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368401.0000Minimal selective concentrations of tetracycline in complex aquatic bacterial biofilms. Selection pressure generated by antibiotics released into the environment could enrich for antibiotic resistance genes and antibiotic resistant bacteria, thereby increasing the risk for transmission to humans and animals. Tetracyclines comprise an antibiotic class of great importance to both human and animal health. Accordingly, residues of tetracycline are commonly detected in aquatic environments. To assess if tetracycline pollution in aquatic environments promotes development of resistance, we determined minimal selective concentrations (MSCs) in biofilms of complex aquatic bacterial communities using both phenotypic and genotypic assays. Tetracycline significantly increased the relative abundance of resistant bacteria at 10 μg/L, while specific tet genes (tetA and tetG) increased significantly at the lowest concentration tested (1 μg/L). Taxonomic composition of the biofilm communities was altered with increasing tetracycline concentrations. Metagenomic analysis revealed a concurrent increase of several tet genes and a range of other genes providing resistance to different classes of antibiotics (e.g. cmlA, floR, sul1, and mphA), indicating potential for co-selection. Consequently, MSCs for the tet genes of ≤ 1 μg/L suggests that current exposure levels in e.g. sewage treatment plants could be sufficient to promote resistance. The methodology used here to assess MSCs could be applied in risk assessment of other antibiotics as well.201626938321
368510.9999Selective concentration for ciprofloxacin resistance in Escherichia coli grown in complex aquatic bacterial biofilms. There is concern that antibiotics in the environment can select for and enrich bacteria carrying acquired antibiotic resistance genes, thus increasing the potential of those genes to emerge in a clinical context. A critical question for understanding and managing such risks is what levels of antibiotics are needed to select for resistance in complex bacterial communities. Here, we address this question by examining the phenotypic and genotypic profiles of aquatic communities exposed to ciprofloxacin, also evaluating the within-species selection of resistant E. coli in complex communities. The taxonomic composition was significantly altered at ciprofloxacin exposure concentrations down to 1 μg/L. Shotgun metagenomic analysis indicated that mobile quinolone resistance determinants (qnrD, qnrS and qnrB) were enriched as a direct consequence of ciprofloxacin exposure from 1 μg/L or higher. Only at 5-10 μg/L resistant E.coli increased relative to their sensitive counterparts. These resistant E. coli predominantly harbored non-transferrable, chromosomal triple mutations (gyrA S83 L, D87N and parC S80I), which confer high-level resistance. In a controlled experimental setup such as this, we interpret effects on taxonomic composition and enrichment of mobile quinolone resistance genes as relevant indicators of risk. Hence, the lowest observed effect concentration for resistance selection in complex communities by ciprofloxacin was 1 μg/L and the corresponding no observed effect concentration 0.1 μg/L. These findings can be used to define and implement discharge or surface water limits to reduce risks for selection of antibiotic resistance in the environment.201829704804
740620.9999Relationship between antibiotic resistance genes and metals in residential soil samples from Western Australia. Increasing drug-resistant infections have drawn research interest towards examining environmental bacteria and the discovery that many factors, including elevated metal conditions, contribute to proliferation of antibiotic resistance (AR). This study examined 90 garden soils from Western Australia to evaluate predictions of antibiotic resistance genes from total metal conditions by comparing the concentrations of 12 metals and 13 genes related to tetracycline, beta-lactam and sulphonamide resistance. Relationships existed between metals and genes, but trends varied. All metals, except Se and Co, were related to at least one AR gene in terms of absolute gene numbers, but only Al, Mn and Pb were associated with a higher percentage of soil bacteria exhibiting resistance, which is a possible indicator of population selection. Correlations improved when multiple factors were considered simultaneously in a multiple linear regression model, suggesting the possibility of additive effects occurring. Soil-metal concentrations must be considered when determining risks of AR in the environment and the proliferation of resistance.201727822686
368130.9999A closer look at the antibiotic-resistant bacterial community found in urban wastewater treatment systems. The conventional biological treatment process can provide a favorable environment for the maintenance and dissemination of antibiotic-resistant bacteria and the antibiotic resistance genes (ARG) they carry. This study investigated the occurrence of antibiotic resistance in three wastewater treatment plants (WWTP) to determine the role they play in the dissemination of ARGs. Bacterial isolates resistant to tetracycline were collected, and tested against eight antibiotics to determine their resistance profiles and the prevalence of multiple antibiotic resistance. It was found that bacteria resistant to tetracycline were more likely to display resistance to multiple antibiotics compared to those isolates that were not tetracycline resistant. Polymerase chain reaction (PCR) was used to identify the tetracycline resistance determinants present within the bacterial communities of the WWTPs and receiving waters, and it was found that ARGs may not be released from the treatment process. Identification of isolates showed that there was a large diversity of species in both the tetracycline-resistant and tetracycline-sensitive populations and that the two groups were significantly different in composition. Antibiotic resistance profiles of each population showed that a large diversity of resistance patterns existed within genera suggesting that transmission of ARG may progress by both horizontal gene and vertical proliferation.201829484827
367640.9999Diversity of antibiotic resistance genes and encoding ribosomal protection proteins gene in livestock waste polluted environment. The rapid development and increase of antibiotic resistance are global phenomena resulting from the extensive use of antibiotics in human clinics and animal feeding operations. Antibiotics can promote the occurrence of antibiotic resistance genes (ARGs), which can be transferred horizontally to humans and animals through water and the food chain. In this study, the presence and abundance of ARGs in livestock waste was monitored by quantitative PCR. A diverse set of bacteria and tetracycline resistance genes encoding ribosomal protection proteins (RPPs) from three livestock farms and a river were analyzed through denaturing gradient gel electrophoresis (DGGE). The abundance of sul(I) was 10(3) to 10(5) orders of magnitude higher than that of sul(II). Among 11 tet-ARGs, the most abundant was tet(O). The results regarding bacterial diversity indicated that the presence of antibiotics might have an evident impact on bacterial diversity at every site, particularly at the investigated swine producer. The effect of livestock waste on the bacterial diversity of soil was stronger than that of water. Furthermore, a sequencing analysis showed that tet(M) exhibited two genotypes, while the other RPPs-encoding genes exhibited at least three genotypes. This study showed that various ARGs and RPPs-encoding genes are particularly widespread among livestock.201829469609
368350.9999Small and large-scale distribution of four classes of antibiotics in sediment: association with metals and antibiotic resistance genes. Antibiotic chemicals and antibiotic resistance genes enter the environment via wastewater effluents as well as from runoff from agricultural operations. The relative importance of these two sources, however, is largely unknown. The relationship between the concentrations of chemicals and genes requires exploration, for antibiotics in the environment may lead to development or retention of resistance genes by bacteria. The genes that confer resistance to metal toxicity may also be important in antibiotic resistance. In this work, concentrations of 19 antibiotics (using liquid chromatography tandem mass spectrometry), 14 metals (using inductively coupled plasma-mass spectrometry), and 45 metal, antibiotic, and antibiotic-resistance associated genes (using a multiplex, microfluidic quantitative polymerase chain reaction method) were measured in 13 sediment samples from two large rivers as well as along a spatial transect in a wastewater effluent-impacted lake. Nine of the antibiotics were detected in the rivers and 13 were detected in the lake. Sixteen different resistance genes were detected. The surrounding land use and proximity to wastewater treatment plants are important factors in the number and concentrations of antibiotics detected. Correlations among antibiotic chemical concentrations, metal concentrations, and resistance genes occur over short spatial scales in a lake but not over longer distances in major rivers. The observed correlations likely result from the chemicals and resistance genes arising from the same source, and differences in fate and transport over larger scales lead to loss of this relationship.201830043816
343360.9999Effect of subinhibitory concentrations on the spreading of the ampicillin resistance gene bla(CMY-2) in an activated sludge microcosm. As the problem of multi-resistant bacteria grows a better understanding of the spread of antibiotic resistance genes is of utmost importance for society. Wastewater treatment plants contain subinhibitory concentrations of antibiotics and are thought to be hotspots for antibiotic resistance gene propagation. Here we evaluate the influence of sub-minimum inhibitory concentrations of antibiotics on the spread of resistance genes within the bacterial community in activated sludge laboratory-scale sequencing batch reactors. The mixed communities were fed two different ampicillin concentrations (500 and 5000 µg/L) and the reactors were run and monitored for 30 days. During the experiment the β-lactamase resistance gene bla(CMY-2) was monitored via qPCR and DNA samples were taken to monitor the effect of ampicillin on the microbial community. The relative copy number of bla(CMY-2) in the reactor fed with the sub-minimum inhibitory concentration of 500 µg/L ampicillin was spread out over a wider range of values than the control and 5000 µg/L ampicillin reactors indicating more variability of gene number in the 500 µg/L reactor. This result emphasises the problem of sub-minimum inhibitory concentrations of antibiotics in wastewater. High-throughput sequencing showed that continuous exposure to ampicillin caused a shift from a Bacteroidetes to Proteobacteria in the bacterial community. The combined use of qPCR and high-throughput sequencing showed that ampicillin stimulates the spread of resistance genes and leads to the propagation of microbial populations which are resistant to it.202539215485
368070.9999Metagenomic Insights Into the Contribution of Phages to Antibiotic Resistance in Water Samples Related to Swine Feedlot Wastewater Treatment. In this study, we examined the types of antibiotic resistance genes (ARGs) possessed by bacteria and bacteriophages in swine feedlot wastewater before and after treatment using a metagenomics approach. We found that the relative abundance of ARGs in bacterial DNA in all water samples was significantly higher than that in phages DNA (>10.6-fold), and wastewater treatment did not significantly change the relative abundance of bacterial- or phage-associated ARGs. We further detected the distribution and diversity of the different types of ARGs according to the class of antibiotics to which they confer resistance, the tetracycline resistance genes were the most abundant resistance genes and phages were more likely to harbor ATP-binding cassette transporter family and ribosomal protection genes. Moreover, the colistin resistance gene mcr-1 was also detected in the phage population. When assessing the contribution of phages in spreading different groups of ARGs, β-lactamase resistance genes had a relatively high spreading ability even though the abundance was low. These findings possibly indicated that phages not only could serve as important reservoir of ARG but also carry particular ARGs in swine feedlot wastewater, and this phenomenon is independent of the environment.201830459724
710180.9999Tetracycline residues and tetracycline resistance genes in groundwater impacted by swine production facilities. Antibiotics are used at therapeutic levels to treat disease; at slightly lower levels as prophylactics; and at low, subtherapeutic levels for growth promotion and improvement of feed efficiency. Over 88% of swine producers in the United States gave antimicrobials to grower/finisher pigs in feed as a growth promoter in 2000. It is estimated that ca. 75% of antibiotics are not absorbed by animals and are excreted in urine and feces. The extensive use of antibiotics in swine production has resulted in antibiotic resistance in many intestinal bacteria, which are also excreted in swine feces, resulting in dissemination of resistance genes into the environment. To assess the impact of manure management on groundwater quality, groundwater samples have been collected near two swine confinement facilities that use lagoons for manure storage and treatment. Several key contaminant indicators - including inorganic ions, antibiotics, and antibiotic resistance genes - were analyzed in groundwater collected from the monitoring wells. Chloride, ammonium, potassium, and sodium were predominant inorganic constituents in the manure samples and served as indicators of groundwater contamination. Based on these analyses, shallow groundwater has been impacted by lagoon seepage at both sites. Liquid chromatography-mass spectroscopy (LC-MS) was used to measure the dissolved concentrations of tetracycline, chlortetracycline, and oxytetracycline in groundwater and manure. Although tetracyclines were regularly used at both facilities, they were infrequently detected in manure samples and then at relatively trace concentrations. Concentrations of all tetracyclines and their breakdown products in the groundwater sampled were generally less than 0.5 microg/L. Bacterial tetracycline resistance genes served as distinct genotypic markers to indicate the dissemination and mobility of antibiotic resistance genes that originated from the lagoons. Applying PCR to genomic DNA extracted from the lagoon and groundwater samples, four commonly occurring tetracycline (tet) resistance genes - tet(M), tet(O), tet(Q), and tet(W) - were detected. The detection frequency of tet genes was much higher in wells located closer to and down-gradient from the lagoons than in wells more distant from the lagoons. These results suggested that in the groundwater underlying both facilities tetracycline resistance genes exist and are somewhat persistent, but that the distribution and potentially the flux for each tet gene varied throughout the study period.200617127527
741090.9999The effects of subtherapeutic antibiotic use in farm animals on the proliferation and persistence of antibiotic resistance among soil bacteria. The use of antibiotics at subtherapeutic concentrations for agricultural applications is believed to be an important factor in the proliferation of antibiotic-resistant bacteria. The goal of this study was to determine if the application of manure onto agricultural land would result in the proliferation of antibiotic resistance among soil bacteria. Chlortetracycline-resistant bacteria were enumerated and characterized from soils exposed to the manure of animals fed subtherapeutic concentrations of antibiotics and compared to the chlortetracycline-resistant bacteria from soils at farms with restricted antibiotic use (dairy farms) and from non-agricultural soils. No significant differences were observed at nine different study sites with respect to the numbers and types of cultivated chlortetracycline-resistant bacteria. Genes encoding for tetracycline resistance were rarely detected in the resistant bacteria from these sites. In contrast, soils collected from a tenth farm, which allowed manure to indiscriminately accumulate outside the animal pen, had significantly higher chlortetracycline-resistance levels. These resistant bacteria frequently harbored one of 14 different genes encoding for tetracycline resistance, many of which (especially tet(A) and tet(L)) were detected in numerous different bacterial species. Subsequent bacterial enumerations at this site, following the cessation of farming activity, suggested that this farm remained a hotspot for antibiotic resistance. In conclusion, we speculate that excessive application of animal manure leads to the spread of resistance to soil bacteria (potentially by lateral gene transfer), which then serve as persistent reservoir of antibiotic resistance.200718043630
7395100.9999Impacts of anthropogenic activity on the ecology of class 1 integrons and integron-associated genes in the environment. The impact of human activity on the selection for antibiotic resistance in the environment is largely unknown, although considerable amounts of antibiotics are introduced through domestic wastewater and farm animal waste. Selection for resistance may occur by exposure to antibiotic residues or by co-selection for mobile genetic elements (MGEs) which carry genes of varying activity. Class 1 integrons are genetic elements that carry antibiotic and quaternary ammonium compound (QAC) resistance genes that confer resistance to detergents and biocides. This study aimed to investigate the prevalence and diversity of class 1 integron and integron-associated QAC resistance genes in bacteria associated with industrial waste, sewage sludge and pig slurry. We show that prevalence of class 1 integrons is higher in bacteria exposed to detergents and/or antibiotic residues, specifically in sewage sludge and pig slurry compared with agricultural soils to which these waste products are amended. We also show that QAC resistance genes are more prevalent in the presence of detergents. Studies of class 1 integron prevalence in sewage sludge amended soil showed measurable differences compared with controls. Insertion sequence elements were discovered in integrons from QAC contaminated sediment, acting as powerful promoters likely to upregulate cassette gene expression. On the basis of this data, >1 × 10(19) bacteria carrying class 1 integrons enter the United Kingdom environment by disposal of sewage sludge each year.201121368907
7407110.9999Impact of salmon farming in the antibiotic resistance and structure of marine bacterial communities from surface seawater of a northern Patagonian area of Chile. BACKGROUND: Aquaculture and salmon farming can cause environmental problems due to the pollution of the surrounding waters with nutrients, solid wastes and chemicals, such as antibiotics, which are used for disease control in the aquaculture facilities. Increasing antibiotic resistance in human-impacted environments, such as coastal waters with aquaculture activity, is linked to the widespread use of antibiotics, even at sub-lethal concentrations. In Chile, the world's second largest producer of salmon, aquaculture is considered the primary source of antibiotics residues in the coastal waters of northern Patagonia. Here, we evaluated whether the structure and diversity of marine bacterial community, the richness of antibiotic resistance bacteria and the frequency of antibiotic resistance genes increase in communities from the surface seawater of an area with salmon farming activities, in comparison with communities from an area without major anthropogenic disturbance. RESULTS: The taxonomic structure of bacterial community was significantly different between areas with and without aquaculture production. Growth of the culturable fraction under controlled laboratory conditions showed that, in comparison with the undisturbed area, the bacterial community from salmon farms displayed a higher frequency of colonies resistant to the antibiotics used by the salmon industry. A higher adaptation to antibiotics was revealed by a greater proportion of multi-resistant bacteria isolated from the surface seawater of the salmon farming area. Furthermore, metagenomics data revealed a significant higher abundance of antibiotic resistant genes conferring resistance to 11 antibiotic families in the community from salmon farms, indicating that the proportion of bacteria carrying the resistance determinants was overall higher in salmon farms than in the undisturbed site. CONCLUSIONS: Our results revealed an association between bacterial communities and antibiotic resistance from surface seawater of a coastal area of Chile. Although the total bacterial community may appear comparable between sites, the cultivation technique allowed to expose a higher prevalence of antibiotic resistant bacteria in the salmon farming area. Moreover, we demonstrated that metagenomics (culture-independent) and phenotypic (culture-dependent) methods are complementary to evaluate the bacterial communities' risk for antibiotic resistance, and that a human-influenced environment (such as salmon farms) can potentiate bacteria to adapt to environmental stresses, such as antibiotics.202439523335
3678120.9999Abundance and dynamics of antibiotic resistance genes and integrons in lake sediment microcosms. Antibiotic resistance in bacteria causing disease is an ever growing threat to the world. Recently, environmental bacteria have become established as important both as sources of antibiotic resistance genes and in disseminating resistance genes. Low levels of antibiotics and other pharmaceuticals are regularly released into water environments via wastewater, and the concern is that such environmental contamination may serve to create hotspots for antibiotic resistance gene selection and dissemination. In this study, microcosms were created from water and sediments gathered from a lake in Sweden only lightly affected by human activities. The microcosms were exposed to a mixture of antibiotics of varying environmentally relevant concentrations (i.e., concentrations commonly encountered in wastewaters) in order to investigate the effect of low levels of antibiotics on antibiotic resistance gene abundances and dynamics in a previously uncontaminated environment. Antibiotic concentrations were measured using liquid chromatography-tandem mass spectrometry. Abundances of seven antibiotic resistance genes and the class 1 integron integrase gene, intI1, were quantified using real-time PCR. Resistance genes sulI and ermB were quantified in the microcosm sediments with mean abundances 5 and 15 gene copies/10(6) 16S rRNA gene copies, respectively. Class 1 integrons were determined in the sediments with a mean concentration of 3.8 × 10(4) copies/106 16S rRNA gene copies. The antibiotic treatment had no observable effect on antibiotic resistance gene or integron abundances.201425247418
3677130.9999Prevalence of sulfonamide-resistant bacteria, resistance genes and integron-associated horizontal gene transfer in natural water bodies and soils adjacent to a swine feedlot in northern Taiwan. Antibiotics are commonly used in swine feed to treat and prevent disease, as well as to promote growth. Antibiotics released into the environment via wastewater could accelerate the emergence of antibiotic-resistant bacteria and resistance genes in the surrounding environment. In this study, we quantified the occurrence of sulfonamides, sulfonamide-resistant microorganisms and resistance genes in the wastewater from a swine farm in northern Taiwan and its surrounding natural water bodies and soils. Sulfonamide levels were similar in the receiving downstream and upstream river water. However, the prevalence of sulfonamide-resistant bacteria and resistance genes, as analyzed by cultivation-dependent and -independent molecular approaches, was significantly greater in the downstream compared to the upstream river water samples. Barcoded-pyrosequencing revealed a highly diverse bacterial community structure in each sample. However, the sequence identity of the sulfonamide resistance gene sul1 in the wastewater and downstream environment samples was nearly identical (99-100%). The sul1 gene, which is genetically linked to class 1 integrons, was dominant in the downstream water bodies and soils. In conclusion, the increased prevalence of sulfonamide resistance genes in the wastewater from a swine farm, independent of the persistent presence of sulfonamides, could be a potential source of resistant gene pools in the surrounding environment.201424637153
7106140.9999The swine waste resistome: Spreading and transfer of antibiotic resistance genes in Escherichia coli strains and the associated microbial communities. The overuse of antimicrobials in livestock farming has led to the development of resistant bacteria and the spread of antibiotic-resistant genes (ARGs) among animals. When manure containing these antibiotics is applied to agricultural fields, it creates a selective pressure that promotes the acquisition of ARGs by bacteria, primarily through horizontal gene transfer. Most research on ARGs focuses on their role in clinical antibiotic resistance and their transfer from environmental sources to bacteria associated with humans, such as Escherichia coli. The study investigates the spread of antibiotic-resistant genes (ARGs) through class 1 integrons in 27 Escherichia coli strains from pig manure. It focuses on six common ARGs (ermB, cmlA, floR, qnrS, tetA, and TEM) and the class 1 integron gene, assessing their prevalence in manure samples from three pig farms. The study found correlations and anticorrelations among these genes, indicating a predisposition of the integron in spreading certain ARGs. Specifically, cmlA and tetA genes were positively correlated with each other and negatively with int1, suggesting they are not transferred via Int1. Farm B had the highest int1 counts and a higher abundance of the TEM gene, but lower levels of cmlA and tetA genes. The results underscore the complexity of predicting ARG spread in agricultural environments and the associated health risks to humans through the food chain. The study's results offer valuable insights into the antibiotic-resistant genes (ARGs) profile in swine livestock, potentially aiding in the development of methods to trace ARGs in the environment.202439053184
3471150.9999The prevalence of ampicillin-resistant opportunistic pathogenic bacteria undergoing selective stress of heavy metal pollutants in the Xiangjiang River, China. The emergence of clinically relevant β-lactam-resistant bacteria poses a serious threat to human health and presents a major challenge for medical treatment. How opportunistic pathogenic bacteria acquire antibiotic resistance and the prevalence of antibiotic-resistant opportunistic pathogenic bacteria in the environment are still unclear. In this study, we further confirmed that the selective pressure of heavy metals contributes to the increase in ampicillin-resistant opportunistic pathogens in the Xiangjiang River. Four ampicillin-resistant opportunistic pathogenic bacteria (Pseudomonas monteilii, Aeromonas hydrophila, Acinetobacter baumannii, and Staphylococcus epidermidis) were isolated on Luria-Bertani (LB) agar plates and identified by 16S rRNA sequencing. The abundance of these opportunistic pathogenic bacteria significantly increased in the sites downstream of the Xiangjiang River that were heavily influenced by metal mining activities. A microcosm experiment showed that the abundance of β-lactam resistance genes carried by opportunistic pathogenic bacteria in the heavy metal (Cu(2+) and Zn(2+)) treatment group was 2-10 times higher than that in the control. Moreover, heavy metals (Cu(2+) and Zn(2+)) significantly increased the horizontal transfer of plasmids in pathogenic bacteria. Of particular interest is that heavy metals facilitated the horizontal transfer of conjugative plasmids, which may lead to the prevalence of multidrug-resistant pathogenic bacteria in the Xiangjiang River.202133035873
7115160.9999Antibiotics, antibiotic resistance and associated risk in natural springs from an agroecosystem environment. This study investigates the occurrence, transport, and risks associated to antibiotic residues, antibiotic resistance genes (ARGs) and antibiotic resistant Escherichia coli (AR-E. coli) in eleven natural springs in an agroecosystem environment with intense livestock production, where groundwater nitrate concentration usually sets above 50 mg L(-1). Out of 23 multiple-class antibiotics monitored, tetracycline and sulfonamide residues were the most ubiquitous, and they were detected at concentrations ranging from ng L(-1) to μg L(-1). Five ARGs were monitored, conferring resistance to the antibiotic classes of major use in livestock production. Thus, genes conferring resistance to sulfonamides (sul1 and sul2) and tetracyclines (tetW) as well as a gene proxy for anthropogenic pollution (intI1) were present in most springs. sul1 was the most abundant, with absolute concentrations ranging from 4 × 10(2) to 5.6 × 10(6) gene copies L(-1) water. AR-E. coli showing resistance to sulfonamides and tetracyclines was also detected, with a prevalence up to approximately 40 % in some sites but with poor correlations with the concentration of antibiotic residues and ARGs. The occurrence of antibiotics, ARGs and AR-E. coli was characterized by large seasonal variations which were mostly associated to both hydrological factors and reactive transport processes. Finally, a risk assessment approach pointed out towards low risk for both the groundwater environment and human health, when spring water is used for direct human consumption, associated with the occurrence of antibiotics, ARGs and AR-E. coli. However, long-term effects cannot be neglected, and proper actions must be taken to preserve groundwater quality.202336208750
5293170.9999Tetracycline-Resistant Bacteria Selected from Water and Zebrafish after Antibiotic Exposure. The emergence of antibiotic-resistant pathogens due to worldwide antibiotic use is raising concern in several settings, including aquaculture. In this work, the selection of antibiotic-resistant bacteria (ARB) and antibiotic resistance genes (ARGs) was evaluated after exposure of zebrafish to oxytetracycline (OTC) for two months, followed by a recovery period. The selection of ARB in water and fish was determined using selective media. The abundance of tetA genes was estimated through qPCR. Higher prevalence of ARB was measured in all samples exposed to the antibiotic when compared to control samples, although statistical significance was only achieved five days after exposure. Isolates recovered from samples exposed to the antibiotic were affiliated with Pseudomonas and Stenotrophomonas. Various antibiotic susceptibility profiles were detected and 37% of the isolates displayed multidrug resistance (MDR). The selection of the tetA gene was confirmed by qPCR at the highest OTC concentration tested. Two MDR isolates, tested using zebrafish embryos, caused significant mortality, indicating a potential impact on fish health and survival. Overall, our work highlights the potential impact of antibiotic contamination in the selection of potential pathogenic ARB and ARGS.202133804606
7414180.9999Structure of the manure resistome and the associated mobilome for assessing the risk of antimicrobial resistance transmission to crops. In this study, the impact of bovine and poultry manure on the quantitative and qualitative composition of antibiotic resistance genes (ARGs) and the environmental mobilome associated with antimicrobial resistance in soil and crops was determined with the use of next generation sequencing methods. The aim of the study was to perform a metagenomic analysis of manure to estimate the risk of the transmission of ARGs and bacterial drug resistance carriers to fertilized soil and crops. The total copy number of ARGs was nearly four times higher in poultry manure (555 ppm) than in bovine manure (140 ppm), and this relationship was also noted in fertilized soil. Poultry manure induced a much greater increase in the concentrations of ARGs in the soil environment (196.4 ppm) than bovine manure (137.8 ppm) immediately after supplementation. The application of poultry manure led to the highest increase in the abundance of genes encoding resistance to tetracyclines (9%), aminoglycosides (3.5%), sulfonamides (3%), bacitracin (2%), chloramphenicol (2%), and macrolide-lincosamide-streptogramin antibiotics (1%). Heavy metals were stronger promoters of antibiotic resistance in the environment than antibiotics. Antibiotics exerted a greater influence on maintaining the diversity of ARGs than on increasing their abundance in soil. Large quantities of insertion sequences (IS), including those associated with the mobility of ARGs in the population of ESKAPEE pathogens, are introduced to soil with manure. These IS remain stable for up to several months, which indicates that manure, in particular poultry manure, significantly increases the risk of rapid ARG transfer to the environment. Manure also largely contributes to an increase in the diversity of the resistome and mobilome in the metagenome of bacteria isolated from crops. Bacteria of the phylum Proteobacteria appear to play a major role in the transmission of multiple ARGs in crops grown for human and animal consumption.202234864022
3844190.9999Effects of Nutrient Level and Growth Rate on the Conjugation Process That Transfers Mobile Antibiotic Resistance Genes in Continuous Cultures. Bacteria in the effluent of wastewater treatment plants (WWTPs) can transfer antibiotic resistance genes (ARGs) to the bacteria in receiving water through conjugation; however, there is a lack of quantitative assessment of this phenomenon in continuous cultures. Our objective was to determine the effects of background nutrient levels in river water column and growth rates of bacteria on the conjugation frequency of ARGs from effluent bacteria to river bacteria, as well as on the resulting resistance level (i.e., MICs) of the river bacteria. Chemostats were employed to simulate the discharge points of WWTPs into rivers, where effluent bacteria (donor cells) meet river bacteria (recipient cells). Both donor and recipient cells were Escherichia coli cells, and the donor cells were constructed by filter mating with bacteria in the effluent of a local WWTP. Results showed that higher bacterial growth rate (0.45 h(-1) versus 0.15 h(-1)) led to higher conjugation frequencies (10(-4) versus 10(-6) transconjugant per recipient). The nutrient level also significantly affected the conjugation frequency, albeit to a lesser extent than the growth rate. The MIC against tetracycline increased from 2 mg/L in the recipient to 64 to 128 mg/L in transconjugants. In comparison, the MIC only increased to as high as 8 mg/L in mutants. Whole-genome sequencing showed that the tet-containing plasmid in both the donor and the transconjugant cells also occur in other fecal bacterial genera. The quantitative information obtained from this study can inform hazard identification related to the proliferation of wastewater-associated ARGs in surface water. IMPORTANCE WWTPs have been regarded as an important hot spot of ARGs. The discharge point of WWTP effluent, where ARGs may be horizontally transferred from bacteria of treated wastewater to bacteria of receiving water, is an important interface between the human-dominated ecosystem and the natural environment. The use of batch cultures in previous studies cannot adequately simulate the nutrient conditions and growth rates in receiving water. In this study, chemostats were employed to simulate the continuous growth of bacteria in receiving water. Furthermore, the experimental setup allowed for separate investigations on the effects of nutrient levels (i.e., simulating background nutrients in river water) and bacterial growth rates on conjugation frequencies and resulting resistance levels. The study generates statistically sound ecological data that can be used to estimate the risk of wastewater-originated ARGs as part of the One Health framework.202236094214