Abstract: INTRODUCTION: Before 2004, the occurrence of acute Chagas disease (ACD) by oral transmission associated with food was scarcely known or investigated. Originally sporadic and circumstantial, ACD occurrences have now become frequent in the Amazon region, with recently related outbreaks spreading to several Brazilian states. These cases are associated with the consumption of açai juice by waste reservoir animals or insect vectors infected with Trypanosoma cruzi in endemic areas. Although guidelines for processing the fruit to minimize contamination through microorganisms and parasites exist, açai-based products must be assessed for quality, for which the demand for appropriate methodologies must be met. METHODS: Dilutions ranging from 5 to 1,000 T. cruzi CL Brener cells were mixed with 2mL of acai juice. Four Extraction of T. cruzi DNA methods were used on the fruit, and the cetyltrimethyl ammonium bromide (CTAB) method was selected according to JRC, 2005. RESULTS: DNA extraction by the CTAB method yielded satisfactory results with regard to purity and concentration for use in PCR. Overall, the methods employed proved that not only extraction efficiency but also high sensitivity in amplification was important. CONCLUSIONS: The method for T. cruzi detection in food is a powerful tool in the epidemiological investigation of outbreaks as it turns epidemiological evidence into supporting data that serve to confirm T. cruzi infection in the foods. It also facilitates food quality control and assessment of good manufacturing practices involving acai-based products.

The Brazilian government has approved many transgenic maize lines for commercialization and has established a threshold of 1% for food labeling, which underscores need for monitoring programs. Thirty four samples including flours and different types of nacho chips were analyzed by conventional and real-time PCR in 2011 and 2012. The events MON810, Bt11, and TC1507 were detected in most of the samples, and NK603 was present only in the samples analyzed in 2012. The authorized lines GA21, T25, and the unauthorized Bt176 were not detected. All positive samples in the qualitative tests collected in 2011 showed a transgenic content higher than 1%, and none of them was correctly labeled. Regarding the samples collected in 2012, all positive samples were quantified higher than the threshold, and 47.0% were not correctly labeled. The overall results indicated that the major genetically modified organisms detected were MON810, TC1507, Bt11, and NK603 events. Some industries that had failed to label their products in 2011 started labeling them in 2012, demonstrating compliance with the current legislation observing the consumer rights. Although these results are encouraging, it has been clearly demonstrated the need for continuous monitoring programs to ensure consumers that food products are labeled properly.

O uso de materiais de referência (MR) é uma das principais ferramentas utilizadas para garantia e controle da qualidade de laboratórios de microbiologia de alimentos. No Brasil, a RDC n.º 12/01 da Anvisa prevê como um dos parâmetros para a avaliação da qualidade de queijos a enumeração de estafilococos coagulase positiva (ECP). O grande desafio na produção de MR destinados a ensaios microbiológicos é a instabilidade natural dos micro-organismos, o que dificulta o desenvolvimento e a manutenção desses MR. O objetivo deste estudo foi produzir um MR quantitativo destinado ao ensaio de enumeração de ECP em matriz queijo. Uma amostra de queijo ultrafiltrado com contagem de ECP <10 UFC/g e número de aeróbios viáveis de 1,22 × 10³ UFC/g foi utilizada como matriz para produção do MR. A matriz foi distribuída em frascos, contaminada com a bactéria alvo em concentração específica e submetida à liofilização. Como crioprotetor, foi utilizada sacarose. O MR produzido foi considerado homogêneo e estável à temperatura < -70 ºC durante todo o período estudado (dez meses). O material apresentou estabilidade a 4, 25, 30 e 35 ºC durante quatro dias; contudo, os resultados indicam que, a 35 ºC, ocorre um decréscimo na concentração celular. A -20 ºC, o MR apresentou-se estável durante 48 dias. Conclui-se que o material apresentou todos os requisitos necessários de um MR de qualidade e poderia ser transportado aos laboratórios participantes de um ensaio de proficiência a temperaturas de até 35 ºC por até quatro dias, uma vez que os resultados indicaram a manutenção da concentração celular neste período. Esse foi o primeiro trabalho a descrever uma metodologia de produção de MR contendo ECP em matriz queijo.

The use of reference materials (RM) is one of the principal tools used for assurance and quality control in food microbiology laboratories. In Brazil, Anvisa RDC nº 12/01 specifies the enumeration of coagulase-positive staphylococcus (CPS) as one of the parameters for evaluating cheese quality. The main challenge in the production of RM destined for microbiological assays is the natural instability of the microorganisms, which makes it difficult to develop and maintain them. This study aimed to produce a quantitative RM for use in enumeration assays of CPS in cheese matrixes. A sample of an ultra-filtered cheese with a CPS count of <10 CFU/g and a total nº of viable aerobes of 1.2 × 10³ CFU/g was used as the matrix to produce the RM. The matrix was distributed in flasks, contaminated with specific concentrations of the target bacteria and submitted to freeze-drying. Sucrose was used as the cryo-protector. The RM produced was considered homogeneous and stable at < -70ºC during the entire study period (10 months). The material was shown to be stable at 4, 25, 30 and 35 ºC for 4 days, although the results indicated a decrease in cell concentration at 35 ºC. At -20 ºC the RM was stable for 48 days. It was concluded that the material showed all the necessary requirements for a quality RM and could be transported to the laboratories taking part in a proficiency test at up to 35 ºC for up to 4 days, since the results indicated maintenance of the cell concentrations during this period. This is the first study to describe a methodology for producing MR containing CPS in a cheese matrix.

A extração de DNA é uma etapa crítica na análise de organismos geneticamente modificados em alimentos por PCR em tempo real. Neste trabalho, os métodos CTAB e DNeasy forneceram preparações de DNA em quantidade e com qualidade a partir de amostras de proteína texturizada de soja, fórmula infantil e extrato de soja. Em relação ao Material de Referência Certificado contendo 5% de soja RoundupReady® , nenhum dos métodos forneceu DNA de boa qualidade. Entretanto, o teste de diluição realizado nos extratos de CTAB não demonstrou interferência de substâncias inibidoras. As eficiências das amplificações do alvo lectin não foram estatisticamente diferentes e os coeficientes de correlação (R²) demonstraram alto grau de correlação entre o número de cópias e os valores de Ct. A ANOVA demonstrou ajuste das curvas de regressão e ausência de desvios significativos. As eficiências das amplificações do alvo p35S, utilizando extratos DNeasy, não foram estatisticamente diferentes e os valores de R² ficaram acima de 0,98, sem desvios de linearidade. Dois valores de R², utilizando extratos CTAB, foram menores que 0,98, correspondendo a menor grau de correlação edesvio de linearidade em uma corrida. A análise comparativa dos Cts dos alvos p35S e lectin não demonstrou diferenças significativas entre as curvas analíticas da amplificação de cada alvo.

The DNA extraction is a critical step in Genetically Modified Organisms analysis based on real-time PCR. In this study, the CTAB and DNeasy methods provided good quality and quantity of DNA from the texturized soy protein, infant formula, and soy milk samples. Concerning the Certified Reference Material consisting of 5% Roundup Ready® soybean, neither method yielded DNA of good quality. However, the dilution test applied in the CTAB extracts showed no interference of inhibitory substances. The PCR efficiencies of lectin target amplification were not statistically different, and the coefficients of correlation (R²) demonstrated high degree of correlation between the copy numbers and the threshold cycle (Ct) values. ANOVA showed suitable adjustment of the regression and absence of significant linear deviations. The efficiencies of the p35S amplification were not statistically different, and all R² values using DNeasy extracts were above 0.98 with no significant linear deviations. Two out of three R² values using CTAB extracts were lower than 0.98, corresponding to lower degree of correlation, and the lack-of-fit test showed significant linear deviation in one run. The comparative analysis of the Ct values for the p35S and lectin targets demonstrated no statistical significant differences between the analytical curves of each target.