Use of RT-PCR on oral fluid samples to assist the identification of measles cases during an outbreak.

This study investigated the occurrence of mild modified measles cases during an outbreak in Niterói, RJ, Brazil by using RT-PCR on oral fluid samples. From August to December 1997 a total of 76 patients with rash were seen at the study sites. Confirmed diagnosis by serology was achieved in 47 cases: measles (39.5%), rubella (13.2%), HHV-6 (3.9%), human parvovirus B19 (3.9%), dengue fever (3%). For 19 of the 29 patients without a conclusive diagnosis paired serum and saliva samples were available for further tests. In four of them, measles virus RNA was detected by RT-PCR in saliva samples in the absence of specific IgM in serum samples. Vaccination histories obtained from three of the RT-PCR positive cases showed that individuals previously immunized can still be infected and contribute to the circulation of measles virus. This study demonstrated the usefulness of RT-PCR on non-invasive clinical samples for the investigation of measles cases.

Seroprevalence of parvovirus B19 in urban Chilean children and young adults, 1990 and 1996.

An immunofluorescence test for detecting parvovirus B19 IgG was developed by infecting insect cells with recombinant baculovirus expressing the capsid protein VPI. The test was used to study the prevalence of antibodies in 725 healthy children and young adults living in Santiago, Chile. In total, 248 sera were taken in 1990 and 477 in 1996. The seroprevalence was low in children less than 5 years old (3% in 1990 and 21% in 1996). It rose during school age to a prevalence around 50%, reaching 60% in young adults. No differences were found between genders. There was a statistically significant higher seroprevalence in the low socioeconomic status group in 1990 samples, but this was not observed in 1996. The higher prevalence observed in children less than 5 years of age in 1996 compared with 1990 could be explained by the occurrence of intervening epidemics of parvovirus B19 infection.

Human parvovirus B19 infection in HIV-positive patients.

Parvovirus B19 infects predominantly erythroid cells, leading to transient inhibition of erythropoiesis. Immunocompromised patients may be unable to produce neutralizing antibodies and may develop severe chronic anemia. Epidemiological studies done on Niterói population showed that B19 infection occurs periodically in late spring and summer. We report a study from 55 HIV infected patients attending an infectious diseases outpatient clinic in this city during a 5-month period in which B19 circulation was well documented. All patients were under anti-retroviral therapy. No anti-B19 IgM was found, but a high prevalence of IgG anti-B19 (91%) was observed. In six patients, B19 DNA was found by dot-blot hybridization techniques, but this was not confirmed by PCR. None of these 6 patients manifested anemia and only one had CD4 cell count below 200 x 10(7)/L. We conclude that persistent infection causing anemia is an infrequent finding in our HIV positive patients under drug therapy.

Human parvovirus B19 infection in HIV-positive patients

Parvovirus B19 infects predominantly erythroid cells, leading to transient inhibition of erythropoiesis. Immunocompromised patients may be unable to produce neutralizing antibodies and may develop severe chronic anemia. Epidemiological studies done on Niterói population showed that B19 infection occurs periodically in late spring and summer. We report a study from 55 HIV infected patients attending an infectious diseases outpatient clinic in this city during a 5-month period in which B19 circulation was well documented. All patients were under anti-retroviral therapy. No anti-B19 IgM was found, but a high prevalence of IgG anti-B19 (91%) was observed. In six patients, B19 DNA was found by dot-blot hybridization techniques, but this was not confirmed by PCR. None of these 6 patients manifested anemia and only one had CD4 cell count below 200 x 10(7)/L. We conclude that persistent infection causing anemia is an infrequent finding in our HIV positive patients under drug therapy.

O parvovírus B19 infecta predominantemente células eritróides, causando inibição transitória da eritropoiese. Pacientes imunocomprometidos podem ser incapazes de produzir anticorpos neutralizantes, evoluindo com grave anemia crônica. Estudos epidemiológicos da população de Niterói mostraram que a infecção ocorre periodicamente no final da primavera e no verão. Descrevem-se 55 pacientes infectados pelo HIV atendidos num ambulatório de doenças infecciosas nesta cidade num período de cinco meses, no qual a circulação do parvovírus B19 foi documentada. Todos os pacientes estavam sob terapia anti-retroviral. Não se encontrou IgM anti-B19, mas notou-se uma prevalência alta de IgG anti-B19 (91%). Em seis pacientes verificou-se a presença de DNA do B19 por hibridização em dot-blot, o que não se confirmou por PCR. Nenhum destes seis pacientes tinha anemia, e apenas um tinha células CD4 abaixo de 200 x 107/L. Conclui-se que infecção persistente causando anemia é um achado infreqüente em nossos pacientes HIV positivos sob terapia medicamentosa.

Diagnosis of rubella infection by detecting specific immunoglobulin M antibodies in saliva samples: a clinic-based study in Niterói, RJ, Brazil.

This study was designed to investigate whether saliva could be a feasible alternative to serum for the diagnosis of recent rubella infection in a clinic setting. Forty-five paired blood and saliva samples collected 1 to 29 days after onset of illness were tested for specific immunoglobulin (Ig) M by antibody-capture radioimmunoassay (MACRIA). Rubella IgM was detected in all serum samples and in 38 (84.4%) saliva specimens. Forty-six serum and saliva samples from other patients with rash diseases were tested by MACRIA for control purposes and two saliva specimens were reactive. The saliva test had specificity of 96%. These results indicate that salivary IgM detection may be a convenient non-invasive alternative to serum for investigation of recent rubella cases, especially for disease surveillance and control programmes.

Atypical measles in a patient twice vaccinated against measles: transmission from an unvaccinated household contact.

Described are two cases within the same household that were involved in an outbreak of measles in Niterói, RJ. Measles diagnosis was confirmed serologically by specific IgM detection in Case 1 (classic measles) who was unvaccinated, and rising measles specific IgG in the absence of IgM in Case 2 (mild modified measles) who had a history of two vaccinations with measles-containing vaccines. Measles virus was detected by reverse transcriptase polymerase chain reaction (RT-PCR) in saliva samples from both cases. The nucleic acid amplified by RT-PCR was sequenced and showed identical measles sequence in the two cases. This study highlights the difficulty of diagnosing nonclassical measles infection on clinical and serological grounds, and the usefulness of PCR for viral RNA sequencing from noninvasive specimens for confirming epidemiologic links.

Salivary diagnosis of measles for surveillance: a clinic-based study in Niterói, state of Rio de Janeiro, Brazil.

This study was designed to investigate whether saliva could be a feasible alternative to serum for the diagnosis of recent measles infection in a clinic setting. Forty-two paired blood and saliva samples collected 1 to 16 days after onset of illness from 29 patients with clinical measles were tested for specific immunoglobulin (Ig) M by antibody-capture radioimmunoassay. Measles IgM was detected in all serum samples and in 39 (92.9%) saliva specimens. Between 1 and 3 weeks after illness onset, virus-specific IgM was detected in 100% of saliva samples. Measles IgM was also detected in 17 saliva specimens, not paired with blood samples, collected from study patients 5 days to 3 weeks after onset. Our results indicate that salivary IgM detection is a suitable non-invasive method for investigation of notified cases under conditions of routine clinic use.

Human parvovirus B19 infection and hydrops fetalis in Rio de Janeiro, Brazil.

Formalin-fixed paraffin embedded lung and liver tissue from 23 cases of non immune hydrops fetalis and five control cases, in which hydrops were due to syphilis (3) and genetic causes (2), were examined for the presence of human parvovirus B19 by DNA hybridisation. Using in situ hybridisation with a biotynilated probe one positive case was detected. Using 32P-labelled probes in a dot blot assay format, five further positives were obtained. These were all confirmed as positive by a nested polymerase chain reaction assay. Electron microscopy revealed virus in all these five positive cases. The six B19 DNA positive cases of hydrops fetalis were from 1974, 1980, 1982, 1987 and 1988, four of which occurred during the second half of the year, confirming the seasonality of the disease.

Diagnosis of parvovirus B19 infection by detection of specific immunoglobulin M antibody in saliva.

Serum and saliva samples were simultaneously collected from patients with B19 infection. Specimens were collected in a period of 1 to 18 days after the onset of symptoms. Saliva samples were collected with a commercial device, OraSure. The quality of these samples was evaluated by determining the concentration of total immunoglobulin G (IgG) by an enzyme immunoassay. The concentration of IgG in these samples ranged from 4.8 to > 250 mg/liter. B19 infection was confirmed for 20 patients by testing sera in a 1: 100 dilution by an IgM capture enzyme immunoassay (MACEIA) and an IgM capture hemadherence test (MACHAT). Saliva samples from these IgM-positive patients were tested neat by MACEIA and MACHAT. IgM could be detected in 11 of 20 (55%) samples by MACEIA and in 15 of 18 (83%) samples by MACHAT. Serum and saliva samples from a further 17 patients with rash were also tested. All of these specimens were unreactive by both assays. These results show that saliva may be a convenient alternative to serum for the diagnosis of recent B19 infection.

Salivary antibody detection in epidemiological surveys: a pilot study after a mass vaccination campaign against rubella in São Paulo, Brazil.

The sensitivity and specificity of salivary rubella antibody detection was investigated using samples collected from 301 children after a mass vaccination campaign in the state of São Paulo, Brazil. Saliva samples were collected by 2 different methods: directly dribbling into a container or using a commercial collecting device. Corresponding finger-prick blood samples were collected on filter paper. Rubella specific immunoglobulin G (IgG) was measured in saliva by antibody capture radioimmunoassay and in blood samples by indirect enzyme-linked immunosorbent assay. The detection of salivary rubella specific IgG showed good correlation with the detection of rubella antibody in the blood samples. For both collecting techniques the predictive value for a positive saliva test was > 99% compared with the results from the blood tests. However, the predictive value for a negative saliva test was only 58.3% for a dribbled sample, compared to 100% for saliva collected using the commercial device. Moreover, collecting saliva by dribbling from children less than 4 years old was difficult. The detection of rubella specific IgG in saliva collected using a commercial device proved to be sensitive and specific in this epidemiological study, encouraging its more widespread application as a means of surveillance after mass vaccination.

IgM antibody capture haemadherence test (MACHAT) for the detection of measles specific IgM.

An antibody capture haemadherence test (MACHAT) for detecting measles-specific IgM is described. The assay is based on the antibody capture principle with rhesus monkey erythrocytes as detector system in place of labelled antisera. MACHAT was compared with a commercial indirect enzyme immunoassay (EIA) for measles-specific IgM using 382 sera from patients notified as measles. There was good agreement between the two tests; 106 sera were found to contain measles IgM by both tests, 7 further sera were positive only in the commercial EIA and 9 only in MACHAT. One sera gave an equivocal result in MACHAT and another in the commercial EIA. Twelve of the 18 sera with discrepant results were also tested by MACRIA; in 7 MACRIA gave the same results as MACHAT, in 3 the MACRIA results agreed with the commercial test and in 2 the MACRIA results were equivocal. Specificity was established by a lack of MACHAT reactivity in sera collected from blood donors (n = 83) and from cases of recent rubella, dengue and parvovirus B19 infection (n = 51). The MACHAT is a simple, cheap test that can be read by eye and is suitable for measles surveillance programmes in the developing world.

Application to immunoglobulin M capture hemadherence assays of hemagglutination of monkey erythrocytes by native and recombinant human parvovirus B19 antigens.

Human parvovirus B19 recently was shown to agglutinate baboon and human erythrocytes. We have now demonstrated that both recombinant and native B19 antigens agglutinate rhesus, cynomolgus, and Saimiri monkey erythrocytes. Using cynomolgus erythrocytes and the recombinant antigen, we developed an immunoglobulin M (IgM) antibody capture hemadherence test (MACHAT) for the detection of specific B19 IgM antibodies in human sera. The results obtained with MACHAT were compared with those obtained with an IgM capture enzyme immunoassay (MACEIA) employing the native antigen routinely used in our laboratory. For 229 patient serum samples, we found 96% agreement between the results of the two assays. There was some evidence that MACHAT was slightly more sensitive than MACEIA. Our results add to the range of erythrocytes that can be agglutinated by B19 virus and show that native as well as recombinant antigens may be used in MACHAT.

The prevalence of antibody to human parvovirus B19 in Rio de Janeiro, Brazil.

During 1985 and 1986 serum samples were collected from the Rio de Janeiro population and examined for the presence of IgG antibody to human parvovirus B19. No difference in prevalence was found between males and females. Antibody prevalence rose from 35% in children less than five years old to almost 80% in children aged eleven to fifteen years. The antibody prevalence in individuals over 50 years old was over 90%.

Human parvovirus (B19) and erythema infectiosum.

An outbreak of erythema infectiosum ("fifth disease") was studied in Fukuoka, Japan, in 1980-1981. Human parvovirus (HPV) antigen was not detected in any patients, but anti-HPV, measured by countercurrent immunoelectrophoresis, was found in 33 of 34 affected children and in 21 (15%) of 141 children of the same ages without the disease. Immunoglobulin M class anti-HPV was present in all 25 children with erythema infectiosum tested. In a survey of hospital patients, the prevalence of anti-HPV detected by CIE was 12% in the cohort 5 to 9 years of age, 19% in the cohort 10 to 14 years, and 32 to 55% in the cohorts greater than or equal to 30 years. The antibody reactions in the cases of erythema infectiosum, which were already well established at the onset of disease, indicate that HPV was the cause of the outbreak.