Marie-Louise Scippo

Investigation of the genotoxicity of substances migrating from polycarbonate replacement baby bottles to identify chemicals of high concern

Due to the worldwide concern that bisphenol A might act as an endocrine disruptor, alternative materials for polycarbonate (PC) have been introduced on the European market. However, PC-replacement products might also release substances of which the toxicological profile--including their genotoxic effects--has not yet been characterized. Because a thorough characterization of the genotoxic profile of all these substances is impossible in the short term, a strategy was developed in order to prioritize those substances for which additional data are urgently needed. The strategy consisted of a decision tree using hazard information related to genotoxicity. The relevant information was obtained from the database of the European Chemicals Agency (ECHA), in silico prediction tools (ToxTree and Derek Nexus(TM)) and the in vitro Vitotox(®) test for detecting DNA damage. By applying the decision tree, substances could be classified into different groups, each characterized by a different probability to induce genotoxic effects. Although none of the investigated substances could be unequivocally identified as genotoxic, the presence of genotoxic effects could neither be excluded for any of them. Consequently, all substances require more data to investigate the genotoxic potential. However, the type and the urge for these data differs among the substances.

Development of an LC-MS/MS analytical method for the simultaneous measurement of aldehydes from polyunsaturated fatty acids degradation in animal feed

Knowing that polyunsaturated fatty acids can lead to the formation of potentially toxic aldehydes as secondary oxidation products, an analytical method using liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) detection was developed to measure the concentration of eight aldehydes in animal feed: malondialdehyde (MDA), 4-hydroxy-2-nonenal (4-HNE), 4-hydroxy-2-hexenal (4-HHE), crotonaldehyde (CRT), benzaldehyde (BNZ), hexanal (HXL), 2,4-nonadienal, and 2,4-decadienal. The developed method was validated according to the criteria and procedure described in international standards. The evaluated parameters were specificity/selectivity, recovery, precision, accuracy, uncertainty, limits of detection and quantification, using the concept of accuracy profiles. These parameters were determined during experiments conducted over three different days with ground Kelloggs® Corn Flakes® cereals as model matrix for animal feed and spiked at different levels of concentration. Malondialdehyde, 4-HHE, 4-HNE, crotonaldehyde, benzaldehyde, and hexanal can be analyzed in the same run in animal feed with a very good accuracy, with recovery rates ranging from 86 to 109% for a working range going from 0.16 to 12.50 mg/kg. The analysis of 2,4-nonadienal and 2,4-decadienal can also be performed but in a limited range of concentration and with a limited degree of accuracy. Their recovery rates ranged between 54 and 114% and coefficient of variation for the intermediate precision between 11 and 25% for these two compounds. Copyright

Performances of local poultry breed fed black soldier fly larvae reared on horse manure

In poultry, feed based on maggots, like larvae of black soldier fly (Hermetia illucens) is an attractive option to substitute current ingredients which are expensive and often in direct or indirect competition with human food. Little information is currently available on the utility of these larvae in poultry feed, so goals of this study were to determine whether larvae could be reared on horse manure under traditional farming conditions and to evaluate the growth performances of a local poultry fed these larvae and the fatty acids profiles of their meat. After freezing and thawing, larvae were introduced in the feed of Ardennaise chickens between 30 and 80 days of age. Birds in the control group received a commercial standard feed, while those in the treatment group received the same commercial feed in which 8% was substituted with whole fresh larvae corresponding to 2% on a dry matter basis. Means ± standard errors of larval length and weight were 20.67 ± 2.21 mm and 0.14 ± 0.02 g, respectively. Mean larval percentages of dry matter and of substances extractable in diethyl ether were 24.6% and 23.1%, respectively. Larval fatty acids profiles were predominantly composed of lauric acid (28.1%) and palmitic acid (22.0%). Least squares means of weekly weights of chicken, adjusted for the effects of sex, replication and initial weights, were significantly higher (P < 0.05) by 77.03 ± 53.37 g in larvae-fed than in control chickens. All other measurements were not statistically different between larvae-fed and control chicken, including fatty acid profiles, protein content and ω6/ω3 ratio.

Development of an HPTLC method for determination of hypoglycin A in aqueous extracts of seedlings and samaras of Acer species.

Hypoglycin A (HGA) is a toxin contained in seeds of the sycamore maple tree (Acer pseudoplatanus). Ingestion of this amino acid causes equine atypical myopathy (AM) in Europe. Another variety, A. negundo, is claimed to be present where AM cases were reported in the US. For unknown reasons, occurrence of this disease has increased. It is important to define environmental key factors that may influence toxicity of samaras from Acer species. In addition, the content of HGA in seedlings needs to be determined since AM outbreaks, during autumn period when the seeds fall but also during spring when seeds are germinating. The present study aims to validate a reliable method using high performance thin layer chromatography for determination and comparison of HGA in samaras and seedlings. The working range of the method was between 20 μg HGA to 408 μg HGA per ml water, corresponding to 12 - 244 mg/kg fresh weight or 40 - 816 mg/kg dry weight, taking into account of an arbitrary average dry matter content of 30%. Instrumental limit of detection and limit of quantification were of 10 μg HGA/ml and 20 μg HGA/ml water, respectively. Instrumental precision was 4% (RSD on 20 repeated measurements) while instrumental accuracy ranged between 86% and 121% of expected value. The HGA recovery of the analytical method estimated from spiked samaras and seedlings samples ranged between 63 and 103%. The method was applied to 9 samples of samaras from Acer pseudoplatanus, A. platanoides and A. campestre and 5 seedlings samples from A. pseudoplatanus. The results confirm detection of HGA in samaras from A. pseudoplatanus and the absence of detection in samaras of other tested species. They also suggest that detected levels of HGA are highly variable. This confirmed the suitability of the method for HGA detection in samaras or seedling.

Sources and fate of antimicrobials in integrated fish-pig and non-integrated tilapia farms

Antimicrobial contamination in aquaculture products constitutes a food safety hazard, but little is known about the introduction and accumulation of antimicrobials in integrated fish-pig aquaculture. This study, conducted in 2013, aimed to determine the residues of 11 types of antimicrobials by UPLC-MS/MS analysis in fish feed (n=37), pig feed (n=9), pig manure (n=9), pond sediment (n=20), fish skin (n=20) and muscle tissue (n=20) sampled from integrated tilapia-pig farms, non-integrated tilapia farms and fish feed supply shops. There was a higher occurrence of antimicrobial residues in fish skin from both integrated and non-integrated farms, and in pig manure. Enrofloxacin (3.9-129.3μg/kg) and sulfadiazine (0.7-7.8μg/kg) were commonly detected in fish skin and muscle, pig manure and pond sediment from integrated farms, with different types of antimicrobials found in pig manure and tilapia samples. In non-integrated farms, sulfadiazine (2.5-89.9μg/kg) was the predominant antimicrobial detected in fish skin and muscle, fish feed and pond sediment. In general, antimicrobials seemed not to be commonly transmitted from pig to fish in tilapia-pig integrated farms, and fish feed, pig feed and pond sediment did not seem as important sources of the antimicrobials found in fish from both systems. The frequent findings of antimicrobial residues in fish skin compared with fish muscle was probably due to different pharmacokinetics in different tissue types, which have practical food safety implications since antimicrobial residues monitoring is usually performed analyzing mixed skin and fish muscle samples.

Bioconcentration and half-life of quinalphos pesticide in rice-fish integration system in the Mekong Delta, Vietnam

ABSTRACT In order to determine the distribution and enable the elimination of quinalphos, a popular active pesticide compound used in the Mekong Delta, an experiment was set up in a rice-fish integration system in Can Tho City, Vietnam. Fish was stocked into the field when the rice was two-months old. Quinalphos was applied twice in doses of 42.5 g per 1000 m2. Water, fish and sediment samples were collected at time intervals and analyzed by a Gas Chromatography Electron Capture Detector system. The results show that quinalphos residues in fish muscles were much higher than those of the water and the bioconcentration factor (logBCF) was above 2 for the fish. The half-life of first and second quinalphos applications were 12.2 and 11.1 days for sediment, 2.5 and 1.1 days for silver barb, 1.9 and 1.3 days for common carp, and 1.1 and 1.0 days for water, respectively.

The use of antibiotics in cattle in North-East Benin: pharmaceutical inventory and risk practices of cattle breeders

This study’s aim is to inventory antibiotics used in cattle in North-East Benin and assess risk practices that could be the cause of both food chain contamination by antibiotic residues and selection of antibiotic-resistant bacteria in animals and humans. A survey was conducted among 98 cattle breeders in the districts of Banikoara, Kandi, Bembereke, and Kalale in North Benin. Semi-structured interviews were conducted, covering breeder status, breeding system, and antibiotic use. Multiple correspondence analysis and hierarchical classification analysis were conducted to establish a breeder typology. Breeders mainly belonged to the Fulani ethnic group (71.4 ± 8.9%) and almost all of them received “no formal education” (96.9 ± 3.4%). Cattle herds were mainly composed of a single breed, the Borgou (76.4 ± 8.1%) or the Fulani Zebu (16.0 ± 7.0%). Some herds were mixed. Antibiotics groups used in cattle breeding were tetracyclines, beta-lactams, sulfonamides, aminoglycosides, and macrolides, used by respectively 100%, 69.4 ± 9.1%, 56.1 ± 9.8%, 44.9 ± 9.8%, and 34.7 ± 9.4% of breeders. These drugs were purchased in local markets (59.0 ± 15.4%) and veterinary pharmacy (41.0 ± 15.4%). They were mainly used against respiratory diseases, lameness, mastitis, omphalitis and neonatal enteritis, and skin diseases. Only 49.0 ± 9.9% of breeders seek veterinary services to treat animals and 92.9 ± 5.1% of them did not respect antibiotic withdrawal times. These practices suggest that both contamination of bovine meat with antibiotic residues and selection of resistant bacteria are to be expected, resulting in adverse health effects on consumers.

Safety of ready-to-eat chicken in Burkina Faso: Microbiological quality, antibiotic resistance, and virulence genes in Escherichia coli isolated from chicken samples of Ouagadougou

Abstract In Burkina Faso, flamed/grilled chickens are very popular and well known to consumers. The aim of this study was to evaluate the microbiological quality, the antibiotic resistance, and the virulence gene from Escherichia coli isolated from these chickens in Ouagadougou. A total of 102 grilled, flamed, and fumed chickens were collected in Ouagadougou and analyzed, using standard microbiological methods. All E. coli isolates were checked with the antimicrobial test and also typed by 16‐plex PCR. The mean of aerobic mesophilic bacteria (AMB) and thermo‐tolerant coliforms (TTC) was found respectively between 6.90 ± 0.12 × 107 CFU/g to 2.76 ± 0.44 × 108 CFU/g and 2.4 ± 0.82 × 107 CFU/g to 1.27 ± 0.9 × 108 CFU/g. E. coli strains were found to 27.45%. Forty samples (38.24%) were unacceptable based on the AMB load. Fifty‐nine samples (57.85%) were contaminated with TTCs. Low resistance was observed with antibiotics of betalactamin family. Diarrheagenic E. coli strains were detected in 21.43% of all samples. This study showed that flamed/grilled chickens sold in Ouagadougou could pose health risks for the consumers. Need of hygienic practices or system and good manufacturing practices is necessary to improve the hygienic quality of flamed/grilled chickens. Our results highlight the need of control of good hygiene and production practices to contribute to the improvement of the safety of the products and also to avoid antibiotic resistance. Slaughter, scalding, evisceration, plucking, bleeding, washing, rinsing, preserving, grilling, and selling may be the ways of contamination.

Physiological and proteomic responses to corticosteroid treatments in Eurasian perch, Perca fluviatilis: Investigation of immune-related parameters

The comparative effects of cortisol and 11-deoxycorticosterone (DOC), two major corticosteroids in fish, have yet received little attention in teleosts. We evaluated the proteomic and immune responses of Eurasian perch to chronic corticosteroid treatments. We implanted immature perch with cortisol (80mg/kg) or DOC (4mg/kg) and measured the proportions of blood leucocytes, immune indices in the plasma, spleen and liver (complement and lysozyme activity, total immunoglobulin and immune gene expression in the tissues) and differential proteome expression (corticosteroid versus control) in the liver and the spleen on days 2, 4 and 14 post-treatment. Implantation of cortisol decreased the ratio of blood leucocytes and depressed Ig levels in both organs while DOC modulated the proportion of leucocyte sub-populations (increase in lymphocytes and decrease in granulocytes). In contrast, the innate humoral immunity was not strongly influenced by any of corticosteroid implants. The only immune parameter that was significantly affected was lysozyme, after DOC treatment. A number of proteins were differentially regulated by these hormones and some were identified in the liver (21 for cortisol and 8 for DOC) and in the spleen (10 for cortisol and 10 for DOC). None of the proteins was directly linked to immunity, except the natural killer enhancing factor, which was repressed by cortisol in the spleen. Our results also confirm that the proteins involved in energetic and glucose metabolism are affected by corticosteroids. Furthermore, these corticosteroids differently regulate immune status in Eurasian perch and they primarily impact leucocytes, as opposed to innate immune function.

Assessment of methods of detection of water estrogenicity for their use as monitoring tools in a process of estrogenicity removal

ABSTRACT Methods of monitoring of estrogenicity in water were gathered, compared, and tested within the context of their practical use as measurement and design tools, in the development of a process of degradation of estrogenic endocrine disruptors. In this work, the focus was put on in vitro assays, with the use of analytical techniques as additional analysis when possible. Practically, from a literature review, four methods that seemed most suitable to practical use required in a process development were tested: the Yeast Estrogen Screen assay, the Lyticase-assisted Yeast Estrogen Screen assay (LYES), the MMV-LUC assay and the HPLC-UV analytical method. Dose–response curves in response to estrogenic standard 17β-estradiol were compared. Bisphenol A estrogenicity was measured by the methods as well. The model for the calculation of estradiol equivalents as measurements units was adapted. The methods were assessed in terms of ranges of detection, time of experiment, cost, ease of the experiment, reproducibility, etc. Based on that assessment, the LYES assay was selected and successfully applied to the monitoring of estrogenicity removal from 17β-estradiol and bisphenol A. More precisely, the bioassay allowed the acquisition of kinetic curves for a laboratory-scaled process of estrogenicity removal by immobilized enzymes in a continuous packed-bed reactor. The LYES assay was found to have a real methodological potential for scale-up and design of a treatment process. The HPLC-UV method showed good complementarity with the LYES assay for the monitoring of bisphenol A concentrations in parallel with estrogenicity, reporting no significant estrogenicity from degradation byproducts, among others.