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Amostras de soro sanguineo (n: 110) de individuos de comportamento de risco e doadores de sangue da cidade de Cordoba, na Argentina, com perfis de reactividade para HIV-1 e HTLV-I/II indeterminada por Western blot (Wb), foram estudadas para anticorpos especificos contra HTLV-I/II e HIV-1 por meio do Ensaio de Imunofluorescencia Indireta (IFI). Este estudo foi realizado para caracterizar as reacoes putativas com proteinas HIV-1 e HTLV-I/II e resolver o estado da infeccao por retrovirus destes individuos. Os resultados mostram que os soros dos doadores sanguineos que apresentam padroes indeterminados para HTLV-I/II e HIV-1 no Wb nao sao reagentes contra HTLV-I/II e HIV-1 por IFI. Mas, um alto indice de amostras de individuos com alto risco com resultado indeterminado no Wb para HIV-1 e HTLV-I/II apresentaram resultados positivos para HTLV-I/II e HIV-1 por IFI, respectivamente. Nosso estudo sugere que a reatividade indeterminada para HTLV-HIV na populacao de baixo risco deve-se a uma reacao cruzada contra antigenos nao virais; e que na populacao de alto risco as amostras indeterminadas apresentam reacao cruzada entre as proteinas HIV-1 e HTLV-I/II no Wb. Estes resultados indicam que se faz preciso pesquisar a reatividade de HTLV-I/II nas amostras indeterminadas de HIV-1 e vice-versa, para confirmar o diagnostico. Por ultimo, este trabalho mostra a utilidade potencial da IFI para determinar o estado de infeccao HIV-1 e HTLV-I/II dos individuos com perfis indeterminados por Wb, permitindo assim, a resolucao do estado real de infeccao por retrovirus.

To evaluate the prevalence of enteric viruses and their possible association with diarrhea, 244 stool samples were collected from HIV-infected and uninfected patients with or without diarrhea (subgroups I-a, Ib, II-a, and II-b, respectively). Subjects were screened by polyacrylamide gel electrophoresis, latex agglutination, and enzyme immunoassays for rotaviruses, adenoviruses, picobirnaviruses, and astroviruses. Enteric viruses were found significantly more often in specimens from HIV patients (20%) than in specimens from uninfected HIV patients (0%) (p0.05). Picobirnavirus was detected in 14.63% of 82 HIV-infected patients with diarrhea, but it was detected neither in those without diarrhea (0%) (p0.05) nor in the groups of uninfected HIV subjects (0%) (p0.05). Nor could astrovirus (subgroups I-a [4.00%] versus subgroup I-b [5.26%],p0.05) or enteric adenovirus (subgroup I-a [1.22%] versus subgroup I-b [0%], p0.05) be linked to the diarrhea disorder in HIV-infected patients. Rotaviruses were not detected in any of the clinical subgroups studied. Enteric viruses were detected in 15 of 93 (16.13%) of the HIV-infected patients with CD4+ T cell count200/microl and 3 of 19 (15.79%) of those HIV-infected individuals with a CD4+ T cell count 200-499/microl, showing no significant difference (p0.05). According to our data, unusual enteric viruses such as picobirnavirus, astrovirus, and enteric adenovirus occur in HIV-infected population in Córdoba, Argentina. However, only picobirnaviruses could be significantly associated with diarrhea in these patients.

We compared the indirect immunofluorescence assay (IFA) with Western blot (Wb) as a confirmatory method to detect antibodies anti retrovirus (HIV-1 and HTLV-I/II). Positive and negative HIV-1 and HTLV-I/II serum samples from different risk populations were studied. Sensitivity, specificity, positive, negative predictive and kappa index values were assayed, to assess the IFA efficiency versus Wb. The following cell lines were used as a source of viral antigens: H9 ( HTLV-III b); MT-2 and MT-4 (persistently infected with HTLV-I) and MO-T (persistently infected with HTLV-II). Sensitivity and specificity rates for HIV-1 were 96.80% and 98.60% respectively, while predictive positive and negative values were 99.50% and 92.00% respectively. No differences were found in HIV IFA performance between the various populations studied. As for IFA HTLV system, the sensitivity and specificity values were 97.91% and 100% respectively with positive and negative predictive values of 100% and 97.92%. Moreover, the sensitivity of the IFA for HTLV-I/II proved to be higher when the samples were tested simultaneously against both antigens (HTLV-I-MT-2 and HTLV-II-MO-T). The overall IFA efficiency for HIV-1 and HTLV-I/II-MT-2 antibody detection probed to be very satisfactory with an excellent correlation with Wb (Kappa indexes 0.93 and 0.98 respectively). These results confirmed that the IFA is a sensitive and specific alternative method for the confirmatory diagnosis of HIV-1 and HTLV-I/II infection in populations at different levels of risk to acquire the infection and suggest that IFA could be included in the serologic diagnostic algorithm.

Culture amplification in colon adenocarcinoma cell line (CaCo-2) combined with enzyme immunoassay (Pathfinder ELISA) was developed as a supplementary tool for rotavirus diagnosis. One hundred and thirty stools in which results by polyacrylamide gel electrophoresis (PAGE) were in agreement with those obtained by ELISA were amplified in the CaCo-2 cell line. After the first passage 100% specimens were revealed as positive by ELISA. This result was confirmed by PAGE and direct electron microscopy (EM) which increased the rates of rotavirus detection up to 100% after the third and fifth cell passages, respectively. All of the amplified negative stools were confirmed as negative. Among discordant results, three of the eight specimens positive by ELISA but negative by PAGE were confirmed as true positive after the third cell passage. False positive ELISA results could be discarded when the samples were culture amplified and retested by the same ELISA. Using the CaCo-2 amplification-ELISA as supplementary assay, sensitivity and specificity were 1.000 and 0.953 for ELISA and 0.917 and 1.000 for PAGE, respectively. The combined CaCo-2 cell line amplification-immunoassay method proved to be suitable both to evaluate increase in sensitivity of newly developed rotavirus assays and for rotaviral amplification before antigen assays.

Indirect immunofluorescence assay (IFA) is a well-accepted assay for the confirmation of human retrovirus infection. Fluctuations in HIV-1 antigen expression in infected E-B2 cells depending on several factors have been reported. Cells kept in log phase expressed the highest levels of viral antigen. Thus, we studied the time kinetics of IFA positivity in MT-2 (HTLV-I), MO-T (HTLV-II), CEM, and H9 (HIV-1) cell lines. Uninfected T cell line, HT, was used as nonspecific control. Reference HTLV-I/II and HIV-1 serum panels from the Centers for Disease Control and Prevention (CDC) were tested by conventional IFA procedure on slides of each cell line made on different days. On the second day after subculture, HTLV-I strongly positive sera reacted on MT-2 and MO-T cells with a bright pericytoplasmic fluorescence pattern. Weakly positive sera showed a faint staining from the fifth day on, when all the sera showed the highest degree of fluorescence. With HIV-1 cell lines, sera predominantly reacted with a diffuse cytoplasmic pattern, although some sera showed a granular and pericytoplasmic capping staining. The highest degree of fluorescence was found at 3-5 days after subculture. We demonstrated that the sensitivity of the IFA for the detection of antibodies against human retroviruses depended on the day when the slides were assayed and on the serum antibody titer. The fifth day was the most appropriate for HTLV-I/II and HIV-1/H9 systems, whereas for HIV-1/CEM, the fourth day was better. Furthermore, the intensity of the immunofluorescence pattern differed with the antibody titers and the level of antigens expressed on the four cell lines studied. The IFA, improved in our laboratory, proved to be very sensitive, specific, and rapid and could be used as a supplementary/confirmatory assay for retrovirus infection.

Human rotaviruses (HRV) are the most important etiologic agents of acquired diarrhea in infants and young children worldwide. Therefore, the early diagnosis is essential for effective patient management and infection control. We have developed a rapid, simple technique for the diagnosis of rotavirus based on the sensitive detection of rotavirus double-stranded RNA genome segments separated in vertical agarose gels and developed by silver staining (AGE-SS). This method also has the ability to detect differences in the electrophoretic mobility of RNA bands among group C rotaviruses, reovirus, and group A rotaviruses. The results indicate that this assay is as sensitive and specific as the polyacrylamide gel electrophoresis and silver stain method (PAGE/SS) and it could be applied on large scale for the screening of stool suspected of rotaviral diarrhea. This assay does not need sophisticated equipment and the cost per sample is minimal compared with other available assays.

The neuroinvasiveness of Junin virus depends on the viral strain, animal species, and age. The role of infected blood cells in hematogenous Junin virus spread to the central nervous system (CNS) was studied by determining the growth in pheripheral mononuclear cells and brain tissue of Candid 1 and XJCL3 laboratory strains, in Calomys musculinus and guinea pigs. The present study demonstrated that Junin virus replicates in circulating peripheral lymphocytes and macrophages of 11-day-old guinea pigs and 6 +/- 1-day-old Calomys musculinus. Moreover, the observation that mononuclear phagocyte depletion did block Junin virus neuroinvasion firmly indicates that the cellular viremia (circulating monocytes) is one of the mechanisms of Junin virus attenuated strains spread to the CNS in animal hosts.

This study aimed at implementing a Nested-polymerase chain reaction (Nested-PCR) for the molecular diagnosis of human T-cell lymphotropic virus type I/II (HTLV-I and HTLV-II) infections in peripheral blood mononuclear cells of infected subjects in Argentina. The sensitivity and specificity of the assay for the detection of regional strains were assessed by comparing them with the molecular assay of reference PCR-hybridization. The Nested-PCR detected 1 MT-2 cell (³ 8 proviral copies)/1x106 non-infected cells showing high sensitivity for provirus detection. While both molecular assays showed high specificity (100%) for HTLV-I and HTLV-II detection, the sensitivity values differed: 100% for Nested-PCR and 67% for PCR-hybridization assay. Moreover, this technique showed less sensitivity for the detection of DNA sequences of HTLV-II (33%) than for the detection of DNA sequences of HTLV-I (75%). The high sensitivity and specificity of the Nested-PCR for regional strains and its low costs indicate that this assay could replace the PCR-hybridization assay for the molecular diagnosis of HTLV-I/II infections. It will be interesting to assess the usefulness of this assay as a tool for the molecular diagnosis of HTLV-I/II infections in other developing countries. Other studies that include a greater number of samples should be conducted.

Background: Human herpesvirus-6 (HHV-6) infection is widespread throughout the world. No data are available in Argentina about the loss of maternally derived HHV-6 immunity and natural infection in infants.Methods: A population of 100 pregnant women and 407 children between 1 and 15 months of age were assayed by indirect immunofluorescence to detect and quantify specific IgG anti-human herpesvirus-6 (anti-HHV-6) antibodies in Córdoba City, Argentina.Results: There was no significant difference in the positive rate between infants aged 1 to 9 months (range, 43.6–35.5%) and pregnant women (37%). Seropositive ratio dropped in the 10-month group (23.33% seropositive) and rose sharply in the 11-month group (38.89%), 12-month (60.61 %), and 13- to 15-month group (63.46%). The geometric mean titer (GMT) for infants in the 12 to 15 months age group (23.4–41.64) was significantly higher than the GMT for infants 10 months of age (11.04) (P < 0.05 with the Tukey-HSD test).Conclusions: This study shows a significant association between loss of passive HHV-6 antibody and age among infants. The results support evidence that HHV-6 enters the susceptible population at 11 months, leading to a high prevalence of antibodies in children between 13 and 15 months of age.

Diarrhea due to enteric pathogens is an important complication of advanced HIV infection. Picobirnaviruses are agents recently linked with human enteritis. In total, 197 fecal samples collected from HIV-infected and noninfected patients with and without diarrhea were investigated for the presence of rotavirus and picobirnavirus by polyacrylamide gel electrophoresis. Picobirnavirus was detected in 8.8% of 57 HIV-infected patients with diarrhea, but it was detected in neither those without diarrhea (p.018) nor in the group of subjects uninfected with HIV (p.022). All genomic electropherotypes of picobirnavirus strains had a wide pattern. Picobirnavirus genome segments varied in size between 2.4 and 2.7 and 1.6 and 1.9 kbp for the slow and fast migrating bands, respectively. Rotaviruses were not detected in any of the clinical groups studied. Two methods for the extraction of nucleic acid-phenol/chloroform and guanidinium thiocynate (GTC)/silica-were compared. Detection of picobirnavirus by polyacrylamide gel electrophoresis was 2.5 times more sensitive following guanidinium thiocynate RNA extraction. This investigation offers preliminary results about the circulation of picobirnavirus in HIV-infected patients in Córdoba, Argentina.In 1988, a new group of viruses containing a bisegmented double-stranded RNA genome was described and named "picobirnavirus" (PBV). Viruses with similar properties have subsequently been found in fecal specimens collected from HIV-infected and noninfected patients with gastrointestinal symptoms in several countries. The present study used polyacrylamide gel electrophoresis (PAGE) to examine fecal specimens from 197 HIV infected and noninfected adults, with and without diarrhea, from Cordoba, Argentina, for rotavirus and PBV. PBVs were detected in the stools of 5 HIV-infected patients with diarrhea (8.8%), but in none of the other subgroups (HIV-positive patients without diarrhea, HIV-negative patients with diarrhea, HIV-negative patients without diarrhea). 3 of the 5 stool samples positive for PBV were also positive for intestinal parasites (mixed infection), but these parasites were found with equal frequency in HIV-infected patients without diarrhea. Rotaviruses were not detected in any of the subgroups. PBV genome segments varied in size between 2.4-2.7 and 1.6-1.9 kbp for the slow and fast migrating bands, respectively. PBV detection by the PAGE technique was 2.5 times more sensitive after guanidinium thiocyanate RNA extraction. Further research is required to determine the duration of excretion of PBVs in HIV-infected patients with diarrhea and understand the immune response to infection.

Background. Measles immunization of children at 1 year of age with a single dose of the current vaccine has successfully reduced measles incidence in Argentina. However, the optimal schedule of measles vaccination of young infants would balance the risk of early loss of maternal antibody in the majority of infants with the risk of primary vaccine failure because of passive measles immunity. This study is the first to document a significant association between loss of passive measles antibody and age among infants born in 1995 and 1996 in Cordoba City, Argentina. Methods. This is a seroprevalence study of 340 infants to investigate the duration of transplacentally derived measles antibody, assayed by a neutralization test, during the first 8 months of age in C6rdoba City, Argentina. Results. The proportion with detectable neutralizing measles antibodies decreased from 85% at 1 month of age to 8% at 8 months of age. The simple logistic model with age (in weeks) as the only variable showed that the decline in the proportion of infants with a positive antibody titer was sharpest during the second and fifth months of age (6.6 and 6.8% per week during a 4-week period, respectively). Conclusions. These findings suggest that 80% of infants are susceptible to measles infection for at least 3 months before routine immunization at 12 months of age.

Evaluation of the measles virus ELISA kit (Merck) to detect specific IgM as an indicator of primary measles antibody response was carried out. A modification of the manufacturer's cutoff value interpretation was introduced to allow for equivocal results in addition to positive and negative ones. With this modification, the test assayed gave an overall reproducibility of 96.16%. The IgM seropositivity rate for seroneutralization-confirmed measles cases was 100% for naturally infected measles subjects and 90% for primary measles vaccinated subjects. Individuals with positive neutralizing antimeasles antibodies in close contact with a confirmed measles case gave the following measles IgM ELISA results: 54.54% negative, 9.09% positive, and 36.36% equivocal, showing a booster with IgM antibody response on reexposure to the virus. Positive subjects with neutralizing antimeasles antibodies without recent contact with a measles case gave negative IgM results. IgM seropositivity was strongly associated with IgG seroconversion and clinical measles (p < 0.0001). The technique assayed performed adequately for the confirmation of both measles natural infection and primary vaccination and for the differentiation of primary and secondary antibody response, taking into account the modification in the cutoff value interpretation introduced and providing that the serum samples are obtained between days 5 and 30 after onset of rash.

Summary The neuroinvasiveness of Candid 1 and XJCL3 laboratory strains and CbalV4454 and CbaFHA5069 wild strains of Junin virus was studied in albino mice, guinea pigs, and a South American wild rodent, Calomys musculinus (Cm), of different ages inoculated by a non-neural route. Infectivity in brain, blood and organs, as well as lethality, were determined. The results with the 3 hosts indicate that Junin virus neuroinvasiveness is virus-strain-dependent, host species- and age-dependent, with the Candid 1 strain proving to be the least neuroinvasive of the strains studied. The lethal efficiency index (log PFU/LD 50 ) in 2-day old albino mice and the neuroinvasiveness index (Log PFU/ND 50 ) in 6 ± 1 day-old Cm of the various strains using the intraperitoneal (ip) route could therefore be useful markers of Junin virus neuroinvasasiveness. Moreover, different patterns of infection were established using the results of the presence of infectious virus in brain and viraemia in the 3 hosts. In nearly all cases, virus neuroinvasion was present without detectable viraemia (virus in plasma). Current evidence leads to the assumption that virus might reach the brain associated with the white cells in blood (undetectable by conventional isolation methods) or by another possible mechanism of neuroinvasion which is not haematogenous.

The neurovirulence of Candid #1 and XJCL3 laboratory strains and CbalV4454 and CbaFHA5069 wild strains of Junin virus was studied in albino mice, guinea pigs, and a South American wild rodent, Calomys musculinus, of different ages inoculated by the intracerebral route. Infectivity in brain and organs, lethality, and neuropathological lesions were determined. The laboratory and wild strains showed similar neurovirulence only in 2-day-old mice. The neurovirulence of laboratory strains decreased with the age of the animal, and the Candid #1 strain affected only 2-day-old mice. In guinea pigs, the 2 wild strains and XJCL3 laboratory strain were neurovirulent for 11-day-old and adult animals giving moderate lymphocytic infiltration in the brain and mild lesions in the spinal cord. Virus titres from the brain and the spinal cord were lower with the XJCL3 and CbalV4454 strains than with the CbaFHA5069 strain; with the latter, virus was recovered only from the lymph nodes, the lung, kidney, liver, and spleen. The Candid #1 strain was not neurovirulent even for 11-day-old animals. In contrast, the laboratory strains were neurovirulent for Calomys musculinus, depending on the age of the animal. Virus was recovered from the brains showing lymphocyte infiltration but not from other organs. The CbaFHA5069 strain was not neurovirulent, although virus was recovered from the brain, spleen, liver, lymph nodes, and salivary glands. These results with the 3 hosts indicate that Junin virus neurovirulence is virus strain-dependent, and host species and age-dependent, with the Candid #1 strain proving the least neurovirulent of the strains studied.

Recent epidemiologic studies in Buenos Aires suggest the presence of HTLV-I/II in blood banks and in the population at risk for HIV-1 infection with seroprevalence of 4% among IV drug users 0.5% in homosexual men and 0.1% in blood donors. However neither blood nor blood derivatives in Argentina are screened for these agents. Findings are presented from an HTLV-I/II seroprevalence survey conducted in Cordoba City Argentina. 312 serum samples collected during 1985-95 from HIV-1-positive subjects and others exposed to HIV through blood transfusion were screened for antibodies to HTLV-I/II. Of 10 sera reactive to Serodia HTLV-I particulate agglutination (PA) assay 6 were confirmed HTLV-I-positive and 1 was confirmed HTLV-II-positive by indirect immunofluorescence assay (IFA). Of 6 HTLV-I-positive samples on IFA 5 were confirmed positive by Western blot (WB) but 1 sample was not tested. The IFA HTLV-II-positive sample was indeterminate with a weak reaction to the p24 and p19 HTLV-I proteins. Blood banks should screen for the presence of HTLV-I/II infection in blood supplies in order to prevent their widespread dissemination in the population.