Tetracycline Resistance Genes Research Articles

An Expanded Genetic Toolbox to Accelerate the Creation of Acholeplasma laidlawii Driven by Synthetic Genomes.

We have developed genetic tools for the atypical bacterium Acholeplasma laidlawii. A. laidlawii is a member of the class Mollicutes, which lacks cell walls, has small genomes, and has limited metabolic capabilities, requiring many metabolites from their hosts. Several of these traits have facilitated the development of genome transplantation for some Mollicutes, consequently enabling the generation of synthetic cells. Here, we propose the development of genome transplantation for A. laidlawii. We first investigated a donor-recipient relationship between two strains, PG-8A and PG-8195, through whole-genome sequencing. We then created multihost shuttle plasmids and used them to optimize an electroporation protocol. We also evolved a superior strain for DNA uptake via electroporation. We created a PG-8A donor strain with a Tn5 transposon carrying a tetracycline resistance gene. These tools will enhance Acholeplasma research and accelerate the effort toward creating A. laidlawii strains with synthetic genomes.

The prevalence of Staphylococcus aureus and the emergence of livestock-associated MRSA CC398 in pig production in eastern China.

Livestock-associated Staphylococcus aureus (LA-MRSA) has been of increasing concern due to its potential risk to humans. This study investigated the prevalence of MRSA in pig production in Eastern China and determined the genomic characteristics of pig-associated MRSA isolates by whole-genome sequencing (WGS). A total of 1,318 samples were collected from pig farms and pig slaughterhouses, and 150 S. aureus were identified, including 63 MRSA isolates and 87 MSSA isolates. MRSA was detected in all pig farms and pig slaughterhouses. The antimicrobial susceptibility test revealed that all MRSA isolates were multidrug-resistant. The WGS and MLST analysis demonstrated that 56 MRSA isolates belonged to clonal complex (CC) 398, and seven MRSA isolates belonged to CC9. All LA-MRSA isolates were absent of phiSa3 phage containing immune evasion cluster (IEC) and possessed an intact hlb gene. In addition, genes associated with Panton-Valentine leukocidin, typically indicative of human adaptation, were not detected. The analysis of antibiotic resistance genes (ARGs) demonstrated that all MRSA isolates contained multiple ARGs. All MRSA isolates had Plthe mecA gene and at least one tetracycline resistance gene. Both tetM and tetK were detected in all MRSA CC398 isolates, while tetL was detected in all MRSA CC9 isolates. The phenicol resistance gene fexA was detected in 51 MRSA isolates, while the linezolid resistance gene cfr was detected in 60 MRSA isolates. The emergence of LA-MRSA CC398 in four pig farms and one slaughterhouse in this study indicates the spread of this clonal complex in the pig production sector in Eastern China. Further investigations are required to understand the potential transmission routes of LA-MRSA CC398 within the pork production chain in China and to assess the potential risks to humans.

Antibiotic resistance profiles of gut microbiota across various primate species in Guangxi.

Understanding the gut microbiota and antibiotic resistance gene (ARG) profiles in non-human primates (NHPs) is crucial for evaluating their potential impact on human health and the environment. In this study, we performed metagenomic analysis of 203 primate fecal samples, including nine NHP species and humans, to comprehensively characterize their gut microbiota and ARGs. Our study reveals the prevailing phyla in primates as Firmicutes, Bacteroidetes, Euryarchaeota, and Proteobacteria. The captive NHPs exhibited higher ARG abundance compared to their wild counterparts, with tetracycline and beta-lactam resistance genes prevailing. Notably, ARG subtypes in Trachypithecus leucocephalus (T. leucocephalus) residing in karst limestone habitats displayed a more dispersed distribution compared to other species. Interestingly, ARG profiles of NHPs clustered based on geographic location and captivity status. Co-occurrence network analysis revealed intricate correlations between ARG subtypes and bacterial taxa. Procrustes analysis unveiled a significant correlation between ARGs and microbial phylogenetic community structure. Taxonomic composition analysis further highlighted differences in microbial abundance among NHPs and humans. Our study underscores the impact of lifestyle and geographical location on NHP gut microbiota and ARGs, providing essential insights into the potential risks posed by NHPs to antibiotic resistance dissemination. This comprehensive analysis enhances our understanding of the interplay between NHPs and the gut resistome, offering a critical reference for future research on antibiotic resistance and host-microbe interactions.

Transmission of antimicrobial resistance in the gut microbiome of gregarious cockroaches: the importance of interaction between antibiotic exposed and non-exposed populations.

Antimicrobial resistance (AMR) is a major global health concern, further complicated by its spread via the microbiome bacterial members. While mathematical models discuss AMR transmission through the symbiotic microbiome, experimental studies are scarce. Herein, we used a gregarious cockroach, Pycnoscelus surinamensis, as an in vivo animal model for AMR transmission investigations. We explored whether the effect of antimicrobial treatment is detectable with metagenomic sequencing, and whether AMR genes can be spread and established in unchallenged (not treated with antibiotics) individuals following contact with treated donors, and under various frequencies of interaction. Gut and soil substrate microbiomes were investigated by metagenomic sequencing for bacterial community composition and resistome profiling. We found that tetracycline treatment altered the treated gut microbiome by decreasing bacterial diversity and increasing the abundance of tetracycline resistance genes. Untreated cockroaches that interacted with treated donors also had elevated tetracycline resistance. The levels of resistance differed depending on the magnitude and frequency of donor transfer. Additionally, treated donors showed signs of microbiome recovery due to their interaction with the untreated ones. Similar patterns were also recorded in the soil substrate microbiomes. Our results shed light on how interacting microbiomes facilitate AMR gene transmission to previously unchallenged hosts, a dynamic influenced by the interaction frequencies, using an in vivo model to validate theoretical AMR transmission models.IMPORTANCEAntimicrobial resistance is a rising threat to human and animal health. The spread of resistance through the transmission of the symbiotic gut microbiome is of concern and has been explored in theoretical modeling studies. In this study, we employ gregarious insect populations to examine the emergence and transmission of antimicrobial resistance in vivo and validate modeling hypotheses. We find that antimicrobial treatment increases the levels of resistance in treated populations. Most importantly, we show that resistance increased in untreated populations after interacting with the treated ones. The level of resistance transmission was affected by the magnitude and frequency of population mixing. Our results highlight the importance of microbial transmission in the spread of antimicrobial resistance.

Effect of Antimicrobial Use in Conventional Versus Natural Cattle Feedlots on the Microbiome and Resistome.

Antimicrobial use (AMU) in the livestock industry has been associated with increased levels of antimicrobial resistance. Recently, there has been an increase in the number of "natural" feedlots in the beef cattle sector that raise cattle without antibiotics. Shotgun metagenomics was employed to characterize the impact of AMU in feedlot cattle on the microbiome, resistome, and mobilome. Sequenced fecal samples identified a decline (q < 0.01) in the genera Methanobrevibacter and Treponema in the microbiome of naturally vs. conventionally raised feedlot cattle, but this difference was not (q > 0.05) observed in catch basin samples. No differences (q > 0.05) were found in the class-level resistome between feedlot practices. In fecal samples, decreases from conventional to natural (q < 0.05) were noted in reads for the antimicrobial-resistant genes (ARGs) mefA, tet40, tetO, tetQ, and tetW. Plasmid-associated ARGs were more common in feces from conventional than natural feedlot cattle. Interestingly, more chromosomal- than plasmid-associated macrolide resistance genes were observed in both natural and conventional feedlots, suggesting that they were more stably conserved than the predominately plasmid-associated tetracycline resistance genes. This study suggests that generationally selected resistomes through decades of AMU persist even after AMU ceases in natural production systems.

Exogenous mobile genetic elements and their associated integrons drive the enrichment of antibiotic-resistant genes in the river of a valley basin city (Lanzhou, China).

River is a unique source of drinking water in valley-type cities, affecting local urban development and human lifestyles. However, the key driving factors for dissemination of antibiotic-resistant genes (ARGs) in valley-type urban environments remain unclear. This study aimed to investigate the distribution of ARGs in the Yellow River and to clarify the driving factors of ARGs in a typical valley basin city (Lanzhou, China). The seven selected ARGs with higher abundances including tetracycline resistance genes (tetM, tetX), macrolide resistance genes (ermB, ermF, ereA), and sulfonamide resistance genes (sul1, sul2) were detected. The results showed that the total absolute abundance of all the selected ARGs varied from 9.97 × 1012 to 1.04 × 1015 copies/L in the water body, with higher abundances in the wet season, relative to the dry season. Among these, sulfonamide resistance genes (sul1, sul2) displayed the highest absolute abundance in the river and soil. The ARGs and mobile genetic elements (MGEs) were significantly correlated with bacterial abundance, dissolved organic carbon (DOC), ammonia nitrogen (NH4+), and total nitrogen (TN) levels in the water environment (Mantel test, P < 0.01). Structural equation modeling revealed the direct input of point-source and nonpoint-source ARGs in this area contributed less to the overall level of the ARGs in the water. Among the multiple drivers, the MGEs derived from wastewater treatment plant and anthropogenic nonpoint area positively and directly affected the ARG profiles in water (P < 0.01), rather than the factors of bacterial abundance and physicochemical properties. According to this study, the exogenous MGEs from anthropogenic activities are the main driver for the enrichment of ARGs in the valley-type urban river environment.

Unveiling antimicrobial resistance in Chilean fertilized soils: a One Health perspective on environmental AMR surveillance.

Antimicrobial resistance (AMR) poses a significant threat to humans and animals as well as the environment. Within agricultural settings, the utilization of antimicrobial agents in animal husbandry can lead to the emergence of antimicrobial resistance. In Chile, the widespread use of animal-derived organic amendments, including manure and compost, requires an examination of the potential emergence of AMR resulting from their application. The aim of this research was to identify and compare AMR genes found in fertilized soils and manure in Los Andes city, Chile. Soil samples were collected from an agricultural field, comprising unamended soils, amended soils, and manure used for crop fertilization. The selected genes (n = 28) included genes associated with resistance to beta-lactams, tetracyclines, sulfonamides, polymyxins, macrolides, quinolones, aminoglycosides, as well as mobile genetic elements and multidrug resistance genes. Twenty genes were successfully identified in the samples. Tetracycline resistance genes displayed the highest prevalence, followed by MGE and sulfonamides, while quinolone resistance genes were comparatively less abundant. Notably, blaOXA, sulA, tetO, tetW, tetM, aac (6) ib., and intI1, exhibited higher frequencies in unamended soils, indicating their potential persistence within the soil microbiome and contribution to the perpetuation of AMR over time. Given the complex nature of AMR, it is crucial to adopt an integrated surveillance framework that embraces the One Health approach, involving multiple sectors, to effectively address this challenge. This study represents the first investigation of antimicrobial resistance genes in agricultural soils in Chile, shedding light on the presence and dynamics of AMR in this context.

A Novel IncX Plasmid Mediates High-Level Oxytetracycline and Streptomycin Resistance in Erwinia amylovora from Commercial Pear Orchards in California.

Isolates of the fire blight pathogen Erwinia amylovora with high-level resistance to oxytetracycline (minimal inhibitory concentration [MIC] > 100 μg/ml) and to streptomycin (MIC > 100 μg/ml) were recovered from four commercial pear orchards in California between 2018 and 2020. The two representative oxytetracycline- and streptomycin-resistant (OxyTcR-SmR) strains 32-10 and 33-1 were as virulent as the antibiotic susceptible strain 13-1 in causing blossom blight of pear and were recovered more than 50% of the time 7 days after co-inoculation to pear flowers with strain 13-1. In the field, inoculation of strain 32-10 to pear flowers that were pretreated with oxytetracycline at 200 μg/ml did not reduce disease compared with an untreated control. Four OxyTcR-SmR strains were subjected to draft genome sequencing to identify the genetic determinants of antibiotic resistance and their location. A 43.6-kb IncX plasmid, designated pX11-7, was detected in each of the four strains, and this plasmid encoded the tetracycline-resistance gene tetB and the streptomycin-resistance gene pair strAB within a large putatively mobile genetic element consisting of the transposon Tn10 that had inserted within the streptomycin-resistance transposon Tn6082. We also determined that pX11-7 was conjugative and was transferred at a rate that was 104 to 105 higher into an E. amylovora strain isolated in California compared with an E. amylovora strain that was isolated in Michigan. The occurrence of high levels of resistance to both oxytetracycline and streptomycin in E. amylovora strains from commercial pear orchards in California significantly limits the options for blossom blight management in these locations.

Tracking the extracellular and intracellular antibiotic resistance genes across whole year in wastewater of intensive dairy farm

Monitoring the annual variation of antibiotic resistance genes (ARGs) in livestock wastewater is important for determining the high-risk period of transfer and spread of animal-derived antibiotic resistance into the environment. However, the knowledge regarding the variation patterns of ARGs, especially intracellular ARGs (iARGs) and extracellular ARGs (eARGs), over time in livestock wastewater is still unclear. Herein, we conducted a year-round study to trace the profiles of ARGs at a Chinese-intensive dairy farm, focusing on the shifts observed in different months. The results showed significant differences in the composition and variation between iARGs and eARGs. Tetracycline, sulfonamide, and macrolide resistance genes were the major types of iARGs, while cfr was the major type of eARG. The environmental adaptations of the host bacteria determine whether ARGs appear as intracellular or extracellular forms. The total abundance of ARGs was higher from April to September, which can be attributed to the favorable climatic conditions for bacterial colonization and increased antibiotic administration during this period. Integron was found to be highly correlated with most iARGs, potentially playing a role in the presence of these genes within cells and their similar transmission patterns in wastewater. The intracellular and extracellular bacterial communities were significantly different, primarily because of variations in bacterial adaptability to the high salt and anaerobic environment. The intracellular co-occurrence network indicated that some dominant genera in wastewater, such as Turicibacter, Clostridium IV, Cloacibacillus, Subdivision5_genera_incertae_sedis, Saccharibacteria_genera_incertae_sedis and Halomonas, were potential hosts for many ARGs. To the best of our knowledge, this study demonstrates, for the first time, the annual variation of ARGs at critical points in the reuse of dairy farm wastewater. It also offers valuable insights into the prevention and control of ARGs derived from animals.

Antimicrobial resistance and genomic characteristics of Salmonella from broilers in Shandong Province

IntroductionThe emergence of multidrug-resistant (MDR) strains of Salmonella, which is a genus of important zoonotic pathogens, has aroused great public health concern worldwide.MethodsIn this study, 167 strains of Salmonella were isolated from 947 samples from broiler farms, slaughterhouses, and markets in Shandong Province. Antibiotic sensitivity testing was performed, and 70 strains of Salmonella were screened out by whole-genome sequencing (WGS) to evaluate serotypes, antimicrobial resistance genes (ARGs), the prevalence of class 1 integrons, the plasmid carriage rate, and phylogenetic characteristics and for multilocus sequence typing (MLST).ResultsThe results showed that the 167 isolates showed the highest resistance to ampicillin (AMP, 87.4%), sulfamethoxazole (SF, 87.4%), compound sulfamethoxazole (SXT, 81.4%), nalidixic acid (NAL, 80.2%), and amoxicillin/clavulanic acid (A/C, 77.8%). All the strains were sensitive to meropenem (MEM), and 91.0% of the isolates were MDR strains. We screened a total of 45 ARGs, with the highest detection rate observed for the tetracycline (TET) resistance gene tet (A) (81.4%). A total of 21 types of plasmid replicons were detected in Salmonella, of which IncX1 was the most common (74.3%), and 62.9% of the isolates carried a class 1 integron. In addition, a total of 11 different serotypes were detected, with S. enteritidis as the predominant serovar., followed by S. infantis and S. Newport. Twelve different sequence types (STs) were detected, among which ST11 was the main type. There was a strong correspondence between serotypes and STs. We also found that S. Indiana and S. Kentucky had extremely high rates of resistance to ciprofloxacin (CIP) and third-generation cephalosporins. System-wide genome analysis showed the occurrence of long-distance transmission across fields.ConclusionIn conclusion, the detection of multidrug resistance and isolates carrying multidrug resistance genes is the main problem, and emergency strategies should be implemented to address this issue.

Effects of chicken manure‐modified biochar on the adsorption capacity of tetracycline and abundance of antibiotic resistance genes in soil

AbstractLarge quantities of livestock manure not only occupy farmland but also serve as the primary source of antibiotics and antibiotic resistance genes (ARGs) in agricultural soil. To enhance the resource utilization of animal manure and mitigate ecological risks associated with antibiotics in agroecosystems, chicken manure was used as a raw material for biochar production. The manure derived biochar (MB) was modified with an alkali–acid two‐step modification process. The results showed that the total pore volume and surface area of alkali–acid modified MB (AMMB) was significantly increased about three and five times, respectively after modification. The SEM images showed that the surface of AMMB had a more regular and clearer structure with fewer impurities. The improvement of surface properties of AMMB significantly increased its adsorbing capacity of tetracycline in the soil. The adsorption kinetics of tetracycline by AMMB were better fitted by the pseudo‐second‐order kinetic model, suggesting that the adsorption of tetracycline on biochar followed the process of chemisorption. The adsorption mechanisms mainly involved the pore filling, hydrophobic effect, and π–π EDA interaction. Moreover, after a 60 days amendment of AMMB, the total relative abundances (RAs) of tested tetracycline resistance genes (TRGs) in soil decreased significantly, which suggested that AMMB can be a potential material to reduce risks of antibiotics and its resistance genes.

Investigation of the antimicrobial susceptibility patterns of marine cyanobacteria in Bohai Bay: Cyanobacteria may be important hosts of antibiotic resistance genes in marine environment

Marine cyanobacteria, as widely distributed and photosynthetically autotrophic bacteria in the ocean, may contribute to the global dissemination of antibiotic resistance genes (ARGs) and develop a different antimicrobial susceptibility pattern from heterotrophic bacteria and cyanobacteria from freshwater environments. However, studies on antimicrobial susceptibility and the carriage of ARGs in marine cyanobacteria are still very limited. In this study, the antibiotic resistance characteristics of cyanobacteria in nearshore waters were examined through field monitoring and laboratory investigations, which included PCR detection and ARG transformation. The results showed a positive correlation between marine cyanobacteria and some ARGs in the nearshore waters of Bohai Bay. Moreover, most screened cyanobacteria showed high minimum inhibitory concentration (MIC) values for polymyxins, tetracyclines, kanamycin, and sulfonamides, moderate MIC values for streptomycin, chloramphenicol, rifampicin, and norfloxacin, and low MIC values for roxithromycin and cephalosporins. The blaTEM, blaKPC, sul1, sul2, strA, tetA, tetB, tetC, tetM, mdfA, and intI1 genes were detected in the screened marine cyanobacteria. The highest detection rates were observed for blaTEM (93.3 %), sul1 (56.6 %), sul2 (90 %), and strA (73.3 %). The detection rate of tetA (33.3 %) was the highest among the tetracycline resistance genes, and mdfA, a multidrug-resistant pump gene with resistance to tetracycline, also showed a high detection level (23.3 %). Overall, most of the screened marine cyanobacteria were found to tolerate multiple antibiotics in seawater, and the condition of the ARGs carriage was serious. Furthermore, the screened marine Synechocystis sp. C12-2 demonstrated the ability to accept ARGs on the RP4 plasmid through natural transformation and showed reduced sensitivity to ampicillin, suggesting the possibility that some marine cyanobacteria could acquire ARGs from the environment through horizontal gene transfer. Thus, marine cyanobacteria may play an important role in the propagation of marine ARGs.

One-pot hydrothermal preparation of rich-oxygen vacant Bi2SiO5/CuBi2O4 Z-Scheme heterojunction for visible light-driven photocatalytic removal of antibiotic-resistant bacteria

Bi2SiO5/CuBi2O4 Z-Scheme heterojunction enriched with oxygen vacancies were prepared by a simple one-pot hydrothermal method. The photocatalytic activity of Bi2SiO5/CuBi2O4 nanocomposites was evaluated under visible light irradiation using tetracycline, antibiotic-resistant Escherichia coli (AR E. coli), and tetracycline-resistant genes as the target pollutants. The results showed that Bi2SiO5/CuBi2O4 nanocomposites had the best photocatalytic activity, which could degrade 83.3 % of tetracycline within 2 h, completely inactivate AR E. coli within 2.5 h, and almost completely destroy tetracycline-resistant genes within 3 h. In addition, the scavenging experiments and ESR experiments not only showed that the active substances of O2-, h+ and OH played a dominant role and 1O2 played a secondary role in the photocatalytic system of Bi2SiO5/CuBi2O4 nanocomposites, but also confirmed that the Z-Scheme heterojunction was successfully constructed between Bi2SiO5 and CuBi2O4. Oxygen vacancies were detected by EPR experiments. The formation of oxygen vacancies and Z-Scheme heterojunction enhanced the light absorption properties of the Bi2SiO5/CuBi2O4 nanocomposites and improved the separation efficiency of the photogenerated electron-hole pairs compared to the single Bi2SiO5 and CuBi2O4 materials. This work showed that Bi2SiO5/CuBi2O4 was a new and efficient photocatalyst for the removal of tetracycline, AR E. coli and tetracycline-resistant genes, which provided a feasible idea for the management of water environment pollution.

The distribution profiles of tetracycline resistance genes in rice: Comparisons using four genotypes

The potential transmission of antibiotic resistance genes (ARGs) from the rhizosphere to plants and humans poses a significant concern. This study aims to investigate the distribution of tetracycline resistance genes (TRGs) in rice using four genotypes and identify the primary source of TRGs in grains. Quantitative polymerase chain reaction (qPCR) was employed to determine the abundance of seven TRGs and intI1 in four rice varieties and three partitions during the jointing and heading stages, respectively. The analysis of the bacterial community was conducted to elucidate the underlying mechanism of the profiles of TRGs. It was observed that tetZ was predominantly present in the rhizosphere and endoroot, whereas tetX became dominant in grains. The relative abundances of TRGs and intI1 exhibited significant variations across both the variety and partition. However, no significant differences were observed in grains, where the abundances of TRGs were several orders of magnitude lower compared to those in the rhizosphere. Nevertheless, the potential risk of the dissemination of TRGs to humans, particularly those carried by potential pathogens in grains, warrants attention. The increased likelihood of TRGs accumulation in the rhizosphere and endoroot of hybrid rice varieties, as opposed to japonica varieties, may be attributed to the heightened metabolic activities of their roots. The significant associations observed between intI1 and TRGs, coupled with the substantial alterations in potential hosts for intI1 across various treatments, indicate that intI1-mediated horizontal gene transfer plays a role in the diverse range of bacterial hosts for TRGs. The study also revealed that rhizosphere bacteria during the jointing stage serve as the primary contributors of TRGs in grains through the endoroot junction. The findings indicate that Japonica rice varieties exhibit superior control over TRGs compared to hybrid varieties, emphasizing the need for early interventions throughout the entire growth period of rice.

380 The Sow Milk Microbiome and Resistome

Abstract In addition to oligosaccharides, proteins (e.g., immunoglobulins), lipids, vitamins, and other nutrients vital for piglet health, sow’s milk contains bacteria believed to be important in gut health. Therefore, certain sow milk isolates with potentially beneficial properties may be of interest for use as probiotic strains. Furthermore, given the relatively large reservoir of antimicrobial-resistant bacteria carried by commercial pigs even in the absence of direct antimicrobial exposure, it is possible that sow’s milk may also transfer antimicrobial-resistant bacteria to their piglets. The objective of this study was to characterize the bacterial microbiome of sow’s milk using both culturing and shotgun metagenomics. To do this, colostrum (within 24 h of farrowing) and milk samples (n = 42) were collected from 14 sows at d 0, 7, and 21 during the suckling phase. We recovered 323 isolates on four different culture media, and 135 of these were chosen for whole-genome sequencing based on their 16S rRNA gene sequence identity to maximize isolate diversity. The majority (n = 90) of the sequenced isolates belonged to either the Staphylococcus or Streptococcus genera. Twelve isolates (Lactiplantibacillus plantarum, Lactobacillus amylovorus, Ligilactobacillus salivarius, Limosilactobacillus reuteri, and Weissella paramesenteroides) were members of lactic acid-producing bacterial species noted for their beneficial properties in mammals. Several potentially pathogenic bacterial species were also isolated including Clostridium perfringens, Escherichia coli, Klebsiella pneumonia, Streptococcus dysgalactiae, Staphylococcus hyicus, and Streptococcus suis. In addition, seven isolates could not be assigned to a specific species (&amp;lt; 95% average nucleotide identity) and thus may represent potentially novel bacterial species. We also mapped the shotgun metagenomic reads back to the sequenced isolate genomes and found Rothia spp. and L. amylovorus to be relatively most abundant ( &amp;gt; 0.25%). Nearly all of the isolates (n = 130) carried at least one antimicrobial resistance gene and many strains, including S. suis and S. dysgalactiae isolates, encoded genes conferring resistance to more than three antimicrobial classes. The tetracycline resistance genes tet(W), tet(Q), tet(W/N/W), tet(K), and tet(O) were the relatively most abundant antimicrobial resistance genes in the colostrum and milk metagenomes. These results show that sow’s milk may be a source of both beneficial and potentially pathogenic bacteria as well as a significant source of antimicrobial-resistant bacteria.

Characteristics and diversity of Vibrio parahaemolyticus causing acute hepatopancreatic necrosis disease in Vietnam

Acute hepatopancreatic necrosis disease (AHPND) is a contagious disease in shrimp with extremely high mortality rate, causing an estimated $23.6 billion loss to the global shrimp industry during 2009–2016. A binary toxin, PirAB, is the causative agent, which is encoded by the pirABvp genes on a virulent plasmid typically found in Vibrio parahaemolyticus. Here, seventeen strains of V. parahaemolyticus were isolated from the Mekong Delta of Vietnam, of which 15 harboured identical pirABvp genes. The remaining two strains, which were PCR-negative for pirAB, and nine positive strains were selected for whole genome sequencing. Multi-locus sequence typing revealed five sequence types (STs), including one novel, that were associated with sites of isolation. A maximum likelihood tree of core genomes revealed that our strains formed three clusters and were closely related to strains from Thailand, South Korea, China, and Philippines. All sequenced strains carried β-lactam (including blaOXA-10 and blaNDM-1) and tetracycline resistance genes, with a range of 3–13 resistance genes. The pirAB-positive strains shared large regions of homology with the prototypical AHPND virulent plasmid. In summary, this work revealed the characteristics of AHPND-causing strains in the Mekong Delta of Vietnam and suggested possible transmission routes among Vietnam and other shrimp farming countries.

Synergistic effects of polymyxin and vancomycin combinations on carbapenem- and polymyxin-resistant Klebsiella pneumoniae and their molecular characteristics.

The emergence and spread of polymyxin resistance, especially among Klebsiella pneumoniae isolates, threaten the effective management of infections. This study profiled for polymyxin resistance mechanisms and investigated the activity of polymyxins plus vancomycin against carbapenem- and polymyxin-resistant K. pneumoniae. The entire genome sequences of seven isolates were profiled for resistance and virulence determinants. The effects of combination therapy were evaluated using the checkerboard technique, time-kill assay, and population profile analysis. Protein profiles of the isolates treated with monotherapy were compared to that of combination therapy. The whole-genome sequencing data revealed that the isolates harbored β-lactams, carbapenems, aminoglycoside, fluoroquinolones, macrolides, and tetracycline resistance genes, with several virulence-associated genes. The plasmids including the bla OXA-232-bearing ColKP3 plasmid were also identified in our isolates. Profiling for polymyxin resistance mechanism revealed a missense mutation in the crrB gene that resulted in a Q180L variant that conferred a deleterious effect on protein function. The combination assay indicated fractional inhibitory concentration index ranging from 0.31 to 1.13, whereas the time-kill assay demonstrated synergistic log reduction in colony-forming units per milliliter. Furthermore, population analysis profiling using dual antibiotics indicated enhancement in bacterial log reduction at lower antibiotics concentrations, compared to higher concentrations of single polymyxins. For protein profiling, 796 proteins were identified, and 56 and 94 of them were increased and decreased in the combined drug treatment groups, respectively, while other differentially produced proteins were detected in all treatment groups, except for the control group. The results demonstrated that the vancomycin combination might benefit the antimicrobial activities of polymyxins. IMPORTANCE This study provides insights into the mechanisms of polymyxin resistance in K. pneumoniae clinical isolates and demonstrates potential strategies of polymyxin and vancomycin combinations for combating this resistance. We also identified possible mechanisms that might be associated with the treatment of these combinations against carbapenem- and polymyxin-resistant K. pneumoniae clinical isolates. The findings have significant implications for the development of alternative therapies and the effective management of infections caused by these pathogens.

Pet cats may shape the antibiotic resistome of their owner’s gut and living environment

BackgroundCompanion animals can contribute to the physical and mental health of people and often live in very close association with their owners. However, the antibiotic resistome carried by companion animals and the impact they have on their owners and living environment remain unclear. In this study, we compared the ARG profiles of cats, humans, and their living environments using metagenomic analysis to identify the core ARGs in the cat and human gut and explore the potential impact of cats on ARGs in the human gut through the environment.ResultsResults showed that the abundance of ARGs in the cat gut was significantly higher than that in the human gut (P < 0.0001), with aminoglycoside and tetracycline resistance genes being the dominant ARGs in the cat gut. There was no significant difference in the abundance of total ARGs in the guts of cat owners and non-owners (P > 0.05). However, the abundance of aminoglycoside resistance genes including APH(2'')-IIa and AAC(6')-Im was significantly higher in cat owners than that in non-cat owners (P < 0.001). Also, ARG abundance was positively correlated with the frequency of cat activity in the living environment. Enterobacteriaceae was the dominant ARG host co-occurring in the cat gut, human gut, and living environment.ConclusionsOur results show that cats may shape the living environment resistome and thus the composition of some ARGs in the human gut, highlighting the importance of companion animal environment health.CA_8GzYC3SY3RzvHMSVpYQVideo

Isolation, antimicrobial resistance and virulence characterization of Salmonella spp. from fresh foods in retail markets in Hangzhou, China.

Salmonella can cause severe foodborne diseases. This study investigated the prevalence of Salmonella spp. in fresh foods in Hangzhou market and their harborage of antibiotic resistance and virulence genes, antibiotic susceptibility, and pathogenicity. A total of 500 samples (pork, n = 140; chicken, n = 128; vegetable, n = 232) were collected over a one-year period. Salmonella was found in 4.2% (21) of samples with the detection rate in pork, chicken and vegetables as 4.3% (6), 6.3% (8), and 3% (7), respectively. One Salmonella strain was recovered from each positive sample. The isolates were identified as six serotypes, of which S. Enteritidis (n = 7) and S. Typhimurium (n = 6) were the most predominant serotypes. The majority of isolates showed resistance to tetracycline (85.7%) and/or ciprofloxacin (71.4%). Tetracycline resistance genes showed the highest prevalence (90.5%). The occurrence of resistance genes for β-lactams (blaTEM-1, 66.7%; and blaSHV, 9.5%) and aminoglycosides (aadA1, 47.6%; Aac(3)-Ia, 19%) was higher than sulfonamides (sul1, 42.9%) and quinolones (parC, 38.1%). The virulence gene fimA was detected in 57.1% of isolates. Gene co-occurrence analysis implied that resistance genes were associated with virulence genes. Furthermore, selected S. Typhimurium isolates (n = 4) carrying different resistance and virulence genes up-regulated the secretions of cytokines IL-6 and IL-8 by Caco-2 cells in different degrees, suggesting that virulence genes may play a role in inflammatory transcription. In in vivo virulence test, microbiological counts in mouse feces and tissues showed that all included S. Typhimurium were able to infect mice, with one strain showing significantly higher virulence than others. In conclusion, this study indicates Salmonella contamination in fresh foods in Hangzhou market poses a risk to public health and it should be closely monitored to prevent and control foodborne diseases.

High prevalence of antibiotic-resistant and metal-tolerant cultivable bacteria in remote glacier environment

Studies of antibiotic-resistant bacteria (ARB) have mainly originated from anthropic-influenced environments, with limited information from pristine environments. Remote cold environments are major reservoirs of ARB and have been determined in polar regions; however, their abundance in non-polar cold habitats is underexplored. This study evaluated antibiotics and metals resistance profiles, prevalence of antibiotic resistance genes (ARGs) and metals tolerance genes (MTGs) in 38 ARB isolated from the glacier debris and meltwater from Baishui Glacier No 1, China. Molecular identification displayed Proteobacteria (39.3%) predominant in debris, while meltwater was dominated by Actinobacteria (30%) and Proteobacteria (30%). Bacterial isolates exhibited multiple antibiotic resistance index values > 0.2. Gram-negative bacteria displayed higher resistance to antibiotics and metals than Gram-positive. PCR amplification exhibited distinct ARGs in bacteria dominated by β-lactam genes blaCTX-M (21.1–71.1%), blaACC (21.1–60.5%), tetracycline-resistant gene tetA (21.1–60.5%), and sulfonamide-resistant gene sulI (18.4–52.6%). Moreover, different MTGs were reported in bacterial isolates, including mercury-resistant merA (21.1–63.2%), copper-resistant copB (18.4–57.9%), chromium-resistant chrA (15.8–44.7%) and arsenic-resistant arsB (10.5–44.7%). This highlights the co-selection and co-occurrence of MTGs and ARGs in remote glacier environments. Different bacteria shared same ARGs, signifying horizontal gene transfer between species. Strong positive correlation among ARGs and MTGs was reported. Metals tolerance range exhibited that Gram-negative and Gram-positive bacteria clustered distinctly. Gram-negative bacteria were significantly tolerant to metals. Amino acid sequences of blaACC,blaCTX-M,blaSHV,blaampC,qnrA, sulI, tetA and blaTEM revealed variations. This study presents promising ARB, harboring ARGs with variations in amino acid sequences, highlighting the need to assess the transcriptome study of glacier bacteria conferring ARGs and MTGs.