Abundance Of Antibiotic Resistance Genes Research Articles

Multi-omics strategy reveals potential role of antimicrobial resistance and virulence factor genes responsible for Simmental diarrheic calves caused by Escherichia coli.

Escherichia coli (E. coli) is reported to be an important pathogen associated with calf diarrhea. Antibiotic resistance genes (ARGs) and virulence factor genes (VFGs) pose a considerable threat to both animal and human health. However, little is known about the characterization of ARGs and VFGs presented in the gut microbiota of diarrheic calves caused by E. coli. In this study, we used multi-omics strategy to analyze the ARG and VFG profiles of Simmental calves with diarrhea caused by E. coli K99. We found that gut bacterial composition and their microbiome metabolic functions varied greatly in diarrheic calves compared to healthy calves. In total, 175 ARGs were identified, and diarrheal calves showed a significantly higher diversity and abundance of ARGs than healthy calves. Simmental calves with diarrhea showed higher association of VFGs with pili function, curli assembly, and ferrienterobactin transport of E. coli. Co-occurrence patterns based on Pearson correlation analysis revealed that E. coli had a highly significant (P < 0.0001) correlation coefficient (>0.8) with 16 ARGs and 7 VFGs. Metabolomics analysis showed that differentially expressed metabolites in Simmental calves with diarrhea displayed a high correlation with the aforementioned ARGs and VFGs. Phylotype analysis of E. coli genomes showed that the predominant phylogroup B1 in diarrheic Simmental calves was associated with 10 ARGs and 3 VFGs. These findings provide an overview of the diversity and abundance of the gut microbiota in diarrheic calves caused by E. coli and pave the way for further studies on the mechanisms of antibiotic resistance and virulence in the calves affected with diarrhea.IMPORTANCESimmental is a well-recognized beef cattle breed worldwide. They also suffer significant economic losses due to diarrhea. In this study, fecal metagenomic analysis was applied to characterize the antibiotic resistance gene (ARG) and virulence factor gene (VFG) profiles of diarrheic Simmental calves. We identified key ARGs and VFGs correlated with Escherichia coli isolated from Simmental calves. Additionally, metabolomics analysis showed that differentially expressed metabolites in Simmental calves with diarrhea displayed a high correlation with the aforementioned ARGs and VFGs. Our findings provide an insight into the diversity and abundance of the gut microbiota in diarrheic calves caused by Escherichia coli and pave the way for further studies on the mechanisms of antibiotic resistance and virulence in the diarrheal calves from cattle hosts.

[Characteristics of Microorganisms and Antibiotic Resistance Genes of the Riparian Soil in the Lanzhou Section of the Yellow River].

Riparian soil is a critical area of watersheds. The characteristics of biological contaminants in riparian soil affect the pollution control of the watershed water environment. Thus, the microbial community structure, antibiotic resistance genes (ARGs), and mobile genetic elements (MGEs) in the riparian soil of the Lanzhou section of the Yellow River were investigated by analyzing the characteristics of soil samples collected from farmland, mountains, and industrial land. The results showed that the Proteobacteria, Bacteroidetes, and Actinobacteria were the dominant phyla in the riparian soil of Lanzhou section of the Yellow River. The microbial structure in the riparian soil was significantly correlated with the land use type (P &lt; 0.05). The α diversity index of bacterial communities in land types was in the order of farmland &gt; mountain &gt; industry. Sulfonamide-typed ARGs were the most dominant genes in the soil of the Lanzhou section of the Yellow River Basin, among which the sul1 gene had the highest abundance, 20-36 000 times that of other detected ARGs. Moreover, the total absolute abundance of ARGs in industrial soil was the highest. Principal coordinate analysis (PCoA) displayed that the ARGs characteristics had a significant correlation with land types (P &lt; 0.05), and intl1 and tnpA-04 drove the diffuseness of sulfonamide and tetracycline ARGs, respectively. Redundancy analysis (RDA) demonstrated that the content of inorganic salt ions and total phosphorus in the soil of the riparian zone of the Yellow River Lanzhou section were the main environmental factors, modifying the distribution of the microbial structure. Halobacterota and Acidobacteriota were the main microflora that drove the structural change in ARGs.

Impact of earthworms on antibiotic resistance genes removal in ampicillin-contaminated soil through bacterial community alteration.

The emergence of antibiotic resistance genes (ARGs) as contaminants in soil poses a significant threat to public health. Earthworms (Eisenia foetida), which are common inhabitants of soil, have been extensively studied for their influence on ARGs. However, the specific impact of earthworms on penicillin-related ARGs remains unclear. In this study, we investigate the role of earthworms in mitigating ARGs, specifically penicillin-related ARGs, in ampicillin-contaminated soil. Utilizing high-throughput quantitative PCR (HT-qPCR), we quantified a significant reduction in the relative abundance of penicillin-related ARGs in soil treated with earthworms, showing a decrease with a p-value of <0.01. Furthermore, high-throughput 16S rRNA gene sequencing revealed that earthworm intervention markedly alters the microbial community structure, notably enhancing the prevalence of specific bacterial phyla such as Proteobacteria, Firmicutes, Chloroflexi, and Tenericutes. Our findings not only demonstrate the effectiveness of earthworms in reducing the environmental load of penicillin-related ARGs but also provide insight into the alteration of microbial communities as a potential mechanism. This research contributes to our understanding of the role of earthworms in mitigating the spread of antibiotic resistance and provides valuable insights for the development of strategies to combat this global health issue.

Entry pathways determined the effects of MPs on sludge anaerobic digestion system: The views of methane production and antibiotic resistance genes fates

Sludge is one of the primary reservoirs of microplastics (MPs), and the effects of MPs on subsequent sludge treatment raised attention. Given the entry pathways, MPs would exhibit different properties, but the entry pathway-dependent effect of MPs on sludge treatment performance and the fates of antibiotic resistance genes (ARGs), another high-risk emerging contaminant, were seldom documented. Herein, MPs with two predominant entry pathways, including wastewater-derived (WW-derived) and anaerobic digestion-introduced (AD-introduced), were used to investigate the effects on AD performance and ARGs abundances. The results indicated that WW-derived MPs, namely the MPs accumulated in sludge during the wastewater treatment process, exhibited significant inhibition on methane production by 22.8%–71.6%, while the AD-introduced MPs, being introduced in the sludge AD process, slightly increased the methane yield by 4.7%–17.1%. Meanwhile, MPs were responsible for promoting transmission of target ARGs, and polyethylene terephthalate MPs (PET-MPs) showed a greater promotion effect (0.0154–0.0936) than polyamide MPs (PA-MPs) (0.0013–0.0724). Compared to size, entry pathways and types played more vital roles on MPs influences. Investigation on mechanisms based on microbial community structure revealed characteristics (aging degree and types) of MPs determined the differences of AD performance and ARGs fates. WW-derived MPs with longer aging period and higher aging degree would release toxics and decrease the activities of microorganisms, resulting in the negative impact on AD performance. However, AD-introduced MPs with short aging period exhibited marginal impacts on AD performance. Furthermore, the co-occurrent network analysis suggested that the variations of potential host bacteria induced by MPs with different types and aging degree attributed to the dissemination of ARGs. Distinctively from most previous studies, the MPs with different sizes did not show remarkable effects on AD performance and ARGs fates. Our findings benefited the understanding of realistic environmental behavior and effect of MPs with different sources.

Spread of plasmids carrying antibiotic resistance genes in soil-lettuce-snail food chain.

Fertilization can change the composition of antibiotic resistance genes(ARGs) and their host bacteria in agricultural fields, while complex microbial activities help ARGs into crops and transmit them to humans through agricultural products.Therefore, this study constructed a farmland food chain with soil-lettuce-snail as a typical structure, added genetically engineered Pseudomonas fluorescens containing multidrug-resistant plasmid RP4 to track its spread in the farmland food chain, and used different fertilization methods to explore its influence on the spread and diffusion of ARGs and intl1 in the farmland food chain. It was found that exogenous Pseudomonas can enter plants from soil and pass into snails' intestines, and there is horizontal gene transfer phenomenon of RP4 plasmid in bacteria. At different interfaces of the constructed food chain, the addition of exogenous drug-resistant bacteria had different effects on the total abundance of ARGs and intl1. Fertilization, especially manure, not only promoted the spread of Pseudomonas aeruginosa and the transfer of RP4 plasmid levels, but also significantly increased the total abundance of ARGs and intl1 at all interfaces of the constructed food chain. The main ARGs host bacteria in the constructed food chain include Proteobacteria, Bacteroides, and Firmicutes, while Flavobacterium of Bacteroides is the unique potential host bacteria of RP4 plasmid. In conclusion, this study provides a reference for the risk assessment of ARGs transmitted to the human body through the food chain, and has important practical significance to reduce the antibiotic resistance contamination of agricultural products and ensure the safety of vegetable basket.

Transmission mechanism of antibiotic resistance genes and their differences between water and sediment in the Weihe River Basin

Antibiotic resistance genes (ARGs) are emerging microbial pollutants that are regulated by many factors and pose potential threats to aquatic environments. In this study, we used network analysis, correlation analysis, and constructed models based on metagenomic sequencing results to explore the spatial patterns, impact mechanisms, transmission risks and differences in ARGs in the water and sediment of the Weihe River Basin. The findings revealed notable disparities in ARGs, mobile genetic elements (MGEs), and bacterial communities. In the sediment, the abundance of ARGs was considerably greater than that in water. Moreover, the percentage of ARGs shared by the two components reached a value of 85.8%. Through network analysis, it was determined that the presence of 16 MGEs and 20 bacterial phyla was strongly associated with ARGs (R2>0.7, P<0.05). The Mantel test showed that abiotic factors including DO, pH, nutrients, and heavy metals played important roles in the distribution of ARGs (P<0.05). A structural equation model revealed that the key factors influencing the distribution of ARGs in water were bacterial diversity and environmental parameters (standardized effects of -0.730 and -0.667), and those in sediment were bacterial diversity and MGEs (standardized effects of -0.751 and 0.851). Neutral modeling indicated that deterministic processes played an important role in the assembly of ARGs in the water of the Weihe River Basin, and stochastic processes were dominant in the sediment. There was a highly significant positive linear correlation between ARGs and pathogens, and there was more complex co-occurrence in the water than in the sediment (R2>0.9, P<0.05), with stronger migration and transmission occurring. Exploring ARGs in large-scale watersheds is immensely important for elucidating their traits and transmission mechanisms and consequently paving the way for the formulation of efficient strategies to mitigate resistance threats.

Phenacetin promoted the rapid start-up and stable maintenance of partial nitrification: Responses of nitrifiers and antibiotic resistance genes

Phenacetin (PNCT) belongs to one of the earliest synthetic antipyretics. However, impact of PNCT on nitrifying microorganisms in wastewater treatment plants and its potential microbial mechanism was still unclear. In this study, PN could be initiated within six days by PNCT anaerobic soaking treatment (8 mg/L). In order to improve the stable performance of PN, 21 times of PNCT aerobic soaking treatment every three days was conducted and PN was stabilized for 191 days. After PN was damaged, ten times of PNCT aerobic soaking treatment every three days was conducted and PN was recovered after once soaking, maintained over 88 days. Ammonia oxidizing bacteria might change the dominant oligotype to gradually adjust to PNCT, and the increase of abundance and activity of Nitrosomonas promoted the initiation of PN. For nitrite-oxidizing bacteria (NOB), the increase of Candidatus Nitrotoga and Nitrospira destroyed PN, but PN could be recovered after once aerobic soaking illustrating NOB was not resistant to PNCT. KEGG and COG analysis suggested PNCT might disrupt rTCA cycle of Nitrospira, resulting in the decrease of relative abundance of Nitrospira. Moreover, PNCT did not lead to the sharp increase of absolute abundances of antibiotic resistance genes (ARGs), and the risk of ARGs transmission was negligible.

Leaching of antibiotic resistance genes and microbial assemblages following poultry litter applications in karst and non-karst landscapes

Antibiotic resistance is increasingly recognized as a critical challenge affecting human, animal, and environmental health. Yet, environmental dynamics and transport of antibiotic resistance genes (ARGs) and microbial communities in karst and non-karst leachate following poultry litter land applications are not well understood. This study investigates impacts of broiler poultry litter application on the proliferation of ARGs (tetW, qnrS, ermB, sulI, and blaCTX-M-32), class 1 integron (intI1 i), and alterations in microbial communities (16S rRNA) within karst derived soils, which are crucial and under-researched systems in the global hydrological cycle, and non-karst landscapes. Using large, intact soil columns (45 cm diam. × 100 cm depth) from karst and non-karst landscapes, the role of preferential flow and ARG transport in leachate was enumerated following surface application of poultry litter and simulated rain events. This research demonstrated that in poultry litter amended karst soils, ARG (i.e., ermB and tetW) abundance in leachate increased 1.5 times compared to non-karst systems (p < 0.05), highlighting the influence of geological factors on ARG proliferation. Notably, microbial communities in karst soil leachate exhibited increased diversity and abundance, suggesting a potential linkage between microbial composition and ARG presence. Further, our correlation and network analyses identified relationships between leachate ARGs, microbial taxa, and physicochemical properties, underscoring the complex interplay in these environmentally sensitive areas. These findings illuminate the critical role of karst systems in shaping ARG abundance and pollutant dispersal and microbial community dynamics, thus emphasizing the need for landscape-specific approaches in managing ARG dissemination to the environment. This study provides a deeper understanding of hydrogeological ARG dynamics but also lays the groundwork for future research and targeted strategies to mitigate ARG dissemination through targeted manure applications across agricultural landscapes.

Could the effect of antimicrobials on antimicrobial resistance be saturated at high antimicrobial consumption? A comparison of the MORDOR and ResistAZM studies

Background and objectivesAntimicrobial resistance poses a considerable threat in high-antimicrobial-consumption populations, such as men who have sex with men (MSM) taking HIV pre-exposure prophylaxis. While the ResistAZM trial found no increase in macrolide resistance genes in MSM with gonorrhea after azithromycin treatment, the MORDOR trial observed an increase in these genes after mass azithromycin distribution. We hypothesized that this could be due to saturation of the resistome. To test this hypothesis, we compared the abundance of macrolide resistance determinants in anorectal samples between the baselines of the two trials. MethodsShotgun metagenome reads from the anorectal baseline samples from the ResistAZM (n=42) and MORDOR (n=30) trials were analyzed using AMRPlusPlus. Non-host reads were mapped to the MEGARes database to detect antibiotic resistance genes (ARG). AMR was normalized using cumulative sum scaling, and ARG abundance was estimated. ResultsMacrolide, lincosamides and streptogramins (MLS) determinants were approximately 10-fold more abundant in the ResistAZM than the MORDOR samples (p = <0.001). ConclusionsThe findings are compatible with our hypothesis. Thus, in populations with high antimicrobial use, the relationship between antimicrobial consumption and AMR may be diminished due to saturation. These findings are vital for future studies investigating the resistogencity of novel interventions, such as doxycycline post-exposure prophylaxis, in populations with high preceding consumption of antimicrobials.

Critical insights into the Hormesis of antibiotic resistome in saline soil: Implications from salinity regulation

Soil is recognized as an important reservoir of antibiotic resistance genes (ARGs). However, the effect of salinity on the antibiotic resistome in saline soils remains largely misunderstood. In this study, high-throughput qPCR was used to investigate the impact of low-variable salinity levels on the occurrence, health risks, driving factors, and assembly processes of the antibiotic resistome. The results revealed 206 subtype ARGs across 10 categories, with medium-salinity soil exhibiting the highest abundance and number of ARGs. Among them, high-risk ARGs were enriched in medium-salinity soil. Further exploration showed that bacterial interaction favored the proliferation of ARGs. Meanwhile, functional genes related to reactive oxygen species production, membrane permeability, and adenosine triphosphate synthesis were upregulated by 6.9%, 2.9%, and 18.0%, respectively, at medium salinity compared to those at low salinity. With increasing salinity, the driver of ARGs in saline soils shifts from bacterial community to mobile gene elements, and energy supply contributed 28.2% to the ARGs at extreme salinity. As indicated by the neutral community model, stochastic processes shaped the assembly of ARGs communities in saline soils. This work emphasizes the importance of salinity on antibiotic resistome, and provides advanced insights into the fate and dissemination of ARGs in saline soils. Environmental ImplicationsSaline soil is a widely distributed soil type in which contaminants can directly and extensively affect the health of a huge population. In contrast, as an emerging contaminant, ARGs have not received enough attention for a long time in terms of their occurrence and health risks in saline soils. In this study, the deterministic effect of salinity on the antibiotic resistomein saline soil and its potential mechanisms were deeply explored. This work expanded our understanding of the neglected health risks of antibiotic resistance, and provided advanced insights for maintaining ecological security in saline soils.

Removal of antibiotic resistant bacteria by the coupled system of ferrous ion activated peroxymonosulfate (PMS) and sodium percarbonate (SPC): Performance and mechanisms

Antibiotic resistance has turned into a focus of universal public health concern. Wastewater treatment plants are considered as reservoirs of antibiotic resistant bacteria (ARB) and antibiotic resistant genes (ARGs), which can be transmitted to the environment. In this research, an effective method for inactivating Escherichia coli DH5α (E. coli DH5α) carrying resistance genes was developed utilizing dual oxidant system of peroxymonosulfate (PMS) and sodium percarbonate (SPC) in the presence of ferrous ions (Fe(II)). The results indicated that E. coli DH5α could be inactivated 6.36 log under the conditions of 0.50 mM PMS, 0.50 mM SPC and 0.50 mM Fe(II) after 30 min. Besides, the removal of different types of ARB and pollutants could also be achieved by Fe(II)/PMS/SPC system. The quenching experiment and EPR analysis demonstrated that reactive species (•OH, SO4•−, O2•−, 1O2 and CO3•−) involved in the inactivation of E. coli DH5α. Bacterial damage mechanisms were systematically studied in terms of cell structure and morphology, enzyme activity, malondialdehyde and intracellular reactive oxygen species levels. The inactivation of E. coli DH5α in complex water matrix (including coexisting of anions and natural organic matter) and real wastewater was inhibited. The abundance of intracellular ARGs decreased by 1.15 log, whereas extracellular ARGs increased by 0.32 log. This research supplied a prospective approach for inhibiting the dissemination of antibiotic resistance.

Microplastics in anoxic/aerobic membrane bioreactor (A/O-MBR): Characteristics, biofilms, degradation and carrier for antibiotic resistance genes

Wastewater treatment plants (WWTPs) are proposed as one of the ways for microplastics (MPs) to enter the water environment. MPs as the carrier of other pollutants, the inevitable aging process will change the characteristics and fate of MPs. Herein, the physicochemical property alteration, microbial community evolution and antibiotic resistance genes (ARGs) occurrence of the aged MPs surface were analyzed in A/O-MBR system. Remarkable alteration in MPs surface properties, which included the decrease in hydrophobicity and the increase in oxygen-containing functional groups, led to aging of MPs. Aging MPs are more likely to grow biofilms, which further increased the probability of microbial colonization. The genera Hydrogenophaga, Exiguobacterium, Enterobacter, Vibrio, Flavobacterium and Bacillus on MPs surface showed higher degradation potential. ARGs determination results indicated that MPs could enrich ARGs on their surface, and the relative abundance of β-lactam ARGs on different MPs surfaces was the highest. The above research results indicated that MPs in sewage treatment systems not only undergo aging, but also carry ARGs, causing further harm to the environment.

Microbiologically induced calcite precipitation (MICP) in situ remediated heavy metal contamination in sludge nutrient soil

Microbiologically induced calcite precipitation (MICP), as a newly developing bioremediation technology, could redeem heavy metal contamination in diverse scenarios. In this study, MICP bacterium Sporosarcina ureilytica ML-2 was employed to suppress the pollution of Pb, Cd and Zn in municipal sludge nutrient soil. After MICP remediation, the exchangeable Cd and Zn in sludge nutrient soil were correspondingly reduced by 31.02% and 6.09%, while the carbonate-bound Pb, Cd and Zn as well as the residual fractions were increased by 16.12%, 6.63%, 13.09% and 6.10%, 45.70%, 3.86%, respectively. In addition, the extractable Pb, Cd and Zn either by diethylenetriaminepentaacetic acid (DTPA) or toxicity characteristic leaching procedure (TCLP) in sludge nutrient soil were significantly reduced. These results demonstrated that the bio-calcite generated via MICP helped to immobilize heavy metals. Furthermore, MICP treatment improved the abundance of functional microorganisms related to urea cycle, while reduced the overall abundance of metal resistance genes (MRGs) and antibiotic resistance genes (ARGs). This work confirmed the feasibility of MICP in remediation of heavy metal in sludge nutrient soil, which expanded the application field of MICP and provided a promising way for heavy metal pollution management. Environmental ImplicationMunicipal sludge is a repertoire of various pollutants, in which heavy metals are typical hazardous pollutants with significant biological toxicity. Our work applied the emerging microbiologically induced calcite precipitation (MICP) technology to reduce the toxicity of Pb, Cd and Zn in municipal sludge nutrient soil before land disposal. The results showed that the MICP mediated by Sporosarcina ureilytica ML-2 could convert the heavy metals from bioavailable form to insoluble carbonate-bound and residual forms, significantly reducing the bio-toxicity. This work expanded the application field of MICP and provided a promising way for heavy metal contamination management.

Characterisation of microbial communities and quantification of antibiotic resistance genes in Italian wastewater treatment plants using 16S rRNA sequencing and digital PCR

The spread of antibiotic resistant bacteria (ARB) and antibiotic resistance genes (ARGs) in humans, animals and environment is a growing threat to public health. Wastewater treatment plants (WWTPs) are crucial in mitigating the risk of environmental contamination by effectively removing contaminants before discharge. However, the persistence of ARB and ARGs even after treatment is a challenge for the management of water system.To comprehensively assess antimicrobial resistance dynamics, we conducted a one-year monitoring study in three WWTPs in central Italy, both influents and effluents. We used seasonal sampling to analyze microbial communities by 16S rRNA, as well as to determine the prevalence and behaviour of major ARGs (sul1, tetA, blaTEM, blaOXA-48, blaCTX-M-1 group, blaKPC) and the class 1 Integron (int1). Predominant genera included in order: Arcobacter, Acinetobacter, Flavobacterium, Pseudarcobacter, Bacteroides, Aeromonas, Trichococcus, Cloacibacterium, Pseudomonas and Streptococcus. A higher diversity of bacterial communities was observed in the effluents compared to the influents. Within these communities, we also identified bacteria that may be associated with antibiotic resistance and pose a significant threat to human health. The mean concentrations (in gene copies per liter, gc/L) of ARGs and int1 in untreated wastewater (absolute abundance) were as follows: sul1 (4.1 × 109), tetA (5.2 × 108), blaTEM (1.1 × 108), blaOXA-48 (2.1 × 107), blaCTX-M-1 group (1.1 × 107), blaKPC (9.4 × 105), and int1 (5.5 × 109). The mean values in treated effluents showed reductions ranging from one to three log. However, after normalizing to the 16S rRNA gene (relative abundance), it was observed that in 37.5 % (42/112) of measurements, the relative abundance of ARGs increased in effluents compared to influents. Furthermore, correlations were identified between ARGs and bacterial genera including priority pathogens. This study improves our understanding of the dynamics of ARGs and provides insights to develop more effective strategies to reduce their spread, protecting public health and preserving the future efficacy of antibiotics.

Accelerated dissemination of antibiotic resistant genes via conjugative transfer driven by deficient denitrification in biochar-based biofiltration systems

Biofiltration systems harbored and disseminated antibiotic resistance genes (ARGs), when confronting antibiotic-contained wastewater. Biochar, a widely used environmental remediation material, can mitigate antibiotic stress on adjoining microbes by lowering the availability of sorbed antibiotics, and enhance the attachment of denitrifiers. Herein, bench-scale biofiltration systems, packed with commercial biochars, were established to explore the pivotal drivers affecting ARG emergence. Results showed that biofiltration columns, achieving higher TN removal and denitrification capacity, showed a significant decrease in ARG accumulation (p < 0.05). The relative abundance of ARGs (0.014 ± 0.0008) in the attached biofilms decreased to 1/5-folds of that in the control group (0.065 ± 0.004). Functional analysis indicated ARGs' accumulation was less attributed to ARG activation or horizontal gene transfer (HGT) driven by sorbed antibiotics. Most denitrifiers, like Bradyrhizobium, Geothrix, etc., were found to be enriched and host ARGs. Nitrosative stress from deficient denitrification was demonstrated to be the dominant driver for affecting ARG accumulation and dissemination. Metagenomic and metaproteomic analysis revealed that nitrosative stress promoted the conjugative HGT of ARGs mainly via increasing the transmembrane permeability and enhancing the amino acid transport and metabolism, such as cysteine, methionine, and valine metabolism. Overall, this study highlighted the risks of deficient denitrification in promoting ARG transfer and transmission in biofiltration systems and natural ecosystems.

Metagenomic and Culturomics Analysis of Microbial Communities within Surface Sediments and the Prevalence of Antibiotic Resistance Genes in a Pristine River: The Zaqu River in the Lancang River Source Region, China

Microbial communities inhabiting sedimentary environments in river source regions serve as pivotal indicators of pristine river ecosystems. While the correlation between antibiotic resistome and pathogenicity with core gut bacteria in humans is well established, there exists a significant knowledge gap concerning the interaction of antibiotic resistance genes (ARGs) and human pathogenic bacteria (HPB) with specific microbes in river source basins, often referred to as “terrestrial gut”. Understanding the microbial composition, including bacteria and resident genetic elements such as ARGs, HPB, Mobile Genetic Elements (MGEs), and Virulence Factors (VFs), within natural habitats against the backdrop of global change, is imperative. To address this gap, an enrichment-based culturomics complementary along with metagenomics was conducted in this study to characterize the microbial biobank and provide preliminary ecological insights into profiling the dissemination of ARGs in the Lancang River Source Basin. Based on our findings, in the main stream of the Lancang River Source Basin, 674 strains of bacteria, comprising 540 strains under anaerobic conditions and 124 under aerobic conditions, were successfully isolated. Among these, 98 species were identified as known species, while 4 were potential novel species. Of these 98 species, 30 were HPB relevant to human health. Additionally, bacA and bacitracin emerged as the most abundant ARGs and antibiotics in this river, respectively. Furthermore, the risk assessment of ARGs predominantly indicated the lowest risk rank (Rank Ⅳ) in terms of endangering human health. In summary, enrichment-based culturomics proved effective in isolating rare and unknown bacteria, particularly under anaerobic conditions. The emergence of ARGs showed limited correlation with MGEs, indicating minimal threats to human health within the main stream of the Lancang River Source Basin.

Arctic plasmidome analysis reveals distinct relationships among associated antimicrobial resistance genes and virulence genes along anthropogenic gradients.

Polar regions are relatively isolated from human activity and thus could offer insight into anthropogenic and ecological drivers of the spread of antibiotic resistance. Plasmids are of particular interest in this context given the central role that they are thought to play in the dissemination of antibiotic resistance genes (ARGs). However, plasmidomes are challenging to profile in environmental samples. The objective of this study was to compare various aspects of the plasmidome associated with glacial ice and adjacent aquatic environments across the high Arctic archipelago of Svalbard, representing a gradient of anthropogenic inputs and specific treated and untreated wastewater outflows to the sea. We accessed plasmidomes by applying enrichment cultures, plasmid isolation and shotgun Illumina sequencing of environmental samples. We examined the abundance and diversity of ARGs and other stress-response genes that might be co/cross-selected or co-transported in these environments, including biocide resistance genes (BRGs), metal resistance genes (MRGs), virulence genes (VGs) and integrons. We found striking differences between glacial ice and aquatic environments in terms of the ARGs carried by plasmids. We found a strong correlation between MRGs and ARGs in plasmids in the wastewaters and fjords. Alternatively, in glacial ice, VGs and BRGs genes were dominant, suggesting that glacial ice may be a repository of pathogenic strains. Moreover, ARGs were not found within the cassettes of integrons carried by the plasmids, which is suggestive of unique adaptive features of the microbial communities to their extreme environment. This study provides insight into the role of plasmids in facilitating bacterial adaptation to Arctic ecosystems as well as in shaping corresponding resistomes. Increasing human activity, warming of Arctic regions and associated increases in the meltwater run-off from glaciers could contribute to the release and spread of plasmid-related genes from Svalbard to the broader pool of ARGs in the Arctic Ocean.

Effects of earthworms on antibiotic resistance genes in different soil-plant systems.

Earthworms play an important role in the soil environment. To explore the difference in earthworms influence on various media in different soil-plant systems, the abundance of tetracycline, sulfonamide and quinolone resistance genes and the structure of the bacterial community were analysed from five different media including non-rhizosphere soil, rhizosphere soil, phyllosphere, root endophytes and earthworm intestine by real-time quantitative PCR and high-throughput 16S rRNA sequencing. Studies have shown that earthworms can reduce the absolute abundance of antibiotic resistance genes (ARGs) in non-rhizosphere soil. Root endophytes in the soil-cabbage system and rhizosphere soil in the soil-setaria system had the same findings. Earthworms can change the bacterial community structure, especially that of Proteobacteria and Cyanobacteria in the phyllosphere and root endophytes. Redundancy analysis (RDA) results that bacterial community change was the main factor affecting ARGs. In addition, earthworms increased the proportion of Cyanobacteria in root endophytes, and Cyanobacteria was significantly positively correlated with sul3. This study provides a scientific basis for controlling the migration and diffusion of ARGs and reducing environmental risks in soil-plant systems in the future.

Body size: A hidden trait of the organisms that influences the distribution of antibiotic resistance genes in soil

Body size is a key life-history trait of organisms, which has important ecological functions. However, the relationship between soil antibiotic resistance gene (ARG) distribution and organisms' body size has not been systematically reported so far. Herein, the impact of organic fertilizer on the soil ARGs and organisms (bacteria, fungi, and nematode) at the aggregate level was analyzed. The results showed that the smaller the soil aggregate size, the greater the abundance of ARGs, and the larger the body size of bacteria and nematodes. Further analysis revealed significant positive correlations of ARG abundance with the body sizes of bacteria, fungi, and nematodes, respectively. Additionally, the structural equation model demonstrated that changes in soil fertility mainly regulate the ARG abundance by affecting bacterial body size. The random forest model revealed that total phosphorus was the primary soil fertility factor influencing the body size of organisms. Therefore, these findings proposed that excessive application of phosphate fertilizers could increase the risk of soil ARG transmission by increasing the body size of soil organisms. This study highlights the significance of organisms' body size in determining the distribution of soil ARGs and proposes a new disadvantage of excessive fertilization from the perspective of ARGs.

Microbial-mediated conversion of soil organic carbon co-regulates the evolution of antibiotic resistance

The influence of organic carbon on the proliferation of antibiotic resistance genes (ARGs) in the soil has been widely documented. However, it is unclear how soil organic carbon (SOC) interacts with the evolution of antibiotic resistance in bacteria. Here, we examined the variations in ARGs abundance during SOC mineralization and explored the microbiological mechanisms and key metabolic pathways involved in their coevolution. The results showed that the SOC mineralization rate was closely correlated with ARGs abundance (p < 0.05). High organic carbon (OC) mineralization was conducive to the occurrence of multidrug resistance genes. For example, multidrug_transporter and mexB increased 2.26 and 7.83 times from the initial level. The competitor (stress) evolutionary strategy model revealed that higher OC inputs drive environmental microorganisms to evolve from stress tolerant to high resistance and strong adaptation. Meta-genomic and transcriptomic analyses revealed that the conversion process of pyruvate to acetyl-CoA to acetate was the critical metabolic pathway for the co-regulation of antibiotic resistance. Gene deletion validation trials have demonstrated that the key functional genes (ackA and pta) involved in this process can modulate the development of vancomycin and multidrug resistance. This outcome provides a preliminary framework for microbial mechanisms that target the co-regulation of microbial OC conversion and the evolution of antibiotic resistance.