Detection of human polyomaviruses JC and BK in liver pretransplant patients.

OBJECTIVE: This study aimed to identify and quantify polyomaviruses (BKPyV and JCPyV) in the saliva, mouthwash, blood and urine of liver pretransplant patients. MATERIALS AND METHODS: A case-control study was performed using a convenience sample of 21 end-stage liver disease patients (EG = experimental group) and 20 normoreactive controls (CG = control group). In total, 162 samples were collected. Detection and quantification of polyomaviruses were performed using real-time PCR method. RESULTS: In the EG, 21 samples (25%) were positive for BKPyV and 10 (11.90%) for JCPyV, while in the CG, 27 samples (34.61%) were positive for BKPyV and six (7.69%) for JCPyV. With regard to the number of samples positive for BKPyV and JCPyV, there was no statistically significant difference between EG and CG (p = .52 and p = .25). In the EG, we observed a panorama similar to that of the CG regarding the presence of polyomaviruses in mouthwash, blood and urine. The greatest difference between the samples was that regarding the identification of BKPyV in saliva. CONCLUSION: Cirrhotic patients on the liver transplant waiting list did not show higher prevalence of BKPyV and JCPyV compared to normoreactive controls.

Pre-transplant shedding of BK virus in urine is unrelated to post-transplant viruria and viremia in kidney transplant recipients.

BK virus-(BKV) associated nephropathy (BKVN) is a major cause of allograft injury in kidney transplant recipients. In such patients, subclinical reactivation of latent BKV infection can occur in the pre-transplant period. The purpose of this study was to determine whether urinary BKV shedding in the immediate pre-transplant period is associated with a higher incidence of viruria and viremia during the first year after kidney transplantation. We examined urine samples from 34 kidney transplant recipients, using real-time quantitative polymerase chain reaction to detect BKV. Urine samples were obtained in the immediate pre-transplant period and during the first year after transplant on a monthly basis. If BKV viruria was detected, blood samples were collected and screened for BKV viremia. In the immediate pre-transplant period, we detected BKV viruria in 11 (32.3%) of the 34 recipients. During the first year after transplantation, we detected BKV viruria in all 34 patients and viremia in eight (23.5%). We found no correlation between pre-transplant viruria and post-transplant viruria or viremia (p = 0.2). Although reactivation of latent BKV infection in the pre-transplant period is fairly common among kidney transplant recipients, it is not a risk factor for post-transplant BKV viruria or viremia.

JC virus DNA in cerebrospinal fluid samples from Brazilian AIDS patients with focal brain lesions without mass effect.

OBJECTIVE: To evaluate the presence of JC virus DNA in CSF samples from Brazilian AIDS patients with focal lesions of CNS white matter without mass effect compatible with progressive multifocal leukoencephalopathy (PML). METHODS: CSF samples from AIDS patients with neurological symptoms and a CT scan showing focal lesions of CNS white matter without mass effect suggestive of PML, and from AIDS and non-AIDS patients with non-PML neurological diseases were tested for JC virus DNA by PCR. The primers used to amplify the T antigen region of the JC virus resulted in a 173-bp fragment. The presence of the JC virus was confirmed by digestion of the PCR product using BamH1. RESULTS: The PCR for JCV DNA was negative in 119/120 non-PML CSF samples (specificity =99.2%). Of 56 CSF samples from AIDS patients with focal lesions of CNS white matter without mass effect, JCV DNA was positive in 48.2% (27/56). In 23/29 (79.3%) JCV DNA-negative cases, other causes for the encephalitic lesions were found. No JCV DNA-positive cases showed other diagnoses. CONCLUSIONS: The prevalence of JCV DNA by PCR in CSF samples from Brazilian AIDS patients with focal brain lesions, without mass effect was 48.2%. In these patients, a negative JCV PCR is highly suggestive of other neurological conditions.

[Alternative diagnoses among suspected herpes simplex encephalitis patients with negative polymerase chain reaction]. / Diagnósticos alternativos em pacientes com suspeita de encefalite por Herpes simplex e negativos à reação em cadeia por polimerase (PCR).

The aim of this study was to analyze the diagnosis found in a series of patients in which the diagnosis of Herpes simplex encephalitis (HSE) was ruled out by a negative polymerase chain reaction (PCR) result for HSV DNA in cerebrospinal fluid (CSF) samples. Forty three out of 61 HSE suspected patients had negative PCR. An alternative diagnosis was established in 41.9% of these patients. These patients were diagnosed as having viral (2 cases-11.1%) and non viral (5 cases-27.2%) CNS infections, vascular (4 cases-22.2%) and demyelinating diseases (3 cases-16.7%), metabolic disturbances (3 cases-16.7%), and CNS tumor (1 case-5.6%). The non specific clinical presentation of this disease and the availability of an efficient treatment for HSE explain why several patients with other diseases were initially treated with acyclovir. The early use of PCR in CSF was considered essential for the evaluation of the acute encephalitis cases in this study.

Cytomegalovirus infection in children with Down syndrome in a day-care center in Brazil.

This study evaluates the transmission of CMV infection in 120 children aged 1 to 15 years with Down syndrome who attended a day-care center for handicapped children in São Paulo, Brazil. A blood sample was obtained from each children at the beginning of the study for detection of IgG and IgM cytomegalovirus (CMV) antibodies by an immunofluorescence assay. Samples of saliva and urine were obtained every 3 months from the children with CMV antibodies to detect shedding of the virus by culture in human foreskin fibroblasts, by detection of pp65 CMV-antigen and by a nested PCR assay. The prevalence of anti CMV-IgG antibodies was 76.6% (92/120), and IgM anti-CMV antibodies were detected in 13% (12/92) of the seropositive children. During the first viral evaluation, CMV was detected in the urine and/or saliva in 39/90 (43.3%) of the seropositive children. In the second and third evaluations, CMV was detected in 41/89 (46%) and in 35/89 (39.3%) children, respectively. Detection of CMV was shown both in urine and saliva in 28/39 (71.8%), 19/41(46.3%) and 20/35 (57.1%) of the children excreting the virus, respectively. Additionally, in 3(3/4)9 (67.4%) of the excreters CMV could be demonstrated in urine or saliva in at least two out of the three virological evaluations carried out sequentially in a six month period. Of the 28 initially seronegative children, 26 were re-examined for anti-CMV IgG antibodies about 18 months after the negative sample; seroconversion was found in 10/26 (38.5%). Taking all 536 samples of urine or saliva examined by virus culture and pp65 antigen detection during the study into account, 159 (29.6%) were positive by virus culture and 59 (11%) gave a positive result with the pp65 assay. These data demonstrate the high prevalence of CMV shedding and the high risk of CMV infection in children with Down syndrome attending a day-care center for mentally handicapped patients. The virus culture was more sensitive than the pp65 CMV antigen assay for CMV detection in both urine and saliva samples.

Diagnosis of herpes simplex encephalitis by magnetic resonance imaging and polymerase chain reaction assay of cerebrospinal fluid.

The early diagnosis of herpes simplex encephalitis (HSE) is essential because early introduction of antiviral therapy can significantly reduce the mortality of this disease. Herpes simplex virus (HSV) DNA detection in cerebrospinal fluid (CSF) samples is a rapid, noninvasive, specific, and highly sensitive method for HSE diagnosis. Neurodiagnostic methods have also been studied for noninvasive diagnosis of HSE. Magnetic resonance imaging (MRI) seems to be the most sensitive of them but it has not been compared to PCR in terms of efficacy for HSE diagnosis. In this study, 17 patients with focal encephalitis were prospectively evaluated by PCR analysis of CSF samples and MRI examination. MRI lesions involving the inferomedial region of one or both temporal lobes were observed in all PCR-positive patients but one. No PCR-negative patient presented with the same pattern of MRI lesions. MRI was also important for the establishment of an alternative diagnosis in three of eight PCR-negative patients. Both methods should be routinely applied in the evaluation of presumed HSE cases.

Molecular detection and isolation of human T-cell lymphotropic virus type I (HTLV-I) from patients with HAM/TSP in São Paulo, Brazil.

BACKGROUND: Infection with HTLV-I is etiologically linked with HTLV-I-associated myelopathy/tropical spastic paraparesis (HAM/TSP). However some patients with chronic progressive paraparesis resembling HAM/TSP have been shown to be infected with HTLV-II. OBJECTIVE: To clarify the role of each of these human retroviruses in the etiology of HAM/TSP in São Paulo, Brazil. STUDY DESIGN: A detailed serological and molecular analysis of HTLV-I/II infection was performed in a cohort of 19 patients with HAM/TSP attending a neurological clinic. RESULTS: Plasma samples analyzed for anti-HTLV-I/II antibodies using a Western blot assay, comprising HTLV-I (rgp46I)- and HTLV-II (rgp46II)-specific recombinant env epitopes, demonstrated reactivity to rgp46I and hence were typed as seropositive for HTLV-I. Presence of HTLV genomic sequences in peripheral blood mononuclear cells (PBMC) was sought after by PCR using consensus primers SK 110 and SK 111 for the pol region of HTLV proviral DNA followed by hybridization with type-specific probes--SK 112 (HTLV-I) and SK 188 (HTLV-II). Southern blots from all individuals hybridized with SK 112 but not with SK 188, further confirming HTLV-I infection. Cocultivation of PBMC from eight of these patients with activated lymphocytes from normal individuals resulted in active viral production, detected as presence of soluble p24gag antigen in culture supernatants. Investigation of risk factors for HTLV-I infection in these individuals revealed that five out of 19 patients studied (26.3%) had received blood transfusions previous to disease onset.

Randomised, double blind, placebo controlled clinical trial of efficacy of vitamin A treatment in non-measles childhood pneumonia.

OBJECTIVE: To evaluate the impact on clinical recovery and severity of the addition of large doses of vitamin A to the standard treatment for childhood pneumonia. DESIGN: A randomised, double blind, placebo controlled trial. SETTING: Study children were recruited at a public hospital in Recife, north east Brazil, an area of marginal vitamin A deficiency. SUBJECTS: 472 children aged 6 to 59 months with clinical diagnosis of pneumonia. INTERVENTIONS: 200,000 IU (infants) or 400,000 IU (1-4 year olds) of vitamin A in oil or similar capsules of placebo divided into two daily oral doses, in addition to the standard treatment. MAIN OUTCOME MEASURES: Duration of the episode and incidence of adverse outcomes. RESULTS: The groups were similar with respect to overall duration of pneumonia and incidence of adverse outcomes. Children who received vitamin A, however, were less likely to have fever by day 3 (P = 0.008) and were 29% less likely to fail to respond to the first line antibiotic (P = 0.054). CONCLUSION: There was little evidence for an effect of vitamin A treatment on the immediate outcome of the pneumonia episode.

Advantage of polymerase chain reaction in the diagnosis of herpes simplex encephalitis: presentation of 5 atypical cases.

Four case of herpes encephalitis (HSVE) are described. The diagnosis was established by polymerase chain reaction (PCR) assay of cerebrospinal fluid (CSF). These reports illustrate different situations in the clinical management of this disease. PCR was considered useful in confirming the HSVE diagnosis in 3 atypical cases, and in the differentiation between virologic failure and postinfectious encephalitis in a patient with recurrence of symptoms. A case with typical HSVE clinical findings is also reported where PCR was negative and a temporal lobe lymphoma was diagnosed at autopsy. This last case is representative of the utility of PCR in the management of other diseases mimicking HSVE.

Prevalence of rubella antibodies in a non-immunized urban population, São Paulo, Brazil. The Division of Immunization, CVE.

The prevalence of rubella antibodies was evaluated through a random seroepidemiological survey in 1400 blood samples of 2-14 year old children and in 329 samples of umbilical cord serum. Rubella IgG antibodies were detected by ELISA, and the sera were collected in 1987, five years before the mass vaccination campaign with measles-mumps-rubella vaccine carried out in the city of São Paulo in 1992. A significant increase in prevalence of rubella infection was observed after 6 years of age, and 77% of the individuals aged from 15 to 19 years had detectable rubella antibodies. However, the seroprevalence rose to 90.5% (171/189) in cord serum samples from children whose mothers were 20 to 29 years old, and reached 95.6% in newborns of mothers who were 30 to 34 years old, indicating that a large number of women are infected during childbearing years. This study confirms that rubella infection represents an important Public Health problem in São Paulo city. The data on the seroprevalence of rubella antibodies before the mass vaccination campaign reflects the baseline immunological status of this population before any intervention and should be used to design an adequate vaccination strategy and to assess the seroepidemiological impact of this intervention.

Infecção perinatal pelo citomegalovirus em hospital publico do municipio de Sao Paulo: estudo prospectivo / Perinatal infection due to cytomegaloviruses in a public hospital of the municipality of Säo Paulo: a prospective study

Com o objetivo de se avaliar a magnitude da infeccao perinatal pelo citomegalovirus em hospital publico do municipio de Sao Paulo, os autores acompanharam prospectivamente 98 recem-nascidos ete o quarto mes de vida. Amostras de urina foram coletadas ao nascimento e posteriormente a cada mes, para inoculacao em tubos contendo fibroblastos humanos. Amostras de sangue foram coletadas ao nascimento, no segundo e quarto mes de vida para pesquisa de anticorpos IgM especificos para o CMV, pelo metodo de imunofluorescencia indireta. Dos 37 recem-nascidos que foram acompanhados ate o quarto mes de vida, 9 se infectaram neste periodo, com diagnostico feito pelo isolamento do CMV. O risco de aquisicao da infeccao pelo citomegalovirus no periodo perinatal estimado pela tabua de sobrevivencia foi de 30,9 por cento. A pesquisa de anticorpos IgM por imunofluorescencia so permitiu tal diagnostico em 2 casos (8,1 por cento). A diferenca observada entre os dois metodos foi estatisticamente significante (p = 0,015). O estudo da prevalencia de anticorpos IgG pelo ensaio imunoenzimatico nas maes das criancas mostrou taxas de 92,7 por cento. Nao se isolou CMV nas amostras de leite materno, coletadas mensalmente ate o terceiro mes de lactacao...

[Perinatal infection due to cytomegaloviruses in a public hospital of the municipality of São Paulo: a prospective study]. / Infecção perinatal pelo citomegalovírus em hospital público do município de São Paulo: estudo prospectivo.

In order to demonstrate the occurrence of CMV perinatal infection in a middle socioeconomic class population, the authors conducted a 8-month prospective study in 37 children, not infected congenitally, born in a public hospital of São Paulo city, Prevalence of CMV-IgG antibodies in mothers, detected by immunoenzymatic assay (ELISA), was 92.7%. Survival analysis showed that the risk of acquiring CMV perinatal infection diagnosed by virus isolation in human fibroblasts was 30.9%. When the diagnostic method was detection of IgM class antibodies by indirect immunofluorescence the risk was 8.1% (p < 0.05). Milk samples inoculated in human fibroblasts failed to demonstrate the presence of virus. The infected children did not present any signal of disease in a 4-month follow-up.

[Serological evaluation of immunization against measles with 2 doses administered at 6 and 11 months of age: a prospective study]. / Avaliação sorológica de imunização contra o sarampo com duas doses administradas aos 6 e 11 meses de idade: estudo prospectivo.

A prospective study was designed to evaluate the serologic efficacy of a two dose measles vaccination schedule, at 6 months and 11 months of age. Infants were given a further attenuated measles virus vaccine (BIKEN CAM 70, Fundação Oswaldo Cruz) and serum samples were tested for measles antibodies using the indirect immunofluorescence technique (IFA) and ELISA. Seroconversion rates 6 to 12 months (mean of 8.0 +/- 1.7 months) following the second dose of measles vaccine were 88.5% (85/96) by IFA and 96.8% (93/96) by ELISA. No measles cases were reported during the study period. In regions where a significant proportion of measles cases occurs before nine months of age, vaccination with a two doses schedule, at 6 and 11 months of age, may represent an alternative for measles control.

Cytomegalovirus and other herpesviruses infections in heart and bone marrow transplant recipients.

From January 1988 to January 1989 all the heart transplant and bone marrow recipients at the Instituto do Coração of the Hospital das Clínicas of the University of São Paulo Medical School were studied for the incidence and morbidity associated with herpesviruses infections after transplantation. Five bone marrow and 5 heart transplant recipients were followed for a mean of 4.2 months post-transplantation. All the patients were seropositive for cytomegalovirus (CMV) before admission and 80% experienced one or more recurrences during the observation period. Of the 12 episodes of CMV infection, that were identified in this study, 83% were accompanied by clinical or laboratory abnormalities. However, there was only one case of severe disease. The overall incidence of infection for herpes simplex (HSV) was 50%. Although most of HSV reactivations were oral or genital, one case of HSV hepatitis occurred. One of the 6 episodes of HSV infections that were treated with acyclovir showed an unsatisfactory response and was successfully managed with ganciclovir. All the individuals had anti-varicella zoster virus antibodies, but none of them developed infection. The study emphasizes the importance of active diagnostic surveillance of herpesvirus infections in transplant patients. Both CMV and HSV reactivations showed high incidence and important morbidity and thus, deserve prophylactic therapy.

Enzyme linked immunosorbent assay: determination of anti-adenovirus antibodies in an infant population.

In order to define an accurate assay for anti-adenovirus antibody detection, a recently developed ELISA was compared with IFA and CF. On 58 sera, the ELISA was more sensitive than both CF and IFA, which showed relative sensitivities of 63% and 94%, respectively. It was not possible to determine the exact specificity of the tests because of the lack of a gold standard. Furthermore, the ELISA was used to define the prevalence of adenovirus antibodies in 116 infants between 1 and 24 months old (mean 7.28). The data showed that maternal antibodies waned by the age of 5 to 6 months and that more than 80% of the children had been infected by adenoviruses by the age of 10 months.