Key Antibiotic Resistance Genes Research Articles

Hydraulic conditions control the abundance of antibiotic resistance genes and their potential host microorganisms in a frequently regulated river–lake system

Antibiotic resistance genes (ARGs) are a growing problem that is widespread in river–lake ecosystems, where they pose a threat to the aquatic environment's health and public safety. These systems serve as critical nodes in water management, as they facilitate the equitable allocation of water resources through long-term and frequent water diversions. However, hydrological disturbances associated with water-regulation practices can influence the dynamics of their potential host microorganisms and associated resistance genes. Consequently, identifying the key ARGs and their resistance mechanisms in heavily regulated waters is vital for safeguarding human health and that of river–lake ecosystems. In this study, we examined the impact of water-regulation factors on ARGs and their hosts within a river–lake continuum using 16S rRNA and metagenomic sequencing. We found that a significant increase in ARG abundance during regulation periods (p < 0.05), especially in the aquatic environment. Key resistance genes were macB, tetA, evgS, novA, and msbA, with increased efflux pinpointed as their principal resistance mechanism. Network analysis identified Flavobacteriales, Acinetobacter, Pseudomonas, Burkholderiaceae, and Erythrobacter as key potential host microorganisms, which showed increased abundance within the water column during regulation periods (p < 0.05). Flow velocity and water depth both drove the host microorganisms and critical ARGs. Our findings underscore the importance of monitoring and mitigating the antibiotic resistance risk during water transfers in river–lake systems, thereby supporting informed management and conservation strategies.

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.

Wastewater surveillance for antibiotics and resistance genes in a river catchment: Spatiotemporal variations and the main drivers

Wastewater-based epidemiology (WBE) is used for mining information about public health such as antibiotics resistance. This study investigated the distribution profiles of six types of antibiotic resistance genes (ARGs) in wastewater and rivers in Wuhu City, China. The levels of ARGs found in the Qingyijiang River were significantly higher than other rivers, and were comparable to effluent levels. Among the ARGs, sulfonamides ARGs and intI1 were the predominant in both wastewaters and rivers. Additionally, the concentrations of ARGs were higher on weekends as opposed to weekdays. Their distribution patterns remained consistent inter-week and inter-season using linear regression analysis (p < 0.001). Interestingly, the occurrence levels of ARGs in wastewaters during spring were significantly higher than in autumn, although insignificant in rivers. The apparent removal rate of ARGs in domestic wastewater sources ranged from 61.52–99.29%, except for qepA (-1.91% to 81.09%), whereas the removal rates in mixed domestic and industrial wastewaters showed a marked decrease (-92.94% to 76.67%). A correlation network analysis revealed that azithromycin and erythromycin were key antibiotics, while blaNDM-1, tetM, tetB, and ermB were identified as key ARGs. Sulfonamide and fluoroquinolone antibiotics, and tetracycline and macrolide ARGs were the primary contributors. Linear mixed models demonstrated that socio-economic variables positively impacted the occurrence levels of ARGs, whereas wastewater flow and river runoff were the negative drivers for their concentrations in wastewaters and surface waters, respectively. Overall, this WBE study contributes to the understanding of spatiotemporal profiles and main drivers of the occurrence of ARGs in wastewater and receiving water.

Loratadine Combats Methicillin-Resistant Staphylococcus aureus by Modulating Virulence, Antibiotic Resistance, and Biofilm Genes.

Methicillin-resistant Staphylococcus aureus (MRSA) has evolved to become resistant to multiple classes of antibiotics. New antibiotics are costly to develop and deploy, and they have a limited effective lifespan. Antibiotic adjuvants are molecules that potentiate existing antibiotics through nontoxic mechanisms. We previously reported that loratadine, the active ingredient in Claritin, potentiates multiple cell-wall active antibiotics in vitro and disrupts biofilm formation through a hypothesized inhibition of the master regulatory kinase Stk1. Loratadine and oxacillin combined repressed the expression of key antibiotic resistance genes in the bla and mec operons. We hypothesized that additional genes involved in antibiotic resistance, biofilm formation, and other cellular pathways would be modulated when looking transcriptome-wide. To test this, we used RNA-seq to quantify transcript levels and found significant effects in gene expression, including genes controlling virulence, antibiotic resistance, metabolism, transcription (core RNA polymerase subunits and sigma factors), and translation (a plethora of genes encoding ribosomal proteins and elongation factor Tu). We further demonstrated the impacts of these transcriptional effects by investigating loratadine treatment on intracellular ATP levels, persister formation, and biofilm formation and morphology. Loratadine minimized biofilm formation in vitro and enhanced the survival of infected Caenorhabditis elegans. These pleiotropic effects and their demonstrated outcomes on MRSA virulence and survival phenotypes position loratadine as an attractive anti-infective against MRSA.

Multiple Antibiotic-Resistant Bacteria Resistant to Electrochemical Disinfection with Variation of Key Antibiotic Resistance Genes

Electrochemical disinfection (ED) is effective in removal of antibiotic-resistant bacteria (ARB) and antibiotic resistance genes (ARGs) during wastewater treatment. However, the inner relationship of antibiotic-resistant phenotypes with their genotypes of ARB is still unclear in the ED process. This study explored the disinfection mechanism at the microcosmic level using four Escherichia coli (E. coli) strains with different antibiotic-resistant phenotypes. Results showed that bacteria with multiple antibiotic resistance tended to have stronger viability in disinfection compared with those resistant to fewer antibiotics. Interestingly, antibiotic-resistant phenotypes changed little in the disinfection process, while genotypes closely responded to bacterial resistance of different E. coli. In detail, beta-lactamase resistance genes played a leading role in the cross-resistance between antibiotics and electrochemical oxidation. They were proved to reduce oxidative destruction by increasing the strength of the cell wall and membrane. In addition, diaminopyrimidine and macrolide-lincosamide-streptogramin B resistance genes are also closely related to the bacterial resistance to ED. Combined with the quantitative determination of ARGs, the abundance of IntI1 and aminoglycoside resistance genes can be regarded as indicators for the risk of cross-resistance to antibiotics and electrochemical oxidation. This study provided important references for in depth understanding and accurate control of antibiotic resistance with advanced oxidation processes.

Antibiotic resistance of Riemerella anatipestifer and comparative analysis of antibiotic-resistance gene detection methods

Riemerella anatipestifer is an important pathogen in waterfowl, and is generally multidrug resistant. This study assessed the current status of Riemerella anatipestifer antibiotic resistance and antibiotic-resistance genes (ARGs), compared the results of different detection methods, and evaluated a new method of studying the association between antibiotic resistance and ARGs in Riemerella anatipestifer. In this study, 51 strains of Riemerella anatipestifer were isolated from ducks on several farms, their resistance to 28 antibiotics was assessed, and the isolates were subjected to whole-genome sequencing. The number of ARGs carried by Riemerella anatipestifer was predicted, compared, and analyzed, and the consistency between ARGs and antibiotic-resistance phenotypes was assessed. The potential for loss of resistance genes during the sequencing and assembly of genome-wide framework map was assessed, and a new ARG detection method was pilot tested. The 51 strains of Riemerella anatipestifer were multidrug resistant (MDR) and had high level of resistance to aminoglycosides, trimethoprim, lincosamides, polypeptides, and macrolides. Based on the genome-wide framework map of the 51 strains, 3 local databases of ABRicate software and 1 online database of CARD website were used to detect ARGs, and a mean of 4 to 5 ARGs were identified per isolate. Although the detection results differed according to the database used, the general performance was consistent. The online website detected more types of ARGs than the ABRicate software. The association between ARGs and antibiotic-resistance phenotypes was assessed, and the ermF gene was identified as a possible key ARGs regulating macrolide resistance of Riemerella anatipestifer. The method used to investigate and detect Riemerella anatipestifer ARGs was convenient and rapid, and had strong accuracy and pertinence. The ARGs detection method reported here combined the advantages of PCR and genome detection, and could greatly reduce workload and detect ARGs more precisely.

Captivity and geography influence the antibiotic resistome of non-human primates.

Antibiotic resistance poses a serious threat for animals and humans health worldwide. Yet a comprehensive exploration of the influence of captivity and geography on non-human primate (NPH) gut antibiotic resistance remains incomplete. In this study, 131 metagenomic sequencing datasets of five species of NHPs included different regions and lifestyles were selected to perform the antibiotic resistance analysis. Nineteen related resistance antibiotics and 325 antibiotic resistance genes (ARGs) were obtained. A significantly higher abundance and diversity index of ARGs in the captive NHPs than in the wild was found but not for all of the samples. The biomarker-tracking of ARGs analysis identified key ARGs related to aminoglycoside resistance genes and tetracycline resistance genes. These results suggest that captivity and geography changes associated with human activities can lead to marked changes in the ecology of the NHP gut flora ARGs.

Antimicrobial Activity of the Iron-Chelator, DIBI, against Multidrug-Resistant Canine Methicillin-Susceptible Staphylococcus pseudintermedius: A Preliminary Study of Four Clinical Strains.

Staphylococcus pseudintermedius is an important opportunistic pathogen causing various infections in dogs. Furthermore, it is an emerging zoonotic agent and both multidrug-resistant methicillin-resistant S. pseudintermedius (MRSP) as well as methicillin-susceptible (MSSP) strains represent an important therapeutic challenge to veterinary medicine and pose a potential threat to human health. We tested representative S. pseudintermedius clinical strains from dogs suffering from otitis externa for their susceptibilities to a panel of 17 antimicrobials compared to DIBI. DIBI, unlike antibiotics, is a novel water-soluble hydroxypyridinone-containing iron-chelating agent that deprives microbes of growth-essential iron and has been previously shown to inhibit methicillin-resistant Staphylococcus aureus (MRSA). We also characterised the strains according to whether they harbour key antibiotic resistance genes. The strains each displayed multiple antimicrobial resistance patterns; all were negative for the mecA gene and possessed the tetK and tetM genes, but they varied as to their possession of the ermB gene. However, all the isolates had similar susceptibility to DIBI with low MICs (2 µg/mL or 0.2 µM). Because the four MSSPs were equally susceptible to DIBI, subject to confirmation with additional strains, this could provide a potential non-antibiotic, anti-infective alternative approach for the treatment of antimicrobial-resistant canine S. pseudintermedius otitis.

Multiplexed detection of bacterial nucleic acids using Cas13 in droplet microarrays

Rapid and accurate diagnosis of infections is fundamental to individual patient care and public health management. Nucleic acid detection methods are critical to this effort, but are limited either in the breadth of pathogens targeted or by the expertise and infrastructure required. We present here a high-throughput system that enables rapid identification of bacterial pathogens, bCARMEN, which utilizes: (1) modular CRISPR-Cas13-based nucleic acid detection with enhanced sensitivity and specificity; and (2) a droplet microfluidic system that enables thousands of simultaneous, spatially multiplexed detection reactions at nanoliter volumes; and (3) a novel preamplification strategy that further enhances sensitivity and specificity. We demonstrate bCARMEN is capable of detecting and discriminating 52 clinically relevant bacterial species and several key antibiotic resistance genes. We further develop a simple proof of principle workflow using stabilized reagents and cell phone camera optical readout, opening up the possibility of a rapid point-of-care multiplexed bacterial pathogen identification and antibiotic susceptibility testing.

Abundance and diversity of antibiotic resistance genes and bacterial communities in the western Pacific and Southern Oceans

This study investigated the abundance and diversity of antibiotic resistance genes (ARGs) and the composition of bacterial communities along a transect covering the western Pacific Ocean (36°N) to the Southern Ocean (74°S) using the Korean icebreaker R/V Araon (total cruise distance: 14,942 km). The relative abundances of ARGs and bacteria were assessed with quantitative PCR and next generation sequencing, respectively. The absolute abundance of ARGs was 3.0 × 106 ± 1.6 × 106 copies/mL in the western Pacific Ocean, with the highest value (7.8 × 106 copies/mL) recorded at a station in the Tasman Sea (37°S). The absolute abundance of ARGs in the Southern Ocean was 1.8-fold lower than that in the western Pacific Ocean, and slightly increased (0.7–fold) toward Terra Nova Bay in Antarctica, possibly resulting from natural terrestrial sources or human activity. β-Lactam and tetracycline resistance genes were dominant in all samples (88–99%), indicating that they are likely the key ARGs in the ocean. Correlation and network analysis showed that Bdellovibrionota, Bacteroidetes, Cyanobacteria, Margulisbacteria, and Proteobacteria were positively correlated with ARGs, suggesting that these bacteria are the most likely ARG carriers. This study highlights the latitudinal profile of ARG distribution in the open ocean system and provides insights that will help in monitoring emerging pollutants on a global scale.

Antimicrobial resistance genes are enriched in aerosols near impacted urban surface waters in La Paz, Bolivia

Antibiotic resistance poses a major global health threat. Understanding emergence and dissemination of antibiotic resistance in environmental media is critical to the design of control strategies. Because antibiotic resistance genes (ARGs) may be aerosolized from contaminated point sources and disseminated more widely in localized environments, we assessed ARGs in aerosols in urban La Paz, Bolivia, where wastewater flows in engineered surface water channels through the densely populated urban core. We quantified key ARGs and a mobile integron (MI) via ddPCR and E. coli spp. as a fecal indicator by culture over two years during both the rainy and dry seasons in sites near wastewater flows. ARG targets represented major antibiotic groups—tetracyclines (tetA), fluoroquinolines (qnrB), and beta-lactams (blaTEM)—and an MI (intI1) represented the potential for mobility of genetic material. Most air samples (82%) had detectable targets above the experimentally determined LOD: most commonly blaTEM and intI1 (68% and 47% respectively) followed by tetA and qnrB (17% and 11% respectively). ARG and MI densities in positive air samples ranged from 1.3 × 101 to 6.6 × 104 gene copies/m3 air. Additionally, we detected culturable E. coli in the air (52% of samples <1 km from impacted surface waters) with an average density of 11 CFU/m3 in positive samples. We observed decreasing density of blaTEM with increasing distance up to 150 m from impacted surface waters. To our knowledge this is the first study conducting absolute quantification and a spatial analysis of ARGs and MIs in ambient urban air of a city with contaminated surface waters. Environments in close proximity to urban wastewater flows in this setting may experience locally elevated concentrations of ARGs, a possible concern for the emergence and dissemination of antimicrobial resistance in cities with poor sanitation.

Characterization of IncC Plasmids in Enterobacterales of Food-Producing Animals Originating From China.

Incompatibility group C (IncC) plasmids have received attention due to their broad host range and because they harbor key antibiotic resistance genes. Because these resistance genes can spread from food-producing animals to human, the proliferation of these plasmids represents a public health risk. In this study, a total of 20 IncC plasmids were collected from food-producing animals in China, and characterized by Oxford Nanopore Technologies long-read sequencing. Based on four key differences of the IncC backbone, 4 IncC plasmids were classified as type 1, 15 were classified as type 1/2 hybrid, and one was classified as type 2. The 15 type 1/2 hybrids were further divided into 13 type 1/2a and 2 type 1/2b, based on sequence differences arising from different homologous recombination events between type 1 and type 2 IncC backbones. Genome comparison of accessory resistance modules showed that different IncC plasmids exhibited various phenotypes via loss and gain of diverse modules, mainly within the blaCMY-carrying region, and two antibiotic resistance islands designated ARI-A and ARI-B. Interestingly, in addition to insertion and deletion events, IS26 or IS1294-mediated large sequence inversions were found in the IncC genome of the 4 type1/2a plasmids, suggesting that insertion sequence-mediated rearrangements also promote the diversity of the IncC genome. This study provides insight into the structural diversification and multidrug resistance of IncC plasmids identified from food-producing animals in China.

Metagenomic profiling of antibiotic resistance genes and their associations with bacterial community during multiple disinfection regimes in a full-scale drinking water treatment plant

For comprehensive insights into the effects of multiple disinfection regimes on antibiotic resistome in drinking water, this study utilized metagenomic approaches to reveal the changing patterns of antibiotic resistance genes (ARGs) and bacterial community as well as their associations. A total of 297 ARGs within 17 types were detected in the drinking water, and their total relative abundance ranged from 195.49 ± 24.85 to 626.31 ± 38.61 copies of ARGs per cell. The total ARG abundance was significantly increased after the antimicrobial resin and ultraviolet (AR/UV) disinfection while significantly decreased after the ozone and chlorine (O3/Cl2) disinfection and remained stable after AR/Cl2 disinfection. Overall, 18 ARGs including bacA, mexT, and blaOXA-12, mainly affiliated to bacitracin, multidrug, and beta-lactam, were persistent and discriminative during all the disinfection strategies in drinking water, and they were considered as key ARGs that represent the antibiotic resistome during drinking water disinfection. Additionally, possible hosts of 50% key ARGs were revealed based on co-occurrence network. During multiple disinfection processes, the change of Fusobacteriales and Aeromonadaceae in abundance mainly contributed to the abundance shift of bacA, and Pseudomonas mainly increased the abundance of mexT. These findings indicated that bacterial community shift may be the key factor driving the change of antibiotic resistome during disinfection. The strong association between antibiotic resistome alteration and bacterial community shift proposed in this study may enhance our understanding of the underlying mechanism of the disinfection effects on antibiotic resistance and benefit effective measures to improve safety of drinking water.

Structural features of the genome of methicillin-resistant Staphylococcus aureus (MRSA), a ceftaroline-resistant representative of ST239 epidemic clone isolated in Russia

Genomic analysis of methicillin-resistant ceftaroline-resistant Staphylococcus aureus and identification of features of the genetic diversity in representatives of the epidemic ST239 clone circulating in hospitals of the Russian Federation. Species identification was performed using Vitek 2 (bioMerieux). Determination of the sensitivity to antibiotics was carried out using a BD Phoenix semiautomatic bacterial analyzer (United States), and manually to ceftaroline. Whole genome sequencing was performed on the Illumina HiSeq 2500 platform. Data processing and analysis were performed using the following resources: CLC Genomics Workbench, SPAdes, RAST Server, NCBI Prokaryotic Genome Annotation Pipeline, PlasmidFinder, Phaster, BLASTp NCBI, and OrthoMCL. The main characteristics of the SA943 genome, including clusters of resistance genes, factors of pathogenicity, adhesion, invasion, and regulation, were revealed. When analyzing the genome structure, the closest similarity to the representatives of the Eurasian ST239 subclone identified in Turkey, as well as a number of differences from 16K and OC3 strains isolated in Russia earlier, were noted. A unique SSR sequence of the spa gene, which was placed into the database as a new spa type t‑18470, was identified. Substitutions in the amino acid sequence of the gene products of the mec complex, ccrC recombinase, and PBP2 protein, which distinguish SA943 from most ST239 representatives, were revealed. Representatives of several ST239 subclones, including ceftaroline-resistant SA943 belonging to the Eurasian subclone, were identified in hospitals of the Russian Federation. The Eurasian subclone representatives are not a homogeneous group, but differ significantly in the structure of SCCmec and mutations in key antibiotic resistance genes. Whole genome sequencing not only allows one to reveal the features of genome microevolution of epidemic strains in distinct regions, but also to improve the quality of epidemiological analysis and to optimize infection control measures.

Successful Identification of Pathogens by Polymerase Chain Reaction (PCR)-Based Electron Spray Ionization Time-of-Flight Mass Spectrometry (ESI-TOF-MS) in Culture-Negative Periprosthetic Joint Infection

The diagnosis of periprosthetic joint infection poses many challenges, one of which is the difficulty of isolating the infecting organism. Recently, a sophisticated modality (the Ibis Biosciences T5000 biosensor system) has been introduced that uses pan-domain primers in a series of polymerase chain reactions (PCRs) to identify and speciate essentially all bacteria and fungi as well as to identify key antibiotic resistance genes. We investigated the role of the Ibis in identifying infecting organisms in cases of known and suspected periprosthetic joint infection. Synovial fluid specimens were collected prospectively from eighty-two patients undergoing eighty-seven arthroplasty procedures (sixty-five knee revisions, fifteen hip revisions, and seven primary knee arthroplasties) and were sent for both conventional culture and Ibis analysis. The surgeon's clinical determination of the cause for revision arthroplasty was failure due to infection in twenty-three cases and noninfectious failure in fifty-seven cases. In the twenty-three cases that were considered on clinical grounds to involve a periprosthetic joint infection, the Ibis detected the same pathogen isolated by conventional culture in seventeen of eighteen cases and also detected one or more organisms in four of the five culture-negative cases. In addition, the Ibis detected organisms in fifty (88%) of the fifty-seven cases in which revision arthroplasty was performed for a presumed noninfectious failure. The Ibis technology was not only effective at detecting organisms in cases of suspected periprosthetic joint infection in which cultures were negative, but it also suggested that many of the revision arthroplasty cases that have previously been considered to be purely aseptic may have a component of unrecognized, subclinical infection.