting the general population. The sera were collected in 1991 and 2002, and screened for CMV-specific IgG by ELISA. Antibody prevalence increased with age from approximately 15% in those aged 1-4 years to approximately 80% in those aged > or = 65 years with no association with gender or region. Analysing by common birth cohort demonstrated that between 1991 and 2002 incidence was highest in children born 1985-1989 (1.62% per year, 95% CI 0.86-2.35), lower in older children and younger adults born 1950-1984 (0.75% per year, 95% CI 0.29-1.19) with little evidence of infection in older adults born pre-1950 (0% per year, 95% CI 0-0.64). Application to population and live-birth estimates for England and Wales suggested that between 1991 and 2002, 159 996 (95% CI 67922-278277) CMV infections occurred annually with an annual average of 2133 (95% CI 816-3435) infections affecting pregnant females.

Estimating hepatitis C infection acquired in England, 1986-2000.

Hepatitis C is a global health problem and in the UK seroprevalence studies have mainly concentrated on specific high-risk groups. The aim of this study was to determine changes in the prevalence of antibody to hepatitis C virus in England using residual specimens collected between 1986 and 2000 reflecting the general population. A cross-sectional study design using a convenience collection of serum specimens from adult patients submitted to laboratories in the years 1986, 1991, 1996 and 2000 from a total of 19 laboratories around England were investigated. The main outcome was to determine anti-HCV prevalence and the average incidence occurring between 1986 and 2000 and factors associated with infection. Multivariable analysis of results from all years showed there was a significant difference in prevalence between males and females (P < 0.001), birth cohort (P < 0.001) and by health region (P < 0.001). An average of 0.72% (95% CI 0-1.65%) of those susceptible to HCV born between 1950 and 1970 were estimated to have acquired the infection between 1986 and 2000. Analysis of this convenience serum collection suggests that HCV prevalence is low in the general population, and is associated with period of birth, gender and health region. There was evidence to support a low incidence of HCV infection in those born between 1950 and 1970 over the period 1986-2000 which, at the population level, equated to a substantial burden of infection (approximately 106,000 persons). Continued surveillance and prevention targeted at injecting drug users are essential for the control of hepatitis C in the UK.

The burden of parvovirus B19 infection in women of childbearing age in England and Wales.

A serological survey has been used to investigate the epidemiology of parvovirus B19 infection in England and Wales. A total of 2835 sera representing the complete age range were selected from a convenience collection obtained in 1996 that reflects the general population and screened for parvovirus B19-specific IgG. Antibody prevalence rose nonlinearly with age from 21% in those aged 1-4 years to >75% in adults aged > or = 45 years. Force-of-infection estimates were similar to those previously made in 1991, being highest in those aged <15 years. There was no association between evidence of previous infection and sex or region. Quantitatively strongest antibody responses were found in those aged 15-34 years and IgG levels in females were 28.5% higher than those found in males (P=0.004, 95% CI 8.2-52.6). Applying the upper 95% confidence interval for the force of infection to maternity estimates for England and Wales in 1996, parvovirus infection in pregnancy was estimated to occur on average in up to 1 in every 512 pregnancies each year. This represents 1257 maternal infections, causing up to an estimated 59 fetal deaths and 11 cases of hydrops fetalis annually. An analysis of all available laboratory-confirmed parvovirus infections found a mean of 944 infections per year in women aged 15-44 years highlighting a need for enhanced surveillance of maternal parvovirus B19 infection in England and Wales, including information on both pregnancy and outcome of pregnancy.

Interpreting serological surveys using mixture models: the seroepidemiology of measles, mumps and rubella in England and Wales at the beginning of the 21st century.

A mixture modelling technique is applied to age-specific frequency distributions of quantitative results from serological surveys for measles, mumps and rubella using samples collected across the age range in England and Wales in 2000. In accordance with previous studies the analysis suggests that the antibody response to natural infection is stronger than that produced by vaccination, that vaccine-induced antibody levels wane with time and that levels of vaccine-induced antibody response vary for each virus infection being strongest for rubella and weakest for mumps. The current mumps epidemic in the United Kingdom is focused in cohorts born during 1982-1987 who were too old to have received routine MMR vaccination. In the cohort born in 1981-1985 the model estimates that 7.5% have no evidence of mumps specific IgG and 24.9% have the lowest level of detectable antibody. The similar proportions of mumps antibody in these categories among cohorts with opportunity for 1 or 2 doses of vaccine is a concern, as the degree to which these individuals are protected is unclear. Investigations into the efficacy of two doses of a mumps containing vaccine should be a priority during the current epidemic.

An RT-PCR assay using oral fluid samples to detect rubella virus genome for epidemiological surveillance.

A reverse transcription nested polymerase chain reaction (RT-PCR) method was developed for detecting rubella virus (RV) RNA using primer pairs which targeted a variable region of the E1 gene. RV genome was detected in oral fluid, throat swabs, serum and tissue samples. This is the first report to show that RV genome can be detected in oral fluid samples, including acute cases < or = 2 days after onset of symptoms, which have previously only been used for antibody testing. This suggests that PCR is useful for assisting with early diagnosis when a sufficient IgM response may not have been mounted. The PCR amplicon of 553 nucleotides was also useful for molecular genotyping, which contributes to RV epidemiological surveillance.

A comparison of oral fluid collection devices for use in the surveillance of virus diseases in children.

Three oral fluid collection devices (OraSure, Omni-SAL and Oracol) were compared in terms of the quality of oral fluid collected by each device for antibody testing and their acceptability to participants. Participants (143 children aged 3.5-5 y from North Hertfordshire, UK, who had recently received DTaP and MMR vaccination) were randomised to use one of the three types of collection device. Oral fluid was collected by a parent who completed a short questionaire recording information on ease of use and willingness to use the device again. A matching serum sample was collected by a nurse. Oral fluid samples were screened for total IgG and IgM by ELISA and for rubella specific IgG and parvovirus specific IgG by radioimmunoassay. Serum samples were screened for rubella specific IgG and parvovirus B19 specific IgG by ELISA. 87.4% (125) of participants provided a matching oral fluid and serum sample. Of these, 100% (125) and 10.4% (13) had serum IgG specific for rubella and parvovirus B19, respectively. The Oracol device provided oral fluid samples with the highest geometric mean titres of total IgG and IgM and with rubella specific IgG results which correlated most closely with those of matching sera. A higher proportion of parents found the Oracol and OraSure devices easier to use than the Omni-SAL (P<0.001) and the proportion who would not take another test was higher for the Omni-SAL than for the Oracol or Orasure. Oral fluid samples collected by each of the devices gave qualitative results acceptable for surveillance and epidemiological studies of rubella and parvovirus B19. The highest quality oral fluid sample for antibody testing in terms of total IgG and IgM concentration and rubella specific IgG concentration was collected by the Oracol. The acceptability to participants of both the Oracol and OraSure was high. As the cheapest device available, the Oracol is the preferred oral fluid collection device for studies involving children in the UK.

Improving sensitivity of oral fluid testing in IgG prevalence studies: application of mixture models to a rubella antibody survey.

A method for the analysis of age-stratified antibody prevalence surveys is applied to a previously reported survey of antibody to rubella virus using oral fluid samples in which the sensitivity of the assay used was shown to be compromised. The age-specific distribution of the quantitative results of antibody tests using oral fluids is modelled as a mixture of strong positive, weak positive and negative components. This yields maximum likelihood estimates of the prevalence at each age and demonstrates that, when used in conjunction with mixture modelling techniques, the results of antibody prevalence studies using oral fluids accurately reflect those obtained using sera.

Seroprevalence of antibody to varicella zoster virus in England and Wales in children and young adults.

This is the first large-scale study to investigate the seroprevalence of varicella zoster (VZV) in the general population of England and Wales. The study focused on those aged 1-20 years, that age group in whom most infections occur. Prevalence rose rapidly with age, with 53% of children showing evidence of prior infection by the age of 5 years and most young adults having experienced infection. In addition to using a fixed cut-off recommended by the manufacturer, a mixture modelling technique was also used to define the proportion of the population seropositive in each age group. This was shown to be a more accurate approach to categorizing data from an epidemiological perspective.

Detection of rubella virus-specific immunoglobulin G in saliva by an amplification-based enzyme-linked immunosorbent assay using monoclonal antibody to fluorescein isothiocyanate.

An immunoglobulin G (IgG)-capture enzyme-linked immunosorbent assay (ELISA) for rubella virus is described. The assay uses a fluorescein isothiocyanate (FITC)-anti-FITC amplification system. The detection limit of the ELISA was approximately 7 IU of rubella virus-specific IgG per ml of serum sample. For saliva samples the performances of the capture ELISA and previously described radioimmunoassay were assessed, and the results of those two assays were compared to the rubella virus-specific IgG result obtained by a commercial ELISA (Behring Enzygnost) with a panel of paired serum and saliva samples. This comparison showed that the capture ELISA with saliva was more sensitive than the radioimmunoassay and that the results correlated better with the serum IgG result than the results of the radioimmunoassay did, with an overall sensitivity of 82% and a rank correlation of 0.68, whereas the sensitivity and rank correlation for the radioimmunoassay were 74% and 0.45, respectively. For subjects of 10 years of age or younger, the ELISA with saliva had a sensitivity of 94% and a specificity of 100% compared to the results of the ELISA (Behring Enzygnost) for rubella virus-specific IgG with corresponding serum samples. The sensitivity was much lower for subjects ages 17 years or older. The assay may have wider epidemiological use with saliva specimens, particularly those from children.

Susceptibility of young offenders to measles and rubella: an antibody prevalence study using oral fluid samples.

Three hundred and two oral fluid samples collected in January 2001 from male young offenders aged 18 to 21 years at HMP Hindley were screened for measles and rubella specific IgG, using 'in-house' amplified ELISA assays. MMR (measles/mumps/rubella) vaccine, offered at the same time, was accepted by 68.1% of the prisoners (92.7% of those agreeing to oral fluid antibody testing). Antibody prevalences were 92.1% for rubella and 80.8% for measles. This may, however, underestimate true prevalence as the sensitivity of the oral fluid assays used may be relatively poor in adults. Susceptibility levels are theoretically high enough to result in outbreaks should either infection enter this closed institutional environment. Since a measles outbreak would carry significant morbidity and rubella would represent a risk to associated non-immune women of childbearing age, it is suggested that prison health services should offer MMR to young offenders on entry.

The burden of infection with HSV-1 and HSV-2 in England and Wales: implications for the changing epidemiology of genital herpes.

OBJECTIVE: To measure the burden of infection with herpes simplex type 1 (HSV-1) and herpes simplex type 2 (HSV-2) in the general population of England and Wales and to assess temporal changes in the incidence of HSV-1 infection in childhood. METHODS: 4930 residual blood samples taken from people aged 0-69 years and submitted to 15 public health laboratories in England and Wales between January 1994 and June 1995, and 500 samples taken from people aged 10-14 years between November 1986 and December 1987, were screened for IgG antibody to HSV-1 and HSV-2 using type specific ELISA assays. RESULTS: The prevalence of antibody to HSV-1 in 10-14 year olds declined from 34% in samples collected in 1986-7 to 24% in samples collected in 1994-5 (p < 0.001). HSV-1 antibody prevalence in adults increased with age and was higher in females than males, reaching 54% in females aged 25-30 years in 1994-5. In samples collected in 1994-5 from people aged 16-69 years HSV-2 antibody was detected in sera from 3.3% of men and 5.1% of women. CONCLUSIONS: The incidence of HSV-1 infection in childhood is falling in England and Wales. The prevalence of HSV-2 infection in the general population is low, with the rate of infection significantly lower than that described for the general population in the United States and developing countries. The falling rate of HSV-1 infection in childhood may be one factor contributing to the increasing incidence of genital HSV-1 infection.

Has oral fluid the potential to replace serum for the evaluation of population immunity levels? A study of measles, rubella and hepatitis B in rural Ethiopia.

OBJECTIVE: To assess the suitability of using oral-fluid samples for determining the prevalence of immunity to vaccine-preventable infections. METHODS: Paired blood and oral-fluid samples were obtained from 853 individuals of all ages from a rural Ethiopian community. Oral fluid around the gums was screened for measles- and rubella-specific antibodies using enhanced IgG antibody capture (GAC) enzyme-linked immunosorbent assays (ELISAs), and for anti-HBc antibodies using a prototype GACELISA. IgG antibodies in serum to measles, rubella and HBc were determined using commercial ELISAs. FINDINGS: Relative to serum, oral fluid assay sensitivity and specificity were as follows: 98% and 87% for measles, 79% and 90% for rubella, and 43% and 87% for anti-HBc. These assay characteristics yielded population prevalence estimates from oral fluid with a precision equal to that of serum for measles (all ages) and rubella (ages < 20 years). CONCLUSION: Our results suggest that oral fluid could have the potential to replace serum in IgG antibody prevalence surveys. Further progress requires assessment of variation in assay performance between populations as well as the availability of standardized, easy to use assays.

The burden of Helicobacter pylori infection in England and Wales.

The prevalence of active infection with Helicobacter pylori in the general population of England and Wales was estimated using high reactivity for specific IgG in serum ELISA as a marker. A total of 10,118 anonymized residues of serum samples collected in 1986 and 1996 from persons aged 1-84 years were used. Estimated prevalence of active infection varied by region and was highest in London. Prevalence was related to decade of birth and increased from 4-3% in those born during the 1980s to 30% in those born before 1940. An estimated total of 7.5 million people living in England and Wales have an active infection and analysis by decade of birth showed no significant difference between samples collected in 1986 and 1996. These data suggest H. pylori infection is becoming less common, is acquired at an early age and is unlikely to be resolved unless suitable antimicrobial treatment is sought.