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Investigating the occurrence of antibiotic-resistance genes (ARGs) in sedimentary archives provides opportunities for reconstructing the distribution and dissemination of historical (i.e., non-anthropogenic origin) ARGs. Although ARGs in freshwater environments have attracted great attention, historical variations in the diversity and abundance of ARGs over centuries to millennia remain largely unknown. In this study, we investigated the vertical change patterns of bacterial communities, ARGs and mobile genetic elements (MGEs) found in sediments of Lake Chenghai spanning the past 600 years. Within resistome preserved in sediments, 177 ARGs subtypes were found with aminoglycosides and multidrug resistance being the most abundant. The ARG abundance in the upper sediment layers (equivalent to the post-antibiotic era since the 1940s) was lower than those during the pre-antibiotic era, whereas the ARG diversity was higher during the post-antibiotic era, possibly because human-induced lake eutrophication over the recent decades facilitated the spread and proliferation of drug-resistant bacteria. Statistical analysis suggested that MGEs abundance and the bacterial community structure were significantly correlated with the abundance and diversity of ARGs, suggesting that the occurrence and distribution of ARGs may be transferred between different bacteria by MGEs. Our results provide new perspectives on the natural history of ARGs in freshwater environments and are essential for understanding the temporal dynamics and dissemination of ARGs.

To investigate the effects of different manures in combination with fulvic acid on the abundance of N-cycling functional genes in greenhouse soils, Chinese cabbage was planted for three growing seasons. A total of six treatments—pig manure (P), pig manure + fulvic acid (PH), chicken manure (C), chicken manure + fulvic acid (CH), sheep manure (S), sheep manure + fulvic acid (SH) and no fertilization (CK)—were set up. The abundance of 13 soil N-cycling functional genes (gdhA, amoA-1, amoA-2, amoB, narG, nirK-1, nirK-2, nirK-3, nirS-1, nirS-2, nirS-3, nosZ and nifH) were investigated after the harvest of the third growing season using a gene chip approach. The results showed that fertilization treatments increased the abundance of most N-cycling functional genes in the soil, such as nitrification genes amoA-2 and amoB as well as denitrification genes narG, nirK-1, nirS-1 and nirS-2, with the stronger influence of sheep and pig manure than chicken manure. Fortunately, the additional fulvic acid reduced the increasing effect resulting from pig, chicken and sheep manure application. The abundance of functional genes for nitrogen cycling in soil was positively correlated with the content of soil organic matter, available phosphorus and NO3−-N, and negatively correlated with electrical conductivity. Overall, fertilization treatments increased soil nitrification and denitrification genes abundance, with a risk of increasing soil nitrogen loss, but the supplementary fulvic acid could limit the increase. In this study, it was concluded that the sheep manure (31.3 t/ha) + fulvic acid (7.5 kg/ha) treatment was more powerful in regulating the abundance of N-cycling functional genes in soil. [ABSTRACT FROM AUTHOR]

Antimicrobial resistance (AMR) is one of the biggest threats to human health worldwide. The World Health Organization (WHO, Geneva, Switzerland) has launched the "One-Health" approach, which encourages assessment of antibiotic-resistant genes (ARGs) within environments shared by human-animals-plants-microbes to constrain and alleviate the development of AMR. Aerosols as a medium to disseminate ARGs, have received minimal attention. In the present study, we investigated the distribution and abundance of ARGs in indoor and outdoor aerosols collected from an urban location in Kuwait and the interior of three hospitals. The high throughput quantitative polymerase chain reaction (HT-qPCR) approach was used for this purpose. The results demonstrate the presence of aminoglycoside, beta-lactam, fluoroquinolone, tetracycline, macrolide-lincosamide-streptogramin B (MLSB), multidrug-resistant (MDR) and vancomycin-resistant genes in the aerosols. The most dominant drug class was beta-lactam and the genes were IMP-2-group (0.85), Per-2 group (0.65), OXA-54 (0.57), QnrS (0.50) and OXA-55 (0.55) in the urban non-clinical settings. The indoor aerosols possessed a richer diversity (Observed, Chao1, Shannon's and Pielou's evenness) of ARGs compared to the outdoors. Seasonal variations (autumn vs. winter) in relative abundances and types of ARGs were also recorded (R2 of 0.132 at p < 0.08). The presence of ARGs was found in both the inhalable (2.1 µm, 1.1 µm, 0.7 µm and < 0.3 µm) and respirable (>9.0 µm, 5.8 µm, 4.7 µm and 3.3 µm) size fractions within hospital aerosols. All the ARGs are of pathogenic bacterial origin and are hosted by pathogenic forms. The findings present baseline data and underpin the need for detailed investigations looking at aerosol as a vehicle for ARG dissemination among human and non-human terrestrial biota. [ABSTRACT FROM AUTHOR]

Introduction: The establishment of a high-throughput quantification approach for waterborne pathogenic protozoa and helminths is crucial for rapid screening and health risk assessment., Methods: We developed a high-throughput quantitative polymerase chain reaction (HT-qPCR) assay targeting 19 waterborne protozoa and 3 waterborne helminths and validated its sensitivity, specificity, and repeatability. The assay was then applied to test various environmental media samples., Results: The HT-qPCR assay's limit of detection (LOD) was 5×10 2 copies/μL DNA, and its specificity was confirmed using Giardia and Cryptosporidium standards. Repeatability, assessed through intra- and inter-group experiments, yielded a coefficient of variation ( CV ) of 1.0%-4.6% and 1.2%-6.4% at concentrations of 1×10 5 and 1×10 4 copies/μL, respectively. The R 2 values of the 22 standard curves ranged from 0.983 to 0.998, with amplification efficiencies between 80% and 107%. In drinking water sources, sludge from municipal wastewater treatment plants (MWTPs), and livestock manure samples, 17 of 22 targets were detected, with Acanthamoeba genus (50.0%), Acanthamoeba castellanii (11.8%), and Enterocytozoon bieneusi (11.8%) showing high prevalence. Cryptosporidium spp., Enterocytozoon bieneusi , and Cyclospora cayetanensis were simultaneously found in all three sample types., Discussion: This study presents a useful tool for the rapid detection of waterborne protozoa and helminths in complex environmental microbiomes, providing scientific data for monitoring cross-media transmission and controlling microbial risk from a One Health perspective., Competing Interests: No conflicts of interest., (Copyright © 2025 by Chinese Center for Disease Control and Prevention.)

To investigate the effects of different manures in combination with fulvic acid on the abundance of N-cycling functional genes in greenhouse soils, Chinese cabbage was planted for three growing seasons. A total of six treatments—pig manure (P), pig manure + fulvic acid (PH), chicken manure (C), chicken manure + fulvic acid (CH), sheep manure (S), sheep manure + fulvic acid (SH) and no fertilization (CK)—were set up. The abundance of 13 soil N-cycling functional genes (gdhA, amoA-1, amoA-2, amoB, narG, nirK-1, nirK-2, nirK-3, nirS-1, nirS-2, nirS-3, nosZ and nifH) were investigated after the harvest of the third growing season using a gene chip approach. The results showed that fertilization treatments increased the abundance of most N-cycling functional genes in the soil, such as nitrification genes amoA-2 and amoB as well as denitrification genes narG, nirK-1, nirS-1 and nirS-2, with the stronger influence of sheep and pig manure than chicken manure. Fortunately, the additional fulvic acid reduced the increasing effect resulting from pig, chicken and sheep manure application. The abundance of functional genes for nitrogen cycling in soil was positively correlated with the content of soil organic matter, available phosphorus and NO3−-N, and negatively correlated with electrical conductivity. Overall, fertilization treatments increased soil nitrification and denitrification genes abundance, with a risk of increasing soil nitrogen loss, but the supplementary fulvic acid could limit the increase. In this study, it was concluded that the sheep manure (31.3 t/ha) + fulvic acid (7.5 kg/ha) treatment was more powerful in regulating the abundance of N-cycling functional genes in soil.

Background: Therapeutic and growth-promoting antibiotics are frequently used in broiler production. Indirect evidence indicates that these practices are linked to the proliferation of antimicrobial resistance (AMR), the spread of antibiotic-resistant bacteria from food animals to humans, and the environment, but there is a lack of comprehensive experimental data supporting this. We investigated the effects of growth promotor (bacitracin) and therapeutic (enrofloxacin) antibiotic administration on AMR in broilers for the duration of a production cycle, using a holistic approach that integrated both culture-dependent and culture-independent methods. We specifically focused on pathogen-harboring families (Enterobacteriaceae, Enterococcaceae, and Staphylococcaceae). Results: Antibiotic-resistant bacteria and antibiotic resistance genes were ubiquitous in chicken cloaca and litter regardless of antibiotic administration. Environment (cloaca vs. litter) and growth stage were the primary drivers of variation in the microbiomes and resistomes, with increased bacterial diversity and a general decrease in abundance of the pathogen-harboring families with age. Bacitracin-fed groups had higher levels of bacitracin resistance genes and of vancomycin-resistant Enterococcaceae (total Enterococcaceae counts were not higher). Although metagenomic analyses classified 28–76% of the Enterococcaceae as the commensal human pathogens E. faecalis and E. faecium, culture-based analysis suggested that approximately 98% of the vancomycin-resistant Enterococcaceae were avian and not human-associated, suggesting differences in the taxonomic profiles of the resistant and non-resistant strains. Enrofloxacin treatments had varying effects, but generally facilitated increased relative abundance of multidrug-resistant Enterobacteriaceae strains, which were primarily E. coli. Metagenomic approaches revealed a diverse array of Staphylococcus spp., but the opportunistic pathogen S. aureus and methicillin resistance genes were not detected in culture-based or metagenomic analyses. Camphylobacteriaceae were significantly more abundant in the cloacal samples, especially in enrofloxacin-treated chickens, where a metagenome-assembled C. jejuni genome harboring fluoroquinolone and β-lactam resistance genes was identified. Conclusions: Within a "farm-to-fork, one health" perspective, considering the evidence that bacitracin and enrofloxacin used in poultry production can select for resistance, we recommend their use be regulated. Furthermore, we suggest routine surveillance of ESBL E. coli, vancomycin-resistant E. faecalis and E. faecium, and fluoroquinolone-resistant C. jejuni strains considering their pathogenic nature and capacity to disseminate AMR to the environment. E2eQV6Q9BhTz4idhLosSeg Video Abstract [ABSTRACT FROM AUTHOR]

Landfills are so far the most common practice for the disposals of municipal solid waste (MSW) worldwide. Since MSW landfill receives miscellaneous wastes, including unused/expired antibiotics and bioactive wastes, it gradually becomes a huge potential bioreactor for breeding antibiotic resistance. Antibiotic resistance genes (ARGs) in landfill can flow to the environment through leakage of landfill leachate and pose a risk to public health. Using high throughput quantitative Polymerase Chain Reaction (HT-qPCR), we investigated the prevalence, diversity of ARGs and its association with various mobile genetic elements (MGEs) in MSW landfill groundwater. Totally 171 unique ARGs (belonging to 9 ARG types, encompassing 3 major resistance mechanisms) and 8 MGEs (6 transposase genes, and 2 integron-integrase genes) were identified. The normalized abundance of ARG was ranging from 0.24 to 5.66 copies/cell with multidrug, beta-lactams and tetracycline resistance genes being the most abundant ARG types. The co-occurrence pattern and significant correlation between MGEs and ARGs, indicated that MGEs may play an important role in the persistence and proliferation of ARGs. A Mantel test and Procrustes analysis suggested that ARG profiles were significantly correlated with bacterial community. Variation partitioning analysis (VPA) further demonstrated that bacterial community shifts contribute 65.8% of the total ARG variations. Additionally network analysis revealed that 15 bacterial taxa at family level might be the potential hosts of ARGs. These findings provide evidence that groundwater near MSW landfill is an underappreciated hotspot of antibiotic resistance and contribute to the spread of ARGs via the flowing contaminated groundwater. [ABSTRACT FROM AUTHOR]

Manure application is a common practice that not only adds nutrients and organic matter to arable soils for crop growth, but also introduces antibiotic resistance genes (ARGs), posing a potential risk to human health. To investigate the mechanisms underlying the spread of ARGs in manured soil, especially the impact of manure-borne and indigenous soil microorganisms, a microcosm experiment with four specially designed treatments over a period of two months was conducted, including soil, soil with irradiated pig manure, irradiated soil with pig manure, and soil with pig manure. A total of 240 unique ARGs were detected via a high-throughput quantitative PCR (HT-qPCR) targeting almost all major classes of ARGs. Manure application significantly increased the diversity and abundance of ARGs in soil ( P < 0.01), and also markedly shifted the bacterial composition that was significantly correlated with ARGs profiles. Manure-borne microorganisms contributed largely to the elevation of ARGs due to both the addition of manure-borne antibiotic resistant bacteria (ARB) in soil and potential horizontal gene transfer (HGT) via mobile genetic elements (MGEs) from manure-borne ARB to indigenous soil microorganisms. In contrast, indigenous soil microorganisms were demonstrated to prevent the dissemination of ARGs from manure to soil. The reason could be due to that indigenous soil microorganisms prevented the invasion and establishment of manure-borne ARB in soil. The abundance of ARG in manured soil decreased over time, but was still higher than that in control soil, indicating the persistence of ARGs in manured soil. These findings may shed light on the mechanisms underlying the spread and fate of ARGs in manured soil and also clues for ARGs mitigation. [ABSTRACT FROM AUTHOR]

Manure application is a significant agriculture practice that can affect the microbial community structure in both topsoil and subsoil. However, how the abundant and rare taxa in the soil profile respond to surface manure application and their ecological roles at different soil depths remain unknown. Based on a soil mesocosm experiment, we used amplicon sequencing and high-throughput quantitative-PCR array to characterize the bacterial community composition and functional traits across soil profiles. Results showed that the relative abundance and diversity of rare taxa increased in the topsoil but decreased in the 40–60 cm soil, where the relative abundance of abundant taxa increased after manure application. Evidence from network analysis, niche width and 16S rRNA operon (rrn) copy number suggest that abundant taxa in subsoils had a greater competitivity for resource mining than rare taxa. Regarding resource utilization strategies, microbial communities under manure treatment preferred to utilize recalcitrant carbon (C) and inorganic phosphorus (P) in the topsoil and labile carbon and organic phosphorus in the subsoil compared with non-manure application. Random Forest regression analysis indicated the relatively greater contribution of abundant taxa to C degradation and fixation in subsoils. These results imply that the manure application effect on microbial resource mining in subsoils is mainly driven by abundant bacteria due to the lower competitiveness of rare taxa for the C sources leached above. Overall, This study linked community functions with the ecological role of abundant and rare taxa in the context of environmental disturbances at different soil depths and shed light on the potential ecological risks of diversity reduction of subsoil rare bacteria. • Manure addition decreased the diversity of rare bacteria in the subsoil. • Manure addition altered the resource utilization strategy of microbial communities. • Abundant taxa are more competitive for subsoil carbon resources than rare taxa. • Abundant taxa initiate the changes in subsoil bacterial communities. [ABSTRACT FROM AUTHOR]

Landfill leachate, a highly concentrated organic wastewater containing diverse microorganisms and various heavy metals, has become an important reservoir of antibiotic resistance genes (ARGs). In this study, a total of 203 unique ARGs and 10 mobile genetic elements (MGEs) were identified from collected landfill leachate and groundwater. The number and abundance (normalized and absolute) of antibiotic resistome in effluent of leachate treatment plants decreased significantly compared to influent. The abundance of ARGs in groundwater increased as the distance from the leachate basin decreased. Fast expectation-maximization microbial source tracking (FEAST) showed that up to 96 % of ARGs in groundwater (GW3) may originate from nearby leachate, suggesting that ARGs in leachate can penetrate and spread into the groundwater environment. A significant correlation between ARGs and bacterial communities was identified. Together with network analysis showing the 12 bacterial taxa co-occurring with seven classes of antibiotic-associated ARGs, our results revealed the diverse potential microbial hosts of ARGs in water samples around the landfill sites. Heavy metals, bacterial community and MGEs were the driving factors shaping the ARGs patterns in the water samples, with their interactions explaining 57 % of ARGs variations. Our results provide an understanding of the distribution and dissemination of ARGs from landfill leachate to the nearby groundwater and suggest a comprehensive impact assessment of ARGs in aquatic environments of landfills. [Display omitted] • Quantified the antibiotic resistome from landfill leachate and groundwater. • Demonstrate that leachate is a major source of antibiotic resistome in groundwater. • Contaminated groundwater can carry antibiotic resistome downstream. • Leachate treatment plants are essential for reducing antibiotic resistome. [ABSTRACT FROM AUTHOR]

We assessed antimicrobial resistance (AMR) potential to seven major classes of antibiotics in Central Thailand's coastal aquaculture region using high-throughput qPCR targeting 295 antibiotic resistance genes (ARGs) and mobile genetic elements (MGEs). In parallel, we used MinION next generation sequencing (NGS) of 16S rRNA gene amplicons, qPCR of faecal pollution makers, and conventional plate count methods for the comprehensive characterization of environmental microbiomes. We tested the hypothesis that aquaculture increases environmental AMR by comparing ARGs and MGEs in water and sediment samples from five aquaculture sites and their associated canals. There was no evidence from the ARG and MGE data that aquaculture is a major driver of environmental AMR in Central Thailand. Instead, the highest relative prevalence of resistance traits was found in Hua Krabue canal water influenced by urban pollution from Bangkok at the inland edge of the coastal aquaculture region. The sum of ARGs and MGEs, relative to 16S rRNA genes used as markers for overall bacterial abundance, was between 0.495 ± 0.011 and 0.498 ± 0.013 in Hua Krabue canal water, compared with at most 0.132 ± 0.005 in all the other environmental samples. Corresponding patterns were observed for most faecal pollution markers, which were also elevated in Hua Krabue canal water. [Display omitted] • Canal and aquaculture water and sediment were investigated in Central Thailand. • Antimicrobial resistance (AMR) was studied with HT-qPCR and NGS. • High AMR prevalence was associated with high fecal indicator bacteria prevalence. • AMR prevalence was highest in peri-urban canal water. • Urban water pollution appears to drive environmental AMR. [ABSTRACT FROM AUTHOR]

• Manure application impacted the resistome in soil, earthworms and phyllosphere. • Manure with reduced antibiotic burden reduced the spread of the antibiotic resistome. • Shared ARGs and OTUs were founded within soil-earthworm-phyllosphere system. The overuse of antibiotics in animal husbandry is widespread and believed to significantly contribute to the selection of antibiotic resistance genes (ARGs) in animals. Thus, there is a global drive to reduce antibiotic use in the agricultural sector. However, it has not been established whether a reduction in the use of antibiotics in livestock production would be effective in reducing the spread of ARGs. A microcosm approach was used to determine how the addition of manure with either reduced antibiotic levels or with typical antibiotic levels could affect the spread of antibiotic resistance genes between soil, earthworms and the phyllosphere. When compared to the control soil, earthworm and phyllosphere samples had the greater increase in ARG abundance in conventional manure treatments (P < 0.05). Reduced antibiotic manure also enriched the abundance of ARGs in the phyllosphere and soil but not earthworm guts when compared to the control (P < 0.05). In both soil and earthworm guts, the enrichment of ARGs was lower in reduced antibiotic manure than in conventional manure. This study has identified bacterial transfer through the soil-earthworm-phyllosphere system as a potential means to spread ARGs between habitats after fertilization with livestock derived manures. [ABSTRACT FROM AUTHOR]