Development of an enzyme-linked immunosorbent assay for diagnosis of human herpesvirus-7 infection.

INTRODUCTION: Human herpesvirus (HHV)-7 establishes a latent infection during the lifetime of the host and can reactivate after the primary infection, leading to lytic replication in immunosuppressed patients. METHODS: This study aimed to develop an enzyme-linked immunosorbent assay (ELISA) to identify HHV-7 serum antibodies and compare its performance with that of an indirect immunofluorescence assay (IFA). RESULTS: Serum samples (n=102) were tested by IgG-IFA and by ELISA. IFA and ELISA showed IgG-positive results in 77 and 73 samples, respectively. Qualitative concordance of 96% was demonstrated between the two techniques. CONCLUSIONS: ELISA may be useful to diagnose HHV-7 infection.

Development of an enzyme-linked immunosorbent assay for diagnosis of human herpesvirus-7 infection

Abstract INTRODUCTION: Human herpesvirus (HHV)-7 establishes a latent infection during the lifetime of the host and can reactivate after the primary infection, leading to lytic replication in immunosuppressed patients. METHODS: This study aimed to develop an enzyme-linked immunosorbent assay (ELISA) to identify HHV-7 serum antibodies and compare its performance with that of an indirect immunofluorescence assay (IFA). RESULTS: Serum samples (n=102) were tested by IgG-IFA and by ELISA. IFA and ELISA showed IgG-positive results in 77 and 73 samples, respectively. Qualitative concordance of 96% was demonstrated between the two techniques. CONCLUSIONS: ELISA may be useful to diagnose HHV-7 infection.

Detección y respuesta inmune de virus herpes humano 6 (VHH-6) y virus herpes humano 7 (VHH-7) en población sana / Detection and immune response of human herpes virus 6 (HHV-6) and human herpesvirus 7 (HHV-7) in healthy population

El Herpesvirus Humano 6 (VHH-6) y el Herpesvirus humano 7 (VHH-7) pertenecen a la familia Herpesviridae con una amplia distribución en la población humana. La infección primaria ocurre a muy temprana edad, antes de los 2 años de vida, y el virus establece, en el hospedero susceptible, un modelo persistente de infección. La latencia del virus se produce en las células linfoides y en las glándulas salivales. La reactivación puede suceder en reiteradas ocasiones a lo largo de la vida de un infectado, ya sea éste inmunocompetente o inmunocomprometido, pudiendo el virus ser detectado en saliva y /o plasma. En individuos sanos, inmunocompetentes, la historia natural de la infección es poco conocida. El objetivo de este trabajo es contribuir a la comprensión de los modelos de persistencia viral de HHV-6 y de HHV-7 y de la respuesta inmune que la actividad viral desencadena en el hospedero sano, la cual se pone en evidencia por el perfil de isotipos específco de Inmunoglobulina G (IgG) específicos.(AU)

ABSTRACT: Human herpesvirus 6 (HHV-6) and Human herpesvirus 7 (HHV-7) belong to the Herpesviridae family and are widely distributed among the human population. The human primary infection occurs early, before the two years of age, and the virus establishes a persistent model of infection in the susceptible host. The virus remains latent mainly in lymph cells and salivary glands. Reactivation may occur several times throughout the life of infected individual, whether immune competent or immune compromised, with the virus being detected in saliva and/or plasma. In healthy immune competent individuals, the natural history of the infection is little known. The aim of this research was to contribute to the understanding of the models of viral persistence of HHV-6 and HHV-7 and the immune response that viral activity triggers in the healthy host, manifested by the profile of specific IgG isotypes (IgG1, IgG2, IgG3 and IgG4).(AU)

Correlation of cytomegalovirus and human herpesvirus 7 with CD3+ and CD3+ CD4+ cells in chronic periodontitis patients.

BACKGROUND AND OBJECTIVE: Human chronic periodontitis is an inflammatory process characterized by dense accumulation of immune cells in the periodontal tissue. The periodontitis can lead to loss of teeth in the patient and the pathogenesis of this disease is not completely known. This study tested the hypothesis that chronic periodontitis-affected sites can harbor betaherpesviruses and that viruses are linked to a profile of the inflammatory infiltrate. MATERIAL AND METHODS: Biopsies of periodontal tissue were taken from periodontitis-affected patients and from healthy subjects. Immunohistochemistry was performed to count CD19(+) B cells, CD3(+) total T cells, T-CD4(+) and T-CD8(+) cell subsets, and PCR was performed to detect cytomegalovirus and human herpesvirus 6 and 7 in the samples. One slide of each sample was stained with Giemsa for histopathological examination and to evaluate the quality of the cellular infiltrate. RESULTS: As expected, tissues collected from healthy subjects presented no significant level of inflammatory infiltration and were therefore excluded from immunostaining procedures. Results showed that CD19(+) B cells were in higher number than CD3(+) T cells in the periodontitis-affected tissue, but this was not statistically significant. The T-CD4(+) lymphocyte subset was significantly higher than the T-CD8(+) lymphocyte subset (p = 0.004) in the samples. Cytomegalovirus and human herpesvirus 7 were found at periodontitis-affected sites, but not in tissue collected from healthy subjects (p = 0.04 and p = 0.04, respectively). Human herpesvirus 6 was rarely detected. We found a correlation between cytomegalovirus and lower CD19(+) /CD3(+) ratios (ratio < 0.9, p = 0.003) and between human herpesvirus 7 and lower CD19(+) /CD3(+) ratios (ratio < 0.9, p = 0.003) and higher CD4(+) /CD8(+) ratios (ratio > 1.1, p = 0.002). CONCLUSION: This study shows that cytomegalovirus and human herpesvirus 7 can be present at periodontitis-affected sites but are uncommon at healthy periodontal sites. Moreover, our data suggest that cytomegalovirus can be related to an inflammatory infiltrate with predominance of CD3(+) T cells, whereas human herpesvirus 7 can be associated with an infiltrate with predominance of T-CD4(+) cells. However, further studies are necessary to support this hypothesis. Herpesviruses could play a role in human chronic periodontitis by modulation of the T cell response.

Cytomegalovirus, human herpesvirus-6, and human herpesvirus-7 in adult liver transplant recipients: diagnosis based on antigenemia.

Human herpesvirus (HHV)-6, HHV-7, and cytomegalovirus (CMV) that remain latent after primary infection can be reactivated during immunosuppression following organ transplantation in liver transplant recipients. The aim of this study was to monitor active infections for HHV-6, HHV-7, and CMV among adult liver transplantation recipients using antigenemia detected by an immunoperoxidase staining. Twenty-eight adult liver transplant patients were monitored using antigenemia in blood samples obtained at the time of transplantation, as well as weekly in the first month and once a month for 6 months. Of these patients, 28.5% showed positive CMV antigenemia; 39.2%, HHV-6 antigenemia; and 14.2%, HHV-7 antigenemia. The detection of the three viruses was considered to be independent of one another (P>.05). The results described above showed that few patients remain free of beta herpesviruses after liver transplantation. Most patients were infected sequentially and not concurrently. Antigenemia has been considered useful to detect active HHV-6 and HHV-7 infections. Antigenemia can be more efficiently interpreted when compared with polymerase chain reaction results, although other studies are necessary to establish the reference of HHV-6 and HHV-7 antigenemia.

IgG subclasses and DNA detection of HHV-6 and HHV-7 in healthy individuals.

Human herpesvirus 6 (HHV-6) and 7 (HHV-7) are common opportunistic agents in immunocompromised hosts, although infection with HHV-6 and HHV-7 can also be observed in immunocompetent hosts. Despite similar biology and epidemiology, this study evaluated differences in the IgG subclass distribution associated with HHV-6 and HHV-7 in seropositive, healthy persons. The identified subclasses were also compared with the detection of HHV-6 and HHV-7 DNA. For these assays, sera, plasma, and saliva samples were obtained from 40 healthy blood donors in Argentina who were seropositive for both HHV-6 and HHV-7. HHV-6 and HHV-7 DNA were detected in saliva and plasma samples using nested PCR, and specific IgG subclasses were determined using immunofluorescent assays of sera samples. HHV-7 DNA was detected in 90% of all plasma samples and in 100% of saliva samples. In contrast, HHV-6 DNA was not detected in any of the plasma samples, and it was detected in only 6 of 40 saliva samples. Determination of IgG subclass distributions showed that HHV-6 was restricted to IgG1, whereas HHV-7 IgG subclasses included two groups, one restricted only to IgG1 and the other to IgG1 and IgG3. These results demonstrate the differences between HHV-6 and HHV-7 DNA range detection in saliva and plasma samples, as well as the IgG subclass patterns for each virus type, in healthy persons in Argentina.

Human herpesvirus-7 in Brazilian liver transplant recipients: a follow-up comparison between molecular and immunological assays.

Human herpesvirus-6 and -7 (HHV-6, HHV-7) remain latent after primary infection and can reactivate after transplantation. HHV-6 active infection has been related to some clinical manifestation, but the role of HHV-7 remains unclear. The clinical significance of HHV-7 DNAemia is not completely known and the immune response against HHV-7 has been poorly studied in transplantation. In this study, we investigated HHV-7 DNAemia in liver transplant recipients and evaluated the immunoglobulin (Ig) G and IgM response against HHV-7. A total of 22 adult liver transplant recipients were followed up for 90 days. HHV-7 DNAemia was detected by nested polymerase chain reaction (PCR) in DNA extracted from sera. IgG and IgM detection was performed by immunofluorescent assay using HHV-7-infected cord blood mononuclear cells. A significant virus antibody response was defined as either a positive IgM or a > or =4-fold rise in the virus IgG antibody. All patients had pre-transplant HHV-7-positive serostatus. Nine of 22 (40.9%) patients presented HHV-7 DNAemia during follow-up. All these patients had anti-HHV-7-positive IgM and/or significant increase in IgG titers with concurrent or subsequent DNAemia. In patients without DNAemia and low persistent IgG antibody titers, IgM was not detected. Correlation between nested PCR and IgM detection was statistically significant (P=0.01). Our study indicates that nested PCR in DNA extraction from serum can be useful to detect and monitor HHV-7 active infection in liver transplant recipients. IgM antibody detection also can be useful as a first immunological technique to detect active infection, especially if combined with PCR.

Loss of maternally-derived human herpesvirus-7 immunity and natural infection in Argentinian infants.

BACKGROUND: Human herpes virus-7 (HHV-7) infection is widespread throughout the world. No data are available in Argentina about loss of maternally-derived HHV-7 immunity and natural infection. OBJECTIVES: The objective of this study was to characterize the time when children lose maternal antibodies and become susceptible to natural infection. METHODS: Sera from 39 pregnant women and 207 infants between 2 and 29 months of age were tested. Determination of IgG antibodies was made by indirect immunofluorescence. RESULTS: The seropositive ratio fell in the 2-4 month group (15% seropositive) and increased between 5 months (47% seropositive) and 23 months (67%). Geometric mean titers (GMT) of the infants aged 2-4 months (GMT = 60) were statistically different (p < 0.0001, Student's t-test) to those from the group of pregnant women (GMT = 83) and those from the other infant groups (p < 0.001, least significant difference (LSD) test). The GMT of the groups between 5 and 23 months did not show significant differences whereas those of infants between 24 and 29 months (GMT = 179, 79% seropositive) were different from all the groups studied (p < 0.0001, LSD test). CONCLUSIONS: This study shows a significant association between the loss of passive HHV-7 antibody and age. HHV-7 enters the susceptible population at 5 months, leading to the high prevalence of antibodies between 24 and 29 months of age. This study also shows that natural infection by HHV-7 in children during their first years of life follows the infection pattern found in developing countries.

Human herpesvirus-7 as a cause of exanthematous ilnesses in Belém, Pará, Brazil.

We screened sera from 370 patients suffering from exanthematous illnesses in Belém, North Brazil, for the presence of human herpesvirus-7 (HHV-7) IgM and IgG antibodies. Samples were obtained from January 1996 to December 2002 and were processed by a HHV-7-specific indirect immunofluorescence assay (IFA). HHV-7-specific IgM and/or IgG antibodies were found in 190 (51.4%) of these patients, with similar prevalence rates (IgM+ and IgG+ subgroups taken together) for female and male subjects: 52.5% and 50.3%, respectively. Serological status as defined by IgG was identified in 135 (36.5%) patients. In 55 (14.9%) of the patients HHV-7 IgM antibodies were detected. HHV-7 IgM- and- IgG antibody rates were similar (p > 0.05) when male and female subjects are compared: 14.4% versus 15.3% and 38.1% versus 35.0%, respectively. Statistically significant difference (p = 0.003) was noted when HHV-7-IgM-positive female and male patients aged 5-8 months are compared. Prevalence rates ranging from 4.6% (female, 5-8 months of age) to 93.3% (female, > 10 years of age) and 12.2% (male, 5-8 months) to 80.0% (male, 8-10 years of age) were noted in the IgG- positive subgroups. A subgroup (n = 131) of patients with IgM or IgG HHV-7 antibodies were examined for the presence of DNA using a polymerase chain reaction/nested PCR. Recent/active HHV-7 infection occurred at a rate of 11.0% (6/55) among patients whose samples presented IgM+ specific antibodies. In a subgroup (n = 76) of patients with high HHV-7-IgG antibody levels (titre > 1:160) DNA could not be detected in sera examined by PCR/nested PCR. Of the six recent/active infections, four subjects with less than 1 year and two with 3 and 6 years of age, presented typical exanthem subitum (E.S), as defined by higher fever (> 38.0 degrees C) with duration of 24 to 72 hours, followed by a maculopapular skin rash. Our results underscore the need for searching HHV-7 infection in patients with exanthematous diseases, particularly those presenting with typical E.S. HHV-7 appears therefore to emerge as a newly recognized pathogen of exanthem in our region.

Comparison of seroprevalences of human herpesvirus-6 and -7 in healthy blood donors from nine countries.

BACKGROUND AND OBJECTIVES: The purpose of the study was to register antibody prevalences of HHV-7 in various locations of the world in comparison to the closely related HHV-6. MATERIALS AND METHODS: Sera of healthy blood donors from nine countries in five continents were titered by indirect immunofluorescent assays using HHV-6 infected HSB2 and HHV-7 infected SupT1 cells. RESULTS: Antibody prevalence for HHV-7 is high (75-98%) in practically all countries except for Northern Japan (44%), with no simple correlation to elevated HHV-6 antibody titers. There were regions of low, intermediate and high mean antibody titers against HHV-7 such as 78.5-91.3 for Belgium, Israel, Japan, USA and Australia, 175.4-182.6 for Mexico and Cologne/Germany, and 389.2 for South Africa for which geographic characteristics may be responsible. CONCLUSION: HHV-7, similar to HHV-6, is a widespread human herpesvirus with elevated antibody titers in the healthy human population essentially everywhere. The data warrant further studies to evaluate its possible pathologic potential, preferentially in persons with defective immune responses.

Primary human herpesvirus 7 infection: a comparison of human herpesvirus 7 and human herpesvirus 6 infections in children.

OBJECTIVE: To define the clinical and virologic characteristics of primary human herpesvirus 7 (HHV-7) infection and to compare these characteristics with those of primary human herpesvirus 6 (HHV-6) infection. STUDY DESIGN: A prospective convenience sample study of 496 children < or =3 years old. HHV-7 and HHV-6 infections were identified by viral isolation. Polymerase chain reaction and serology for HHV-7 and HHV-6 were performed. Clinical and laboratory characteristics of patients were obtained from medical records and follow-up interviews. RESULTS: Children with primary HHV-7 infection (n = 8) were identified and compared with children with primary HHV-6 infection (n = 29) detected during the same time period. All children were febrile (mean temperature 39.8 degrees C) with no difference in the degree of fever, frequency of rash, or gastrointestinal complications between the groups. The median age of children with primary HHV-7 infection was 26 months, significantly older than that of children with primary HHV-6 infection (median, 9 months). Children with primary HHV-7 infection were also more likely than those with primary HHV-6 infection to have seizures associated with the illness (P = .004). CONCLUSION: Primary infection with HHV-7 can cause a highly febrile illness in childhood, complicated by seizures. The serologic diagnosis of primary HHV-6 and HHV-7 infections may be confounded by cross-reacting antibodies.

Seroconversion to human herpesvirus 6 and human herpesvirus 7 among Brazilian children with clinical diagnoses of measles or rubella.

We collected acute-phase and convalescent-phase serum samples from Brazilian patients who presented with exanthem of unknown origin and evaluated these samples by means of an immunoblot assay for seroconversion to human herpesvirus 6 (HIV-6) or human herpesvirus 7 (HIV-7). Measles or rubella had been clinically diagnosed in all these patients, but their sera were negative for antibodies to both measles virus and rubella virus. Twenty percent of the patients clearly seroconverted to HHV-6 after manifestation of the exanthem, and 8% seroconverted to HHV-7. All seroconversions to HHV-6 occurred in children aged < or = 5 years; a 41% frequency of seroconversion to HHV-6 was noted among children between 3 months and 23 months of age, whereas seroconversions to HHV-7 were detected during infancy and through adulthood. Our data indicate that primary infections due to HHV-6 or HHV-7 can be misdiagnosed as measles or rubella.

Human herpesvirus 7 infection associated with central nervous system manifestations.

The clinical features of infection with human herpesvirus 7 (HHV-7) are not well described. Exanthem subitum is the only illness that is confirmed to be caused by HHV-7. We report two children who had exanthem subitum associated with central nervous system manifestations. Two strains of HHV-7 were isolated sequentially from peripheral blood mononuclear cells and saliva of the some child who had exanthem subitum complicated with acute hemiplegia in childhood. Two strains were confirmed to be HHV-7 by means of monoclonal antibodies to human herpesvirus 6 (HHV-6) and HHV-7, polymerase chain reaction, and DNA analysis. During the convalescent period, the antibody titer to HHV-7 rose from less than 1:10 to 1:320, whereas the antibody titer to HHV-6 remained less than 1:10. Another child with exanthem subitum complicated by acute hemiplegia had serologic evidence of primary HHV-7 infection. These two cases demonstrate a new relationship between HHV-7 and central nervous system symptoms.

Prevalencia del virus humano herpes 7 en donadores de sangre mexicanos / Prevalence of human herpes viruz 7 in Mexican blood donors

El herpes virus humano 7 (HHV-7) fue aislado recientemente de las células CD4 de individuos sanos; actualmente se realizan estudios de suposible asociación con enfermedades de humanos. Se estudiaron 200 muestras de sangre de candidatos a donadores asintomáticos del banco de sangre del Hospital General de México, utilizando la prueba de inmunofluorescencia indirecta (IFA) en células SupT1 infectadas con el HHV-7 en la Universidad de Colonia, Alemania. El 83.5 por ciento de las muestras fueron de varones y el 16.5 por ciento de mujeres; provenían de 12 diferentes estados de la república mexicana con predominio del Distrito Federal (60.5 por ciento) y del Estado de México (28 por ciento) con edad promedio de 29.2 años. El HHV-7 fue detectado en el 98.5 por ciento de los casos a diferentes titulaciones; el 84.5 por ciento presentó títulos elevados (ò 1:80). El 1 por ciento resultó positivo a hepatitis B, el 2 por ciento a sífilis, y el 0.5 por ciento a brucella. Todos fueron negativos a hepatitis C así como al VIH. La elevada prevalencia de HHV-7 deberá aclararse en investigaciones futuras que permitan determinar la posible asociación de títulos altos con infección activa, así como el significado de ésta en relación a enfermedad

[Prevalence of human herpesvirus 7 in Mexican blood donors]. / Prevalencia del virus humano herpes 7 en donadores de sangre Mexicanos.

The human herpes virus 7 (HHV-7) has been recently isolated from CD4 cells of healthy persons. The present study describes the antibody prevalence of this virus in a healthy Mexican population. Two hundred blood samples from candidates for blood donation at the Hospital General de Mexico were studied with the indirect immunofluorescence test (IFA) in HHV-7 infected SupT1 cells. The testing was done in the University of Cologne, Germany; 167 were males and 33 female; the donors came from 12 of the 31 states in the Mexican republican, predominantly from Mexico City (60.5%) and the State of Mexico (28%). Their mean age was 29.2 years. All but three samples were positive to the HHV-7 (98.5% positivity). Nearly 85% had high titers (> or = 1:80). Other serology testing in the samples revealed 1% positive tests to hepatitis B, 2% to syphilis, and 0.5% to brucella. Hepatitis C and the HIV test were negative in all. The high prevalence of HHV-7 in our donor population should be further studied in order to determine titers indicative of an active infection and of their association with illnesses.