How does active yeast supplementation reduce the deleterious effects of aflatoxins in Wistar rats? A radiolabeled assay and histopathological study.

The aim of this study was to investigate the mechanisms by which yeasts (Saccharomyces cerevisiae) control the toxic effects of aflatoxins, which are not yet fully understood. Radiolabeled aflatoxin B1 (AFB13H) was administered by gavage to Wistar rats fed with aflatoxin (AflDiet) and aflatoxin supplemented with active dehydrated yeast Y904 (AflDiet + Yeast). The distribution of AFB13H and its metabolites were analyzed at 24, 48 and 72 h by tracking back of the radioactivity. No significant differences were observed between the AflDiet and AflDiet + Yeast groups in terms of the distribution of labeled aflatoxin. At 72 h, for the AflDiet group the radiolabeled aflatoxin was distributed as following: feces (79.5%), carcass (10.5%), urine (1.7%), and intestine (7.4%); in the AflDiet + Yeast the following distribution was observed: feces (76%), carcass (15%), urine (2.9%), and intestine (4.9%). These values were below 1% in other organs. These findings indicate that even after 72 h considerable amounts of aflatoxins remains in the intestines, which may play a significant role in the distribution and metabolism of aflatoxins and its metabolites over time. The presence of yeast may not significantly affect this process. Furthermore, histopathological examination of hepatic tissues showed that the presence of active yeast reduced the severity of liver damage caused by aflatoxins, indicating that yeasts control aflatoxin damage through biochemical mechanisms. These findings contribute to a better understanding of the mechanisms underlying the protective effects of yeasts against aflatoxin toxicity.

Modulation of Broiler Intestinal Changes Induced by Clostridium perfringens and Deoxynivalenol through Probiotic, Paraprobiotic, and Postbiotic Supplementation.

Deoxynivalenol (DON) is a predisposing factor for necrotic enteritis. This study aimed to investigate the effects of a DON and Clostridium perfringens (CP) challenge on the intestinal morphology, morphometry, oxidative stress, and immune response of broilers. Additionally, we evaluated the potential of a Lactobacillus spp. mixture as an approach to mitigate the damage induced by the challenge. One-day-old broiler chickens (n = 252) were divided into seven treatment groups: Control, DON, CP, CP + DON, VL (DON + CP + viable Lactobacillus spp. mixture), HIL (DON + CP + heat-inactivated Lactobacillus spp. mixture), and LCS (DON + CP + Lactobacillus spp. mixture culture supernatant). Macroscopic evaluation of the intestines revealed that the CP + DON group exhibited the highest lesion score, while the VL and HIL groups showed the lowest scores. Microscopically, all Lactobacillus spp. treatments mitigated the morphological changes induced by the challenge. DON increased levels of reactive oxygen species (ROS) in the jejunum, and CP increased ROS levels in the jejunum and ileum. Notably, the Lactobacillus spp. treatments did not improve the antioxidant defense against CP-induced oxidative stress. In summary, a Lactobacillus spp. mixture, whether used as a probiotic, paraprobiotic, or postbiotic, exerted a partially protective effect in mitigating most of the intestinal damage induced by DON and CP challenges.

Resistance and Virulence Surveillance in Escherichia coli Isolated from Commercial Meat Samples: A One Health Approach.

Escherichia coli is a key indicator of food hygiene, and its monitoring in meat samples points to the potential presence of antimicrobial-resistant strains capable of causing infections in humans, encompassing resistance profiles categorized as serious threats by the Centers for Disease Control and Prevention (CDC), such as Extended-Spectrum Beta-Lactamase (ESBL)-a problem with consequences for animal, human, and environmental health. The objective of the present work was to isolate and characterize ESBL-producing E. coli strains from poultry, pork, and beef meat samples, with a characterization of their virulence and antimicrobial resistance profiles. A total of 450 meat samples (150 chicken, 150 beef, and 150 pork) were obtained from supermarkets and subsequently cultured in medium supplemented with cefotaxime. The isolated colonies were characterized biochemically, followed by antibiogram testing using the disk diffusion technique. Further classification involved biofilm formation and the presence of antimicrobial resistance genes (blaCTX-M, AmpC-type, mcr-1, and fosA3), and virulence genes (eaeA, st, bfpA, lt, stx1, stx2, aggR, iss, ompT, hlyF, iutA, iroN, fyuA, cvaC, and hylA). Statistical analysis was performed via the likelihood-ratio test. In total, 168 strains were obtained, with 73% originating from chicken, 22% from pork, and 17% from beef samples. Notably, strains exhibited greater resistance to tetracycline (51%), ciprofloxacin (46%), and fosfomycin (38%), apart from ß-lactams. The detection of antimicrobial resistance in food-isolated strains is noteworthy, underscoring the significance of antimicrobial resistance as a global concern. More than 90% of the strains were biofilm producers, and strains carrying many ExPEC genes were more likely to be biofilm formers (OR 2.42), which increases the problem since the microorganisms have a greater chance of environment persistence and genetic exchange. Regarding molecular characterization, bovine samples showed a higher prevalence of blaCTX-M-1 (OR 6.52), while chicken strains were more likely to carry the fosA3 gene (OR 2.43, CI 1.17-5.05) and presented between 6 to 8 ExPEC genes (OR 2.5, CI 1.33-5.01) compared to other meat samples. Concerning diarrheagenic E. coli genes, two strains harbored eae. It is important to highlight these strains, as they exhibited both biofilm-forming capacities and multidrug resistance (MDR), potentially enabling colonization in diverse environments and causing infections. In conclusion, this study underscores the presence of ß-lactamase-producing E. coli strains, mainly in poultry samples, compared to beef and pork samples. Furthermore, all meat sample strains exhibited many virulence-associated extraintestinal genes, with some strains harboring diarrheagenic E. coli (DEC) genes.

Guar gum as galactomannan source induces dysbiosis and reduces performance in broiler chickens and dietary ß-mannanase restores the gut homeostasis.

Galactomannans are abundant nonstarch polysaccharides in broiler feed ingredients. In broilers, diets with high levels of galactomannans have been associated with innate immune response stimulation, poor zootechnical performance, nutrient and lipid absorption, and excessive digesta viscosity. However, data about its effects on the gut microbiome are scarce. ß-Mannanases are enzymes that can hydrolyze ß-mannans, resulting in better nutrient utilization. In the current study, we have evaluated the effect of guar gum, a source of galactomannans, supplemented to broiler diets, either with or without ß-mannanase supplementation, on the microbiota composition, in an attempt to describe the potential role of the intestinal microbiota in ß-mannanase-induced gut health and performance improvements. One-day-old broiler chickens (n = 756) were randomly divided into 3 treatments: control diet, guar gum-supplemented diet (1.7%), or guar gum-supplemented diet + ß-mannanase (Hemicell 330 g/ton). The zootechnical performance, gut morphometry, ileal and cecal microbiome, and short-chain fatty acid concentrations were evaluated at different time points. The guar gum supplementation decreased the zootechnical performance, and the ß-mannanase supplementation restored performance to control levels. The mannan-rich diet-induced dysbiosis, with marked effects on the cecal microbiota composition. The guar gum-supplemented diet increased the cecal abundance of the genera Lactobacillus, Roseburia, Clostridium sensu stricto 1, and Escherichia-Shigella, and decreased Intestinimonas, Alistipes, Butyricicoccus, and Faecalibacterium. In general, dietary ß-mannanase supplementation restored the main microbial shifts induced by guar gum to levels of the control group. In addition, the ß-mannanase supplementation reduced cecal isobutyric, isovaleric, valeric acid, and branched-chain fatty acid concentrations as compared to the guar gum-supplemented diet group, suggesting improved protein digestion and reduced cecal protein fermentation. In conclusion, a galactomannan-rich diet impairs zootechnical performance in broilers and results in a diet-induced dysbiosis. ß-Mannanase supplementation restored the gut microbiota composition and zootechnical performance to control levels.

High-Frequency Detection of fosA3 and bla CTX-M-55 Genes in Escherichia coli From Longitudinal Monitoring in Broiler Chicken Farms.

Considering the worrying emergence of multidrug resistance, including in animal husbandry and especially in food-producing animals, the need to detect antimicrobial resistance strains in poultry environments is relevant, mainly considering a One Health approach. Thus, this study aimed to conduct longitudinal monitoring of antimicrobial resistance in broiler chicken farms, with an emphasis on evaluating the frequency of resistance to fosfomycin and ß-lactams. Escherichia coli was isolated from broiler chicken farms (cloacal swabs, meconium, poultry feed, water, poultry litter, and Alphitobius diaperinus) in northern Paraná from 2019 to 2020 during three periods: the first period (1st days of life), the second period (20th to 25th days of life), and third period (40th to 42nd days of life). Antibiogram tests and the detection of phenotypic extended-spectrum ß-lactamase (ESBL) were performed, and they were confirmed by seaching for genes from the bla CTX-M group. The other resistance genes searched were mcr-1 and fosA3. Some ESBL bla CTX-M-1 group strains were selected for ESBL identification by sequencing and enterobacterial repetitive intergenic consensus-polymerase chain reaction analysis. To determine the transferability of the bla CTX-M-1- and fosA3-carrying plasmids, strains were subjected to conjugation experiments. A total of 507 E. coli were analyzed: 360 from cloacal swabs, 24 from meconium samples, 3 from poultry feed samples, 18 from water samples, 69 from poultry litter samples, and 33 from A. diaperinus samples. Among the strain isolate, 80% (406/507) were multidrug-resistant (MDR), and 51% (260/507) were ESBL-positive, with the bla CTX-M-1 group being the most frequent. For the fosA3 gene, 68% (344/507) of the strains isolated were positive, deserves to be highlighted E. coli isolated from day-old chickens (OR 6.34, CI 2.34-17.17), when compared with strains isolated from other origins (poultry litter, A. diaperinus, water, and poultry feed). This work alerts us to the high frequency of the fosA3 gene correlated with the CTX-M-1 group (OR 3.57, CI 95% 2.7-4.72, p < 0.05), especially the bla CTX-M-55 gene, in broiler chickens. This profile was observed mainly in day-old chicken, with a high percentage of E. coli that were MDR. The findings emphasize the importance of conducting longitudinal monitoring to detect the primary risk points during poultry production.

Tryptophan Attenuates the Effects of OTA on Intestinal Morphology and Local IgA/IgY Production in Broiler Chicks.

Ochratoxin A (OTA) is a mycotoxin produced by species of Penicillium and Aspergillus that can contaminate products of plant origin that are used as animal feed. Through oral exposure, this mycotoxin primarily affects the chicken gastrointestinal system. The present study evaluated the intestinal toxic effects of OTA and the introduction of L-tryptophan to alleviate these effects in chickens. One-day-old chicks were exposed to a single OTA dose (1.4 mg/kg body weight-b.w.) and treated with or without four daily doses of L-tryptophan (100 mg/kg b.w.). Duodenal villus height/crypt depth, fecal immunoglobulin A/immunoglobulin Y (IgA/IgY) levels, and duodenal positive immunoglobulin A cells (IgA+) were evaluated by histology, ELISA, and immunohistochemistry, respectively, on the 14th day. There were significant changes in the duodenal villus height, crypt depth, and levels of fecal IgA/IgY and duodenal IgA+ cells (p < 0.05) in groups exposed to OTA. On the other hand, groups exposed to OTA and treated with L-tryptophan showed similar levels of villus height, IgA/IgY levels, and duodenal IgA+ cells to those of the control group (p > 0.05). In conclusion, exposure to a single dose of OTA orally induces changes in intestinal morphology, levels of IgA/IgY antibodies, and IgA+ cells. Thus, treatment with L-tryptophan may be a valid alternative means to reduce the harmful effects of OTA on the intestinal mucosa, which requires further study.

Proteus mirabilis causing cellulitis in broiler chickens.

Given the need to understand the virulence profile of Proteus mirabilis isolates from cellulitis in broiler chickens and their ability to cause lesions, the present study aimed to characterize genotypically and phenotypically the virulence profiles of two strains of P. mirabilis isolated from cellulitis in broilers, as well as to evaluate their ability to experimentally reproduce the lesions in vivo. The strain with the highest virulence potential (LBUEL-A33) possessed mrpA, pmfA, ucaA, atfA (fimbriae), zapA, ptA (proteases), hpmA (hemolysin), and ireA (siderophore) genes, formed a very strong biofilm, and expressed the pattern of aggregative adhesion and cytotoxicity in Vero cells. The strain with the lowest virulence potential (LBUEL-A34) did not present the pmfA and ucaA genes, but expressed the pattern of aggregative adhesion, formed a strong biofilm, and did not show cytotoxicity. Both strains developed cellulitis in an animal model within 24 h post-inoculation (PI), and the degree of lesions was not significantly altered up to 120 h PI. The LBUEL-A33 strain was also inoculated in combination with an avian pathogenic Escherichia coli (APEC 046), and the lesions showed no significant changes from the individual inoculation of these two strains. Histological analysis showed that the LBUEL-A33 strain developed characteristic cellulitis lesions. Thus, both strains of P. mirabilis isolated in our study have several virulence factors and the ability to develop cellulitis in broilers.

Genotypic and phenotypic profiles of virulence factors and antimicrobial resistance of Proteus mirabilis isolated from chicken carcasses: potential zoonotic risk.

Proteus mirabilis is an opportunistic pathogen often associated with a variety of human infections acquired both in the community and in hospitals. In this context, the present work aimed to evaluate the genotypic and phenotypic characteristics of the virulence factors and antimicrobial resistance determinants of 32 P. mirabilis strains isolated from chicken carcasses in a poultry slaughterhouse in the north of the state of Paraná, Brazil, in order to assess a potential zoonotic risk. The isolates presented a variety of virulence genes that contribute to the development of infection in humans. The mrpA, pmfA, atfA (fimbriae), ireA (siderophores receptor), zapA, ptA (Proteases), and hpmA (hemolysin) genes were found in 32 (100%) isolates and ucaA (fimbriae) in 16 (50%). All isolates showed aggregative adherence in HEp-2 cells and formed biofilms. Of all strains, 27 (84.38%) showed cytotoxic effects in Vero cells. Antimicrobial susceptibility was tested using 20 antimicrobials, in which 25 (78.13%) strains were considered multidrug-resistant. The presence of blaESBL and blaampC genes conferring resistance to ß-lactams and qnr to quinolones were also detected in the isolates after presumption in the phenotypic test, in which 7 (21.88%) isolates contained the CTX-M-2 group, 11 (34.38%) contained CIT group and 19 (59.38%) contained qnrD. Therefore, chicken carcasses contaminated with P. mirabilis may pose a health risk to the consumer, as these isolates have a variety of virulence and antimicrobial resistance characteristics that can be found in P. mirabilis strains isolated from human infections.

Identification and adhesion profile of Lactobacillus spp. strains isolated from poultry.

In the aviculture industry, the use of Lactobacillus spp. as a probiotic has been shown to be frequent and satisfactory, both in improving bird production indexes and in protecting intestine against colonization by pathogenic bacteria. Adhesion is an important characteristic in selecting Lactobacillus probiotic strains since it impedes its immediate elimination to enable its beneficial action in the host. This study aimed to isolate, identify and characterize the in vitro and in vivo adhesion of Lactobacillus strains isolated from birds. The Lactobacillus spp. was identified by PCR and sequencing and the strains and its adhesion evaluated in vitro via BMM cell matrix and in vivo by inoculation in one-day-old birds. Duodenum, jejunum, ileum and cecum were collected one, four, 12 and 24 h after inoculation. The findings demonstrate greater adhesion of strains in the cecum and an important correlation between in vitro and in vivo results. It was concluded that BMM utilization represents an important technique for triage of Lactobacillus for subsequent in vivo evaluation, which was shown to be efficient in identifying bacterial adhesion to the enteric tract.

Identification and adhesion profile of Lactobacillus spp. strains isolated from poultry

In the aviculture industry, the use of Lactobacillus spp. as a probiotic has been shown to be frequent and satisfactory, both in improving bird production indexes and in protecting intestine against colonization by pathogenic bacteria. Adhesion is an important characteristic in selecting Lactobacillus probiotic strains since it impedes its immediate elimination to enable its beneficial action in the host. This study aimed to isolate, identify and characterize the in vitro and in vivo adhesion of Lactobacillus strains isolated from birds. The Lactobacillus spp. was identified by PCR and sequencing and the strains and its adhesion evaluated in vitro via BMM cell matrix and in vivo by inoculation in one-day-old birds. Duodenum, jejunum, ileum and cecum were collected one, four, 12 and 24 h after inoculation. The findings demonstrate greater adhesion of strains in the cecum and an important correlation between in vitro and in vivo results. It was concluded that BMM utilization represents an important technique for triage of Lactobacillus for subsequent in vivo evaluation, which was shown to be efficient in identifying bacterial adhesion to the enteric tract.

Bacteriophage-induced reduction in Salmonella Enteritidis counts in the crop of broiler chickens undergoing preslaughter feed withdrawal.

Salmonella food poisoning is a public health problem. Feed withdrawal from broiler chickens before slaughter can favor the multiplication of Salmonella in the cecum and crop of contaminated animals and subsequently lead to contamination of carcasses in the processing plant. In the present study, a cocktail of lytic bacteriophages isolated from sewage water was orally administered to 45-d-old broiler chickens 1 h after they received an oral dose of 10(7) cfu/mL Salmonella enterica subspecies enterica serotype Enteritidis. Immediately after phage administration and 30 min, 1, 3, 6, and 12 h thereafter, groups of chicken were killed. Ceca and crops were analyzed for the presence of Salmonella. At 3 h posttreatment, there were 10(3) cfu/g and 10(1) cfu/g of cecal and crop suspension, respectively. At 6 h after treatment, the number of Salmonella was 10(3) cfu/g in the cecal suspension, but below the detection limit in the crops. Our results suggest that bacteriophage therapy may be able to reduce the contamination of chicken carcasses by reducing the preslaughter load of Salmonella in the birds.