Reactivation of 2 genetically distinct varicella-zoster viruses in the same individual.

Varicella-zoster viruses recovered from 2 episodes of herpes zoster in an immunocompetent man were found to be different genotypes. The fact that the 2 isolates came from the same individual was confirmed by DNA fingerprinting. Immunity following chickenpox may not always protect against systemic reinfection. This finding raises questions about varicella-zoster virus pathogenesis and may have an impact on public health policy.

Modified latex agglutination test for antibodies to Toxoplasma gondii in eluates from Guthrie cards.

AIMS: To determine whether the Eiken particle agglutination test could be modified to make it sufficiently sensitive to screen blood samples collected on Guthrie cards for the presence of antibodies to Toxoplasma gondii; to evaluate the specificity of the modified system; and to compare seroepidemiological data on the prevalence of T gondii in pregnant women. METHODS: Simulated dried blood spots were prepared from sera from pregnant women booking for antenatal care. Eluates from the simulated dried blood spot cards and sera were tested in parallel using the modified test (1 in 5 dilution of latex) and the standard assay (neat latex particles) and endpoints determined. Guthrie card eluates, from neonates in three Thames regions, were then tested using the modified test. RESULTS: The modified test produced a 4.21-fold increase in antibody titre in 85 sera when tested in parallel with the standard test. Eluates of 168/170 from simulated dried blood spots derived from seropositive patients gave a positive result in the modified test. The two eluates which gave a negative result were derived from patients with an equivocal titre of 1/16 in the standard serum test. Of the eluates derived from serum negative patients all 103 were negative at a dilution of 1 in 4 in the modified test. The seroprevalence of antibodies to T gondii in pregnancy was 21.8% using the standard test. A similar value of 20.5% was obtained when dried blood spots from neonates in a similar region of London were tested by the modified test. CONCLUSIONS: The modified Eiken Toxo-reagent test is sensitive, simple, and economic for screening large numbers of dried blood spots. The procedure could be easily semiautomated and the technique applied to the mass screening of neonatal blood samples collected on Guthrie cards to determine the seroprevalence of T gondii in pregnant women.

Use of dried blood spots for the detection and confirmation of HTLV-I specific antibodies for epidemiological purposes.

AIMS--To modify and evaluate a gelatin particle agglutination test that could provide a sensitive, specific and inexpensive method for the detection of HTLV-I antibody in dried blood spot samples (DBS) collected on filter paper. METHODS--A set of 26 reference samples confirmed as HTLV-I antibody positive were assembled from patients with tropical spastic paraparesis or adult T cell leukaemia and blood donors. Serum samples and simulated antibody positive dried blood spot eluates were tested using the Serodia assay together with two confirmatory tests: HTLV BLOT 2.3, a western blot, and Select-HTLV, an enzyme immunoassay (EIA). Both confirmatory tests use synthetic peptides to differentiate between antibodies to HTLV-I and -II. The modified Serodia assay was then used to test anonymously 10,135 DBS collected from neonates from London. Samples reactive in the modified Serodia test producing a positive result were titrated to an end point and confirmed as before. RESULTS--All 26 eluates made from simulated DBS derived from positive reference samples were identified as positive by the modified Serodia HTLV-I test and were confirmed as anti-HTLV-I positive by EIA. Two eluates derived from relatively low titre reference samples gave indeterminate results on western blotting. Screening of the 10,135 neonatal DBS resulted in six repeat reactives, five of which were confirmed. The remaining reactive sample gave an indeterminate result on western blotting and there was insufficient eluate for testing by EIA. The overall seroprevalence of HTLV-I in this population was 0.05% (five of 10,135). CONCLUSION--The modified Serodia HTLV-I assay provides a sensitive, specific and inexpensive (10 pence/test) method for screening large numbers of DBS. The format of the assay makes it ideally suited for simultaneous screening of antibodies to HIV-1, HIV-2 and HTLV-I using semi-automated equipment.

Impact of rotavirus infection on a paediatric hospital in the east end of London.

AIMS: To study the impact of confirmed rotavirus infection at a paediatric hospital; to use the data to obtain a minimum estimate of the cost of treating reported cases of rotavirus in England and Wales. METHODS: Data were obtained on all patients with rotavirus over a two year period. Information was collected on 386 patients with rotavirus infection who were treated at the 120 bed Queen Elizabeth Hospital for Children in East London. This included the virus serotype, the patient's age, whether they required intravenous infusion, duration of hospital stay, numbers of patients treated in the casualty department, and numbers who had to be admitted. Treatment costs were obtained from the Finance Department of the Hospitals for Sick Children. RESULTS: The minimum cost of treating patients, excluding the cost of medical staff at the hospital, was estimated to be 95,400 pounds a year. One hundred and forty eight (38%) patients were admitted to the wards and a further 49 patients developed symptoms while in hospital. Intravenous infusion was required by 18 patients. The mean duration of hospital stay was 5.5 days. One hundred and eighty nine (49%) patients were treated with oral rehydration solution in casualty, given advice, and sent home. Ninety four per cent of the patients were aged under 2 years. The findings were comparable with those obtained in a study at Texas Children's Hospital, USA. The G serotype (VP7) of rotavirus did not influence the severity of infection. CONCLUSION: Rotavirus infections accounted for a significant number of patients treated in casualty, admissions to hospital, and bed occupancy in a paediatric hospital. The estimated cost of treating reported cases of rotavirus in England and Wales is in excess of 6.3 pounds million a year.

A method for the detection and confirmation of antibodies to hepatitis C virus in dried blood spots.

This study describes the development and evaluation of a cost effective test rationale for the detection of anti-HCV in dried blood spots. Samples were screened using an 'in house' IgG ELISA that incorporated the recombinant proteins c22-3, c200 and NS5. Confirmation of specific antibody to HCV was by a modification of the immunoblot RIBA 3.0. An extensive panel of well evaluated anti-HCV positive and negative samples from the UK and South Africa were used to assess the sensitivity and specificity of the two tests. One third of the anti-HCV positive samples had been typed. All anti-HCV positive samples were detected by the 'in house' screening EIA. Test/negative optical density ratios showed that more than 95% of reactive samples produced values greater than 5.0. Antibodies to HCV could be detected in a wide range of samples derived from asymptomatic and symptomatic patients and of different genotypes, with similar sensitivity. The presence of anti-HCV could be confirmed by RIBA 3.0 in samples with low reactivity but not in anti-HCV negative samples. Furthermore the immunoblot assay successfully increased specificity by screening out false reactive EIA samples that might occur in an epidemiological survey of a multi-ethnic population.

PIP: While hepatitis C virus (HCV) is a major etiological agent of post-transfusion and community acquired non-A, non-B hepatitis, little is known about the epidemiology of HCV in the UK. A cost-effective method using dried blood spots to determine anti-HCV IgG in subjects which could be used in large-scale epidemiological studies is described. Samples were screened using an in-house IgG ELISA incorporating the recombinant proteins c22-3, c200, and NS5, while specific antibody to HCV was confirmed using a modified immunoblot RIBA 3.0. A panel of well evaluated anti-HCV positive and negative samples from the UK and South Africa were used to assess the sensitivity and specificity of the 2 tests. All anti-HCV positive samples were detected by the in-house screening EIA. Test/negative optical density ratios showed that more than 95% of reactive samples produced values greater than 5.0. Antibodies to HCV could be detected in a wide range of samples derived from asymptomatic and symptomatic patients and of different genotypes, with similar sensitivity. The presence of anti-HCV could be confirmed by RIBA 3.0 in samples with low reactivity, but not in anti-HCV negative samples. The immunoblot assay increased specificity by screening out false reactive EIA samples.

Subdural empyema in an HIV positive patient.

A subdural empyema with a Salmonella species as the likely causative organism is presented. We believe that this is the first reported case of such an infection in an HIV positive patient. The difficulties in treatment and diagnosis are discussed.

Effect of music training on monaural perception of pitch and rhythm.

A monaural study of music perception was conducted on 77 right-handed subjects from a university population. The musical ability of each subject was classified in two ways, according to years of training and by total score for three tests of musical achievement [melody (sequence and excerpt), harmony, and rhythm]. Analysis indicated that subjects with more years of training showed a right-car dominance for recognition of excerpts but those with high scores had no such dominance. Both groups with either low scores or no formal training had a right-ear dominance for recognition of sequences. Correlations of scores from each ear within subjects and between tests indicated that perception of pitch tended to be more accurate in the same car. For all classifications of subjects no ear dominance was found for harmony and rhythm tests. These results suggest that the measurement of hemispheric asymmetry of music perception is dependent upon the criteria chosen for classification of subjects, in this case, training and achievement.

Persistence of varicella-zoster virus viraemia in patients with herpes zoster.

BACKGROUND: Herpes zoster is caused by the reactivation of varicella-zoster virus from sensory neurons. The commonest complication following zoster is chronic pain termed post herpetic neuralgia. OBJECTIVES: To investigate the dynamics of VZV viraemia and viral load following the resolution of zoster and its relationship to PHN development. STUDY DESIGN: Blood samples were collected at baseline, 1 month, 3 months and 6 month from a prospective study of 63 patients with active zoster. Quantification of VZV DNA in whole blood was performed using a real-time PCR assay. RESULTS: During acute zoster, all patients had detectable VZV DNA in their blood. VZV DNA remained detectable in the blood of 91% of patients at 6 months although levels declined significantly (p<0.0001). A history of prodromal symptoms (p=0.005) and severity of pain at baseline (p=0.038) as well as taking antivirals (p=0.046) and being immunocompromised (p=0.043) were associated, with longer time to recovery from PHN. Viral DNA loads were consistently higher in patients with risk factors for PHN and higher viral DNA loads over time were associated with longer time to recovery (p=0.058 overall and 0.038 in immunocompetent). CONCLUSIONS: Based on these observations we hypothesise that VZV replication persists following acute shingles and that higher viral DNA loads contribute to the risk factors for PHN.

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.

Genotyping of varicella-zoster virus and the discrimination of Oka vaccine strains by TaqMan real-time PCR.

Single nucleotide polymorphisms (SNPs) in five genes have been used to identify four major subtypes of wild-type varicella-zoster virus (VZV) A, B, C, and J. Additional SNPs, located in the IE62 major transactivating gene can be used to differentiate the Oka vaccine strain (vOka) from wild-type VZV. Primer-probe sets for the detection of the five polymorphic loci were designed by Applied Biosystems for the ABI 7900HT platform. Probes for each allele were labeled with VIC or 6-carboxyfluorescein fluorogenic markers. Each primer-probe set was validated to establish assay sensitivity and specificity using VZV DNA of predetermined copy number and genotype. Further evaluation was carried out using DNA samples from the vesicle fluid or skin swab of the rash of adult patients with herpes zoster or rashes due to vOka. Assay sensitivity ranged from 10 and 10(8) copies/ml of VZV DNA (equivalent to 2 to 20 copies per reaction). Statistical analyses showed that for each genotype, a set of two probes clearly differentiated the nucleotide present (allele) at that locus (P < 0.0001). It was possible to determine the genotype of wild-type VZV using one of four SNP assays and also to differentiate wild type from vOka using a single SNP assay. The assay can be used for diagnostic and epidemiological studies of VZV, including the differentiation of vOka from wild-type strains, investigation of breakthrough infections, and varicella outbreaks following immunization.

Measurement of IgA responses following Norwalk virus infection and other human caliciviruses using a recombinant Norwalk virus protein EIA.

An enzyme immunoassay employing recombinant Norwalk virus capsid protein was evaluated for the measurement of IgA responses. Tests on 23 volunteers and patients known to have been infected with Norwalk virus (NV) showed that 19 developed significant IgA responses, 2 had unchanging levels of IgA and 2 failed to respond. There was no evidence of IgA responses to NV following infection with Hawaii or Snow Mountain-like viruses. Tests on sera from patients involved in outbreaks associated with eating contaminated shellfish suggest that some patients may have been infected with more than one strain of calicivirus. The use of the rNV EIA for measuring IgA and IgG responses in patients involved in a major outbreak of food poisoning affecting hospital staff indicated that the causative agent was probably NV.

Acuity of sound localisation: a topography of auditory space. III. Monaural hearing conditions.

A study is reported in which the acuity of azimuth and elevation discrimination under monaural listening conditions was measured. Six subjects localised a sound source (white noise through a speaker) which varied in position over a range of elevations (-40 degrees to +40 degrees) and azimuths (0 degrees to 180 degrees), at 10 degrees intervals, on the left side of the head. Monaural listening conditions were established by the fitting of an ear defender and one earmuff to the right ear. The absolute and algebraic, azimuth and elevation errors were measured for all subjects at each position of the source. The results indicate that all subjects suffered a marked reduction of azimuth acuity under monaural conditions, although a coarse capacity to discriminate azimuth still remained. Considerable between-subject variability was observed. Front/back discrimination was retained, although it was slightly impaired compared to that observed under normal listening conditions. Elevation discrimination was, on the whole, quite good under monaural conditions. However, a comparison of the performance of these subjects under monaural conditions with that observed under normal listening conditions indicated that some reduction in elevation localisation acuity occurred in the frontal quadrants in the median plane and in the upper quadrants of more lateral source positions. The reduction in acuity seen in these regions is attributed to the loss of information from the pinna of the occluded ear rather than to the observed reduction in azimuth error. The results provide partial support for the binaural pinna disparity model.

Acuity of sound localisation: a topography of auditory space. I. Normal hearing conditions.

Eight subjects were required to localise a sound source (white noise through a speaker) which varied in position on both sides of the head over a range of elevations (-40 degrees to +40 degrees) and azimuths (0 degree to 180 degrees) at 10 degrees intervals. The perceived position of the source was indicated by pointing a special gun. Depression of the trigger activated a photographic system which recorded two views of the subject, the sound source, and the gun. The absolute and algebraic, azimuth and elevation errors were measured for all subjects at each position of the source. The variability of azimuth and elevation error was also computed. In a second experiment, four of the same subjects performed the same task but in this case visually located the sources. This experiment provided an estimate of inherent motor error in the pointing task. No differences in localisation acuity between sides were found, but there were significant differences between front and back regions. Azimuth and elevation error were well matched and low in the front. However, azimuth error increased in the regions behind the head, particularly for azimuth positions 120 degrees to 160 degrees. Larger increases were found for positions in the upper elevations of this region. Elevation error also increased in the upper elevations behind the head. A comparison of the auditory and visual data indicates that this pattern of error is not due to motor factors. The results are discussed in relation to the structural characteristics of the pinnae and modifications that they impose on incoming sound energy.

Acuity of sound localisation: a topography of auditory space. II. Pinna cues absent.

The acuity of azimuth and elevation discrimination was measured under conditions in which the cues to localisation provided by the pinnae were removed. Four subjects localised a sound source (white noise through a speaker) which varied in position over a range of elevations (-40 degrees to +40 degrees) and azimuths (0 degree to 180 degrees), at 10 degrees intervals, on the left side of the head. Pinna cues were removed by the insertion of individually cast moulds in both pinnae. Each mould had an access hole to the auditory canal. The absolute and algebraic, azimuth and elevation errors were measured for all subjects at each position of the source. The variability of azimuth and elevation error was also computed. The performance of the subjects was compared to their performance under normal hearing conditions. Insertion of the pinnae moulds was found to increase substantially elevation error and the number of front/back reversals. The importance of the cues provided by the pinnae in these discriminations was thus confirmed. However, the increase in elevation error did not result in a corresponding increase in azimuth error. These findings provide support for the proposition that azimuth and elevation discrimination are coded independently.

Enzyme immunoassay using baculovirus-expressed human calicivirus (Mexico) for the measurement of IgG responses and determining its seroprevalence in London, UK.

The use of an enzyme immunoassay (EIA) employing a baculovirus-expressed recombinant human calicivirus (Mexico virus, MxV) for the detection of IgG-specific antibodies is described. MxV appeared to be related antigenically to a strain of small round structured virus, SRSV/UK4/Leeds/91, which had previously been shown by solid phase immune electron microscopy (SPIEM) to be related to Snow Mountain agent (SMA). One other outbreak which occurred in San Anita, USA in 1980 and was due to consumption of contaminated water was caused by a virus antigenically related to MxV. Volunteers and patients who developed significant IgG responses to rMxV showed anamnestic IgG responses (2 to 4-fold) in the recombinant Norwalk virus (rNV) IgG assay. Patients and volunteers who were known to have been infected with several other strains of calicivirus/small round spherical viruses (SRSV) including NV and SRSV UK3 showed no significant antibody response to rMxV in the EIA. A seroepidemiological survey of sera from 338 children in London showed that infection with MxV occurred earlier in life than NV. Primary infections with MxV were common after the age of 6 months. Over 70% of children had evidence of infection by the ages of 2 years, whereas only 12% of these children had been infected with NV. High concentrations of maternal antibody were present during the first month of life which was detected in 96% of the neonates. The results suggest that the high sensitivity of the EIA may be detecting maternal antibody throughout the first 8 months of life.

Detection of antibodies to hepatitis C virus in dried blood spot samples from mothers and their offspring in Lahore, Pakistan.

Dried blood spot samples from mothers and their offspring attending the obstetric and pediatric departments of two hospitals in Lahore, Pakistan, were tested for antibodies to hepatitis C virus (HCV). The seroprevalence of HCV in the women was 6.7% (95% confidence interval [CI], 4.3 to 9.1), and that in the children was 1.3% (95% CI, 0.34 to 2.26). Four anti-HCV immunoglobulin G (IgG)-positive children had mothers that were anti-HCV IgG negative, which suggested that their infection was community acquired.

Measurement of human nasal ventilation using an oxygen cannula as a pitot tube.

A pressure transducer connected to a nasal oxygen cannula detects fluctuations of about 0.1 kPa (1 cm H2O) during nose breathing because the prongs act as two one-sided pitot tubes in parallel. We describe a digital technique for converting this signal to flow and volume records without encumbering the head except during calibration. The signal could be calibrated or validated using flow measured with a face mask by intubation of the nostrils or using a respiratory inductive plethysmograph. In many subjects, the calibration gave satisfactory results for as long as 30 min, but in some, more frequent checks were needed. Changes in the relative airflow in the two nostrils may upset the results, but this can be corrected by recording and calibrating the pressure from each side separately. The meaurements can be made at high ventilation rates of when oxygen is flowing along the cannula or when gas is being sucked out for analysis.

Psychomotor, pulmonary and exercise responses to sleep medication.

1 Athletes may use benzodiazepines before events to improve sleep, but these drugs may adversely affect performance. 2 Nitrazepam (10 mg) and temazepam (30 mg) were compared with placebo in 27 physical education students, (14 males, 13 females). Treatments were administered at night, using a double-blind, double dummy protocol, for 9 nights. Observations were made in the morning after night 2 and night 9. At least 2 weeks interval was allowed between each treatment. 3 At each examination lung mechanics were measured, a Leeds Sleep Evaluation Questionnaire completed, recognition reaction time, choice reaction time and the critical flicker fusion threshold test were used to assess psychomotor activity and an exercise test was performed. The subject exercised to exhaustion on a bicycle ergometer while ventilation, gas exchange and heart rate were recorded on an FM tape unit for off-line digital analysis. 4 The questionnaire indicated that both drugs were equally effective in promoting and maintaining sleep, but nitrazepam had a marked 'hangover' effect. The psychomotor activity and lung mechanics however seemed unaffected. On day 2, maximum exercise levels attained using either drug were comparable to placebo whilst on day 9 temazepam and placebo were significantly higher than nitrazepam. 5 Heart rate was significantly increased at each exercise level with both drugs. 6 Although there may be some effect of these drugs on athletic performance this is likely to be small especially with temazepam.