Statement on the toxicological properties and maximum residue levels of acetamiprid and its metabolites.

Acetamiprid is a pesticide active substance with insecticidal action whose approval was renewed by Commission Implementing Regulation (EU) 2018/113. In January 2022, the EFSA PPR Panel published a statement following a request from the European Commission to advise on human health or the environment based on new scientific evidence presented by France during the decision-making phase. In July 2022, by means of a further mandate received from the European Commission, EFSA was requested to provide advice if new information and any other scientific evidence that has become available since the assessment conducted for the renewal in 2018 warrant re-evaluation of (i) toxicological parameters used for the risk assessment of acetamiprid during the renewal process, including toxicological endpoints; (ii) the residue definition for acetamiprid in products of plant origin; and (iii) the safety of existing maximum residue levels (MRLs). Meanwhile, the applicant of acetamiprid in the EU submitted new toxicology studies regarding the toxicological profile of the metabolite IM-2-1. Furthermore, the European Commission was made aware that several recent publications in scientific literature were made available after the literature searches conducted by EFSA. As the new data could affect the advice that EFSA was expected to deliver through the 2022 mandate, EFSA was further requested to consider this information by means of a revised mandate received in September 2023. As regards re-evaluation of point (i) in this statement, this was addressed by an EFSA Working Group integrating all the available evidence. The results of the weight of evidence indicated that there are major uncertainties in the body of evidence for the developmental neurotoxicity (DNT) properties of acetamiprid and further data are therefore needed to come to a more robust mechanistic understanding to enable appropriate hazard and risk assessment. In view of these uncertainties, the EFSA WG proposed to lower the acceptable daily intake (ADI) and acute reference dose (ARfD) from 0.025 to 0.005 mg/kg body weight (per day). A revised residue definition for risk assessment was proposed for leafy and fruit crops as sum of acetamiprid and N-desmethyl-acetamiprid (IM-2-1), expressed as acetamiprid. Regarding pulses/oilseeds, root crops and cereals, the new data received did not indicate a need to modify the existing residue definition for risk assessment, which therefore remains as parent acetamiprid. Regarding the residue definition for enforcement, the available data did not indicate a need to modify the existing definition because acetamiprid is still a sufficient marker of the residues in all crop groups. Considering the new health-based guidance values derived in the present statement, a risk for consumer has been identified for 38 MRLs currently in place in the EU Regulation. Consequently, EFSA recommended to lower the existing MRLs for 38 commodities based on the assessment of fall-back Good Agricultural Practices received within an ad hoc data call. Some fall-back MRLs proposals require further risk management considerations.

Hazard characterization of Alternaria toxins to identify data gaps and improve risk assessment for human health.

Fungi of the genus Alternaria are ubiquitous plant pathogens and saprophytes which are able to grow under varying temperature and moisture conditions as well as on a large range of substrates. A spectrum of structurally diverse secondary metabolites with toxic potential has been identified, but occurrence and relative proportion of the different metabolites in complex mixtures depend on strain, substrate, and growth conditions. This review compiles the available knowledge on hazard identification and characterization of Alternaria toxins. Alternariol (AOH), its monomethylether AME and the perylene quinones altertoxin I (ATX-I), ATX-II, ATX-III, alterperylenol (ALP), and stemphyltoxin III (STTX-III) showed in vitro genotoxic and mutagenic properties. Of all identified Alternaria toxins, the epoxide-bearing analogs ATX-II, ATX-III, and STTX-III show the highest cytotoxic, genotoxic, and mutagenic potential in vitro. Under hormone-sensitive conditions, AOH and AME act as moderate xenoestrogens, but in silico modeling predicts further Alternaria toxins as potential estrogenic factors. Recent studies indicate also an immunosuppressive role of AOH and ATX-II; however, no data are available for the majority of Alternaria toxins. Overall, hazard characterization of Alternaria toxins focused, so far, primarily on the commercially available dibenzo-α-pyrones AOH and AME and tenuazonic acid (TeA). Limited data sets are available for altersetin (ALS), altenuene (ALT), and tentoxin (TEN). The occurrence and toxicological relevance of perylene quinone-based Alternaria toxins still remain to be fully elucidated. We identified data gaps on hazard identification and characterization crucial to improve risk assessment of Alternaria mycotoxins for consumers and occupationally exposed workers.

Determination of the elimination half-life of Glyphosate and its main metabolite, AMPA, in chicken plasma.

Glyphosate-based herbicides (GBHs) are the most-used herbicides worldwide. Concerns about their toxicity and ecotoxicity have motivated scientists to assess their potential effects on animals, as well as their toxicokinetic parameters in rats and humans. However, to our knowledge, such data have not been produced for avian models. In this study, toxicokinetic parameters for glyphosate and AMPA were calculated after one unique dietary exposure (40 mg of glyphosate equivalent per kg) and one unique intravenous injection of a GBH, in hens and roosters respectively. Non compartmental analysis was used to show the evolution of glyphosate and AMPA plasma concentrations over time. After one unique intravenous injection of a glyphosate-based herbicide, glyphosate and AMPA were quickly eliminated from plasma and were poorly distributed (Vssglyphosate = 0.30 L/kg). Their terminal half-lives are 4.7 h and 8.10 h, respectively. After dietary exposure, glyphosate and AMPA followed a 6 h absorption phase followed by a 42 h elimination phase. They were poorly distributed (Vssglyphosate = 0.00562 L/kg), and their maximum concentrations (Cmax) were 21285 µg/L and 108 µg/L, respectively. Their terminal elimination half-lives were 8.94 h and 6.93 h, respectively. Taken together, this study provides new data on the elimination rate and approximate biological half-life range of glyphosate in birds.

PBPK Model To Predict Marbofloxacin Distribution in Edible Tissues and Intestinal Exposure in Pigs.

Marbofloxacin (MAR) is a fluoroquinolone antibiotic used in food-producing animals in European Union, especially in pigs. In this study, MAR concentrations in plasma, comestible tissues, and intestinal segments were determined in pigs injected with MAR. Based on these data and the literature, a flow-limited PBPK model was developed to predict the tissue distribution of MAR and estimate the withdrawal period after label-use in Europe. A submodel describing the different segments of the intestinal lumen was also developed to assess the intestinal exposure of MAR for the commensal bacteria. During model calibration, only four parameters were estimated. Then, Monte Carlo simulations were performed to generate a virtual population of pigs. The simulation results were compared with the observations from an independent data set during the validation step. A global sensitivity analysis was also carried out to identify the most influential parameters. Overall, the PBPK model was able to adequately predict the MAR kinetics in plasma and edible tissues, as well as in small intestines. However, the simulated concentrations in the large intestine were mostly underestimated, highlighting the need for improvements in the field of PBPK modeling to assess the intestinal exposure of antimicrobials in food animals.

Ionophore coccidiostats - disposition kinetics in laying hens and residues transfer to eggs.

Poultry production is linked with the use of veterinary medicinal products to manage diseases. Ionophore coccidiostats have been permitted for use as feed additives within the European Union (EU) for the prevention of coccidiosis in various species of poultry with except of laying hens. The presence of chemical residues in eggs is a matter of major concern for consumers' health. Despite such prohibition of use in laying hens, they were identified as the most common non-target poultry species being frequently exposed to these class of coccidiostats. Many factors can influence the presence of residues in eggs. Carryover of these class of coccidiostat feed additives in the feed of laying hens has been identified as the main reason of their occurrence in commercial poultry eggs. The physicochemical properties of individual compounds, the physiology of the laying hen, and the biology of egg formation are believed to govern the residue transfer rate and its distribution between the egg white and yolk compartments. This paper reviews the causes of occurrence of residues of ionophore coccidiostats in eggs within the EU with special emphasis on their disposition kinetics in laying hens, and residue transfer into eggs. Additional effort was made to highlight future modeling perspectives on the potential application of pharmacokinetic modeling in predicting drug residue transfer and its concentration in eggs.

Permeability of the Cyanotoxin Microcystin-RR across a Caco-2 Cells Monolayer.

Microcystins (MCs) are toxins produced by several cyanobacterial species found worldwide. While MCs have a common structure, the variation of two amino acids in their structure affects their toxicity. As toxicodynamics are very similar between the MC variants, their differential toxicity could rather be explained by toxicokinetic parameters. Microcystin-RR (MC-RR) is the second most abundant congener and induces toxicity through oral exposure. As intestinal permeability is a key parameter of oral toxicokinetics, the apparent permeability of MC-RR across a differentiated intestinal Caco-2 cell monolayer was investigated. We observed a rapid and large decrease of MC-RR levels in the donor compartment. However, irrespective of the loaded concentration and exposure time, the permeabilities were very low from apical to basolateral compartments (from 4 to 15 × 10-8 cm·s-1) and from basolateral to apical compartments (from 2 to 37 × 10-8 cm·s-1). Our results suggested that MC-RR would be poorly absorbed orally. As similar low permeability was reported for the most abundant congener microcystin-LR, and this variant presented a greater acute oral toxicity than MC-RR, we concluded that the intestinal permeability was probably not involved in the differential toxicity between them, in contrast to the hepatic uptake and metabolism.

A PBPK model to study the transfer of α-hexabromocyclododecane (α-HBCDD) to tissues of fast- and slow-growing broilers.

A physiologically based pharmacokinetic (PBPK) model was developed to investigate the production-specific factors involved in the transfer of α-hexabromocyclododecane (α-HBCDD) to broiler meat. The model describes growth and lipid deposition in tissues of fast- (FG) and slow- (SG) growing broilers from hatching to slaughter and simulates the exposure through the ingestion of contaminated feed or expanded polystyrene insulation material. Growth parameters were obtained from the literature while parameters relative to uptake, distribution, and elimination of α-HBCDD were adjusted using results of a previous experiment involving broilers exposed through feed throughout the rearing period or allowed to depurate before slaughter. The model was used to compare the two main edible tissues, breast and leg meat, as well as skin, and to investigate the variability within strain. Between strains and within strain, α-HBCDD assimilation efficiency (AE) is higher when the animals are slaughtered young or heavy. However, increasing slaughter age will lower α-HBCDD concentration in tissues, due to dilution. Based on fresh weight, the concentration of α-HBCDD in breast muscles and skin tends to be lower in SG than in FG broilers (-30 to +10%), while it is 10% to 80% higher in leg muscles. Compared to breast muscles, consuming leg muscles would elicit an exposure 9 and 16 times higher in FG and SG broilers, respectively. The consumption of skin together with muscles would multiply the exposure by up to 36 times compared to breast muscle alone. In case of acute exposure, the α-HBCDD concentration in tissues increased sharply, all the more since the animals are lighter in weight, and then decreased rapidly. In FG broilers, dilution through growth contributed for up to 37%, 28% and 97% to the decontamination of breast muscles, leg muscles and skin, respectively, depending on the duration of depuration before slaughter.

A Population WB-PBPK Model of Colistin and its Prodrug CMS in Pigs: Focus on the Renal Distribution and Excretion.

PURPOSE: The objective was the development of a whole-body physiologically-based pharmacokinetic (WB-PBPK) model for colistin, and its prodrug colistimethate sodium (CMS), in pigs to explore their tissue distribution, especially in kidneys. METHODS: Plasma and tissue concentrations of CMS and colistin were measured after systemic administrations of different dosing regimens of CMS in pigs. The WB-PBPK model was developed based on these data according to a non-linear mixed effect approach and using NONMEM software. A detailed sub-model was implemented for kidneys to handle the complex disposition of CMS and colistin within this organ. RESULTS: The WB-PBPK model well captured the kinetic profiles of CMS and colistin in plasma. In kidneys, an accumulation and slow elimination of colistin were observed and well described by the model. Kidneys seemed to have a major role in the elimination processes, through tubular secretion of CMS and intracellular degradation of colistin. Lastly, to illustrate the usefulness of the PBPK model, an estimation of the withdrawal periods after veterinary use of CMS in pigs was made. CONCLUSIONS: The WB-PBPK model gives an insight into the renal distribution and elimination of CMS and colistin in pigs; it may be further developed to explore the colistin induced-nephrotoxicity in humans.

Lack of experimental evidence to support mcr-1-positive Escherichia coli strain selection during oral administration of colistin at recommended and higher dose given by gavage in weaned piglets.

In this study, we assessed the selective effect of colistin administered orally to healthy weaned piglets harbouring an intestinal mcr-1-positive Escherichia coli strain. Maximum recommended dose and a higher dose often used in European pig farms were given by gavage. No selection of the mcr-1-positive strain was observed in our controlled conditions, irrespective of the dose. Further investigations in real farming conditions seem necessary.

Low in vitro permeability of the cyanotoxin microcystin-LR across a Caco-2 monolayer: with identification of the limiting factors using modelling.

Microcystins (MCs) are toxins produced by several cyanobacteria species found worldwide. MC-LR is the most frequent. Here, we used the human Caco-2 cell line grown on semi-permeable filter supports as an in vitro model for determining MC-LR intestinal bidirectional transport. In this study, there was very low and time-dependent apparent permeability of MC-LR. To identify the limiting factors involved in the low permeability of MC-LR, a mathematical model was constructed to get physiologically relevant and informative parameters. The apical-to-basolateral transport was characterised by a rapid and substantial decrease in apical MC-LR concentrations (24-40% of the initial amount). In the basolateral compartment, the concentrations increased slowly after a lag time, but represented only a small fraction of the loaded concentrations (0.3-1.3%) after 24 h. This weak permeability was mainly due to a low clearance of efflux (from the cellular to the basolateral compartment) and effective secretion (from the cellular to the apical compartment). During the basolateral-to-apical transport, we observed a slow decrease in basolateral concentrations and a rapid increase in apical concentrations. In conclusion, modelling has the potential to highlight the key mechanisms involved in the complex kinetics of toxin transport.

Low or high doses of cefquinome targeting low or high bacterial inocula cure Klebsiella pneumoniae lung infections but differentially impact the levels of antibiotic resistance in fecal flora.

The combination of efficacious treatment against bacterial infections and mitigation of antibiotic resistance amplification in gut microbiota is a major challenge for antimicrobial therapy in food-producing animals. In rats, we evaluated the impact of cefquinome, a fourth-generation cephalosporin, on both Klebsiella pneumoniae lung infection and intestinal flora harboring CTX-M-producing Enterobacteriaceae. Germfree rats received a fecal flora specimen from specific-pathogen-free pigs, to which a CTX-M-producing Escherichia coli strain had been added. K. pneumoniae cells were inoculated in the lungs of these gnotobiotic rats by using either a low (10(5) CFU) or a high (10(9) CFU) inoculum. Without treatment, all animals infected with the low or high K. pneumoniae inoculum developed pneumonia and died before 120 h postchallenge. In the treated groups, the low-inoculum rats received a 4-day treatment of 5 mg/kg of body weight cefquinome beginning at 24 h postchallenge (prepatent phase of the disease), and the high-inoculum rats received a 4-day treatment of 50 mg/kg cefquinome beginning when the animals expressed clinical signs of infection (patent phase of the disease). The dose of 50 mg/kg targeting the high K. pneumoniae inoculum cured all the treated rats and resulted in a massive amplification of CTX-M-producing Enterobacteriaceae. A dose of 5 mg/kg targeting the low K. pneumoniae inoculum cured all the rats and averted an outbreak of clinical disease, all without any amplification of CTX-M-producing Enterobacteriaceae. These findings might have implications for the development of new antimicrobial treatment strategies that ensure a cure for bacterial infections while avoiding the amplification of resistance genes of human concern in the gut microbiota of food-producing animals.

Comparative cytotoxicity, oxidative stress, and cytokine secretion induced by two cyanotoxin variants, microcystin LR and RR, in human intestinal Caco-2 cells.

While MC-LR and MC-RR share significant structural similarity, MC-RR is less cytotoxic than MC-LR. In the current study, we have compared the effects of MC-LR and MC-RR in Caco-2 cells by evaluating cytotoxicity, oxidative stress (reactive oxygen species production), and the cellular proinflammatory response (IL-6 and IL-8 production). Following treatment with 100 µM microcystins (MC), cytotoxicity was two-fold greater with MC-LR as compared to MC-RR after 24 h exposure. Whereas the reactive oxygen species production and IL-6 secretion were similar following a 24-h treatment with either MC, 100 µM MC-LR induced a five-fold greater IL-8 secretion when compared to MC-RR. Our study has demonstrated that, although both MC-LR and MC-RR induced some cytotoxicity in human intestinal cells, a major difference in IL-8 production was observed between the two variants.

Liquid chromatography-electrospray tandem mass spectrometric method for quantification of monensin in plasma and edible tissues of chicken used in pharmacokinetic studies: applying a total error approach.

A liquid chromatography-tandem mass spectrometric (LC-MS/MS) method was developed and validated for use in pharmacokinetic studies in order to determine the concentrations of monensin in plasma and edible tissues of chicken. Two sample preparations were performed, one for determining monensin concentrations in plasma using acetonitrile for protein precipitation and another one for determining monensin concentrations in muscle, liver, and fat using methanol-water followed by a clean up on a solid-phase extraction cartridge. Sample extracts were injected into the LC-MS/MS system, and a gradient elution was performed on a C18 column. Narasin was used as internal standard. The LC-MS/MS method was validated using an approach based on accuracy profiles, and applicability of the method was demonstrated for the determination of monensin in chicken plasma, muscle, liver, and fat in a pharmacokinetic study.