Comprehensive Metagenomic Analysis of Veterinary Probiotics in Broiler Chickens.

Probiotics are widely used in broiler chickens to support the gut microbiome, gut health, and to reduce the amount of antibiotics used. Despite their benefits, there is concern over their ability to carry and spread antimicrobial resistance genes (ARGs), posing a significant public health risk. This study utilized next-generation sequencing to investigate ARGs in probiotics approved for poultry, focusing on their potential to be transferred via mobile genetic elements such as plasmids and phages. We examined the gut microbiome and resistome changes in 60 broiler chickens over their rearing period, correlating these changes with different probiotic treatments. Specific resistance mechanisms against critically important antibiotics were identified, including genes related to fluoroquinolone resistance and peptide antibiotic resistance. We also found genes with significant relevance to public health (aadK, AAC(6')-Ii) and multiple drug-resistance genes (vmlR, ykkC, ykkD, msrC, clbA, eatAv). Only one phage-encoded gene (dfrA43) was detected, with no evidence of plasmid or mobile genetic element transmission. Additionally, metagenomic analysis of fecal samples showed no significant changes corresponding to time or diet across groups. Our findings highlight the potential risks associated with the use of probiotics in poultry, particularly regarding the carriage of ARGs. It is crucial to conduct further research into the molecular genetics of probiotics to develop strategies that mitigate the risk of resistance gene transfer in agriculture, ensuring the safe and effective use of probiotics in animal husbandry.

Monitoring Changes in the Antimicrobial-Resistance Gene Set (ARG) of Raw Milk and Dairy Products in a Cattle Farm, from Production to Consumption.

Raw milk and dairy products can serve as potential vectors for transmissible bacterial, viral and protozoal diseases, alongside harboring antimicrobial-resistance genes. This study monitors the changes in the antimicrobial-resistance gene pool in raw milk and cheese, from farm to consumer, utilizing next-generation sequencing. Five parallel sampling runs were conducted to assess the resistance gene pool, as well as phage or plasmid carriage and potential mobility. In terms of taxonomic composition, in raw milk the Firmicutes phylum made up 41%, while the Proteobacteria phylum accounted for 58%. In fresh cheese, this ratio shifted to 93% Firmicutes and 7% Proteobacteria. In matured cheese, the composition was 79% Firmicutes and 21% Proteobacteria. In total, 112 antimicrobial-resistance genes were identified. While a notable reduction in the resistance gene pool was observed in the freshly made raw cheese compared to the raw milk samples, a significant growth in the resistance gene pool occurred after one month of maturation, surpassing the initial gene frequency. Notably, the presence of extended-spectrum beta-lactamase (ESBL) genes, such as OXA-662 (100% coverage, 99.3% identity) and OXA-309 (97.1% coverage, 96.2% identity), raised concerns; these genes have a major public health relevance. In total, nineteen such genes belonging to nine gene families (ACT, CMY, EC, ORN, OXA, OXY, PLA, RAHN, TER) have been identified. The largest number of resistance genes were identified against fluoroquinolone drugs, which determined efflux pumps predominantly. Our findings underscore the importance of monitoring gene pool variations throughout the product pathway and the potential for horizontal gene transfer in raw products. We advocate the adoption of a new approach to food safety investigations, incorporating next-generation sequencing techniques.

Stability Studies of the Dilution Series of Different Antibiotic Stock Solutions in Culture Medium Incubated at 37 °C.

The long-term stability of antibiotics in culture media remains underexplored in scientific literature. This study evaluated the stability of eight distinct antibiotic stock solutions-amoxicillin, cefotaxime, neomycin, oxytetracycline, florfenicol, enrofloxacin, colistin, and potentiated sulfonamide-and their 10-fold dilution series in tryptone soy broth (TSB) at 37 °C, over 12 days. Samples were collected immediately after preparation and on days 1, 2, 5, 7, 9, and 12, with active substance concentrations measured using ultra-high-performance liquid chromatography (UHPLC) coupled with mass spectrometry. The results indicated that among the ultrapure water stock solutions, neomycin, florfenicol, and potentiated sulfonamide maintained stability (>95%). Within the culture medium, florfenicol showed consistent stability (100%) throughout the study, potentiated sulfonamide experienced minor degradation (>85%), and neomycin underwent significant degradation. Amoxicillin, oxytetracycline, and colistin displayed considerable degradation in both solution types but were more stable in ultrapure water solutions. The stability of cefotaxime and enrofloxacin in ultrapure water solutions and in the medium was very similar when compared; however, 3.6% of the former and 88.7% of the latter remained detectable by day 12. These findings are crucial for minimum inhibitory concentration (MIC) assessments, especially in minimum bactericidal concentration (MBC) studies, and in experiments concerning long-term evolution and co-selection. This study underscores the necessity of stability assessments in culture media to validate future experimental outcomes.

Antimicrobial Susceptibility Profiles of Pasteurella multocida Isolates from Clinical Cases of Waterfowl in Hungary between 2022 and 2023.

The waterfowl industry represents a narrow, yet economically significant, sector within the poultry industry. Although less prominent, the waterfowl sector is nonetheless of equal importance to any other livestock sector in terms of antimicrobial resistance and animal health issues. This study assesses the antimicrobial resistance profile of Pasteurella multocida bacterial strains isolated from clinical cases in Hungary's duck and goose populations, determining the minimal inhibitory concentration (MIC) of 27 samples collected from 15 different locations. The results indicate that the isolated strains were susceptible to most antibiotics, except for notable resistance to enrofloxacin. These findings support that Pasteurella multocida largely retained its susceptibility. However, the observed resistance to enrofloxacin suggests overuse of fluoroquinolones, which indicates the potential need for stricter regulation of their use in the poultry industry.

Resistome analysis of Escherichia coli isolates from layers in Hungary.

The authors aimed to investigate eight strains of Escherichia coli (E. coli) strains from Hungarian layer flocks for antimicrobial resistance genes (ARG), using metagenomic methods. The strains were isolated from cloacal swabs of healthy adult layers. This study employed shotgun sequencing-based genetic and bioinformatic analysis along with determining phenotypic minimum inhibitory concentrations. A total of 59 ARGs were identified in the eight E. coli isolates, carrying ARGs against 15 groups of antibiotics. Among these, 28 ARGs were identified as transferable. Specifically, four ARGs were plasmid-derived, 18 ARGs were phage-derived and an additional six ARGs were predicted to be mobile, contributing to their mobility and potential spread between bacteria.

Determining the In Vivo Efficacy of Plant-Based and Probiotic-Based Antibiotic Alternatives against Mixed Infection with Salmonella enterica and Escherichia coli in Domestic Chickens.

Restrictions on the use of antimicrobial compounds have led to a surge of interest in alternative solutions, such as natural, plant-based compounds. In our study, we investigated the efficacy of three feed supplements containing different additives, namely, probiotics (Lactobacillus spp., "Test substance A"), turmeric (Curcuma longa L., "Test substance B"), and fenugreek (Trigonella foenum graecum, "Test substance C"). In the experiment, we tested 180 birds of the Bábolna Tetra-SL laying hybrid breed that were infected with Salmonella enteritidis strains. The birds were randomly divided into six groups: three groups treated with the different additives, a negative control group, a positive control group, and an antibiotic-treated group using enrofloxacin. We examined the maturation and the time course of shedding of Salmonella; at the end of rearing, pathological and histopathological examinations were performed. When Salmonella was isolated from the cloacal swab samples, the enrofloxacin-treated group had a high number of animals shedding Salmonella by day 9, which was like the group treated with test material C. The greatest reduction in Salmonella shedding was observed in the groups treated with test materials A and B. In terms of pathological parameters, villus length and crypt depth were significantly better in the group treated with test material C compared to the positive and negative controls, and when comparing the body weight of the tested animals, the group treated with test material B had a significantly larger absorption surface area compared to the positive control group. Overall, the supplement with test material C proved to be the most effective. In the future, it is worthwhile to investigate the combination of the tested active substances for their possible synergistic effects and to perform a dose-response study to select the optimal dosage.

Influence of Degree of Severe Plastic Deformation on Thermal Stability of an HfNbTiZr Multi-Principal Element Alloy Processed by High-Pressure Torsion.

Severe plastic deformation (SPD) is an effective route for the nanocrystallization of multi-principal element alloys (MPEAs). The stability of the refined microstructure is important, considering the high temperature applications of these materials. In the present study, the effect of SPD on the stability of a body-centered cubic (bcc) HfNbTiZr MPEA was investigated. SPD was performed using a high-pressure torsion (HPT) technique by varying the number of turns between ½ and 10. The evolution of phase composition and microstructure was studied near the disk centers and edges where the imposed strain values were the lowest and highest, respectively. Thus, the shear strain caused by HPT varies between 3 (½ turn, near the center) and 340 (10 turns, near the edge). It was found that during annealing up to 1000 K, the bcc HfNbTiZr alloy decomposed into two bcc phases with different lattice constants at 740 K. In addition, at high strains a hexagonal close packed (hcp) phase was formed above 890 K. An inhomogeneous elemental distribution was developed at temperatures higher than 890 K due to the phase decomposition. The scale of the chemical heterogeneities decreased from about 10 µm to 30 nm where the shear strain increased from 3 to 340, which is similar to the magnitude of grain refinement. Anneal-induced hardening was observed in the MPEA after HPT for both low and high strains at 740 K, i.e., the hardness of the HPT-processed samples increased due to heat treatment. At low strain, the hardness remained practically unchanged between 740 and 1000 K, while for the alloy receiving high strains there was a softening in this temperature range.

Effect of Heat Treatment on the Microstructure and Performance of Cu Nanofoams Processed by Dealloying.

Cu nanofoams are promising materials for a variety of applications, including anodes in high-performance lithium-ion batteries. The high specific surface area of these materials supports a high capacity and porous structure that helps accommodate volume expansion which occurs as batteries are charged. One of the most efficient methods to produce Cu nanofoams is the dealloying of Cu alloy precursors. This process often yields nanofoams that have low strength, thus requiring additional heat treatment to improve the mechanical properties of Cu foams. This paper provides the effects of heat treatment on the microstructures, mechanical properties, and electrochemical performance of Cu nanofoams. Annealing was conducted under both inert and oxidizing atmospheres. These studies ultimately reveal the underlying mechanisms of ligament coarsening during heat treatment.

Adsorption of Remazol Brilliant Violet-5R Textile Dye from Aqueous Solutions by Using Eggshell Waste Biosorbent.

Based on the well-known excellent adsorbent ability of chicken eggshells, the adsorptive capacity and mechanism of Remazol Brilliant Violet-5R (RBV-5R) dye by eggshell was investigated. Exploiting the high surface-area-to-volume ratio and porous structure of this natural adsorbent, the developed procedure showed to be useful for the efficient adsorption of RBV-5R dye from contaminated water. The protocol was thoroughly optimized by investigating the effect of the dye concentration, biomass-contaminated water ratio, particle size of the adsorbent, pH and temperature, as they are key factors in the efficiency of the dye removal process. The eggshell material was characterized by different types of microscopy techniques (stereo, polarization, SEM) as well as elemental analysis (element distribution mapping, EDX), Raman spectroscopy and BET-surface density measurements. EDX, FTIR and Raman spectroscopy proved the presence of the adsorbed dye on the surface of the biomaterial. It was shown that under optimal conditions, the environmentally friendly and inexpensive eggshell could be a reliable adsorbent for Remazol dye removal from wastewater.

Tracking the occurrence of anthropogenic magnetic particles and potentially toxic elements (PTEs) in house dust using magnetic and geochemical analyses.

The influence of anthropogenic outdoor sources on the geochemical composition of house dust material in large cities is poorly understood. In this study, we investigate the magnetic signature and the concentrations of potentially toxic elements (PTEs) in randomly selected house dust samples from the metropolitan area of Athens, the most populated city in Greece. Environmental magnetic measurements, including isothermal remanent magnetization and thermomagnetism, indicated that the main magnetic mineral is coarse-grained low-coercivity magnetite. Detailed microscopic observations of the magnetically extracted material revealed the presence of three different kinds of Fe-rich particles deriving from both combustion-related and non-exhaust vehicular sources: irregularly-shaped grains and spherules of Fe-oxides, and particles consisting of metallic Fe. Further study of the morphology of single anthropogenic magnetic spherules (size > 30 µm) identified the presence of magnetite spherical particles, typically formed by industrial combustion processes. Enrichment factors (EFs) for the PTEs calculated against the Athens urban soil showed that the house dusts were very highly enriched in Cd, Cu, Zn and significantly enriched in Pb (median EF values of 34.1, 26.2, 25.4 and 10.3, respectively). The oral bioaccessibility of PTEs in the house dust, evaluated using a simulated gastric solution (0.4 M glycine), was in the order Pb > Zn > Mn > Cd > Ni > Cu > Cr > Fe. Concentrations of Pb increased with the house age. Principal component and cluster analysis demonstrated the close association of anthropogenic Cu, Pb and Zn with the magnetic susceptibility of the house dusts. We conclude that both traffic-related and industrial sources trigger the occurrence of magnetic Fe/PTEs- rich particles in house dust. These results reinforce the use of environmental magnetism determinations for assessing anthropogenic contamination of PTEs in the indoor environment in large cities.