We determined the distribution and temporal variation of Carbapenem Resistant Enterobacterales (CRE), carbapenemase-encoding genes and other antibiotic resistance genes (ARGs) in a highly polluted river (Lis River; Portugal), also assessing the potential influence of water quality to this distribution. Water samples were collected in two sampling campaigns performed one year apart (2018/2019) from fifteen sites and water quality was analyzed. CRE were isolated and characterized. The abundance of four ARGs (blaNDM, blaKPC, tetA, blaCTX₋M), two Microbial Source Tracking (MST) indicators (HF183 and Pig-2-Bac) and the class 1 integrase gene (IntI1) was measured by qPCR. confirmed the poor quality of the Lis River water, particularly in sites near pig farms. A collection of 23 CRE was obtained: Klebsiella (n = 19), Enterobacter (n = 2) and Raoultella (n = 2). PFGE analysis revealed a clonal relationship between isolates obtained in different sampling years and sites. All CRE isolates exhibited multidrug resistance profiles. Klebsiella and Raoultella isolates carried blaKPC while Enterobacter harbored blaNDM. Conjugation experiments were successful for only four Klebsiella isolates. All ARGs were detected by qPCR on both sampling campaigns. An increase in ARGs and IntI1 abundances was detected in sites located downstream of wastewater treatment plants. Strong correlations were observed between blaCTX₋M, IntI1 and the human-pollution marker HF183, and also between tetA and the pig-pollution marker Pig-2-bac, suggesting that both human- and animal-derived pollution in the Lis River are a potential source of ARGs. Plus, water quality parameters related to eutrophication and land use were significantly correlated with ARGs abundances. Our findings demonstrated that the Lis River encloses high levels of antibiotic resistant bacteria and ARGs, including CRE and carbapenemase-encoding genes. Overall, this study provides a better understanding on the impacts of water pollution resulting from human and animal activities on the resistome of natural aquatic systems.

Veterinary pharmaceuticals, including antibiotics, are emerging contaminants of concern worldwide. Avian scavengers are exposed to pharmaceuticals through consumption of livestock carcasses used for feeding wildlife for conservation purposes at supplementary feeding stations. Here we tested the hypothesis that griffon vultures (Gyps fulvus) would be more exposed to antibiotics (i.e., quinolones) when feeding on livestock carcasses from intensive farming than when they rely on carcasses from extensive farming or wild animals. We sampled 657 adult griffon vultures captured between 2008 and 2012. In addition, we sampled tissues from domestic livestock supplied at feeding stations in the study area between 2009 and 2019; pig (n = 114), sheep (n = 28), cow (n = 1) and goat (n = 2). Samples were analysed by liquid chromatography with electrospray ionization mass spectrometry (LC-ESI-MS). Quinolones were detected in plasma from 12.9% of the griffon vultures analysed. Quinolone prevalence in griffon vultures varied significantly among feeding stations but was also affected by the total amount of carcasses supplemented, especially the mass of pig carcasses. These results aligned with a 21.1% quinolone prevalence in pig carcasses sampled at feeding stations, wherein enrofloxacin and ciprofloxacin levels of up to 3359 ng/g and 1550 ng/g, respectively, were found. Given enrofloxacin pharmacokinetics in pig tissues, 5.3% of the analysed pigs may have died no more than one day after treatment. Quinolone presence in vultures was negatively associated with blood lead levels, which mostly originates from lead ammunition and indicates a higher consumption of game animal carcasses. Carcass disposal for feeding avian scavengers must always assess and manage the risks posed by veterinary pharmaceuticals, especially when livestock provided may have died soon after treatment.

A high demand exists in bisphenols (BPs) screening studies for quick, reliable and straightforward analytical methods that generate data faster and simultaneously. Herein, we describe a combination of enzymatic probe sonication (EPS) and ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) for quick extraction and simultaneous quantification of eight important BPs in animal-derived foods. Results obtained demonstrated that the ultrasonic probe power could not only enhance the enzymatic hydrolysis efficiency, but also accelerate the liquid-liquid extraction procedure. Under optimized EPS parameters, one sample could be exhaustively extracted within 120 s, as compared with 12 h needed for the conventional enzymatic extraction which is more suitable for high-throughput analysis. The method was successfully applied to analyze residual BPs in animal-derived foods collected from Beijing, China. Widespread occurrence of BPA, BPS, BPF, BPAF, BPP, and BPB were found, with detection frequencies of 65.2%, 42.4%, 33.7%, 29.4%, 28.3%, and 27.2%, respectively. The highest total concentration levels of BPs (sum of the eight BPs analyzed, ΣBPs) were found in chicken liver (mean 12.2 μg/kg), followed by swine liver (6.37 μg/kg), bovine muscle (3.24 μg/kg), egg (2.03 μg/kg), sheep muscle (2.03 μg/kg), chicken muscle (1.45 μg/kg), swine muscle (1.42 μg/kg), and milk (1.17 μg/kg). The estimated daily intake (EDI) of BPs, based on the mean and 95th percentile concentrations and daily food consumptions, was estimated to be 5.687 ng/kg bw/d and 22.71 ng/kg bw/d, respectively. The human health risk assessment in this work suggests that currently BPs do not pose significant risks to the consumers because the hazard index (HI) was <1.

Fipronil, a phenyl-pyrazole insecticide, has a wide range of uses, from agriculture to veterinary medicine. Due to its large-scale applications, the risk of environmental and occupational exposure and bioaccumulation raises concerns. Moreover, relatively little is known about the intracellular mechanisms of fipronil in trophoblasts and the endometrium involved in implantation. Here, we demonstrated that fipronil reduced the viability of porcine trophectoderm and luminal epithelial cells. Fipronil induced cell cycle arrest at the sub-G1 phase and apoptotic cell death through DNA fragmentation and inhibition of DNA replication. These reactions were accompanied by homeostatic changes, including mitochondrial depolarization and cytosolic calcium depletion. In addition, we found that exposure to fipronil compromised the migration and implantation ability of pTr and pLE cells. Moreover, alterations in PI3K-AKT and MAPK-ERK1/2 signal transduction were observed in fipronil-treated pTr and pLE cells. Finally, the antiproliferative and apoptotic effects of fipronil were also demonstrated in 3D cell culture conditions. In summary, our results suggest that fipronil impairs implantation potentials in fetal trophectoderm and maternal endometrial cells during early pregnancy.

Air and water quality at a concentrated animal feeding operation (CAFO) in Eastern North Carolina that uses a covered lagoon and anaerobic digester was evaluated for 2 weeks in August 2020. Real-time PM₂.₅ mass concentrations were determined using a reference ADR-1500 nephelometer and high-frequency measurements of dissolved inorganic nitrogen (DIN) were evaluated using autonomously logging sensors. Air and water quality parameters were assessed before, during and after wastewater from the lagoon was irrigated onto adjacent spray fields. Reference measurements were conducted alongside a HOBO weather station to collect real-time wind speed and direction, temperature, and humidity measurements. PM₂.₅ concentrations varied between 0 and 159 μg/m³ with an average concentration of 11 μg/m³, below EPA standard for secondary aerosols of 15 μg/m³. Higher PM₂.₅ concentrations were observed when wind originated from swine barns but not from covered lagoons. Water quality data showed that DIN concentrations downgradient from the CAFO were elevated relative to upstream concentrations. A groundwater seep that drains a spray field contained the highest average DIN concentration (31.0 ± 12.8 mg L⁻¹), which was 25 times greater than upstream DIN concentrations (1.2 ± 0.8 mg L⁻¹). Average DIN concentration at the downstream station was lower than the seep concentration (8.6 ± 16.2 mg L⁻¹), but approximately 8 times greater than upstream. Air quality data show that the lagoon cover was effective at mitigating air quality degradation, whereas DIN concentrations in water were similar to previous studies on CAFOs using open lagoons. In addition, air and water quality parameters were significantly (p < 0.001) higher after irrigation, indicating possible influence due to ammonia and nitrate elevation. Additional research is needed to compare high-frequency data collected from swine CAFOs using capped and uncapped lagoon systems to better understand spatiotemporal air and water quality trends of this practice.

Due to high global demand, large amounts of abbattoir waste are generated from pork production. Mismanagement of abattoir waste on agricultural lands can result in soil and water contamination with pathogens and contaminants like metals and nutrients. Therefore, possible effects on soil organisms prior to application should be evaluated. Thus, the aim of this study was to determine the effects of fresh pig abattoir waste (PAWf) and waste after stabilization processes on E. andrei through tests of avoidance behavior, acute toxicity and chronic toxicity. In order to do this, the waste was evaluated fresh (i.e., non-treated), and after aerated composting (PAWa), natural composting (PAWn) and vermicomposting (PAWv). In addition, we used a natural soil with no history of agricultural use as control soil. The evaluation was based on avoidance behavior, mortality, initial and final earthworm weight, and reproduction, in addition to a set of enzyme assays formed by acetylcholinesterase, lipid peroxidation, catalase and glutathione S-transferase measured over time. The ecotoxicological results showed that PAWf and PAWa increased AChE activity at different experimental periods, while PAWn decreased activity at 14 days compared to the control. PAWf and PAWa increased TBARS levels at 7 and 14 days, respectively. CAT activity decreased at 3, 7 and 14 days in PAWv, while GST activity increased at 3 days in PAWa and at 3 and 14 days in PAWf compared to the control. In the acute toxicity test, PAWa and PAWn had a toxic effect on E. andrei, resulting in 100% mortality at 14 days of exposure. Based on our findings, pig abattoir waste should undergo vermicomposting prior to agricultural application to soils.

Endocrine-disrupting chemicals (EDCs) are compounds that interfere with the expression, synthesis, and activity of hormones in organisms. They are released into the environment from flame retardants and products containing plasticizers. Persistent pesticides, such as dichlorodiphenyltrichloroethane (DDT) and hexachlorobenzene, also disrupt the endocrine system through interaction with hormone receptors. Endogenous hormones, such as 17β-estradiol (E2), are released in the urine and feces of farm animals and seep into terrestrial and aquatic ecosystems through sewage. Pigs are widely used as animal models to determine the effects of EDCs because they are physiologically, biochemically, and histologically similar to humans. EDCs primarily disrupt the reproductive and nervous systems of pigs. Moreover, embryonic development during the prenatal and early postnatal periods is particularly sensitive to EDCs. Mycotoxins, such as zearalenone, are food contaminants that alter hormonal activities in pigs. Mycotoxins also alter the innate immune system in pigs, making them vulnerable to diseases. It has been reported that farm animals are exposed to various types of EDCs, which accumulate in tissues, such as those of gonads, livers, and intestines. There is a lack of an integrated understanding of the impact of EDCs on porcine reproduction and development. Thus, this article aims to provide a comprehensive review of literature regarding the effects of EDCs in pigs.

The pollution of wastewater with antibiotics and antibiotics resistance genes has attracted public concerns about ecosystem and global health. Swine wastewater can contain high concentrations of antibiotics, especially sulfonamides, even after full-scale wastewater treatment. In this study, mesocosm-scale vertical flow constructed wetlands (VF-CWs) were applied to abate nutrients and antibiotics in swine wastewater containing sulfonamides. VF-CWs performed well in the removal of both nutrients and antibiotics. Sulfonamides did not influence total organic carbon (TOC) and total phosphorus (TP) removal, and even slightly enhanced NH₄⁺–N removal. High removal efficiencies (26.42–84.05%) were achieved for sulfadiazine (SDZ), sulfamethoxazole (SMX) and sulfamethazine (SMZ). Together with lab-scale sorption and biodegradation experiments, microbial degradation was found to be the most important removal mechanism for sulfonamides in VF-CWs. Sulfonamides addition increased bacterial alpha-diversity and changed microbial community structure. Moreover, antibiotics promoted antibiotic-resistant or -degrading bacteria. Bacillus, Geobacter and other seven genera were correlated with sulfonamides reduction under either aerobic or anaerobic condition. In summary, VF-CW is a suitable alternative for swine wastewater treatment, and biodegradation plays the key role in sulfonamides abatement.Main findings of the work.This was the first work to combine bacterial community analysis with microcosm experiments to uncover the major removal mechanism of sulfonamides in constructed wetlands.

The use of the phenicol antibiotic florfenicol in livestock can select for the optrA gene, which also confers resistance to the critically important oxazolidinone antibiotic linezolid. However, the occurrence and dissemination of florfenicol and linezolid cross-resistance genes in anaerobic treatment systems for livestock waste are unknown. Herein, the phenotypes and genotypes (optrA, fexA, fexB, and cfr) of florfenicol and linezolid cross-resistance were investigated in 339 enterococci strains isolated from lab- and full-scale mesophilic anaerobic digestion systems treating swine waste. It was found that optrA, fexA, and fexB were frequently detected in isolated enterococci in both systems by PCR screening, whereas cfr was not detected. The most abundant gene was optrA, which was detected in 73.5% (n = 50) and 38.9% (n = 23) of enterococci isolates in the full-scale influent and effluent, respectively. Most strains carried more than two resistance genes, and the average percentage of co-occurrence of optrA/fexA was 16.6%. Based on minimum inhibitory concentrations of the enterococci strain phenotypes, 85.7%, 77.5%, and 77.5% of strains in influent were resistant to chloramphenicol, florfenicol, and linezolid, respectively, while 56.3%, 65.2%, and 13% in the effluent isolates were found, respectively, which was consistent with the genotype results. The phenotypes and genotypes of florfenicol and linezolid resistance were relative stable in the enterococci isolated from the influent and effluent in lab-scale anaerobic digestion system. The findings signify the enterococci isolates harboring the optrA gene remained in effluents of both full- and lab-scale swine waste anaerobic digestion system; hence, effective management strategies should be implemented to prevent the discharge of antibiotic resistance from the livestock waste treatment systems.