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[High performance liquid chromatography: application in pesticide residue analysis].

Chromatographic techniques allow separation and quantitation of trace levels of organic compounds in different matrices. Chromatography is widely used in many scientific areas, including food safety. Currently, high performance liquid chromatography (HPLC) and related techniques have become the dominant analytical separation tools in such areas as pharmaceutical, chemical and food industries and environmental monitoring. Contrary to gas chromatography (GC), HPLC allows to determine low-volatile and thermolabile compounds. A variety of packings and bonded phases as well as eluents and their combinations make this technique very useful in analysis of food contaminants, including pesticide residues. Plant protection products, called commonly pesticides, contain biologically active substances having wide mode of action, used in crop protection and in many other areas. Unfortunately, besides advantages related to their use, it should be taken into account that they are also toxic for humans and environment. As general population is exposed to pesticides mainly through the food, it is necessary to monitor concentrations of these compounds using sensitive techniques for ppm or even ppb levels. This article discusses the advantages of HPLC technique for the residues analysis of some active substances of plant protection products.

Chromatography, High Pressure Liquid↗

The ethnophysiology of digestion and diarrhoea in a Bangladeshi hospital population.

The results presented in this paper are drawn from a study of the acceptability of the weaning food, ARGC. The study aimed to investigate attitudes and practices surrounding weaning and the dietary management of diarrhoea. One hundred and twenty mothers of children aged between six months and 24 months suffering from mild diarrhoea and admitted to the ICDDR,B treatment centre were randomly selected. Diarrhoea was attributed by mothers to a number of causes; most common were contaminated food and breastmilk. Breastmilk was understood to have been spoiled either by the mother's diet or mystical forces termed batash. Batash was also suspected to directly making children sick in some instances. Thirty-six per cent of mothers attempted to manage diarrhoea at home by withholding normal foods from their children's diets and others modified their own diets. Less than a quarter of the children were normally fed vegetables, dal (lentils) or small fish. It appeared that fish was rarely given to young children and was regarded with some ambivalence and considered potentially attractive as a vehicle for malign forces that might attack young children and their mothers and cause illness. People were unwilling to feed their children fish and other items of the normal family diet because of notions about the digestive system and the concept of "digestive power" and the idea that young children did not have the digestive power to digest certain foods. It was suggested that early weaning might lead to poor and abnormal growth and development.

Adult↗

Dietary exposure of children in lead-laden environments.

Children are the most susceptible population to lead exposure because of three interacting factors; they have more opportunity for contact with lead sources due to their activities, lead absorption occurs more readily in a child compared to an adult, and the child's development is more vulnerable to lead than adults. Low levels of lead in the blood have been shown to cause adverse health effects; the level of concern for children is currently 10 microg/dl. The contribution of dietary exposure of lead to increased blood lead levels (PbB) is not well characterized. This study was conducted to measure potential dietary lead intakes of children 2 to 3 years of age who live in homes contaminated with environmental lead. Objectives were to estimate lead intakes for children consuming food in contaminated environments, recognizing unstructured eating patterns and to investigate if correlations exist between daily dietary exposure and measured PbB. Dietary exposure was evaluated by collecting samples that were typical of the foods the young children ate in their homes. A 24-h duplicate of all foods plus sentinel foods, i.e., individual items used to represent foods contaminated during handling, were collected from 48 children. Ten homes were revisited to obtain information on the variation in daily dietary intakes. Drinking water was evaluated both as part of the segregated beverage sample composite and by itself. Additional information collected included lead concentrations from hand wipes, floor wipes, and venous blood, and questionnaire responses from the caregiver on activities potentially related to exposure. Activities and hygiene practices of the children and contamination of foods in their environment influences total dietary intake. Estimated mean dietary intakes of lead (29.2 microg Pb/day) were more than three times the measured 24-h duplicate-diet levels (8.37 microg Pb/day), which were almost six times higher than current national estimates (1.40 microg Pb/day). Statistically significant correlations were observed between floor wipes and foods contacting contaminated surfaces, hand wipes and foods contacting contaminated hands and surfaces, and hand wipes and floor wipes. This study indicates that the dietary pathway of exposure to lead is impacted by eating activities of children living in lead-contaminated environments and that analysis of foods themselves is not enough to determine excess dietary exposures that are occurring.

Activities of Daily Living↗

Dioxins, dioxin-like PCBs and non-dioxin-like PCBs in foodstuffs: occurrence and dietary intake in The Netherlands.

Data on occurrence of dioxins (polychlorinated dibenzo-p-dioxins [PCDDs] and dibenzofurans [PCDFs]), dioxin-like PCBs (polychlorinated non-ortho and mono-ortho biphenyls) and non-dioxin-like PCBs (as represented by the so-called indicator-PCBs: congeners 28, 52, 101, 118, 138, 153 and 180) in food products consumed in The Netherlands that were collected in measurement programs carried out during 1998 and 1999, and combined with food consumption data to assess the dietary intake of these persistent food contaminants. The estimated median life-long-averaged intake of the sum of dioxins and dioxin-like PCBs in the population is 1.2 pg WHO-TEQ (toxic equivalents) per kg body weight (bw) per day, while the estimated median life-long-averaged intake of indicator-PCBs is 5.6 ng per kg bw per day. The contribution of different food groups to the total intake of both dioxins + dioxin-like PCBs and non-dioxin-like PCBs is fairly uniformly distributed over the foods consumed: meat products (23% and 27%, respectively), dairy products (27% and 17%, respectively), fish (16% and 26%, respectively), eggs (4% and 5%, respectively), vegetable products (13% and 7%, respectively), and industrial oils and fats (17% and 18%, respectively). Compared with earlier intake estimations the present estimation shows a continued reduction in the intake of dioxins as well as PCBs. This reduction is related to the decrease in the concentration of these substances in the majority of foodstuffs. Nevertheless, a small part of the population still has a rather high life-long averaged intake: 8% of the population is exposed to intake levels above the tolerable weekly intake for dioxins and dioxin-like PCBs of 14 pg WHO-TEQ per kg bw per week, as recently derived by the Scientific Committee on Food of the European Commission. For the non-dioxin-like PCBs an internationally accepted maximum intake level is still lacking. However, to provide risk managers with a health-based guideline to prevent health effects of exposure to non-dioxin-like PCBs, the (international) derivation of a tolerable daily intake is recommended. Monitoring the dietary intake of PCBs is just as important as monitoring the intake of dioxins and dioxin-like PCBs, and attempts to decrease the exposure to both compound classes need continuous attention.

Adolescent↗

Influence of dispersants on the bioavailability and trophic transfer of petroleum hydrocarbons to larval topsmelt (Atherinops affinis).

Use of chemical dispersants as oil spill clean-up agents may alter the normal behavior of petroleum hydrocarbons (PH) by increasing their functional water solubility, resulting in increased bioavailability and altered interactions between dispersant, oil, and biological membranes. The objective of this research was to determine the impact of dispersing agents on PH bioavailability and trophic transfer to larval fish from primary levels of a marine food chain. Uptake, bioaccumulation, depuration, and metabolic transformation of a model PH, [14C]naphthalene, were measured and compared for Prudhoe Bay crude oil (PBCO) dispersed with Corexit 9527(R) (DO) and undispersed preparations of the water-accommodated fraction (WAF) of PBCO. The model food chain consisted of a primary producer, Isochrysis galbana; and a primary consumer, the rotifer, Brachionus plicatilis; and larval topsmelt, Atherinops affinis. Direct aqueous (AQ) exposure was compared with combined aqueous and dietary (AQ&D) exposure. Dispersants altered the uptake and depuration processes of naphthalene, independent of aqueous concentrations, in primary trophic species of a marine food chain. The amount of naphthalene taken up by topsmelt was initially significantly (P < or = 0.05) enhanced in the presence of dispersant, reaching a maximum more quickly than undispersed samples. Dispersion treatment significantly increased naphthalene dispension in topsmelt (P < or = 0.05) from both AQ and AQ&D exposures. No significant change in naphthalene uptake by fish was observed with the addition of contaminated food for either WAF or DO medium; however, both uptake and depuration rate constants varied significantly with route of exposure consistent with greater naphthalene turnover. The majority (> or = 72%) of naphthalene-derived radioactivity from fish tissue following all exposures was in the parent form, with smaller quantities of alpha- and beta-naphthols, alpha- and beta-naphthyl sulfates, and an unidentified derivative.

Animals↗

A real-time PCR assay for the detection of Campylobacter jejuni in foods after enrichment culture.

A real-time PCR assay was developed for the quantitative detection of Campylobacter jejuni in foods after enrichment culture. The specificity of the assay for C. jejuni was demonstrated with a diverse range of Campylobacter species, related organisms, and unrelated genera. The assay had a linear range of quantification over six orders of magnitude, and the limit of detection was approximately 12 genome equivalents. The assay was used to detect C. jejuni in both naturally and artificially contaminated food samples. Ninety-seven foods, including raw poultry meat, offal, raw shellfish, and milk samples, were enriched in blood-free Campylobacter enrichment broth at 37 degrees C for 24 h, followed by 42 degrees C for 24 h. Enrichment cultures were subcultured to Campylobacter charcoal-cefoperazone-deoxycholate blood-free selective agar, and presumptive Campylobacter isolates were identified with phenotypic methods. DNA was extracted from enrichment cultures with a rapid lysis method and used as the template in the real-time PCR assay. A total of 66 samples were positive for C. jejuni by either method, with 57 samples positive for C. jejuni by subculture to selective agar medium and 63 samples positive in the real-time PCR assay. The results of both methods were concordant for 84 of the samples. The total time taken for detection from enrichment broth samples was approximately 3 h for the real-time PCR assay, with the results being available immediately at the end of PCR cycling, compared to 48 h for subculture to selective agar. This assay significantly reduces the total time taken for the detection of C. jejuni in foods and is an important model for other food-borne pathogens.

Animals↗

Serum dioxin-like activity is associated with reproductive parameters in young men from the general Flemish population.

BACKGROUND: 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) and some related environmental contaminants are aryl hydrocarbon receptor (AhR) ligands that exert reproductive and developmental toxicity in laboratory animals. In humans, fertility-related effects are less documented. OBJECTIVE: The aim of this study was to investigate the relationship between dioxin-like biological activity in serum and parameters of reproductive status in men from the general population 5 months after a polychlorinated biphenyl and dioxin food-contamination episode in Belgium. DESIGN: In the framework of the cross-sectional Flemish Environment and Health Study (FLEHS), we recruited 101 men 20-40 years of age and evaluated sperm parameters, measured sex hormones, and gathered information on a number of lifestyle factors. In addition, we determined the AhR-mediated enzymatic response elicited by individual serum samples and expressed it as TCDD equivalent concentrations (CALUX-TEQs) using an established transactivation assay. RESULTS: Age (p = 0.04) and the frequency of fish (p = 0.02) and egg (p = 0.001) consumption were independent positive determinants of serum dioxin-like activity. After correcting for possible confounders, we found that a 2-fold increase in CALUX-TEQ > 16 pg/L was associated with a 7.1% and 6.8% (both p = 0.04) decrease in total and free testosterone, respectively. We also observed a more pronounced drop in semen volume of 16.0% (p = 0.03), whereas sperm concentration rose by 25.2% (p = 0.07). No relationship was found with total sperm count or sperm morphology. CONCLUSIONS: These data suggest an interaction of dioxin-like compounds with the secretory function of the seminal vesicles or prostate, possibly indirectly through an effect on testosterone secretion, at levels not affecting spermatogenesis as such.

Adult↗

Outbreak of typhoid fever on the French Riviera.

A case-control study was conducted to establish the source of a community outbreak of typhoid fever in Utelle, France, a village located in the Alpes-Maritimes district of the French Riviera. Thirteen confirmed cases of typhoid fever and 41 confirmed community controls were included in the study. Cases and controls did not differ regarding ingestion of water. Multivariate logistic regression analysis identified consumption of pork meats during a village festival as the only statistically significant risk factor for typhoid fever after adjusting for age and sex (odds ratio, 76.0; 95% confidence interval, 3.5-1660). Assessment of food-handling procedures at the inn where the food had been prepared showed that the refrigeration and cooking facilities were inadequate to maintain a proper sanitary environment during the preparation of a meal for 350 people. Although the exact cause of the epidemic could not be confirmed, food contaminated by a chronic typhi carrier is the most plausible hypothesis.

Animals↗

[Butyltin and its degradation products in the aspect of food toxicology].

The paper briefly reviews and up-dates available data on some aspects of toxicity of organotins and especially of butyltins, their applications, legal aspects as well as aquatic environment, seafood and food contamination. Special attention is paid on assessed rates of daily intake of butyltins with fish diet by some local groups of inhabitants with high consumption rates of fish originating from the region of the Gulf of Gdański.

Conservation of Natural Resources↗

The effects of ingested petroleum on oviposition and some aspects of reproduction in experimental colonies of mallard ducks (Anas platyrhynchos).

Compared to unmated mallard ducks fed an uncontaminated diet, unmated birds given food contaminated with 3 ml South Louisiana crude oil per 100 g dry weight showed an 84% decline in the daily rate of oviposition, a 33% decrease in egg-shell thickness and at autopsy more than 82% of the ovarian mass consisted of atretic follicles. Similar studies on groups of mated females showed that although the addition of 1 ml South Louisiana crude oil/100 g dry food had no effect on the daily rate of oviposition, none of the eggs had been fertilized while a concentration of 3 ml South Louisiana crude oil/100 g dry food suppressed the daily rate of oviposition significantly. Less than 25% of these eggs had been fertilized and only 40% of the fertilized eggs yielded viable ducklings. In both of these groups of mated birds, normal patterns of oviposition, fertilization and hatchability were restored after removal of petroleum from the diet. The addition of 1 ml Kuwait crude oil/100 g dry food had no effect on the rate of oviposition, the incidence of fertility or the hatchability of the fertilized eggs. The addition of 3 ml oil/100 g dry food completely abolished oviposition, but a normal rate of oviposition was restored when the concentration of the crude oil was reduced from 3 to 1 ml/100 g dry food. However, the incidence of fertilization remained low and none of the fertilized eggs gave rise to viable ducklings. Kuwait crude oil had no effect on shell thickness.

Animals↗

Typing of Austrian Listeria monocytogenes isolates by automated laser fluorescence analysis of randomly amplified polymorphic DNA.

We used automated laser fluorescence analysis of randomly amplified polymorphic DNA (RAPD-Alfa) to study the epidemiology of listeriosis in western Austria. There were no discrepancies between RAPD-Alfa patterns and serotypes found in 18 food isolates and 18 clinical isolates. The results of our study suggest that the food isolates typed were not at the origin of the human cases in western Austria. Using RAPD-Alfa, it was possible to link 9 out of 16 "sporadic" Listeria infections (mother-child cases counted as one) to the occurrence of other cases. Our results underline the necessity of epidemiological clarification of listeriosis cases as a prerequisite for specific preventive measures by public health services (e.g. confiscation of contaminated food products, issue of public warnings). To establish the chain of infection, more is needed than just speciation of bacteria in incriminated food products. Automated laser fluorescence analysis of randomly amplified polymorphic DNA seems a suitable, easy and rapid method for the typing of Listeria monocytogenes strains.

Adult↗

A point-source foodborne listeriosis outbreak: documented incubation period and possible mild illness.

Listeria bacteremia occurred in 2 pregnant women whose only common exposure was attendance at a party. The incubation period, the possibility of mild disease due to Listeria infection, and foods associated with risk of disease were evaluated. Ten (28%) of 36 party attenders met a case definition, which included isolation of Listeria monocytogenes from blood or stool or two of the following: fever, musculoskeletal symptoms, nausea, vomiting, diarrhea. One of 25 stool cultures was positive. The 2 blood isolates and 1 stool isolate were serotype 4b and identical by enzyme typing. The incubation periods for illness in the 2 pregnant women were 19 and 23 days. Consumption of large amounts of shrimp, nonalcoholic beverages, Camembert cheese, and cauliflower was significantly associated with illness. Eating shrimp remained a significant risk factor for illness after controlling for consumption of other foods. This study suggests a milder illness may exist in healthy persons who consume foods contaminated with L. monocytogenes and demonstrates a prolonged incubation period for disease.

Adult↗

A resuscitation/selection system for rapid determination of Salmonella in foods.

A resuscitation medium was developed consisting of a trypticase soy broth base supplemented with 0.5% yeast extract, 0.25% sodium pyruvate, 0.01% sodium thioglycollate, and 0.1% chicken fat. After a resuscitation period of 4 h, the medium was made selective by addition of either sodium thiosulfate, bile salts and iodine, or sodium selenite and L-cystine. The now selective medium was incubated for 16 h. The presence or absence of Salmonella was determined by the Salmonella-Tek antibody-based detection kit. The present system was compared with a method of the Bacteriological Analytical Manual (BAM) for naturally contaminated foods. Nineteen egg products were screened; 3/19 were positive using the BAM method, 9/19 were positive using the present system. Seventeen chicken samples were assayed; 10/17 were positive using the BAM method; 13/17 were positive using the present system. Of 8 pepper samples, 4/8 were positive using the BAM method; 6/8 were positive using the present system. Of 8 spice samples, 6/8 were positive using the BAM method, 7/8 were positive using the present system. Of 6 onion products sampled, 5/6 were positive using the BAM method; 6/6 were positive using the present system.

Culture Media↗

Effect of formaldehyde on the immunogenicity of staphylococcal enterotoxin B for Macaca mulatta.

Monkeys were immunized with enterotoxin and enterotoxoid by intracutaneous injection or by feeding. Identical schedules were used to compare the effectiveness of the two antigens and the two routes. Enterotoxin administered intracutaneously was the most effective antigen, whereas oral administration of enterotoxoid was least effective. Intracutaneous injection of toxoid and oral feeding of toxin were intermediate and not too dissimilar in effectiveness. Antibody titers and protection persisted for at least 1 year at a relatively high level. Monkeys that had preimmunization hemagglutinins showed an anamnestic response after immunization. The development of protection and the appearance of antibodies subsequent to feeding toxin or toxoid suggest that ingestion of food contaminated by staphylococci or their metabolites may be one cause for the appearance of antitoxin in the serum of supposedly unexposed animals and man.

Animals↗

PCR-based procedures for detection and quantification of Staphylococcus aureus and their application in food.

AIMS: To evaluate the specificity of nuc targeted primers for PCR detection of Staphylococcus aureus in different food matrices and to establish a RTQ-PCR procedure suitable for the routine detection and quantification of this pathogen in food. METHODS AND RESULTS: Specificity of nuc targeted primers (Pri1-Pri2 and the newly designed RTQ-PCR primers) was tested on a total of 157 strains of genetically confirmed identity, including reference and food isolates. PCR detection on artificially inoculated beef samples by DNA extraction using a DNeasy Tissue Kit (Qiagen GmhH, Hilden, Germany) showed a sensitivity value around 10(3) CFU g(-1). The two RTQ-PCR systems, incorporating SYBR-Green I and TaqMan, respectively, applied in the present work improved the sensitivity of conventional PCR by lowering the detection level to 10 and 100 cells, respectively. Out of 164 naturally contaminated foods tested for the presence of Staph. aureus, 74 were positive by conventional PCR and 69 by the traditional culture method with a high degree of result agreement between both methodologies (93.3%). CONCLUSIONS: PCR approaches, using nuc targeted primers, have proved specific and combined with growth techniques may improve detection of Staph. aureus in different types of food. SIGNIFICANCE AND IMPACT OF THE STUDY: The SYBR-Green I real-time PCR approach established allows the sensitive, automated and quantitative detection of Staph. aureus for routine analysis at a reasonable cost.

Animals↗

Filament formation by Salmonella spp. inoculated into liquid food matrices at refrigeration temperatures, and growth patterns when warmed.

In this study, the formation of multicellular filamentous Salmonella cells in response to low temperatures was investigated by using isolates of Salmonella enterica serovar Enteritidis PT4 and S. enterica serovar Typhimurium DT104 as the inocula. The formation of filamentous cells in two liquid food matrices at the recommended maximum temperature for refrigeration (8 degrees C) was monitored and compared with that in tryptone soya broth. Giemsa staining was performed to locate nuclear material within the filaments. Single filaments were warmed on agar at 37 degrees C, and the subsequent rate of septation was quantified. For all strains tested, > 70% of the Salmonella cells inoculated had become filamentous after 4 days in media at 8 degrees C, indicating that filamentation could occur during the shelf life of most refrigerated foods. Strains with impaired RpoS expression were able to form filaments at 8 degrees C, although these filaments tended to be shorter and less numerous. All strains also formed filamentous cells at 8 degrees C in retail milk or chicken meat extract. Filaments often exceeded 100 microm in length and appeared straight-sided under the microscope in media and in foods, and Giemsa staining demonstrated that regularly spaced nucleoids were present. This phenotype indicates that an early block in cell septation is probably responsible for filamentation. When filaments were warmed on agar at 37 degrees C, there was a rapid completion of septation, and for one filament, a >200-fold increase in cell number was observed within 4 h. There are clear public health implications associated with the filamentation of Salmonella in contaminated foods at refrigeration temperatures, especially when the possibility of rapid septation of filamentous cells upon warming is considered.

Animals↗

Detection of Listeria monocytogenes in food using a combined enrichment/real-time PCR method targeting the prfA gene.

A combined enrichment/real-time PCR method for the detection of Listeria monocytogenes is presented. The method is based on a conventional PCR assay targeting the prfA gene, which has been validated and suggested as an international standard PCR method for identifying L. monocytogenes in food. This real-time PCR assay includes an internal amplification control. Inclusivity and exclusivity were 100% each when testing 100 L. monocytogenes isolates, 30 Listeria spp. isolates other than L. monocytogenes, and 29 non-Listeria isolates. The theoretical detection limit was one copy of the target gene per PCR reaction and the practical detection limit was about 5 copies per PCR. Using the combined enrichment/real-time PCR method, 7.5 CFU/25 ml of artificially contaminated raw milk, and 9, 1, and 1 CFU/15 g of artificially contaminated salmon, pâté, and green-veined cheese, respectively, were detected. When analyzing 76 naturally contaminated food samples of various types and comparing the results with the ISO 11290-1 standard method, the relative accuracy was 96%, the relative specificity 100%, and the relative sensitivity, 76.9%.

Animals↗