ABSTRACT Introduction: The reverse transcriptase polymerase chain reaction (RT-PCR) test has limitations in its diagnostic performance and requires complex equipment. Antibody detection tests can be used as epidemiological screening, but should be evaluated in a regional context. Knowing the profitability of the test, part of the spectrum of humoral immunity evoked by SARS-CoV-2 could be explored. Health personnel, as a population with high commitment and first impact in the pandemic, constitutes the ideal stage. Objective: To evaluate the diagnostic performance of the serological test, antibody response and its correlation with clinical characteristics in health workers with COVID-19 at the General Hospital of Luque. Methodology: To evaluate the diagnostic performance of the serological test, antibody response and its correlation with clinical characteristics in health workers with COVID-19 at the General Hospital of Luque between March and October 2020, a descriptive study was carried out in 107 health workers, selected by convenience between March and October 2020, after having completed ≥14 days from the diagnosis. The RT-PCR test was performed on nasal and pharyngeal swab samples. Antibody detection was performed in plasma sample, analyzed by Elecsys Anti-SARS-CoV-2 electrochemiluminescence immunoassay (<1, negative). Data were also collected: demographic, anthropometric, comorbidities, recent history of dengue, BCG vaccination, symptoms of the acute stage, and time between onset of symptoms and swabbing and onset of symptoms and interview. Descriptive and inferential analysis was used and the sensitivity, specificity, predictive values and accuracy of the test were calculated. Results: Globally, 56 health care workers had a positive diagnosis of COVID-19, of which seven women (asymptomatic) showed levels <1 to the immunoassay for antibodies. Of the 51 workers with a negative RT-PCR result, 3 showed elevated antibody titers. The sensitivity of the test was 89.3%, the specificity 94.1% and the accuracy 91.6%. No correlation was observed between the global level of antibodies and age (r 0.09; p 0.52) as well as with comorbidities. Finally, workers with persistent symptoms had more noticeable antibody levels than those who resolved the disease (p = 0.049). Conclusion: The anti-N antibody detection test for SARS-CoV-2 had a good diagnostic performance, showing some false negatives to RT-PCR, confirming its usefulness for seroprevalence studies. Antibody levels were higher in workers with persistent symptoms.

RESUMEN Introducción: La prueba de reacción en cadena de la polimerasa con transcriptasa inversa (RT-PCR) tiene limitaciones en su desempeño diagnóstico y requiere equipamiento complejo. Las pruebas de detección de anticuerpos pueden ser utilizadas como cribado epidemiológico, pero deberían ser evaluadas en un contexto regional. Conociendo la rentabilidad del test, se podría explorar parte del espectro de la inmunidad humoral que evoca el SARS-CoV-2. El personal de salud, como población de alto compromiso y de primer impacto en la pandemia constituye el proscenio ideal. Objetivo: Evaluar el desempeño diagnóstico de prueba serológica, respuesta de anticuerpos y su correlación con características clínicas en trabajadores de salud con COVID-19 del Hospital General de Luque. Metodología: Evaluar el desempeño diagnóstico de prueba serológica, respuesta de anticuerpos y su correlación con características clínicas en trabajadores de salud con COVID-19 del Hospital General de Luque entre marzo y octubre del año 2020 mediante un estudio observacional con componente analítico realizado en 107 trabajadores de salud, seleccionados por conveniencia entre marzo y octubre del 2020, luego de haber cumplido ≥14 días desde el diagnóstico. La prueba de RT-PCR fue realizada en muestras de hisopado nasal y faríngeo. La detección de anticuerpos fue realizada en muestra de plasma, analizada por inmunoensayo de electro quimioluminiscencia Elecsys Anti-SARS-CoV-2 (<1, negativo). Se recolectaron además datos: demográficos, antropométricos, comorbilidades, historia reciente de dengue, vacunación con BCG, síntomas del estadio agudo y tiempo entre inicio de síntomas y realización del hisopado e inicio de síntomas y entrevista. Se utilizó análisis descriptivo e inferencial y se calculó la sensibilidad, especificidad, valores predictivos y exactitud del test. Resultados: Del total, 56 trabajadores tuvieron diagnóstico de COVID-19 positivo, de los cuales siete mujeres (asintomáticas) mostraron niveles < 1 al inmunoensayo para anticuerpos. De los 51 trabajadores con resultado RT-PCR negativo, 3 mostraron títulos de anticuerpos elevados. La sensibilidad del test fue de 89,3%, la especificidad de 94,1% y la exactitud 91,6%. No se observó correlación entre el nivel global de anticuerpos y la edad (r 0,09; p 0,52) así como tampoco con las comorbilidades. Finalmente, los trabajadores con síntomas persistentes, tuvieron niveles de anticuerpos. Conclusión: La prueba de detección de anticuerpos anti-N para SARS-CoV-2 tuvo un buen desempeño diagnóstico, mostrando algunos falsos negativos a la RT-PCR, confirmando su utilidad para los estudios de seroprevalencia. Los niveles de anticuerpos fueron más elevados en los trabajadores con síntomas persistentes.

ABSTRACT Background: Latent HIV-1 is a major hurdle in obtaining HIV-1 sustained virological remission (SVR). Here we explored histone deacetylation inhibition property of nicotinamide (NAM; n = 17) for the first time in comparison to a combination of methyltransferase inhibitors (MTIs; Chaetocin and BIX01294; n = 25) to reactivate latent HIV ex vivo in CD8-depleted PBMCs from antiretroviral treated aviremic individuals. Results: NAM reactivated HIV-1 from 13/17 (76.4%) samples compared to 20/25 (80.0%) using MTIs with mean viral load (VLs) of 4.32 and 3.22 log10 RNA copies/mL, respectively (p = 0.004). Mean purging time after NAM and MTIs stimulation was 5.1 and 6.75 days, respectively (p = 0.73). Viral purging in autologous cultures exhibited blunted HIV recovery with fluctuating VLs followed by a complete viral extinction when expanded in allogenic system. Electron microscopy from five supernatants revealed anomalous viral particles, with lack of complete viral genomes when characterized by ultradeep sequencing through metagenomics approach (n = 4). Conclusion: NAM alone was more potent HIV-1 activator than combination of MTIs, with potential of clinical use.

ABSTRACT Itajaí is a port city in southern Brazil with one of the highest incidence and mortality rates from AIDS in the country. The prevalence and incidence of HIV infection were investigated in 1085 of 3196 new HIV-1 infection cases evaluated in the counseling and testing center of Itajaí from January 2002 to August 2008. Recent infections were assessed using the BED(tm), and polregion sequencing was performed in 76 samples. The prevalence ranged from 3.08% to 6.17% among women and from 10.26% to 17.36% among men. A total of 17% of infections were classified as recent, with annual incidence varying from 1.6% to 4.8 per 100 patient/year among women and from 2.05% to 8.5 per 100 patient/year among men. Pol sequences were obtained from 38 randomly recent infections selected individuals: 71% were infected by subtype C, 24% B, 2% D, and 2% F1. Among 38 subjects with established infection, 76% were subtype C, and 24% B. Transmitted drug resistance was detected in 18.4% of recent infection subjects (7.8% to nucleoside analog reverse-transcriptase inhibitors, 5.2% to non-nucleoside reverse-transcriptase inhibitors, and 5.2% protease inhibitors) and 5.2% of subjects with established infection had nucleoside analog reverse-transcriptase inhibitors resistance. The high prevalence and incidence of HIV infection in this region is unprecedented in studies involving cases evaluated in the counseling and testing centers in Brazil.

The aims of this study were to compare the automated AxSYM avidity assay index with the BED capture enzyme immunoassay test and to calculate the HIV-1 incidence using the BED capture enzyme immunoassay and AxSYM avidity assay index algorithms within a population seeking the Voluntary Counselling and Testing Centres in two municipalities in the Metropolitan Region of Recife, Northeast of Brazil. An analysis was conducted in 365 samples that tested positive for HIV infection from frozen serum collected during the period 2006-2009. There was a similar proportion of males and females; most patients were heterosexual (86%) with a median age of 29 years. Of the 365 samples, 102 (28%) and 66 (18.1%) were identified as recent infections by BED capture enzyme immunoassay and AxSYM avidity assay index, respectively. The HIV-1 total incidence in the BED capture enzyme immunoassay and AxSYM avidity assay index algorithms were: 0.79 (95% CI: 0.60-0.98) and 0.34 (95% CI: -0.04 to 0.72), respectively. Incidence was higher among men. There was good agreement between the tests, with a kappa of 0.654 and a specificity of 95.8%. AxSYM avidity assay index may be helpful in improving the quality of the estimates of recent HIV infection and incidence, particularly when used in a combined algorithm with BED capture enzyme immunoassay.

Determining the prevalence and type of antiretroviral (ARV) resistance among ARV-naïve individuals is important to assess the potential responses of these individuals to first-line regimens. The prevalence of primary resistance and the occurrence of recent infections among individuals with human immunodeficiency virus (HIV)/acquired immune deficiency syndrome (AIDS) were identified among recently diagnosed patients at five sexually transmitted disease/AIDS testing and counselling centres in the metropolitan region of Recife (RMR), Pernambuco, Brazil, between 2007-2009. One-hundred and eight samples were analysed using the Calypte® BED assay. Males predominated (56%), as did patients aged 31-50 years. Twenty-three percent presented evidence of a recent HIV infection. The median CD4+ T lymphocyte count was 408 cells/mm³ and the median viral load was 3.683 copies/mL. The prevalence of primary resistance was 4.6% (confidence interval 95% = 1-8.2%) based on criteria that excluded common polymorphisms in accordance with the surveillance drug resistance mutation criteria. The prevalence of resistance to non-nucleoside reverse transcriptase, nucleoside/nucleotide reverse transcriptase and protease inhibitors were 3.8%, 1.5% and 0.8%, respectively. Fifty-seven percent of strains were from clade B, 37.7% were clade F and 3.1% were clade C; there were no statistically significant differences with respect to resistance between clades. Recent infection tended to be more common in men (p = 0.06) and in municipalities in the south of the RMR (Jaboatão dos Guararapes and Cabo de Santo Agostinho) (p = 0.046). The high prevalence of recent infection and the high prevalence of non-B strains in this poor Brazilian region merit further attention.

Treatment of HIV-1 infection with highly active antiretroviral therapy has led to sustained viral suppression in the plasma in a large number of children. However, studies have suggested that the integrated provirus in resting CD4+ T lymphocytes could be a source of reactivatable virus and maintain drug-resistant virus. We evaluated the resistance-related mutations in children receiving antiretroviral therapy with prolonged viral suppression. Thirty-two peripheral blood mononuclear cell samples from 16 children with viral loads that had been below detection limits for at least 12 months were obtained at two different time points and the DNAs sequenced. The median CD4 cell count was 1,016 cells/mm³ (347-2,588) and 938 cells/mm³ (440-3,038) at the first and second time points, respectively. The median follow-up time was 15 months (9-27). Six (37.5%) and seven (43.75%) of the 16 patients showed at least one NRTI-associated mutation in the first and second samples, respectively. Two out of 16 (12.5%) had an NNRTI-associated mutation at the first time point and three out of 16 (18.75%) at the second. In addition, 14 out of 16 (87.5%) had at least one PI-associated mutation at both time points. Despite plasma HIV-1 RNA suppression for at least 12 months, resistance-related mutations from previous antiretroviral failures could still be detected in archival virus. Furthermore, viral evolution occurred at the reverse transcriptase region in spite of viral suppression to levels below 400 copies/mL. Persistence of archival resistant virus may be relevant when considering future treatment options.

OBJECTIVE: To investigates how the use of HIV-1 resistance tests influences physician decision-making. METHODS: Ten experienced reference physicians from the Brazilian Network for Drug Resistance each received ten patients' case histories. The selected patients had experienced at least two virological failures. First, reference physicians were asked to empirically select a new regimen for each patient. Second, after genotype report (ViroSeq 2.6) was provided, and physicians were again asked to select a new regimen considering this additional information. Finally, they were asked to select a regimen after receiving a virtual phenotype result (vircoTYPE 3.9.00). RESULTS: In 79% of the cases, physicians changed their empirical choice of regimen after receiving the genotype report, resulting in an increase in the mean number of active drugs from 1.8 to 2.2 (p = 0.0003), while the average number of drugs/regimen remained at 4.0. After receipt of the virtual phenotype report, additional changes were made in 75% of the patient cases, resulting in an increase in the number of active drugs to 2.8 (p < 0.0001), while the average number of drugs/regimen remained at 4.0. After receipt of the genotype report, 48% of the changes were in NRTIs, 29% were in NNRTIs and 60% were in PIs; after consideration of the virtual phenotype, 61%, 10% and 49% of the changes, respectively, were in these categories of drugs. Fourteen percent of the physicians rated the genotype report as "extremely useful", whereas 34% rated the subsequent virtual phenotype report as "extremely useful" (p = 0.0003). CONCLUSIONS: Resistance testing has a significant impact on physicians' choices of antiretroviral salvage therapies, and it promotes the selection of more active drugs

The network of HIV counseling and testing centers in São Paulo, Brazil is a major source of data used to build epidemiological profiles of the client population. We examined HIV-1 incidence from November 2000 to April 2001, comparing epidemiological and socio-behavioral data of recently-infected individuals with those with long-standing infection. A less sensitive ELISA was employed to identify recent infection. The overall incidence of HIV-1 infection was 0.53/100/year (95% CI: 0.31-0.85/100/year): 0.77/100/year for males (95% CI: 0.42-1.27/100/year) and 0.22/100/ year (95% CI: 0.05-0.59/100/year) for females. Overall HIV-1 prevalence was 3.2% (95% CI: 2.8-3.7%), being 4.0% among males (95% CI: 3.3-4.7%) and 2.1% among females (95% CI: 1.6-2.8%). Recent infections accounted for 15% of the total (95% CI: 10.2-20.8%). Recent infection correlated with being younger and male (p = 0.019). Therefore, recent infection was more common among younger males and older females.

We characterized the virologic failure after an initially successful 48-week course of antiretroviral therapy among HIV/AIDS patients in a retrospective cohort study involving patients from Santos, Brazil. Patients with plasma HIV RNA below 500 copies/mL for 48 weeks were included. Variables analyzed included gender, age, level of education, marital status, mode of HIV acquisition, viral load, and CD4 cell count upon admission. There were 4,909 patients registered with the clinic, of which 669 patients met all the inclusion criteria (41.6% female and 58.4% male). Only 27.5% of the patients maintained undetectable viral loads during up to one year of follow-up. After 48 weeks, virologic failure occurred earlier in females and in patients first treated with an antiretroviral regimen other than highly active antiretroviral therapy. Patients who were married or had a steady partner experienced virologic failure later than did those who were separated or widowed. The percentage of public health clinic patients who maintain undetectable viral loads for a period of over a year is much lower than that observed among patients enrolled in clinical trials. Females, individuals in unstable relationships, single individuals and widowed individuals should be given special attention in order to improve durability of viral suppression.

O objetivo deste estudo foi avaliar o perfil de resistência genotípica do HIV-1 em crianças com falha terapêutica ao tratamento anti-retroviral (HAART). Quarenta e uma crianças (idade mediana = 67 meses) em uso de HAART foram submetidas ao teste de genotipagem no momento da detecção de falha ao tratamento. Foi realizada extração de cDNA de células periféricas mononucleares e amplificação do mesmo (regiões da transcriptase reversa e protease do gene pol) através de PCR-nested. O perfil genotípico foi determinado através do seqüenciamnto de nucleotídeos. De acordo com a análise genotípica, 12/36 (33,3%) e 6/36 (16,6%) crianças apresentaram, respectivamente, resistência e possível resistência ao AZT; 5/36 (14%) e 4/36 (11,1%), respectivamente, eram resistentes e possivelmente resistentes ao ddI; 4/36 %11,1%) apresentaram resistência ao 3TC e D4T, e 3/36 (8,3%) eram resistentes ao ABC. Uma alta porcentagem de crianças (54%) apresentou mutações relacionadas à resistência aos inibidores da trancriptase reversa não-análogos de nucleosídeos. As taxas de resistência e possível resistência aos inibidores da protease foram, respectivamente: RTV (12,2%; 7,3%); APV (2,4%; 12,1%); SQV (0%; 12,1%); IDV (14,6%; 4,9%); NFV (22%; 4,9%); LPV/RTV (2,4%; 12,1%). No total, 37/41 (90%) crianças apresentaram vírus com mutações relacionadas à resistência a alguma droga, sendo que 9% delas tinham vírus resistentes às três classes de drogas anti-retrovirais disponíveis.

The aim of this study was to evaluate the genotypic resistance profiles of HIV-1 in children failing highly active antiretroviral therapy (HAART). Forty-one children (median age = 67 months) receiving HAART were submitted to genotypic testing when virological failure was detected. cDNA was extracted from PBMCs and amplified by nested PCR for the reverse transcriptase and protease regions of the pol gene. Drug resistance genotypes were determined from DNA sequencing. According to the genotypic analysis, 12/36 (33.3%) and 6/36 (16.6%) children showed resistance and possible resistance, respectively, to ZDV; 5/36 (14%) and 4/36 (11.1%), respectively, showed resistance and possible resistance to ddI; 4/36 (11.1%) showed resistance to 3TC and D4T; and 3/36 (8.3%) showed resistance to Abacavir. A high percentage (54%) of children exhibited mutations conferring resistance to NNRTI class drugs. Respective rates of resistance and possible resistance to PIs were: RTV (12.2%, 7.3%); APV (2.4%, 12.1%); SQV(0%, 12.1%); IDV (14.6%, 4.9%), NFV (22%, 4.9%), LPV/RTV (2.4%, 12.1%). Overall, 37/41 (90%) children exhibited virus with mutations related to drug resistance, while 9% exhibited resistance to all three antiretroviral drug classes.

Entry of human immunodeficiency type 1 virus (HIV-1) into target cells requires both CD4and one of the chemokine receptors. Viruses predominantly use one, or occasionally both, of the major co-receptors CCR5 and CXCR4, although other receptors, including CCR2B and CCR3, function as minor co-receptors. A 32-nucleotide deletion (delta32) within the beta-chemokine receptor 5 gene (CCR5) has been described in subjects who remain uninfected despite extensive exposition to HIV-1. The heterozygous genotype delays disease progression. This allele is common among Caucasians, but has not been found in people of African or Asian ancestry. A more common transition involving a valine to isoleucine switch in transmembrane domain I of CCR2B (64I), with unknown functional consequences, was found to delay disease progression but not to reduce infection risk. As the Brazilian population consists of a mixture of several ethnic groups, we decided to examine the genotype frequency of these polymorphisms in this country. There were 11.5% CCR5 heterozygotes among the HIV-1 infected population and 12.5% among uninfected individuals, similar to data from North America and Western Europe. The prevalence of CCR2-64I homozygotes and heterozygotes was 0.06 and 15.2%, respectively, also similar to what is known for North America and Western Europe.

Brazil was the first country to provide unrestricted, cost-free access to antiretroviral (ARV) medicine for AIDS treatment. However, there is little data about the benefits of such a policy for these patients. We evaluated the duration of benefit obtained with the introduction of ARVs, defined as the durability of the first ARV regiment. We reviewed the medical charts of patients attended from 1996-2000, at the outpatient clinics of the Federal University of São Paulo, Brazil. A total of 120 drug-naive HIV-1 infected patients were eligible to participate in the study. About half of the individuals (53%) presented with disease symptoms; 59% of them had CD4 count below 200 cells/mm³. Mean estimated duration of the benefit of therapy was 14.1 months. The most used regimen in this cohort was Zidovudine/3TC/Indinavir (26%), followed by Zidovudine/DDI (17%), and Zidovudine/3TC/Nelfinavir (13%). The most frequent cause of interruption of therapy was gastrointestinal intolerance. Use of treatment regimens with three drugs was more effective than with two drugs, but only for patients with CD4<200 cells/mm³ or CV>100,000 copies RNA/mL. However, the use of triple therapy was associated with a significantly higher probability of reaching maximum viral suppression, during a longer period (p<0.05).The patients enrolled in the study benefitted from therapy for a limited time, after the introduction of double or triple antiretroviral therapy. The incidence of adverse events was significantly associated with loss of the benefits provided by the initial therapeutic regimen.