Antibiotic Resistance Genes In Samples Research Articles

Metagenomic analysis of the abundances, diversity, and distribution of antibiotic resistance genes and their potential bacterial hosts in two types of shrimp-rearing farms in South China

Antibiotic resistance genes (ARGs) are attracting increasing concern worldwide. Many previous studies have investigated the occurrence and concentrations of ARGs in aquaculture. However, the sources of ARGs and the links with their potential bacterial hosts have not yet been explored. This study investigated the abundances and diversity of ARGs in two types of shrimp farms in South China through metagenomic sequencing. In total, 14 ARG types were detected. Tetracycline was the dominant ARG type. The abundances of ARGs in samples decreased in the order of duck feces > water source > sediment > shrimp gut > pond water. The samples from the duck–shrimp integrated farm contained 1.29–3.81-fold more abundant ARGs than those from the shrimp monoculture farm (p < 0.05). Several ARGs, that were most predominant in the duck feces samples, were also the most predominant in the shrimp gut samples from the duck–shrimp integrated farm. Redundancy analysis indicated that the abundances and distribution of ARGs formed three clusters: duck feces, water samples, and sediment and shrimp gut samples. The dominant genera in duck feces known as human pathogenic bacteria were potential hosts of ARGs, and were also dominant in the shrimp gut samples in the duck–shrimp integrated farm. Additionally, the abundances of dominant genera in the shrimp gut samples of the duck–shrimp integrated farm were 1.74–35.07-fold higher than those in the shrimp monoculture farm (p < 0.01). The duck–shrimp integrated farm had 3.36-fold and 4.94-fold higher abundances of ARGs and mobile genetic elements in the shrimp gut samples than those from the shrimp monoculture farm, respectively (p < 0.05). The results indicate that duck feces may be a crucial source of diverse and abundant ARGs spreading to reared shrimps in duck–shrimp integrated farms, posing a severe risk to public health.

Metagenomic Analysis of the Long-Term Synergistic Effects of Antibiotics on the Anaerobic Digestion of Cattle Manure

The conversion of cattle manure into biogas in anaerobic digestion (AD) processes has been gaining attention in recent years. However, antibiotic consumption continues to increase worldwide, which is why antimicrobial concentrations can be expected to rise in cattle manure and in digestate. This study examined the long-term synergistic effects of antimicrobials on the anaerobic digestion of cattle manure. The prevalence of antibiotic resistance genes (ARGs) and changes in microbial biodiversity under exposure to the tested drugs was investigated using a metagenomic approach. Methane production was analyzed in lab-scale anaerobic bioreactors. Bacteroidetes, Firmicutes, and Actinobacteria were the most abundant bacteria in the samples. The domain Archaea was represented mainly by methanogenic genera Methanothrix and Methanosarcina and the order Methanomassiliicoccales. Exposure to antibiotics inhibited the growth and development of methanogenic microorganisms in the substrate. Antibiotics also influenced the abundance and prevalence of ARGs in samples. Seventeen types of ARGs were identified and classified. Genes encoding resistance to tetracyclines, macrolide–lincosamide–streptogramin antibiotics, and aminoglycosides, as well as multi-drug resistance genes, were most abundant. Antibiotics affected homoacetogenic bacteria and methanogens, and decreased the production of CH4. However, the antibiotic-induced decrease in CH4 production was minimized in the presence of highly drug-resistant microorganisms in AD bioreactors.

Antibiotic resistance genes in pipe wall biofilm under eight disinfection strategies in domestic hot water system: Occurrence, removal and interactions

For comprehensive insights into the effects of disinfection strategy on antibiotic resistance genes (ARGs) in pipe wall biofilm to ensure the biosafety of domestic hot water system (DHWS), this study investigated the occurrence, removal and interactions of ARGs, as well as their potential hosts. The shifts in biofilm community composition and microbial functions (level three) were observed under different disinfection strategies. A total of 40 ARG subtypes within seven antibiotic types were predicted. The total gene counts ranged from 6.10 × 104 to 2.31 × 105, and decreased after the chlorine, chlorine dioxide, silver ions (Ag+), ultraviolet (UV), and Ag+-UV disinfection. The ARGs in samples under different disinfection strategies were divided into four groups, and the removal and enrichment of the same ARGs in each group were almost consistent. The complex interactions among ARGs were weakened and altered by disinfection (edges decreased from 237 to 178), with yebQ and marC as the hub ARGs, but positive correlations were still dominant. Moreover, 36 nonpathogenic bacteria, including high- (Blastocatella), mid- (Ornatilinea) and low-abundant genera (Bdellovibrio), as well as 18 potential pathogens (Vibrio and Bacillus) were identified as the potential hosts harboring ARGs. Overall, this study provides some new insights into the occurrence of ARGs and their fate under eight disinfection strategies, which is of great significance for a comprehensive understanding and control of ARGs in DHWS.

Antibiotic resistance and antibiotic-resistance genes of Pseudomonas spp. and Escherichia coli isolated from untreated hospital wastewater.

Hospitals are considered an important factor in the spread of antibiotic-resistant bacteria (ARBs) and antibiotic-resistance genes (ARGs). The purpose of this research was to characterize the microbial populations in hospital wastewater and investigated the prevalence of β-lactamase, SulІ and QnrS resistance genes. In the first step, culture method was used to isolate Pseudomonas aeruginosa and Escherichia coli. In the next step, accurate identification of isolated bacteria was carried out using the polymerase chain reaction (PCR) method, then the resistance of the bacteria at different concentrations of antibiotics (8-128 μg/mL) was examined. Finally the ARGs were detected using the PCR method. The averages of heterotrophic plate count (HPC) and ARB concentration in wastewater samples were 1.8 × 108 and 4.3 × 106 CFU/100 mL, respectively. The highest resistance rates were found for sulfamethoxazole and the highest resistance rates in the β-lactamase group were for ceftazidime, while highest sensitivity was for gentamicin and there was no isolate that was sensitive to the studied antibiotics. SulІ and QnrS were the highest and lowest abundance of all ARGs in samples respectively and blaSHV was the highest β-lactam resistance gene. Our results indicated an increase in the resistance of identified bacteria to several antibiotics. So it can be concluded that numerous antibiotic-resistant pathogens and vast numbers of ARGs exist in the human body so that their release from hospitals without effective treatment can cause many dangers to the environment and human health.

Antibiotic resistome profile based on metagenomics in raw surface drinking water source and the influence of environmental factor: A case study in Huaihe River Basin, China

The contamination with antibiotic resistance genes (ARGs) in raw drinking water source may pose a direct threat to human health. In this study, metagenomics sequencing and analysis were applied to investigate the ARG pattern in 12 drinking water sources in upper and middle reach of Huaihe River Basin, China. Based on the redundant analysis and multi-linear regression model, location, specific microbial taxa, number of livestock and health facilities significantly influenced the ARG profile in drinking water sources. Besides the cluster effect of ARG in samples from plain and bedrock mountain areas, the samples from fracture aquifer areas also showed a distinctive biogeographic pattern with that from porous aquifer areas. Putative ARGs host Opitutus and Flavobacterium were the enriched biomarkers in plain and fracture aquifer area respectively, which mainly carried bacitracin, multidrug, beta-lactam and tetracycline ARGs. This result illuminated that both natural background and anthropogenic activities in the watershed influenced the ARG profile in natural freshwater system significantly. The low MGEs abundance and absence of pathogen revealed a low ARG dissemination risk in sampled drinking water sources, while Polynucleobacter was an abundant ARGs host and was significantly related to the ARG profile, which indicated that specific bacteria was responsible for ARGs propagation and accumulation in surface freshwater system. Further researches are needed to assess human exposure to raw drinking water source and the potential risk, as well as the species interaction in microbial community and its impact on ARG propagation under oligotrophic condition.

Selective antibiotic resistance genes in multiphase samples during biofilm growth in a simulated drinking water distribution system: Occurrence, correlation and low-pressure ultraviolet removal

The aim of this study was to gain comprehensive insights into the characteristics of antibiotic resistance genes (ARGs) in multiphase samples from drinking water distribution pipelines using a simulated biofilm reactor. During 120 d of continuous operation, common parameters and six ARGs (ermA, ermB, aphA2, ampC, sulII, and tetO) in samples of three phases (water, particle, and biofilm) from the reactor were investigated, which demonstrated secondary contamination by ARGs. Abundances of the six ARGs in the reactor effluent increased gradually, and in the 120 d effluent, the relative abundances of aphA2 and sulII were the highest, at 9.9 × 10−4 and 1.3 × 10−3, respectively, with a 1.5-fold and 2.8-fold increase, compared with those in the influent. The relative abundances of the six ARGs in the biofilm phase increased significantly (P < 0.05) at 120 d, which was caused by robust bacteria in biofilm that was newly exposed following the detachment of a large piece of aging biofilm. In the particle phase, four of the ARGs did not change significantly during the 120 d period. The six ARGs in the samples of three phases showed a negative correlation with residual chlorine in the pipe water, which demonstrated that low abundance of ARGs in the samples of three phases was related to the improvement of residual chlorine. The proportion of cultivable bacteria illustrated that the robust and active bacteria were negatively correlated with the six ARGs in the biofilm. Total organic carbon (TOC) in the pipeline showed a positive correlation with the proportion of cultivable bacteria in both the water and biofilm phases, which indicated that a TOC reduction in the pipeline contributed to low abundance of ARGs. With low-pressure ultraviolet (LP-UV) irradiation of 20 mJ/cm2, ARGs in the samples of three phases were efficiently controlled, which showed that LP-UV can be used for ARG removal in terminal water for supplemental bactericidal treatment of pipeline effluent.

High-throughput profiling of seasonal variations of antibiotic resistance gene transport in a peri-urban river

The rapid expansion of human activity in a region can exacerbate human health risks induced by antibiotic resistance genes (ARGs). Peri-urban ecosystems serve at the symbiotic interface between urban and rural ecosystems, and investigations into the dissemination of ARGs in peri-urban areas provide a basic framework for tracking the spread of ARGs and potential mitigations. In this study, through the use of high-throughput quantitative PCR and 16S rRNA gene high-throughput sequencing, seasonal and geographical distributions of ARGs and their host bacterial communities were characterized in a peri-urban river. The abundance of ARGs in downstream was 5.2–33.9 times higher than upstream, which indicated distinct antibiotic resistance pollution in the areas where human lives. With the comparison classified based on land use nearby, the abundance of ARGs in samples near farmland and villages was higher than in the background (3.47–5.58 times), pointing to the high load in the river caused by farming and other human activities in the peri-urban areas. With the co-occurrence pattern revealed by network analysis, blaVEB and tetM were proposed to be indicators of ARGs which get together in the same module. Furthermore, seasonal variations in ARGs and the transport of bacterial communities were observed. The effects of seasonal temperature on the dissemination of ARGs along the watershed was also evaluated. The highest absolute abundance of ARGs occurred in summer (2.81 × 109 copies/L on average), the trends of ARG abundances in four seasons were similar with local air temperature. The Linear discriminant analysis effect size (LEfSe) suggested that nine bacterial genera were implicated as biomarkers for the corresponding season. Mobile genetic elements (MGEs) showed significant positive correlation with ARGs (P < 0.01) and MGEs were also identified as the key-contributing factor driving ARG alteration. This study provides an overview of seasonal and geographical variations in ARGs distribution in a peri-urban river and draws attention to controlling pollutants in peri-urban ecosystems.

Detection of Antibiotic Resistance Genes in Source and Drinking Water Samples from a First Nations Community in Canada.

Access to safe drinking water is now recognized as a human right by the United Nations. In developed countries like Canada, access to clean water is generally not a matter of concern. However, one in every five First Nations reserves is under a drinking water advisory, often due to unacceptable microbiological quality. In this study, we analyzed source and potable water from a First Nations community for the presence of coliform bacteria as well as various antibiotic resistance genes. Samples, including those from drinking water sources, were found to be positive for various antibiotic resistance genes, namely, ampC, tet(A), mecA, β-lactamase genes (SHV-type, TEM-type, CTX-M-type, OXA-1, and CMY-2-type), and carbapenemase genes (KPC, IMP, VIM, NDM, GES, and OXA-48 genes). Not surprisingly, substantial numbers of total coliforms, including Escherichia coli, were recovered from these samples, and this result was also confirmed using Illumina sequencing of the 16S rRNA gene. These findings deserve further attention, as the presence of coliforms and antibiotic resistance genes potentially puts the health of the community members at risk. In this study, we highlight the poor microbiological quality of drinking water in a First Nations community in Canada. We examined the coliform load as well as the presence of antibiotic resistance genes in these samples. This study examined the presence of antibiotic-resistant genes in drinking water samples from a First Nations Community in Canada. We believe that our findings are of considerable significance, since the issue of poor water quality in First Nations communities in Canada is often ignored, and our findings will help shed some light on this important issue.

Detection of antibiotic resistance genes in samples from acute and chronic endodontic infections and after treatment

ObjectiveThe purpose of this study was twofold: survey samples from acute and chronic endodontic infections for the presence of genes encoding resistance to beta-lactams, tetracycline and erythromycin, and evaluate the ability of treatment to eliminate these genes from root canals. DesignDNA extracts from samples of abscess aspirates (n=25) and root canals of teeth with asymptomatic apical periodontitis (n=24) were used as template for direct detection of the genes blaTEM, cfxA, tetM, tetQ, tetW, and ermC using real-time polymerase chain reaction (PCR). Bacterial presence was determined using PCR with universal bacterial primers. Root canals of the asymptomatic cases were also sampled and evaluated after chemomechanical procedures using NiTi instruments with 2.5% NaOCl irrigation. ResultsAll abscess and initial root canal samples were positive for bacteria. At least one of the target resistance genes was found in 36% of the abscess samples and 67% of the asymptomatic cases. The most prevalent genes in abscesses were blaTEM (24%) and ermC (24%), while tetM (42%) and tetW (29%) prevailed in asymptomatic cases. The blaTEM gene was significantly associated with acute cases (p=0.02). Conversely, tetM was significantly more prevalent in asymptomatic cases (p=0.008). Treatment eliminated resistance genes from most cases. ConclusionsAcute and chronic endodontic infections harboured resistance genes for 3 classes of widely used antibiotics. In most cases, treatment was effective in eliminating these genes, but there were a few cases in which they persisted. The implications of persistence are unknown. Direct detection of resistance genes in abscesses may be a potential method for rapid diagnosis and establishment of proactive antimicrobial therapy.

Distribution of antibiotic resistance genes in glacier environments

Antibiotic resistance genes are biologically transmitted from microorganism to microorganism in particular micro-environments where dense microbial communities are often exposed to an intensive use of antibiotics, such as intestinal microflora, and the soil microflora of agricultural fields. However, recent studies have detected antibiotic-resistant bacteria and/or antibiotic resistance genes in the natural environment geographically isolated from such areas. Here we sought to examine the prevalence of antibiotic resistance genes in 54 snow and ice samples collected from the Arctic, Antarctic, Central Asia, North and South America and Africa, to evaluate the level of these genes in environments supposedly not affected by anthropogenic factors. We observed a widespread distribution of antibiotic resistance genes in samples from various glaciers in Central Asia, North and South America, Greenland and Africa. In contrast, Antarctic glaciers were virtually free from these genes. Antibiotic resistance genes, of both clinical (i.e. aac(3), blaIMP) and agricultural (i.e. strA and tetW) origin, were detected. Our results show regional geographical distribution of antibiotic resistance genes, with the most plausible modes of transmission through airborne bacteria and migrating birds.

Preparation and application of microarrays for the detection of antibiotic resistance genes in samples isolated from Changchun, China

The emergence of antibiotic-resistant bacteria, especially tetracycline- and beta-lactam-resistant bacteria, poses a great threat to human health. The purpose of this study was to develop and apply a suitable gene microarray for the detection of antibiotic resistance genes. We isolated 463 strains of bacteria from a hospital, a veterinary station, an animal nursery, and living environment of Changchun, China. After screening, it was found that 93.9% of these bacteria were resistant to tetracycline, 74.9% to ampicillin, 55.6% to deoxycycline, and 41.7% to ciprofloxacin. For amplification of antibiotic genes, we designed 28 pairs of primers. In addition, 28 hybridization probes for these genes were developed. The DNA microarray analysis was performed at 42 degrees C for 5 h. We were successful in detecting 12 resistance genes by microarray analysis. After detection, we also evaluated the sensitivity of the microarray analysis. The LDL (Lowest Detection Level) of the microarray was 1 x 10(6) copies/ml of template DNA. It is believed that such microarray-based determination of tetracycline and beta-lactam resistance genes can have a potential application in clinical studies in the future.

Development of a Polymerase Chain Reaction Assay for the Detection of Antibiotic Resistance Genes in Community DNA

Many methods are used to detect antibiotic resistance genes in samples. The objective of the study reported here was to compare polymerase chain reaction (PCR) analysis of community DNA with fecal culturing for detecting antibiotic resistance genes in cattle samples. In the laboratory-based portion of this study, known concentrations of an Escherichia coli strain with 3 antibiotic resistance genes (cmy-2, flo, and cat) were added to feces from dairy cattle. These genes were used to assess the effect of various primer pairs, chromosomally versus plasmid-encoded genes, and gene copy number on the sensitivity of PCR amplification. Gene-specific PCR amplification was performed on the community DNA extracted from the feces. Feces were cultured for the inoculated strain. In the field-based portion of the study, 80 cattle fecal samples of unknown gene status were compared by use of similar methods. Culture and PCR amplification from community DNA extractions produced variable results, and this variability was most noticeable at dilutions that approached the detection limit of the assay. Typically, PCR amplification had a higher sensitivity than did culture for detecting the gene of interest. However, the sensitivity of culture was improved by plating on selective media containing antibiotics. The community DNA approach enables assessment of bacterial communities in complex samples such as feces, a task that can be prohibitive by budget or time constraints associated with culture methods. Through a strategic combination of culture and community DNA approaches, the relationship between specific selection pressures and the persistence and dissemination of specific resistance genes can be elucidated.