An alternative method to recover Toxoplasma gondii from greenery and fruits.

Toxoplasma gondii oocysts are an important form of contamination with a high dispersion in the environment, but their detection is still a challenge. This study evaluated the recovery of oocysts from strawberries and crisphead lettuce. Samples (250 g of strawberries or one head of lettuce) were experimentally inoculated with 10, 10(2), 10(3) and 10(4) T. gondii oocysts, by two separate processes, spot dripping and immersion. Then, 50 g of each sample was washed, filtered through a cellulose ester membrane, and concentrated by centrifugation. Three aliquots were taken for DNA extraction in a direct way, after freeze-thaw (FT) cycles or ultrasound (US), followed by PCR (B22-B23 and Tox4-Tox5 primers). The T. gondii DNA was amplified with the primers B22-B23 in all samples contaminated by dripping and when DNA extraction was carried out after FT or US. These techniques may be useful in epidemiological surveillance in the control of this zoonosis.

Quantitative Risk Assessment of Human Trichinellosis Caused by Consumption of Pork Meat Sausages in Argentina.

In Argentina, there are three known species of genus Trichinella; however, Trichinella spiralis is most commonly associated with domestic pigs and it is recognized as the main cause of human trichinellosis by the consumption of products made with raw or insufficiently cooked pork meat. In some areas of Argentina, this disease is endemic and it is thus necessary to develop a more effective programme of prevention and control. Here, we developed a quantitative risk assessment of human trichinellosis following pork meat sausage consumption, which may be used to identify the stages with greater impact on the probability of acquiring the disease. The quantitative model was designed to describe the conditions in which the meat is produced, processed, transported, stored, sold and consumed in Argentina. The model predicted a risk of human trichinellosis of 4.88 × 10(-6) and an estimated annual number of trichinellosis cases of 109. The risk of human trichinellosis was sensitive to the number of Trichinella larvae that effectively survived the storage period (r = 0.89), the average probability of infection (PPinf ) (r = 0.44) and the storage time (Storage) (r = 0.08). This model allowed assessing the impact of different factors influencing the risk of acquiring trichinellosis. The model may thus help to select possible strategies to reduce the risk in the chain of by-products of pork production.

A PCR procedure for the detection of Giardia intestinalis cysts and Escherichia coli in lettuce.

Giardia intestinalis is a pathogen associated with foodborne outbreaks and Escherichia coli is commonly used as a marker of faecal contamination. Implementation of routine identification methods of G. intestinalis is difficult for the analysis of vegetables and the microbiological detection of E. coli requires several days. This study proposes a PCR-based assay for the detection of E. coli and G. intestinalis cysts using crude DNA isolated from artificially contaminated lettuce. The G. intestinalis and E. coli PCR assays targeted the ß-giardin and uidA genes, respectively, and were 100% specific. Forty lettuces from local markets were analysed by both PCR and light microscopy and no cysts were detected, the calculated detection limit was 20 cysts per gram of lettuce; however, by PCR, E. coli was detected in eight of ten randomly selected samples of lettuce. These data highlight the need to validate procedures for routine quality assurance. These PCR-based assays can be employed as alternative methods for the detection of G. intestinalis and E. coli and have the potential to allow for the automation and simultaneous detection of protozoa and bacterial pathogens in multiple samples. Significance and impact of the study: There are few studies for Giardia intestinalis detection in food because methods for its identification are difficult for routine implementation. Here, we developed a PCR-based method as an alternative to the direct observation of cysts in lettuce by light microscopy. Additionally, Escherichia coli was detected by PCR and the sanitary quality of lettuce was evaluated using molecular and standard microbiological methods. Using PCR, the detection probability of Giardia cysts inoculated onto samples of lettuce was improved compared to light microscopy, with the advantage of easy automation. These methods may be employed to perform timely and affordable detection of foodborne pathogens.

Análisis comparativo de los métodos para la detección de parásitos en las hortalizas para el consumo humano / Comparative analysis of parasite detection methods in vegetables for human consumption

INTRODUCIÓN: la contaminación de los vegetales es un persistente problema de salud. Las diferentes técnicas de diagnóstico y de procesamiento dificultan determinar qué técnica es más eficiente y sensible. OBJETIVO: efectuar una comparación entre la sensibilidad de las técnicas parasitológicas en muestras de vegetales. MÉTODOS: se recogieron 30 muestras de vegetales - lechuga (Lactuca sativa), rúcula (Eruca sativa) y berro (Nasturtium officinale) - y se investigaron por las técnicas de Hoffman, Pons y Janer (HPJ) y Faust (F). Los datos fueron analizados utilizando la prueba de Bland-Altman para evaluar la correlación y la prueba de Mann-Whitney para comparar las medianas. RESULTADOS: de las muestras analizadas, 46,6 por ciento resultó positivo, se observó Balantidium coli (20,0 por ciento de la contaminación), Entamoeba coli (21,6 por ciento), Entamoeba histolityca (5,0 por ciento), Trichuris trichiura (3,3 por ciento) y Strongyloides stercoralis (2,5 por ciento). La prueba de Bland-Altman mostró correlación significativa entre los métodos examinados. Hubo una diferencia significativa (p< 0,05) entre los resultados, al evaluar individualmente el promedio. CONCLUSIÓN: este estudio demostró que la técnica HPJ fue más eficaz en la detección de huevos, larvas de helmintos y quistes de protozoos en las plantas estudiadas.

INTRODUCTION: vegetable contamination is a persistent health problem. The different methods of processing and diagnosis make it difficult to determine the most effective and sensitive technique. OBJECTIVE: a comparative analysis of parasitological technique sensitivity in vegetable samples. METHODS: a total of 30 samples were harvested lettuce (Lactuca sativa), rocket (Eruca sativa) and watercress (Nasturtium officinale) and later analyzed using Hoffman, Pons, and Janer (HPJ) and Faust (f) techniques. Data were analyzed, using the Bland-Altman test to evaluate the correlation and the Mann-Whitney test to compare the medians. RESULTS: of the analyzed samples, 46.6 percent were positive for intestinal parasites; Balantidium coli, accounting for 20 percent of contamination, Entamoeba coli (21.6 percent) and Entamoeba histolityca (5 percent), Trichuris trichiura(3,3 percent) and Strongyloides stercoralis (2.5 percent) The Bland-Altman test showed significant correlation between the analyzed methods. When evaluating the averages separately, there was significant difference (pd¼ 0.05) among the results. CONCLUSIONS: this study proved that the HPJ technique was more effective for the detection of eggs, helminth larvae and protozoan cysts in the plants under study.

[Comparative analysis of parasite detection methods in vegetables for human consumption]. / Análisis comparativo de los métodos para la detección de parásitos en las hortalizas para el consumo humano.

INTRODUCTION: vegetable contamination is a persistent health problem. The different methods of processing and diagnosis make it difficult to determine the most effective and sensitive technique. OBJECTIVE: a comparative analysis of parasitological technique sensitivity in vegetable samples. METHODS: a total of 30 samples were harvested -lettuce (Lactuca sativa), rocket (Eruca sativa) and watercress (Nasturtium officinale)--and later analyzed using Hoffman, Pons, and Janer (HPJ) and Faust (f) techniques. Data were analyzed, using the Bland-Altman test to evaluate the correlation and the Mann-Whitney test to compare the medians. RESULTS: of the analyzed samples, 46.6% were positive for intestinal parasites; Balantidium coli, accounting for 20% of contamination, Entamoeba coli (21.6) and Entamoeba histolityca (5%), Trichuris trichiura (3.3%) and Strongyloides stercoralis (2.5%) The Bland-Altman test showed significant correlation between the analyzed methods. When evaluating the averages separately, there was significant difference (p = 0.05) among the results. CONCLUSIONS: this study proved that the HPJ technique was more effective for the detection of eggs, helminth larvae and protozoan cysts in the plants under study.