Spread Of Antibiotic Resistance Genes Research Articles

Planktonic and Biofilm-Derived Pseudomonas aeruginosa Outer Membrane Vesicles Facilitate Horizontal Gene Transfer of Plasmid DNA.

Outer membrane vesicles (OMVs) produced by Gram-negative bacteria package various cargo, including DNA that can be transferred to other bacteria or to host cells. OMV-associated DNA has been implicated in mediating horizontal gene transfer (HGT) between bacteria, which includes the dissemination of antibiotic resistance genes within and between bacterial species. Despite the known ability of OMVs to mediate HGT, the mechanisms of DNA packaging into OMVs remain poorly characterized, as does the effect of bacterial growth conditions on the DNA cargo composition of OMVs and their subsequent abilities to mediate HGT. In this study, we examined the DNA content of OMVs produced by the opportunistic pathogen Pseudomonas aeruginosa grown in either planktonic or biofilm conditions. Analysis of planktonic growth-derived OMVs revealed their ability to package and protect plasmid DNA from DNase degradation and to transfer plasmid-encoded antibiotic resistance genes to recipient, antibiotic-sensitive P. aeruginosa bacteria at a greater efficiency than transformation with plasmid alone. Comparisons of planktonic and biofilm-derived P. aeruginosa OMVs demonstrated that biofilm-derived OMVs were smaller but were associated with more plasmid DNA than planktonic-derived OMVs. Additionally, biofilm-derived P. aeruginosa OMVs were more efficient in the transformation of competent P. aeruginosa bacteria, compared to transformations with an equivalent number of planktonic-derived OMVs. The findings of this study highlight the importance of bacterial growth conditions for the packaging of DNA within P. aeruginosa OMVs and their ability to facilitate HGT, thus contributing to the spread of antibiotic resistance genes between P. aeruginosa bacteria. IMPORTANCE Bacterial membrane vesicles (BMVs) mediate interbacterial communication, and their ability to package DNA specifically contributes to biofilm formation, antibiotic resistance, and HGT between bacteria. However, the ability of P. aeruginosa OMVs to mediate HGT has not yet been demonstrated. Here, we reveal that P. aeruginosa planktonic and biofilm-derived OMVs can deliver plasmid-encoded antibiotic resistance to recipient P. aeruginosa. Additionally, we demonstrated that P. aeruginosa biofilm-derived OMVs were associated with more plasmid DNA compared to planktonic-derived OMVs and were more efficient in the transfer of plasmid DNA to recipient bacteria. Overall, this demonstrated the ability of P. aeruginosa OMVs to facilitate the dissemination of antibiotic resistance genes, thereby enabling the survival of susceptible bacteria during antibiotic treatment. Investigating the roles of biofilm-derived BMVs may contribute to furthering our understanding of the role of BMVs in HGT and the spread of antibiotic resistance in the environment.

PH drives the spatial variation of antibiotic resistance gene profiles in riparian soils at a watershed scale

Owing to convenient water access, riparian areas are often sites for intensive livestock breeding industries and agriculture, which can increase the spread of antibiotic resistance genes (ARGs). However, studies on ARG profiles in riparian soils are limited and there is little information regarding the factors influencing ARGs at a watershed scale. Here, we analyzed ARG profiles, bacterial communities, and soil properties in riparian soils under different land-use types. A total of 124 ARGs and 25 mobile genetic elements (MGEs) were detected in the riparian soils, which covered almost all major classes of antibiotics. Non-metric multidimensional scaling analysis showed that both the distance to the water reservoir and land-use types played important roles in shaping ARG profiles in riparian soils at a watershed scale. Downstream soils harbored three times the abundance of ARGs compared with upstream and midstream soils. Distance-decay analysis indicated that the similarity of ARG profiles and bacterial community composition decreased significantly with the increase of geographical distance (p < 0.001). When taking the land-use type into consideration, the relative abundance and diversity of ARGs and MGEs in orchard and farmland soils were significantly higher than those in wasteland soils. This indicated that anthropogenic activities can also affect ARG patterns in riparian soils. MGE abundance was identified as major driving factors of ARG profiles. In addition, among all the examined soil properties, soil pH was found to be more important than nutrients and organic carbon in shaping ARG profiles. Our findings provide valuable data on ARG distribution in riparian soils in a reservoir catchment and highlight downstream soils is crucial for ensuring water source security.

Factors and Mechanisms Influencing Conjugation In Vivo in the Gastrointestinal Tract Environment: A Review.

The emergence and spread of antibiotic resistance genes (ARGs) have imposed a serious threat on global public health. Horizontal gene transfer (HGT) via plasmids is mainly responsible for the spread of ARGs, and conjugation plays an important role in HGT. The conjugation process is very active in vivo and its effect on the spreading of ARGs may be underestimated. In this review, factors affecting conjugation in vivo, especially in the intestinal environment, are summarized. In addition, the potential mechanisms affecting conjugation in vivo are summarized from the perspectives of bacterial colonization and the conjugation process.

Granular activated carbon remediates antibiotic resistance propagation and methanogenic inhibition induced by polystyrene nanoplastics in sludge anaerobic digestion

Nano/microplastics (NPs/MPs) in sewage sludge can induce oxidative stress to the anaerobic digestion (AD) and also proliferate antibiotic resistance genes (ARGs). Recently, granular activated carbon (GAC) has been used as an additive to enhance methane production in AD via direct interspecies electron transfer (DIET); however, its impact on AD exposed to NPs/MPs is yet to be studied. This study examined the effect of GAC (5 and 15 g/L) on sludge AD exposed to 150 µg/L of polystyrene nanoplastics (PsNPs). PsNPs decreased methane yield by 32.3% due to elevated levels of reactive oxygen species. However, GAC addition counteracted this adverse effect and improved methane production, attributed to the potential enrichment of DIET-active microbes and the adsorption of PsNPs by GAC. Moreover, GAC reduced the total abundance of ARGs, which was increased by PsNPs exposure. Thus, GAC can provide dual benefits in mitigating methanogenic inhibition caused by PsNPs and ARG spread.

Oxalated zero-valent iron catalyzed ozonation for effective removal of antibiotic resistance genes from municipal sewage sludge

Wastewater treatment plants (WWTPs) are hotspots of antibiotic resistance genes (ARGs), and efficient technologies are needed to prevent the enrichment and spread of ARGs in municipal sludge. Herein, we demonstrated that oxalated zero-valent iron (OA-ZVIbm) catalyzed ozonation could effectively remove typical ARGs, tetG and sul1, from municipal sewage sludge, with the total abundance reduced by 3.02 and 2.00 log per gram sludge, respectively, surpassing the O3 alone and O3/ZVIbm counterparts. Meanwhile, the O3/OA-ZVIbm treatment could simultaneously remove both intracellular and extracellular ARGs from the sludge, moreover, it also eliminated 1.83 log total Class 1 integron (total intI1), thus may suppress the horizontal gene transfer (HGT) potential of ARGs. The good performance of O3/OA-ZVIbm was owing to the FeC2O4·2H2O shell of ZVI, which improved the accessibility of core iron species to catalyze the decomposition of O3, producing more reactive oxygen species (ROSs) including •OH, •O2−, and 1O2, for the removal of ARGs. This study provides a new method to control the dissemination of ARGs in sewage sludge.

Effects of particle size and pyrolytic temperature of biochar on the transformation behavior of antibiotic resistance genes

Rampant use of antibiotics has caused a rapid dissemination of antibiotic resistance genes (ARGs) in environment, posing great threats to ecosystems and human health. Applying biochar (BC) in natural systems to combat the spread of ARGs arises as an attention-grabbing solution. Unfortunately, the effectiveness of BC is still unmanageable due to the incomprehensive knowledge over correlations between BC properties and extracellular ARGs transformation. To pinpoint the crucial factors, we primarily explored transformation behaviors of plasmid-mediated ARGs exposed to BC (in suspensions or extraction solutions), adsorption capacities of ARGs on BC, and growth inhibition of E. coli imposed by BC. Specifically, the effects of BC properties including particle size (large-particulate 150 μm and colloidal 0.45–2 μm) and pyrolytic temperature (300, 400, 500, 600, and 700 °C) on the ARGs transformation were emphasized. Results showed that both large-particulate BC and colloidal BC, irrespective of their pyrolytic temperature, would induce significant inhibitory effects on the ARGs transformation, while the BC extraction solutions showed little effect except BC pyrolyzed at 300 °C. Correlation analysis uncovered that the inhibition effect of BC on ARGs transformation was tightly correlated with its adsorption capacity towards plasmid. Accordingly, greater inhibitory effects from those BCs with higher pyrolytic temperatures and smaller particle sizes mainly originated from their greater adsorption capacities. Intriguingly, E. coli was unable to ingest the plasmid adsorbed on BC, which led to ARGs blocked outside the cell membrane, although this inhibitory effect was partially affected by survival inhibition of BC on E. coli. Particularly, significant plasmid aggregation could occur in the extraction solution of large-particulate BC pyrolyzed at 300 °C, leading to a significant inhibition of ARGs transformation. Overall, our findings complete the insufficient understanding over the effects of BC on ARGs transformation behavior, and potentially provide new insights to scientific communities in mitigating ARGs spreading.

Removal of antibiotics and antibiotic resistance genes in the synthetic oxytetracycline wastewater by UASB-A/O(MBR) process

Antibiotic wastewater contains not only conventional pollutants but also high concentrations of antibiotics and the resulting pollutants, such as antibiotic resistant microorganisms and antibiotic resistant genes (ARGs). If not properly treated, the antibiotic wastewater will further induce the spread of antibiotic resistant microorganisms and ARGs in the environment. In this study, an up-flow anaerobic sludge bed (UASB)-anoxic/aerobic (membrane bioreactor) process (A/O(MBR)) was built to treat oxytetracycline (OTC) wastewater. Results showed that the removal efficiencies of OTC decreased from 72.92% to 59.60% when the OTC concentration increased from 25 mg/L to 50 mg/L. Increasing OTC concentration from 0 to 50 mg/L, the chemical oxygen demand (COD) removal efficiencies decreased from 97.47% to 95.05%. Nevertheless, the total nitrogen removal efficiency increased from 55.53% to 85.33%. Bacteroides and Azospira dominated in UASB and A/O(MBR) respectively with the relative abundance of 20.76% and 18.51% at 50 mg/L OTC. Both ARGs in the UASB and O(MBR) increased with the rise of OTC concentration and reached 3.44 × 109 and 7.61 × 108 copies/mL respectively at 50 mg/L OTC. And the corresponding mobile gene elements (MGEs) reached 4.29 × 108 and 3.58 × 108 copies/mL. The membrane presented an effective pollutants removal efficiency with the interception efficiencies for ARGs and MGEs reaching 90.86% and 87.91%, respectively.

A Novel Phytogenic Formulation, EUBIO-BPSG, as a Promising One Health Approach to Replace Antibiotics and Promote Reproduction Performance in Laying Hens

Gut microbiota play a key role in health maintenance and disease pathogenesis in animals. Dietary phytochemicals are crucial factors shaping gut bacteria. Here, we investigated the function and mechanism of a phytogenic formulation, EUBIO-BPSG (BP), in laying hens. We found that BP dose-dependently improved health and egg production in 54-week-old hens. Furthermore, BP was correlated with increased fecal Lactobacillus, decreased Escherichia coli and Salmonella enterica, and reduced antibiotic resistance (AR) and antibiotic resistance genes (ARG) in chicken stools. The 16S rDNA data showed that BP increased seven genera of probiotics and reduced 13 genera of pathogens in chicken feces. In vitro co-culture experiments showed that BP at 4 µg/mL and above promoted growth of L. reuteri while large 100- and 200-fold higher doses suppressed growth of E. coli and S. enterica, respectively. Mechanistic studies indicated that L. reuteri and its supernatants antagonized growth of E. coli and S. enterica but not vice-versa. Five short-chain fatty acids and derivatives (SCFA) produced from L. reuteri directly killed both pathogens via membrane destruction. Furthermore, BP inhibited conjugation and recombination of ARG via interference with conjugation machinery and integrase activity in E. coli. Collectively, this work suggests that BP promotes host health and reproductive performance in laying hens through regulation of gut microbiota through increasing probiotics and decreasing pathogens and spreading ARG.

Genomic Investigation of Salmonella Typhi in Hong Kong Revealing the Predominance of Genotype 3.2.2 and the First Case of an Extensively Drug-Resistant H58 Genotype.

Typhoid fever is a notable disease in Hong Kong. We noticed two local cases of typhoid fever caused by Salmonella Typhi within a two-week period in late 2022, which had no apparent epidemiological linkage except for residing in the same region of Hong Kong. A phylogenetic study of Salmonella Typhi isolates from Hong Kong Island from 2020 to 2022 was performed, including a whole-genome analysis, the typing of plasmids, and the analysis of antibiotic-resistance genes (ARGs), to identify the dominant circulating strain and the spread of ARGs. A total of seven isolates, from six local cases and an imported case, were identified from positive blood cultures in two hospitals in Hong Kong. Five antibiotic-sensitive strains of genotype 3.2.2 were found, which clustered with another 30 strains originating from Southeast Asia. Whole-genome sequencing revealed clonal transmission between the two index cases. The remaining two local cases belong to genotype 2.3.4 and genotype 4.3.1.1.P1 (also known as the H58 lineage). The genotype 4.3.1.1.P1 strain has an extensively drug-resistant (XDR) phenotype (co-resistance to ampicillin, chloramphenicol, ceftriaxone, ciprofloxacin, and co-trimoxazole). Although the majority of local strains belong to the non-H58 genotype 3.2.2 with a low degree of antibiotic resistance, the introduction of XDR strains with the global dissemination of the H58 lineage remains a concern.

Sulfur: a neglected driver of the increased abundance of antibiotic resistance genes in agricultural reclaimed subsidence land located in coal mines with high phreatic water levels

Due to the shallow burial of groundwater in coal mines with a high phreatic water level, a large area of subsidence lakes is formed after the mine collapses. Agricultural and fishery reclamation activities have been carried out, which introduced antibiotics and exacerbated the contamination of antibiotic resistance genes (ARGs), but this has received limited attention. This study analyzed ARG occurrence in reclaimed mining areas, the key impact factors, and the underlying mechanism. The results show that sulfur is the most critical factor impacting the abundance of ARGs in reclaimed soil, which is due to changes in the microbial community. The species and abundance of ARGs in the reclaimed soil were higher than those in the controlled soil. The relative abundances of most ARGs increased with the depth of reclaimed soil (from 0 to 80 cm). In addition, the microbial structures of the reclaimed and controlled soils were significantly different. Proteobacteria, was the most dominant microbial phylum in the reclaimed soil. This difference is likely related to the high abundance of sulfur metabolism functional genes in the reclaimed soil. Correlation analysis showed that the differences in ARGs and microorganisms in the two soil types were highly correlated with the sulfur content. High levels of sulfur promoted the proliferation of sulfur-metabolizing microbial populations such as Proteobacteria and Gemmatimonadetes in the reclaimed soils. Remarkably, these microbial phyla were the main antibiotic-resistant bacteria in this study, and their proliferation created conditions for the enrichment of ARGs. Overall, this study underscores the risk of the abundance and spread of ARGs driven by high-level sulfur in reclaimed soils and reveals the mechanisms.

Can Antibiotic Resistance Genes in Household Food Waste be Reduced by Earthworm Vermicomposting? Underpinning Mechanisms and Strategies

Hundreds of million tons of food waste (FW) is annually generated from the household sectors. Also, it is regarded as a main conduit for the spread of antibiotic resistance genes (ARGs) in the ‘human–environment’ loop. This paper mainly reviews recent studies on the occurrence and dynamics of ARGs in FW and discusses the ins, outs, and spreads of ARGs by the vermicomposting. Our analysis shows that the concentration of FW-borne ARGs and their major hosts (human pathogens) can be effectively reduced and eliminated in the earthworm guts, respectively, due to the increased bacterial fitness cost for ARG-spreading and earthworm immune responses. Of particular interest, the removal performance could be improved by the measures including agricultural waste co-composting and enforced aeration, which concurrently lead to an elevated vermicomposting loading rate and enhanced quality of compost end-products. Furthermore, our review argues that functional microbial inoculum-feeding possibly results in microbial colonization and stable reproduction in earthworm guts. This proposed optimization approach may be instrumental to contain the spread of ARGs and increase the vermicomposting treatment efficiency at the same time.

Disinfectants facilitate the transformation of exogenous antibiotic resistance genes via multiple pathways

The prevalence and spread of multidrug-resistant (MDR) bacteria pose a global challenge to public health. Natural transformation is one of the essential ways for horizontal transfer of antibiotic resistance genes (ARGs). Although disinfectants are frequently used during COVID-19, little is known about whether these disinfectants are associated with the transformation of plasmid-borne ARGs. In our study, we assessed the effect of some disinfectants on bacterial transformation using resistance plasmids as extracellular DNA and E. coli DH5α as the recipient bacteria. The results showed that these disinfectants at environmentally relevant concentrations, including benzalkonium bromide (BB), benzalkonium chloride (BC) and polyhexamethylene guanidine hydrochloride (PHMG), significantly enhanced the transformation of plasmid-encoded ARGs. Furthermore, we investigated the mechanisms underlying the promotive effect of disinfectants on transformation. We revealed that the addition of disinfectants significantly increased the membrane permeability and promoted membrane-related genes expression. Moreover, disinfectants led to the boosted bacterial respiration, ATP production and flagellum motility, as well as increased expression of bacterial secretion system-related genes. Together, our findings shed insights into the spread of ARGs through bacterial transformation and indicate potential risks associated with the widespread use of disinfectants.

Molecular characterization of class 1 integrons in carbapenem-resistant Enterobacterales isolates

ObjectiveCarbapenem-resistant Enterobacterales (CRE) infections result in higher treatment costs and mortality rates. Integrons play important roles in emergence and spread of antibiotic resistant genes. To get a better understand on the effects of integron on CRE resistance, distribution of common carbapenemase genes and class 1 integron in clinical CRE isolates were investigated. MethodCarbapenemase genes, including blaKPC, blaVIM, blaIMP, blaNDM, blaGES, blaVEB and blaOXA-23, were screened in 161 CRE isolates and subtypes of these genes were confirmed through sequence analysis. Class 1 integron was screened and common promoter and gene cassette arrays were determined by sequencing. The resistant rates to clinical commonly used antibiotics between integron positive and integron negative CRE isolates were compared. ResultsOf 161 CRE isolates, the most prevalent carbapenemase gene was blaKPC-2, which was detected in 139 isolates, including 99 Klebsiella pneumoniae. Class 1 integron was detected in 78 isolates. Twenty different gene cassettes, including two carbapenemase genes blaVEB-1 and blaIMP-4, and nine different gene cassette arrays, including blaVEB-1-aadB-arr-2-cmlA5-blaOXA-10-aadA1, aadB-catB8-blaOXA-10-aadA1-dfrA1-aacA4 and blaIMP-4-qacG-aacA4-catB3, were detected. Five types of common promoters were identified. Relative weak promoter PcH1 was the dominant type. Resistant rates of CRE isolates containing class 1 integrons to ceftazidime, amikacin, trimethoprim/sulfamethoxazole and gentamicin were higher than those without class 1 integrons (P < 0.05). ConclusionClass 1 integrons play important roles in the emergence and spread of CRE resistance. To the best of our knowledge, this is the first report of aadB-catB8-blaOXA-10-aadA1-dfrA1-aacA4 and blaIMP-4-qacG-aacA4-catB3 in the same Providencia rettgeri isolate and blaVEB-1-aadB-arr-2-cmlA5-blaOXA-10-aadA1 in P. rettgeri.

Elevated CO2 alleviated the dissemination of antibiotic resistance genes in sulfadiazine-contaminated soil: A free-air CO2 enrichment study

Climate change affects soil microbial communities and their genetic exchange, and subsequently modifies the transfer of antibiotic resistance genes (ARGs) among bacteria. However, how elevated CO2 impacts soil antibiotic resistome remains poorly characterized. Here, a free-air CO2 enrichment system was used in the field to investigate the responses of ARGs profiles and bacterial communities to elevated CO2 (+200 ppm) in soils amended with sulfadiazine (SDZ) at 0, 0.5 and 5 mg kg−1. Results showed that SDZ exposure induced the co-occurrence of beta-lactamase and tetracycline resistance genes, and SDZ at 5 mg kg−1 enhanced the abundance of aminoglycoside, sulfonamide and multidrug resistance genes. However, elevated CO2 weakened the effects of SDZ at 0.5 mg kg−1 following an observed reduction in the total abundance of ARGs and mobile genetic elements. Additionally, elevated CO2 significantly decreased the abundance of vancomycin resistance genes and alleviated the stimulation of SDZ on the dissemination of aminoglycoside resistance genes. Correlation analysis and structural equation models revealed that elevated CO2 could directly influence the spread of ARGs or impose indirect effects on ARGs by affecting soil properties and bacterial communities. Overall, our results furthered the knowledge of the dissemination risks of ARGs under future climate scenarios.

Surveillance on ESBL-Escherichia coli and Indicator ARG in Wastewater and Reclaimed Water of Four Regions of Spain: Impact of Different Disinfection Treatments

In the present study, the occurrence of indicator antibiotic-resistant bacteria (ARB) and antibiotic resistance genes (ARGs) both in the influent and the effluent of four Spanish wastewater treatment plants (WWTPs) was monitored for 12 months, and the susceptibility profiles of 89 recovered extended spectrum β-lactamase (ESBL)-producing Escherichia coli isolates were obtained against a wide range of antimicrobials. The aim of the study was to better understand whether the current wastewater treatment practices allow us to obtain safe reclaimed water mitigating the spread of ARB and ARGs to the environment. Results showed high concentrations of ESBL-producing E. coli as well as a high prevalence of a range of ARGs in the influent samples. The reclamation treatments implemented in the WWTPs were effective in reducing both the occurrence of ESBL E. coli and ARGs, although significant differences were observed among WWTPs. Despite these reductions in occurrence observed upon wastewater treatment, our findings suggest that WWTP effluents may represent an important source of ARGs, which could be transferred among environmental bacteria and disseminate antimicrobial resistance through the food chain. Remarkably, no major differences were observed in the susceptibility profiles of the ESBL E. coli isolated from influent and effluent waters, indicating that water treatments do not give rise to the emergence of new resistance phenotypes.

Horizontal Gene Transfer, Fitness Costs and Mobility Shape the Spread of Antibiotic Resistance Genes into Experimental Populations of Acinetobacter Baylyi.

Horizontal gene transfer (HGT) is important for microbial evolution, but how evolutionary forces shape the frequencies of horizontally transferred genetic variants in the absence of strong selection remains an open question. In this study, we evolve laboratory populations of Acinetobacter baylyi (ADP1) with HGT from two clinically relevant strains of multidrug-resistant Acinetobacter baumannii (AB5075 and A9844). We find that DNA can cross the species barrier, even without strong selection, and despite substantial DNA sequence divergence between the two species. Our results confirm previous findings that HGT can drive the spread of antibiotic resistance genes (ARGs) without selection for that antibiotic, but not for all of the resistance genes present in the donor genome. We quantify the costs and benefits of horizontally transferred variants and use whole population sequencing to track the spread of ARGs from HGT donors into antibiotic-sensitive recipients. We find that even though most ARGs are taken up by populations of A. baylyi, the long-term fate of an individual gene depends both on its fitness cost and on the type of genetic element that carries the gene. Interestingly, we also found that an integron, but not its host plasmid, is able to spread in A. baylyi populations despite its strong deleterious effect. Altogether, our results show how HGT provides an evolutionary advantage to evolving populations by facilitating the spread of non-selected genetic variation including costly ARGs.

Genomic Insights into CRISPR-Harboring Plasmids in the Klebsiella Genus: Distribution, Backbone Structures, Antibiotic Resistance, and Virulence Determinant Profiles.

CRISPR systems are often encoded by many prokaryotes as adaptive defense against mobile genetic elements (MGEs), but several MGEs also recruit CRISPR components to perform additional biological functions. Type IV-A systems are identified in Klebsiella plasmids, yet the distribution, characterization, and role of these plasmids carrying CRISPR systems in the whole Klebsiella genus remain unclear. Here, we performed large-scale comparative analysis of these plasmids using publicly available plasmid genomes. CRISPR-harboring plasmids were mainly distributed in Klebsiella pneumoniae (9.09%), covering 19.23% of sequence types, but sparse in Klebsiella species outside Klebsiella pneumoniae (3.92%). Plasmid genome comparison reiterated that these plasmids often carried the cointegrates of IncFIB and IncHI1B replicons, occasionally linked to other replicons, such as IncFIA, IncFII, IncR, IncQ, and IncU. Comparative genome analysis showed that CRISPR-carrying Klebsiella plasmids shared a conserved pNDM-MAR-like conjugation module as their backbones and served as an important vector for the accretion of antibiotic resistance genes (ARGs) and even virulence genes (VGs). Moreover, compared with CRISPR-negative IncFIB/IncHIB plasmids, CRISPR-positive IncFIB/IncHIB plasmids displayed high divergences in terms of ARGs, VGs, GC content, plasmid length, and backbone structures, suggesting their divergent evolutionary paths. The network analysis revealed that CRISPR-positive plasmids yielded fierce competitions with other plasmid types, especially conjugative plasmids, thereby affecting the dynamics of plasmid transmission. Overall, our study provides valuable insights into the role of CRISPR-positive plasmids in the spread of ARGs and VGs in Klebsiella genus.

Potential reservoirs of antimicrobial resistance in livestock waste and treated wastewater that can be disseminated to agricultural land

Livestock manure, dairy lagoon effluent, and treated wastewater are known reservoirs of antibiotic resistance genes (ARGs), antibiotic-resistant bacteria (ARB), and virulence factor genes (VFGs), and their application to agricultural farmland could be a serious public health threat. However, their dissemination to agricultural lands and impact on important geochemical pathways such as the nitrogen (N) cycle have not been jointly explored. In this study, shotgun metagenomic sequencing and analyses were performed to examine the diversity and composition of microbial communities, ARGs, VFGs, and N cycling genes in different livestock manure/lagoon and treated wastewater collected from concentrated animal feeding operations (CAFOs) and a municipal wastewater treatment plant along the west coast of the United States. Multivariate analysis showed that diversity indices of bacterial taxa from the different microbiomes were not significantly different based on InvSimpson (P = 0.05), but differences in ARG mechanisms were observed between swine manure and other microbiome sources. Comparative resistome profiling showed that ARGs in microbiome samples belonged to four core resistance classes: aminoglycosides (40–55 %), tetracyclines (30–45 %), beta-lactam-resistance (20–35 %), macrolides (18–30 %), and >50 % of the VFGs that the 24 microbiomes harbored were phyletically affiliated with two bacteria, Bacteroidetes fragilis and Enterobacter aerogenes. Network analysis based on Spearman correlation showed co-occurrence patterns between several genes such as transporter-gene and regulator, efflux pump and involved-in-polymyxin- resistance, aminoglycoside, beta-lactam, and macrolide with VFGs and bacterial taxa such as Firmicutes, Candidatus Themoplasmatota, Actinobacteria, and Bacteroidetes. Metabolic reconstruction of metagenome-assembled genome (MAGs) analysis showed that the most prevalent drug resistance mechanisms were associated with carbapenem resistance, multidrug resistance (MDR), and efflux pump. Bacteroidales was the main taxa involved in dissimilatory nitrate reduction (DNRA) in dairy lagoon effluent. This study demonstrates that the dissemination of waste from these sources can increase the spread of ARGs, ARB, and VFGs into agricultural lands, negatively impacting both soil and human health.

Profiles and key drivers of bacteria/phage co-mediated antibiotic resistance genes during swine manure composting amended with humic acid

Phages can promote the spread of antibiotic resistance genes (ARGs) in agricultural environments through transduction. However, studies on phage-mediated ARG profiles during composting have not been performed. This study investigated the effects of adding humic acid (HA) on the abundances of bacteria/phage co-mediated ARGs (b/pARGs) during swine manure composting and the key factors that affected the transmission of b/pARGs. The results showed that the addition of 5 % HA during composting could effectively reduce the absolute abundances of b/pARGs, inhibit the proliferation of pathogenic microorganisms (e.g., Corynebacterium and Streptococcus) that carried ARGs, and ultimately affect the fate of b/pARGs in the composting process by regulating key environmental factors to change the abundance of co-host bacteria. Overall, the findings of this study provided new information for understanding the main driving factors affecting the b/pARGs profile and provided a reference for the prevention and control of ARGs pollution during composting.

Biosynthetic Gene Clusters from Swine Gut Microbiome.

The abuse of antibiotics has become a serious health challenge in the veterinary field. It creates environmental selection pressure on bacteria and facilitates the rapid spread of antibiotic resistance genes. The speed of discovery and application of cost-effective alternatives to antibiotics is slow in pig production. Natural products from biosynthetic gene clusters (BGCs) represent promising therapeutic agents for animal and human health and have attracted extraordinary passion from researchers due to their ability to participate in biofilm inhibition, stress resistance, and the killing of competitors. In this study, we detected the presence of diverse secondary metabolite genes in porcine intestines through sequence alignment in the antiSMASH database. After comparing variations in microbial BGCs' composition between the ileum and the colon, it was found that the abundance of the resorcinol gene cluster was elevated in the ileal microbiome, whereas the gene cluster of arylpolyene was enriched in the colonic microbiome. The investigation of BGCs' diversity and composition differences between the ileal and colonic microbiomes provided novel insights into further utilizing BGCs in livestock. The importance of BGCs in gut microbiota deserves more attention for promoting healthy swine production.