Analysis of Multiple Pesticides Residues in Market Samples of Okra and Associated Dietary Risk Assessment for Consumers

European Union Reference Laboratory method for Fruits and Vegetable (EURL-FV-2010-M1) for the quantification of pesticide residues was verified for the determination of multiple pesticides residues in okra. The targeted pesticides were extracted using acetonitrile with citrate buffer salts followed by cleanup with primary secondary amine (PSA) and analyzed on LC-MS/MS. The recoveries for all the targeted pesticides were within an acceptable range of 70.1 - 116.6% and precision in terms of RSD was 0.3 - 18.1%, respectively. The limit of quantification ranged from 0.002 mg/kg for carbofuran to 0.5 mg/kg for α-cypermethrin. The status of pesticide residues in okra (n= 21) available to consumers in the main markets of Pakistan has been determined by using this verified method. Sixty-two percent of the tested samples were contaminated out of which three samples were non-compliant with European Union Maximum residue limits (EU-MRL). The pesticides violating the EU-MRL were bifenthrin, thiamethoxam, and triazophos. For all the detected pesticides, estimated daily intake (EDI) ranged between 7.39×10−6 and 1.78×10−4 mg/kg of body weight while the values of Health Index (HI) fluctuated between 8.9×10−5 and 1.8×10−2. Although, residues of some pesticides were reported to be non-compliant with EU-MRLs, yet the risk posed by these pesticides to human health was insignificant.

Challenge Studies to Determine the Ability of Foods to Support the Growth of Listeria monocytogenes

: Listeria monocytogenes is a foodborne pathogen that causes listeriosis, a relatively rare, but potentially fatal, disease, with a mortality rate of 20–30%. In general, European Regulations require the absence of L. monocytogenes in five samples of 25 g before the food has left the producer, but if the food has been demonstrated not to support the growth of L. monocytogenes, up to 100 cfu g-1 are allowed in the food (except for foods for infants or medical purposes) during its shelf-life under reasonably foreseeable storage conditions. It is important for food producers to determine if their food supports the growth of L. monocytogenes. The European Union Reference Laboratory for L. monocytogenes published a Technical Guidance document for conducting shelf-life studies on L. monocytogenes in ready-to-eat foods in June 2014. Primarily based on the EURL guidance document for conducting challenge studies, the ability of cheese (feta and soft goat’s milk cheese), cold-smoked salmon, coleslaw, and pork pate to support the growth of L. monocytogenes was determined using a starting inoculum of approximately 100 cfu g−1. The cheese and pork pate were incubated at 8 °C for 14 days; the smoked salmon was incubated at 6 °C for 5 days and 8°C for 9 days; and the coleslaw was incubated at 8 °C for 7 days and 12 °C for 14 days. The results showed that the smoked salmon and pork pate supported growth, while coleslaw and cheese did not. From this study, it is evident that there are factors in food other than pH, water activity, and total bacterial count (TBC) that can inhibit the ability of L. monocytogenes to grow in food.

Epidemiological study of honeybee pathogens in Europe: The results of Castilla-La Mancha (Spain)

As a part of a Pilot Monitoring Program of honey bee health coordinated by the EURL (European Union Reference Laboratory) and according to the criteria established for Spain, 14 apiaries in Castilla-La Mancha were selected at random and sampled during the autumns of 2012-2014 to identify the most prevalent nosogenic agents, potentially those related to the honey bee colony collapse phenomenon. In all the apiaries studied, Nosema ceranae was the most prevalent pathogen detected over the three years, confirming the worldwide spread of this microsporidian, a pathogen that negatively affects honey bee health at an individual and colony level. Trypanosomatids were also very prevalent in honey bee colonies, although the majority of Trypanosomatids detected were not Crithidia mellificae but rather the genetically distinct Lotmaria passim lineage. We also detected Varroa destructor mites, and the particularly high prevalence in 2014 suggests a possible problem regarding mite control in field conditions that requires attention. In agreement with data from other regions, the BQCV and DWV were the most prevalent viruses in honey bee colonies and thus, the Varroa-DVW interaction may be an important cause of bee colony mortality. While there was little evidence of a relationship between the BQCV virus and N. ceranae under field conditions during 2012, this was not the case in 2013 and 2014. Finally, the AKI-complex or LSV-complex was not detected. The information obtained in this study should help orientate future plans for honey bee disease control.

Collection of Listeria monocytogenes Isolates from Milk, Dairy Products and Food Processing Environments in Slovakia for the Purposes of European Molecular Database

The molecular typing of Listeria monocytogenes isolates is an important tool for monitoring the spread of the strains in food chains, providing evidence for epidemiological investigations and for the detection of out-breaks. The demand of European typing data centralization, collection and sharing stimulated the generation of “EURL L. monocytogenes Database (EURL Lm DB)” in 2012 led by the European Union Reference Laboratory (EURL) for L. monocytogenes (ANSES Maisons-Alfort Laboratory for Food Safety, France) in close collaboration with Applied Maths. This database includes the typing results and epidemiological information on strains isolated from food, environmental or animal samples and it is in connection with human strains database TESSy (The European Surveillance System) led by the ECDC (European Centre for Disease Prevention and Control). In total 147 L. monocytogenes isolates were examined by PFGE (pulsed field gel electrophoresis) in 2014—2015 in VFI Dolny Kubin from different sources. Nearly half (68) of the 147 isolates in the national Slovak database came from milk or dairy products samples and the related manufacturing environment. In this work, 68 isolates associated with milk were selected and divided into 27 clusters (95 % similarity level) after combined comparison analysis (AscI and ApaI) by BioNumerics 6.6 software. Eight clusters included three or more similar PFGE profiles.

A note on challenge trials to determine the growth of Listeria monocytogenes on mushrooms (Agaricus bisporus)

In the EU, food is considered safe with regard toListeria monocytogenesif the number of micro-organisms does not exceed 100 colony forming units (cfu)/g throughout its shelf-life. Therefore, it is important to determine if a food supports growth ofL. monocytogenes. Guidelines for conducting challenge tests for growth assessment ofL. monocytogeneson foods were published by the European Union Reference Laboratory (EURL) in 2014. The aim of this study was to use these guidelines to determine if refrigerated, fresh, whole, closed-cap, prepackaged mushrooms (Agaricus bisporus) support the growth ofL. monocytogenes. Three batches of mushrooms were artificially inoculated at approximately 100 cfu/g with a three-strain mix ofL. monocytogenesand incubated for 2 days at 8°C followed by 4 days at 12°C.L. monocytogenesnumbers were determined (in triplicate for each batch) on days 0, 2 and 6. Water activity, pH and total bacterial counts were also determined. There was no increase in the number ofL. monocytogenesabove the threshold of 0.5 log cfu/g in any of the replicates. In 8 of 9 replicates, the numbers decreased indicating thatA. bisporusdo not support the growth ofL. monocytogenes. As the EU regulations allow < 100 cfu/g if the food cannot support growth ofL. monocytogenes, the significance of this study is that mushrooms with < 100 cfu/g may be within the regulations and therefore, quantitative rather than qualitative determination may be required.