Use of synthetic oligonucleotides for determination of HTLV-1 proviral load by real-time PCR: a helpful alternative approach in the clinical management.

AIMS: To evaluate the potential use of synthetic oligonucleotides as a standard curve for proviral load (PVL) of human T-cell leukaemia virus type 1 (HTLV-1) quantification in peripheral blood mononuclear cells (PBMC) of HTLV-1-infected individuals by quantitative real-time polymerase chain reaction (qPCR) analysis. METHODS AND RESULTS: Synthetic oligonucleotides based on HTLV-1 genome were customized to use as a standard curve. Twelve anti-HTLV-1-positive samples with known HTLV-1 PVL, previously quantified by qPCR assay using TARL-2 cells as a conventional standard curve, were submitted to the new protocol. The proviral quantification levels had a high concordance with qPCR results using a conventional standard curve. The results demonstrate that the conventional standard curve can be replaced by a synthetic standard curve due to its ability to quantification based on the linearity and qPCR efficiency and similar results with a validated qPCR assay using a conventional standard curve. CONCLUSIONS: Synthetic oligonucleotides standard curves could be a very useful tool on HTLV-1 diagnosis and absolute HTLV-1 PVL quantification. SIGNIFICANCE AND IMPACT OF THE STUDY: HTLV-1 PVL determination using synthetic oligonucleotides standard curve by qPCR could be a helpful alternative for the laboratories that monitor infected patients as an important prognostic factor in HTLV-1-associated diseases progression. Also, it can decrease costs and overcome the biological limitations of the plasmid curve.

Detection of East/Central/South African genotype Chikungunya virus during an outbreak in a southeastern state of Brazil.

Chikungunya virus (CHIKV) is an arbovirus transmitted by Aedes mosquitoes that was first identified in Brazil in 2014. It causes a febrile illness characterised by severe arthralgia and rash. Our group investigated a suspected CHIKV outbreak in Governador Valadares, state of Minas Gerais, Brazil and from 25 acute-phase patients, 10 had qRT-PCR positive sera samples and had E1 partial sequence amplified and Sanger sequenced. Samples were identified as East/Central/South African (ECSA) genotype by phylogenetic analysis and clustered with CHIKV sequences isolated in the neighbour state of Bahia. Our findings confirm previous predictions that ECSA genotype would spread through northeast and southeast of Brazil.

Treatment of visceral leishmaniasis in children in the Central-West Region of Brazil.

BACKGROUND: Detailed reports on the treatment of visceral leishmaniasis (VL) are scarce, particularly with regard to the utilization of antimoniate of N-methylglucamine. The aim of this study was to analyze the treatment of children admitted to a reference hospital, focusing in particular on the use of antimoniate of N-methylglucamine and on the supportive measures adopted. MATERIALS AND METHODS: Medical records of children treated for VL from January 1998 to February 2005 in the Hospital of the University of Mato Grosso do Sul, Central-West Region of Brazil, were reviewed retrospectively. RESULTS: A total of 116 children were treated, and 111 received antimoniate as the first therapeutic choice. The drug was highly efficient (96.9%) in patients with no signs of gravity on admission, in cases presenting warning signs of potential evolution to gravity, and even in some severely ill children. The most common adverse effects were increases in transaminase (22.5%) and amylase (17.5%) levels, and generally reversible electrocardiogram changes (18%). Some problems were detected during the treatment, such as inadequate prescription (causing an under- or overdose) or inappropriate change to a second-line scheme. Of the 116 children, 80% were given antibiotics, 71.5% needed a transfusion of red blood cells, 10.3% required a transfusion of platelets, fresh frozen plasma was given to 4.3%, albumin was administered in 3.4, and 8.6% needed intensive care support. The mortality rate was about 2.6%. CONCLUSION: Antimoniate of N-methylglucamine remains highly efficient and well tolerated in pediatric patients, which allows its utilization as a first-line therapy in Brazilian children until a better drug for widespread use becomes available; however, it should be used with caution, and special attention is required during its prescription and for the management of adverse effects. The low mortality rate obtained confirms that, in addition, successful treatment demands the correction of serious anemia and thrombocytopenia, the vigorous use of antibiotics to fight intercurrent bacterial infections, and sometimes the availability of intensive care units to treat more severe patients.

Tuberculin skin testing in HIV-infected patients in Campo Grande, Mato Grosso do Sul State, Brazil

A cross-sectional study on HIV/AIDS was carried out in 108 outpatients from the university hospital of the Federal University of Mato Grosso do Sul, Campo Grande, Brazil, from July to December 2008, to investigate latent tuberculosis infection using the tuberculin skin test (TST). The prevalence of positive results was 13.9 percent. The CD4+ T cell count (p = 0.091) and the diagnosis time (p = 0.010) were statistically significant when compared with TST positivity. In the cohort of HIV/AIDS patients who had latent tuberculosis infection, the median diagnosis time was eight years. Undetectable viral load presented significant association (p = 0.046) with tuberculosis infection. The fact that numerous individuals with HIV/AIDS infection presented a negative reaction to the tuberculin skin test is probably related to alterations in the cellular immune response induced by HIV infection. The tuberculin test is a useful tool for the detection of latent tuberculosis infection and should be performed in all HIV/AIDS individuals at the time of the diagnosis and on a yearly basis, if negative. Both the early identification of the tuberculosis infection by the tuberculin skin test at the moment of immunological restoration and chemoprophylaxis in infected individuals are mechanisms to control HIV/AIDS and tuberculosis coinfection.

[Chagas' disease in a urban population of the health district of Rio Verde, Mato Grosso do Sul State, Brazil]. / Doença de Chagas na população urbana do distrito sanitário de Rio Verde, Mato Grosso do Sul, Brasil.

In order to evaluate some aspects of the Chagas' disease on urban population of the 12 municipalities of the sanitarian district of Rio Verde, Mato Grosso do Sul state, on 1998 we carried out a serological survey investigating anti-T. cruzi IgG by indirect immunofluorescence on blood samples collected on filter paper of 14,709 resident persons, confirmed by indirect hemagglutination and ELISA in serum. The parasitaemia was evaluated by indirect xenodiagnosis on 134 chagasic patients and the morbidity evaluated by anamnesis, physical exam and electrocardiogram in 191 pairs of chagasic/non-chagasic patients. The seropositivity was 1.83% (0.93% in autochtonous, 5.01% in alochtonous), xenodiagnosis was positive in 17.2% of the individuals (12.3% in autochtonous, 20.8% in alochtonous) and 24.6% of the seropositive patients presented chronic chagasic cardiopathy (19.1% in autochtonous, 27.8 in alochtonous). The analysis of the dates showed that alochtonous population is the main responsible for presence of the infection and morbidity of Chagas disease in the studied area.

Dengue epidemic in the State of Rio Grande do Norte, Brazil, in 1997.

During 1997 a large dengue epidemic occurred in Rio Grande do Norte, a State in north-east Brazil. The co-circulation of dengue virus type 1 and dengue virus type 2 was demonstrated by virus isolation in Aedes albopictus clone C6/36 cell-line and by reverse transcriptase-polymerase chain reaction (RT-PCR). IgM capture enzyme-linked immunosorbent assay confirmed 52.3% of the 8105 studied cases and dengue antigen was demonstrated by immunohistochemical reaction on hepatocytes from 2 out of 5 fatal cases studied. Individual risk factors for development of dengue haemorrhagic fever/dengue shock syndrome, such as hypertension, diabetes mellitus and bronchial asthma, are discussed.

Retrospective study on dengue in Fortaleza, state of Ceará, Brazil.

A retrospective serologic study was carried out in Fortaleza, State of Ceará, Brazil, in order to detect the dengue virus activity before recognizing the epidemic of 1994. Mac-Elisa was performed by using a mixture of specific DEN-1 and DEN-2 antigens on serum samples from the Emilio Ribas Laboratory collection. Samples were obtained from 1,224 patients with exanthematic febrile disease and negative serological results for rubella. All specimens were taken during November 1993 to May 1994. The results confirmed dengue infections in Fortaleza by November 1993, approximately six months before the beginning of the epidemic, proving how misleading diagnosis of dengue infection are still troublesome, in spite of the strong dengue activity in Ceará. The authors stress the urgent necessity to implement the active surveillance system in order to prevent another extensive dengue fever epidemics in the state. Epidemiological background of the dengue activity in the State of Ceará is described.

Retrospective study on dengue in Fortaleza, State of Ceará, Brazil

A retrospective serologic study was carried out in Fortaleza, State of Ceará, Brazil, in order to detect the dengue virus activity before recognizing the epidemic of 1994. Mac-Elisa was performed by using a mixture of specific DEN-1 and DEN-2 antigens on serum samples from the Emilio Ribas Laboratory collection. Samples were obtained from 1,224 patients with exanthematic febrile disease and negative serological results for rubella. All specimens were taken during November 1993 to May 1994. The results confirmed dengue infections in Fortaleza by November 1993, approximately six months before the beginning of the epidemic, proving how misleading diagnosis of dengue infection are still troublesome, in spite of the strong dengue activity in Ceará. The authors stress the urgent necessity to implement the active surveillance system in order to prevent another extensive fever epidemics in the state. Epidemiological background of the dengue activity in the State of Ceará is also described.

Dengue infection in Paracambi, State of Rio de Janeiro, 1990-1995.

A seroepidemiological survey was carried out during 1994 in the municipality of Paracambi, state of Rio de Janeiro. Haemagglutination inhibition test positivity was detected in 145 out of 370 (39.2%) schoolchildren. The frequency of positive test by sex was 53.8% (78/145) female and 46.2% (67/145) male. Distribution by age showed the increasing of antibody positivity in older children. Strains of dengue virus type 1 and dengue virus type 2 were isolated before (1990) showing the co-circulation of both serotypes in that area. The house index infestation of Aedes albopictus and Aedes aegypti has been determined.

PIP: Dengue virus type 1 (DEN-1) was first isolated in Rio de Janeiro, Brazil, in 1986. In 1990, another serotype, DEN-2, was introduced. Since then, both strains have co-circulated in the state, especially during the rainy season. This study assessed the prevalence of dengue virus infection in Paracambi--a periurban municipality of Rio de Janeiro with environmental conditions that allow natural breeding of Aedes aegypti and A. albopictus. In 1990-91, 1209 dengue cases were reported in the municipality and laboratory studies confirmed 21 cases of dengue hemorrhagic fever. Serologic samples collected in 1994 from 370 randomly selected schoolchildren 4-16 years of age revealed hemagglutination inhibition test antibody titers equal to or greater than 1/20 to DEN-1 and/or DEN-2 in 145 children (39.2%). The geometric averages of antibody titers were 1/75 for DEN-1 and 1/30 for DEN-2. Entomological surveillance revealed declines in the house index of A. aegypti to 2.0% or under by 1995, while the index of A. albopictus remained at 0.6-7.4%. The decline in A. aegypti was accompanied by a dramatic reduction in dengue virus cases in 1991-94. However, as a result of the interruption of vectors measures control in 1994, the number of dengue cases has shown an upward trend since 1995.

Dengue infection in Paracambi, State of Rio de Janeiro, 1990-1995

Um inquérito soroepidemiológico foi realizado em uma amostra de escolares, em 1994, no município de Paracambi, Estado do Rio de Janeiro. Positividade do teste de Inibição da Hemaglutinação foi detectada em 39,2% (145/370) dos escolares pesquisados. A freqüência de positividade foi de 53,8% (78/145) para o sexo feminino e de 46,2% (67/145) para o sexo masculino. A distribuição por faixa etária mostrou uma positividade crescente com o aumento da idade. Cepas do vírus dengue tipo 1 e vírus dengue tipo 2 foram isoladas anteriormente (1990), mostrando a co-circulação de ambos os sorotipos na área. Os índices de infestação predial pelo Aedes aegypti e pelo Aedes albopictus foram determinados.

A seroepidemiological survey was carried out during 1994 in the municipality of Paracambi, state of Rio de Janeiro. Haemagglutination inhibition test positivity was detected in 145 out of 370 (39.2%) schoolchildren. The frequency of positive test by sex was 53.8% (78/145) female and 46.2% (67/145) male. Distribution by age showed the increasing of antibody positivity in older children. Strains of dengue virus type 1 and dengue virus type 2 were isolated before (1990) showing the co-circulation of both serotypes in that area. The house index infestation of Aedes albopictus and Aedes aegypti has been determined.

[Morbidity of Chagas disease in areas of Sertão da Paraiba and Caatinga do Piauí]. / Morbidade da doença de Chagas em áreas do Sertão da Paraíba e da Caatinga do Piauí.

A clinical and electrocardiographic case control study was carried out with 186 pairs of persons with positive and negative serology for T. cruzi infection from the Sertão Paraíba and in 200 seropositive cases from the region of Caatinga in the State of Piauí, North-eastern Brazil. The predominant clinical manifestations in seropositive cases in both areas were: palpitations, dyspnea on effort, precordial pain, dysphagia, odynophagia, pyrosis and intestinal constipation. The EKG abnormalities rates suggestive of chronic chagasic cardiopathy were respectively in Paraíba and Piauí: AV block 3.8% and 2%, RBBB III 6.4% and 7%, RBBB III+ LAB 10.7% and 10.5%, and multifocal extrasystoles 2.7% and 3%. Xenodiagnosis in a sample of 54 seropositive individuals in the Sert-ao of Para-iba and in 120 in the Caatinga of Piauí was revealed 13% and 34% positive; PCR tests in a sample of 47 seropositives in Paraíba and 101 in Piauí revealed positives in 44.6% and 59.5% respectively. Blood culture in LIT media of 101 seropositive cases from the Caatinga of Piauí was positive for T. cruzi in 25.7%. A triatomine survey carried out in a sample of 132 domiciles and peridomiciles in the Sertão of Paraíba and in a sample of 159 in the Caatinga of Piauí showed the following results: In Paraíba, 16 specimens of T. brasiliensis, not infected with T. cruzi, were captured. In Piauí, 750 triatomines were captured, of these 625 were examined: 49 were T. pseudomaculata, not infected with T. cruzi (19 in peridomiciles and 30 in the domiciles), and 576 were T. brasiliensis (371 in the domiciles and 205 in the peridomiciles) and of this latter specie 32 (5.5%) were infected with T. cruzi (31 in the domiciles and one in the peridomicile).

Secondary dengue infection in schoolchildren in a dengue endemic area in the state of Rio de Janeiro, Brazil.

A seroepidemiologic survey was carried out in schoolchildren from public schools of the Niterói municipality, state of Rio de Janeiro, Brazil, after a period of sequential epidemics by dengue virus type 1 and 2 (DEN-1 and DEN-2). 450 blood samples were obtained by fingertip puncture and collected on filter paper discs. The hemagglutination inhibition (HAI) test was carried out using DEN-1 and DEN-2 antigens. HAI titres were demonstrated in 66% (297/450) of the sera and the geometric means of the titres were 1/182 and 1/71 for DEN-1 and DEN-2, respectively. Secondary infections were observed in 61% (181/297) of positive cases. Among these, 75% (135/181) were under fifteen years old. No dengue haemorrhagic fever (DHF) was reported in these children. Asymptomatic or oligosymptomatic infections were detected in 56% of the studied population. The absolute and relative frequencies of positive tests by age group and sex did not evidence statistically significant difference. The number of individuals infected probably produced a immunologic barrier responsible for the non occurrence of dengue epidemic in the latter years.

[Development of chronic human chagas cardiopathy in the hinterland of the Paraíba State, Brazil, in a 4.5 year period]. / Evolução da cardiopatia chagásica crônica humana no Sertão do Estado da Paraíba, Brasil, no período de 4,5 anos.

Two sectional studies about chronic Chagas' disease were performed at a 4.5 year interval, involving the urban populations of Agua Branca, Catingueira, Emas, Imaculada, Mãe D'Agua, Olho D'Agua, Piancó and São José de Caiana counties in the Sertão region of the State of Paraíba. The evolution of heart disease was evaluated in 125 matched pairs of chronic chagasic and non-chagasic patients of the same sex, age and county of origin through electrocardiograms (ECG) at rest. The following evaluation criteria were considered: unchanged - no change in the original ECG pattern; progressive-changes in ECG pattern from normal to abnormal or deterioration of ECG abnormalities; and ECG normalization. In chagasic patients evolution of the disease was unchanged in 101 (80.8%), progressive in 13 (10.4%) and ECG normalization in 11 (8.8%), while those observed in non-chagasic patients were respectively values 117 (93.6%), 6 (4.8%) and 2%) patients. Findings indicate that the share of Chagas-linked etiological component affecting the development of chronic Chagas cardiopathy was 5.9% with an estimated annual average of 1.3%. There was no significant difference in the frequency of progressive disease between the sexes either in the chagasic or in the non-chagasic group. On the other hand, progression of heart disease occurred earlier among chagasic patients. Lethality caused by heart disease was 1.6% (2 cases) in the chagasic group and zero in the non-chagasic group during the period under survey. These morbidity and mortality rates were significantly lower than those found in endemic areas such as Virgem da Lapa and Pains-Iguatama in the State of Minas Gerais and can probably be attributed to the weaker pathogenic it of human infection by Trypanosoma cruzi in the Paraíba hinterland.