RESUMO
Chagas disease, which is found widely in Latin America and has a great impact on public health, is caused by the parasite Trypanosoma cruzi. It is a neglected parasitic disease that urgently requires rapid diagnostic methods. The objective of this study was to develop a SYBR Green real-time quantitative polymerase chain reaction (qPCR) technique for the direct identification and quantification of T. cruzi from experimentally contaminated açai fruit samples. We used discrete typing units, TcI, containing 3.5 × 104 cells/mL, to infect the pulp of the açai fruit. This was followed by DNA extraction using a standardized procedure. The DNA samples were quantified and amplified at specific time and temperature intervals. The specificity of the oligoinitiators used in the qPCR assays was estimated by calculating the primer dissociation curve (melting curve) along with a detection threshold using different concentrations of DNA. The method used here demonstrated good efficiency and precision for the detection and quantification of T. cruzi DNA, with a detection limit of 2.65 × 10-14 g/µL DNA. The qPCR technique presented here could serve as an important tool for the diagnosis of T. cruzi parasites in açai.
Assuntos
DNA de Protozoário/isolamento & purificação , Euterpe/parasitologia , Sucos de Frutas e Vegetais/parasitologia , Reação em Cadeia da Polimerase em Tempo Real/métodos , Trypanosoma cruzi/genética , Benzotiazóis , Brasil , Doença de Chagas/diagnóstico , Doença de Chagas/parasitologia , Diaminas , Corantes Fluorescentes , Humanos , Quinolinas , Sensibilidade e EspecificidadeRESUMO
INTRODUCTION: Oral transmission of acute Chagas disease is an emerging public health concern. This study aimed to detect insect fragments in experimentally contaminated food, by comparing triatomines with other insects. METHODS: Food samples were experimentally contaminated with insects, processed to recover their fragments by light filth, and analyzed by microscopy and Polymerase Chain Reaction (PCR). RESULTS: Morphological differences between coleopteran and triatomine insects were observed in microscopic images. PCR was efficient in amplifying Triatominae DNA in the experimentally contaminated food. CONCLUSIONS: This methodology could be utilized by food analysts to identify possible insect contamination in food samples.
Assuntos
Besouros/classificação , Parasitologia de Alimentos , Sucos de Frutas e Vegetais/parasitologia , Insetos Vetores/classificação , Saccharum/parasitologia , Triatominae/classificação , Animais , Doença de Chagas/transmissão , Humanos , Reação em Cadeia da PolimeraseRESUMO
In the Brazilian Amazon, the suspected source of infection in an outbreak of acute Chagas disease involving 10 patients was Euterpe oleracea (açaí berry) juice. Patient blood and juice samples contained Trypanosoma cruzi TcIV, indicating oral transmission of the Chagas disease agent.
Assuntos
Doença de Chagas/transmissão , Surtos de Doenças , Euterpe , Sucos de Frutas e Vegetais/parasitologia , Trypanosoma cruzi/fisiologia , Adolescente , Adulto , Idoso , Brasil/epidemiologia , Doença de Chagas/parasitologia , Feminino , Inocuidade dos Alimentos , Humanos , Lactente , Masculino , Pessoa de Meia-Idade , Adulto JovemRESUMO
Chagas disease, caused by Trypanosoma cruzi affects about 6-8 million people worldwide. Although transmission by triatomine insects has been controlled, other means of transmission maintain the infection. These forms of transmission are responsible for introducing Chagas disease in other non-endemic countries of the world. Thus, Chagas disease, nowadays is a worldwide health problem. In Brazil, acai pulp and sugarcane juice have been associated with Chagas disease outbreaks. The difficulties in isolation of the parasite from foods are hampering source tracking which could allow the confirmation of an implicated food commodity in these outbreak investigations. To address this scientific gap, we evaluated the performance of real-time PCR (qPCR) for detecting T. cruzi in acai pulp and sugarcane juice. All experiments were performed with acai pulp and sugarcane juice samples contaminated with different concentrations of T. cruzi. In assays with qPCR, the results showed that the ideal procedure for T. cruzi identification in acai pulp and sugarcane juice consisted of: i. centrifugation; ii. DNA extraction with a commercial kit for stool matrix; and iii. qPCR using a specific molecular marker for T. cruzi. The seeding in LIT medium of experimentally contaminated foods was effective in detecting the parasitic load by qPCR. The efficacy of qPCR was also verified testing food samples crushed with infected Triatomines. In conclusion, this methodology can be used to perform rapid diagnosis in outbreaks, facilitating measures in disease control.
Assuntos
Doença de Chagas/transmissão , Euterpe/parasitologia , Doenças Transmitidas por Alimentos/parasitologia , Sucos de Frutas e Vegetais/parasitologia , Carga Parasitária , Saccharum/parasitologia , Trypanosoma cruzi/genética , Animais , Brasil/epidemiologia , Humanos , Reação em Cadeia da Polimerase em Tempo Real/métodos , Trypanosoma cruzi/isolamento & purificaçãoRESUMO
Chagas disease is a zoonosis caused by protozoan parasite Trypanosoma cruzi, which is most frequently associated with a vectorial transmission. However, in recent years we have observed a significant increase in the oral transmission of the disease, associated mainly with the consumption of drinks made from fruit or other vegetables contaminated with triatomine faeces or secretions from infected mammals. After a latency period of 3 to 22 days after ingestion, the oral infection is characterized by more severe manifestations than those associated with vectorial transmission: prolonged fever, acute myocarditis with heart failure and, in some cases, meningoencephalitis. Mortality can reach up to 33% of those infected. The aim of this paper is to review this matter and to promote prevention practices.
Assuntos
Doença de Chagas/transmissão , Parasitologia de Alimentos , Sucos de Frutas e Vegetais/parasitologia , Trypanosoma cruzi , Animais , Doença de Chagas/diagnóstico , Doença de Chagas/epidemiologia , Doença de Chagas/prevenção & controle , Surtos de Doenças , Reservatórios de Doenças/parasitologia , Vetores de Doenças , Humanos , América do Sul/epidemiologiaRESUMO
Invertebrate-derived ingested DNA (iDNA) is quickly proving to be a valuable, non-invasive tool for monitoring vertebrate species of conservation concern. Using the DNA barcoding locus, we successfully identified both the blood-feeding leech Haementeria acuecueyetzin and its blood meal-the latter is shown to be derived from the Caribbean manatee, Trichechus manatus . DNA amplification was successful despite the fact that the specimen was fixed in Mezcal (a beverage distilled from agave). We report the first confirmed case of a leech feeding on a manatee, the first record of H. acuecueyetzin for the State of Chiapas and, to our knowledge, the first case of successful DNA amplification of a biological sample fixed in Mezcal other than the caterpillar "worms" more commonly found in that beverage.
Assuntos
Agave/parasitologia , Código de Barras de DNA Taxonômico/veterinária , Sucos de Frutas e Vegetais/parasitologia , Sanguessugas/classificação , Trichechus/parasitologia , Animais , Feminino , Fixadores , Sanguessugas/genética , México , Filogenia , Trichechus/sangue , Índias OcidentaisRESUMO
American trypanosomiasis is one of the few native parasites of this continent. As a zoonosis, Trypanosoma cruzi infects about 180 species out of 25 families of mammals. Its regular transmission is through triatomines, which can easily transmit parasites either by the skin route (contamination of mammals skin with their feces) or by oral route (ingestion of food contaminated with complete triatomines or their feces) and additionally through haematogenous via (congenital and transfusional) and by tissues (transplants). The oral route, which seems to be the ancestral form of transmission to wild and domestic mammals, has recently become more important after the success achieved in the control of domicile vectors using residual pesticides. From its initial diagnosis in 1967, tens of oral outbreaks have been diagnosed mostly in the Brazilian Amazon and subsequently in other four countries in South America. Environmental imbalance caused by man through the invasion and deforestation of woodlands, results in reduction of biodiversity of mammals as food source for triatomines, affecting the "dilution effect" of T. cruzi in the nature increasing the risk of human infection. On the other hand, triatomines invade houses looking for new blood sources. One of the consequences of domiciliated triatomines is the food contamination spread, especially in home-made juices, which has been the source of infection of most oral outbreaks. Other biotic and abiotic factors help to explain the recent increase of oral transmission outbreaks of Chagas disease, distributed in nine eco-regions of America.