Serological detection of Plasmodium vivax malaria using recombinant proteins corresponding to the 19-kDa C-terminal region of the merozoite surface protein-1

Abstract Background Serological tests to detect antibodies specific to Plasmodium vivax could be a valuable tool for epidemiological studies, for screening blood donors in areas where the malaria is not endemic and for diagnosis of infected individuals. Because P. vivax cannot be easily obtained in vitro, ELISA assays using total or semi-purified antigens are rarely used. Based on this limitation, we tested whether recombinant proteins representing the 19 kDa C-terminal region of the merozoite surface protein-1 of P. vivax (MSP119) could be useful for serological detection of malaria infection. Methods Three purified recombinant proteins produced in Escherichia coli (GST-MSP119, His6-MSP119 and His6-MSP119-PADRE) and one in Pichia pastoris (yMSP119-PADRE) were compared for their ability to bind to IgG antibodies of individuals with patent P. vivax infection. The method was tested with 200 serum samples collected from individuals living in the north of Brazil in areas endemic for malaria, 53 serum samples from individuals exposed to Plasmodium falciparum infection and 177 serum samples from individuals never exposed to malaria. Results Overall, the sensitivity of the ELISA assessed with sera from naturally infected individuals was 95%. The proportion of serum samples that reacted with recombinant proteins GST-MSP119, His6-MSP119, His6-MSP119-PADRE and yMSP119-PADRE was 90%, 93.5%, 93.5% and 93.5%, respectively. The specificity values of the ELISA determined with sera from healthy individuals and from individuals with other infectious diseases were 98.3% (GST-MSP119), 97.7% (His6-MSP119 and His6-MSP119-PADRE) or 100% (yMSP119-PADRE). Conclusions Our study demonstrated that for the Brazilian population, an ELISA using a recombinant protein of the MSP119 can be used as the basis for the development of a valuable serological assay for the detection of P. vivax malaria.

Factors associated with failure of clinical screening among blood donors who have altered serological results in the Centro Regional de Hemoterapia de Ribeirão Preto

OBJECTIVE: This study aimed to investigate the frequency of positive results for hepatitis B and C, HIV and syphilis in blood donations at the Centro Regional de Hemoterapia de Ribeirão Preto, to describe donors with positive results according to some demographic and socioeconomic variables, to identify risk factors associated to these donors and the reasons that they were not detected during clinical screening. METHODS: A descriptive study was performed between July 1st 2005 and July 31st 2006 by interviewing 106 donors after medical consultations where they were informed of positive results for hepatitis B, hepatitis C, HIV or syphilis. RESULTS: There was a predominance of first-time donors, males, under 50-year olds, married individuals, from Ribeirão Preto, with elementary education, low economic status and of people who donated at the request of friends or relatives. Hepatitis C was the most frequently detected infection (56.6%), followed by hepatitis B (20.7%), HIV (12.3%) and syphilis(10.4%). About 40% of donors had omitted risk factors for different reasons: because they trusted the results of serological tests, did not feel comfortable about talking of risk factors or did not consider them relevant. Other justifications were the duration of the interview, the interviewer was unskilled, embarrassment and doubts about confidentiality. CONCLUSION: The results indicate the need for changes in the approach to clinical screening and a review of methods to attract and guide potential donors.

Fluorescence polarization assay, competitive enzyme-linked immunosorbent assay (ELISA-C) and indirect ELISA for the diagnosis of brucellosis in buffaloes (Bubalus bubalis)

The objective of the present study was to compare the performance of three serological tests for diagnosis of Brucella abortus infections in buffaloes (Bubalus bubalis). Serum samples collected from 696 adult females were submitted to the competitive enzyme-linked immunosorbent assay (ELISAC), (I-ELISA), fluorescence polarization test (FPA), 2-mercaptoethanol test (2-ME) and complement fixation test (CFT). The gold standard was the combination of CFT and 2-ME, considering as positive the reactors in both CFT and 2-ME, and as negative those non-reactors. ROC analyses were done for C-ELISA, I-ELISA and FPA and the Kappa agreement index were also calculated. The best combinations of relative sensitivity (SEr) and relative specificity (SPr) and Kappa were given by C-ELISA (96.9%, 99.1%, and 0.932, respectively) and FPA (92.2%, 97.6 and 0.836, respectively). The C-ELISA and FPA were the most promising confirmatory tests for the serological diagnosis of brucellosis in buffaloes, and for these tests, cut-off values for buffaloes may be the same as those used for bovines.

HTLV-1/2 seroprevalence and coinfection rate in Brazilian first-time blood donors: an 11-year follow-up

The seroprevalence and geographic distribution of HTLV-1/2 among blood donors are extremely important to transfusion services. We evaluated the seroprevalence of HTLV-1/2 infection among first-time blood donor candidates in Ribeirão Preto city and region. From January 2000 to December 2010, 1,038,489 blood donations were obtained and 301,470 were first-time blood donations. All samples were screened with serological tests for HTLV-1/2 using enzyme immunoassay (EIA). In addition, the frequency of coinfection with hepatitis B virus (HBV), hepatitis C virus (HCV), human immunodeficiency virus (HIV), Chagas disease (CD) and syphilis was also determined. In-house PCR was used as confirmatory test for HTLV-1/2. A total of 296 (0.1%) first-time donors were serologically reactive for HTLV-1/2. Confirmatory PCR of 63 samples showed that 28 were HTLV-1 positive, 13 HTLV-2 positive, 19 negative and three indeterminate. Regarding HTLV coinfection rates, the most prevalent was with HBV (51.3%) and HCV (35.9%), but coinfection with HIV, CD and syphilis was also detected. The real number of HTLV-infected individual and coinfection rate in the population is underestimated and epidemiological studies like ours are very informative.

Prevalência de infecção pelo T. cruzi em doadores de sangue nos hemocentros coordenadores do Brasil em 2007

Objetivo: conhecer a prevalência de Trypanosoma cruzi entre os doadores de sangue, analisar a organização da rede de hemoterapia e as normas de segurança do sangue para transfusão no Brasil no ano de 2007. Métodos: estudo descritivo utilizando-se os Regulamentos Técnicos para hemoterapia definidos pela Agência Nacional de Vigilância Sanitária (Anvisa) e o questionário aplicado aos Hemocentros Coordenadores (HC) do Brasil. Resultados: responderam 84% dos hemocentros, onde doaram sangue 3.251.361 indivíduos, sendo 1.192 (0,04%) excluídos na triagem clínica por risco presumido para doença de Chagas; foram realizadas 2.726.668 sorologias; 5.432 (0,20%) foram reagentes para Trypanosoma cruzi e 3.065 (0,11%) inconclusivas. Conclusão: a falta de resposta dos Estados de Pernambuco, Bahia, Paraíba, Goiás e Rondônia constituiu limitação ao estudo; entretanto, os resultados obtidos sugerem baixa prevalência de Trypanosoma cruzi entre os doadores de sangue e indicam o cumprimento dos procedimentos para a segurança do sangue definidos pela Anvisa.

Precipitating factors of porphyria cutanea tarda in Brazil with emphasis on hemochromatosis gene (HFE) mutations. Study of 60 patients

BACKGROUND: Porphyria cutanea tarda is the most common form of porphyria, characterized by the decreased activity of the uroporphyrinogen decarboxylase enzyme. Several reports associated HFE gene mutations of hereditary hemochromatosis with porphyria cutanea tarda worldwide, although up to date only one study has been conducted in Brazil. OBJECTIVES: Investigation of porphyria cutanea tarda association with C282Y and H63D mutations in the HFE gene. Identification of precipitating factors (hepatitis C, HIV, alcoholism and estrogen) and their link with HFE mutations. METHODS: An ambispective study of 60 patients with PCT was conducted during the period from 2003 to 2012. Serological tests for hepatitis C and HIV were performed and histories of alcohol abuse and estrogen intake were investigated. HFE mutations were identified with real-time PCR. RESULTS: Porphyria cutanea tarda predominated in males and alcohol abuse was the main precipitating factor. Estrogen intake was the sole precipitating factor present in 25% of female patients. Hepatitis C was present in 41.7%. All HIV-positive patients (15.3%) had a history of alcohol abuse. Allele frequency for HFE mutations, i.e., C282Y (p = 0.0001) and H63D (p = 0.0004), were significantly higher in porphyria cutanea tarda patients, compared to control group. HFE mutations had no association with the other precipitating factors. CONCLUSIONS: Alcohol abuse, hepatitis C and estrogen intake are prevalent precipitating factors in our porphyria cutanea tarda population; however, hemochromatosis in itself can also contribute to the outbreak of porphyria cutanea tarda, which makes the research for HFE mutations necessary in these patients

Surveillance using serological and molecular methods for the detection of infectious agents in captive Brazilian neotropic and exotic felids

The aim of the current study was to investigate the exposure of captive wild felids to various infectious pathogens using serological and molecular methods. One hundred and fifty-nine neotropic felids and 51 exotic felids from 28 captive settings in Brazil were tested. While antibodies against Feline parvovirus and Feline coronavirus (FCoV), Feline calicivirus and Bartonella spp. were frequently detected by serologic tests, antibodies against Felid herpesvirus 1 or infection with hemotropic mycoplasmas were less prevalent. Serologic evidence of exposure to Ehrlichia spp., Feline immunodeficiency virus, and Feline leukemia virus (FeLV) was detected rarely, and infections with FeLV, Ehrlichia spp., and Cytauxzoon spp. were found infrequently. The detected Bartonella sequence was molecularly similar to B. koehlerae and B. henselae; for Cytauxzoon, the sequence resembled those from domestic cats. No Anaplasma phagocytophilum and Theileria spp. infections were detected. The positive test results varied significantly among different facilities and species. Additionally, FCoV seropositivity was more prevalent in captivity than in free-ranging populations. Results suggest that testing is appropriate prior to relocation of felids.

Evaluation of rapid tests for human immunodeficiency virus as a tool to detect recent seroconversion

The identification of recent HIV infection is important for epidemiological studies and to monitor the epidemic. The objective of this study was to evaluate two rapid tests that are easily available to the Brazilian scientific community for using as markers of recent HIV infection. The Rapid Test - HIV-1/2 Bio-Manguinhos (Bio-Manguinhos/Fiocruz, Brazil) and the Rapid Check HIV 1&2 (NDI-UFES, Center for Infectious Diseases, Universidade Federal do Espirito Santo) were tested, using 489 samples with HIV positive serology, from blood donors, previously classified as recent or long-term infection by serological testing algorithm for recent HIV seroconversion (STARHS) or LS-HIV Vitros assay methods. The samples were diluted prior to testing (1:50 and 1:100 for the Rapid Test - HIV-1/2 Bio-Manguinhos, and 1:500 and 1:600 for the Rapid Check HIV 1&2). Negative samples were considered recent infection, whereas those showing any color intensity were associated with long-term infection. The best dilutions were 1:100 for HIV-1/2 Bio-Manguinhos test (Kappa = 0.840; overall agreement = 0.93), and 1:500 for the Rapid Check HIV 1&2 (Kappa = 0.867; overall agreement = 0.94). The results suggest that both rapid tests can be used to detect recent seroconversion. (C) 2012 Elsevier Editora Ltda. All rights reserved.

Discrepancies and consequences of indirect hemagglutination, indirect immunofluorescence and Elisa tests for the diagnosis of Chagas disease

Using the indirect hemagglutination (IH), indirect immunofluorescence (IIF) and enzyme linked immunosorbent assay (ELISA) tests for the diagnosis of Chagas disease, 4000 serum samples were examined. This study was conducted with different purposes: clinical interest, research support and parasitological monitoring of those patients with Chagas disease who were treated with heart transplantations. The tests occurred without patient selection and in accordance with the medical requests. The results showed discrepancies and brought about several questions, considering the different results that all three methods showed when considered together. What was found brought about concerns and we suggest the adoption of different measures, aiming to avoid these mismatches in the context of this disease.