Transfer Of Antibiotic Resistance Genes Research Articles

Alarming Antibiotic Resistance of Lactobacilli Isolated from Probiotic Preparations and Dietary Supplements.

In this study, we screened eight commercially available brands of Lactobacillus-containing probiotic preparations and dietary supplements for resistance towards commonly administered antibiotics of different classes. According to disc diffusion results, most of the isolates were resistant to vancomycin and susceptible to penicillin-type antibiotics (ampicillin and amoxicillin), carbapenems (imipenem, meropenem, and ertapenem), and inhibitors of protein synthesis (chloramphenicol, erythromycin, tetracycline, clarithromycin, and linezolid). However, based on minimum inhibitory concentration (MIC) values, six strains were reconsidered as resistant to tetracycline. All tested lactobacilli were resistant towards amikacin, ciprofloxacin, and norfloxacin. Resistance to cephalosporins was highly variable and decreased in the following order: ceftazidime/cefepime, ceftriaxone, cefotaxime, cefazolin, and cefoperazone. PCR screening for antibiotic resistance determinants in probiotic lactobacilli revealed a wide occurrence of vancomycin resistance gene vanX, ciprofloxacin resistance gene parC, and extended-spectrum β-lactamase gene blaTEM. We also detected the tetK gene for tetracycline resistance in one isolate. Additionally, we identified discrepancies between the claims of the manufacturers and the identified species composition, as well as the enumerated amount of viable bacteria, for several products. The results of this study raise concerns about the safety of lactobacilli for human consumption as probiotics, as they may act as reservoirs of transferable antibiotic resistance genes.

Elimination and Redistribution of Intracellular and Extracellular Antibiotic Resistance Genes in Water and Wastewater Disinfection Processes: A Review

Antibiotic resistance genes (ARGs) have been identified as emerging contaminants in water and wastewater. Concerns have been raised of whether disinfection processes can effectively eliminate ARGs and thus mitigate their spread. Due to the different existing forms and dissemination pathways, intracellular ARGs (iARGs) and extracellular ARGs (eARGs) have different responses to disinfection processes. This review compares the elimination of iARGs and eARGs in representative disinfection processes applied at water and wastewater treatment plants, including treatment by chlorine, chloramine, chlorine dioxide, ozone, ultraviolet irradiation (UV), and combination of UV with chemicals. Meanwhile, the redistribution of ARGs upon disinfection processes is highlighted. Elimination efficiencies of ARGs generally follow the order: ozone > chlorine ≫ chlorine dioxide > chloramine. The elimination of naked eARGs could be up to 4-log higher than that of iARGs. Notably, these disinfectants and the resulting disinfection byproducts formed in disinfection process can select antibiotic resistant bacteria (ARB) and increase relative abundances of iARGs. Under practical disinfectant exposure, elevated abundances of eARGs may be observed. Low doses of chlorine and chloramine tend to enhance the transfer of ARGs from intracellular to extracellular forms. To selectively control ARGs and associated risks and meanwhile minimize the unintended consequences, the disinfection practice may warrant optimization.

Microplastics affect nitrogen cycling and antibiotic resistance genes transfer of sediment

Microplastics are prevalent in freshwater, coastal, and deep ocean sediments. However, the effects of microplastics on sedimentary microbial ecosystems have not been well reported. Using 16S rRNA gene and metagenomic sequencing, we investigated coastal sedimentary sludge microbial community structure, composition, function, assembly, and propagation of antibiotic resistance genes (ARGs) in response to different microplastics. The nitrifier, denitrifiers and ammonia oxidizer were significantly altered upon microplastics exposure. The concentrations of NO3−, NO2−, and NH4+ in PE and PET treated groups were in consistent with relative abundances of corresponding functional genera. The increased antibiotic efflux ARGs abundance in PE, PET, and PVC groups was directly related to the promoted efflux pump system. Reactive oxygen species (ROS) production, ROS-mediated DNA damage, and increased stress response induced by microplastics exposure might be the reason for the promotion of ARGs transfer. Assembly of dominate nitrogen degrading genus and ARGs hosts tended to be deterministic under the pressure of microplastics, indicating the selective effect of microplastics.

Two sides of the same coin: Meta-analysis uncovered the potential benefits and risks of traditional fermented foods at a large geographical scale.

Traditional fermented foods, which are well-known microbial resources, are also bright national cultural inheritances. Recently, traditional fermented foods have received great attention due to their potential probiotic properties. Based on shotgun metagenomic sequencing data, we analyzed the microbial diversity, taxonomic composition, metabolic pathways, and the potential benefits and risks of fermented foods through a meta-analysis including 179 selected samples, as well as our own sequencing data collected from Hainan Province, China. As expected, raw materials, regions (differentiated by climatic zones), and substrates were the main driving forces for the microbial diversity and taxonomic composition of traditional fermented foods. Interestingly, a higher content of beneficial bacteria but a low biomass of opportunistic pathogens and antibiotic resistance genes were observed in the fermented dairy products, indicating that fermented dairy products are the most beneficial and reliable fermented foods. In contrast, despite the high microbial diversity found in the fermented soy products, their consumption risk was still high due to the enrichment of opportunistic pathogens and transferable antibiotic resistance genes. Overall, we provided the most comprehensive assessment of the microbiome of fermented food to date and generated a new view of its potential benefits and risks related to human health.

Review of the Distribution and Influence of Antibiotic Resistance Genes in Ballast Water

The misuse of antibiotics causes antibiotic resistance genes (ARGs) in bacteria to be gradually enriched by environmental selection, resulting in increased tolerance and resistance in bacteria to antibiotics. Ballast water is a mobile carrier for the global transfer of bacteria and genes, thus posing a certain risk of ARGs spreading into the global ocean. Therefore, it is important to investigate the current status of ARGs in ballast water, as well as control the abundance of ARGs. Herein, we attempt to comprehensively summarize the distribution and abundance of ARGs in ballast water from different sea areas and analyze the influencing factors (such as physical factors, chemical factors, temperature, pH, etc.) on the distribution of ARGs. Furthermore, we seek to review the changes in ARGs after differential disinfection technology treatment in ballast water (including chlorination, ultraviolet, ozone, and free radical technology), especially the enhancing effect of subinhibitory concentrations of disinfectants on ARGs transfer. Overall, we believe this review can serve as a guide for future researchers to establish a more reasonable standard of ballast water discharge that considers the pollution of ARGs and provide new insight into the risk of vertical and horizontal ARG transfer in ballast water after disinfection.

Reductive Stress Boosts the Horizontal Transfer of Plasmid-Borne Antibiotic Resistance Genes: The Neglected Side of the Intracellular Redox Spectrum.

The dissemination of plasmid-borne antibiotic resistance genes (ARGs) among bacteria is becoming a global challenge to the "One Health" concept. During conjugation, the donor/recipient usually encounter diverse stresses induced by the surrounding environment. Previous studies mainly focused on the effects of oxidative stress on plasmid conjugation, but ignored the potential contribution of reductive stress (RS), the other side of the intracellular redox spectrum. Herein, we demonstrated for the first time that RS induced by dithiothreitol could significantly boost the horizontal transfer of plasmid RP4 from Escherichia coli K12 to different recipients (E. coli HB101, Salmonella Typhimurium, and Pseudomonas putida KT2440). Phenotypic and genotypic tests confirmed that RS upregulated genes encoding the transfer apparatus of plasmid RP4, which was attributed to the promoted consumption of intracellular glutamine in the donor rather than the widely reported SOS response. Moreover, RS was verified to benefit ATP supply by activating glycolysis (e.g., GAPDH) and the respiratory chain (e.g., appBC), triggering the deficiency of intracellular free Mg2+ by promoting its binding, and reducing membrane permeability by stimulating cardiolipin biosynthesis, all of which were beneficial to the functioning of transfer apparatus. Overall, our findings uncovered the neglected risks of RS in ARG spreading and updated the regulatory mechanism of plasmid conjugation.

Optimized strategy valorizing unautoclaved cottonseed hull as ruminant alternative feeds via solid-state fermentation: Detoxifying polyphenols, restraining hazardous microflora and antibiotic-resistance gene hosts

Cottonseed hull (CSH), typical agro-industrial byproduct, is widely valorized as ruminant alternative feeds, but many constraints including extensive anti-nutritional factors and indigenous hazardous microflora are still reasonably unsolved. This study proposed a novel strategy that valorizes unautoclaved CSH as ruminant alternative feeds via solid-state fermentation (SSF), where feasibility and microbial mechanism were roundly investigated. The SSF of raw CSH (R group, proposed in this study) was compared with autoclaved CSH (M group, prevalent strategy) based on physicochemical properties, chemical functionalities, and superficial structures. Compared to M group, R group though has exhibited near detoxification effect of polyphenols (81.7% free gossypol (FG) and 62.2% condensed tannin (CT), both below threshold values) after SSF, yet the accumulation of crude protein and fat has been higher (109.3 and 51.0 g/kg dry matter, respectively). Metagenomic analyses indicated the indigenous hazardous microorganisms and antibiotic-resistance genes were prevented from amplification in R group. More importantly, Pleurotus was the keystone contributor to functional gene sets of degrading polyphenols (FG and CT) and lignin during SSF in R group; furthermore, CAZyme gene sets associated with polysaccharide degradation were primarily derived from bacterial communities ( Cellulomonas , and Stenotrophomonas , et al.). Ultimately, microbial SSF of raw CSH can be a feasible strategy with minimum facility investment and energy consumption, thus expediting CSH valorization in feedstock industries and the sustainable bio-economy in developing countries. • SSF valorizing raw cottonseed hull (CSH) as ruminant alternative feeds is feasible. • This strategy facilitates the sustainable bio-economy. • Polyphenols content declined to safe threshold during SSF. • SSF is resilient to animal pathogens and the transfer of antibiotic resistance genes. • Functional gene set about polyphenol and lignin degradation is traced from Pleurotus .

Antibiotic resistance bacteria and antibiotic resistance genes survived from the extremely acidity posing a risk on intestinal bacteria in an in vitro digestion model by horizontal gene transfer

Antibiotic resistance bacteria (ARB) and antibiotic resistance genes (ARGs) are emerging contaminants posing risk to human health. To investigate the pathogenic ARBs and the horizontal gene transfer (HGT) via both extracellular ARGs (eARGs) and intracellular ARGs (iARGs), an in vitro digestion simulation system was established to monitoring the ARB and ARGs passing through the artificial digestive tract. The results showed that ARB was mostly affected by the acidity of the gastric fluid with about 99% ARB (total population of 2.45 × 109–2.54 × 109) killed at pH 2.0 and severe damage of bacterial cell membrane. However, more than 80% ARB (total population of 2.71 × 109–3.90 × 109) survived the challenge when the pH of the gastric fluid was 3.0 and above. Most ARB died from the high acidity, but its ARGs, intI1 and 16 S rRNA could be detected. The eARGs (accounting for 0.03–24.56% of total genes) were less than iARGs obviously. The eARGs showed greater HGT potential than that of iARGs, suggesting that transformation occurred more easily than conjugation. The transferring potential followed: tet (100%) > sul (75%) > bla (58%), related to the high correlation of intI1 with tetA and sul2 (p < 0.01). Moreover, gastric juice of pH 1.0 could decrease the transfer frequency of ARGs by 2–3 order of magnitude compared to the control, but still posing potential risks to human health. Under the treatment of digestive fluid, ARGs showed high gene horizontal transfer potential, suggesting that food-borne ARBs pose a great risk of horizontal transfer of ARGs to intestinal bacteria.

Chlorination (but Not UV Disinfection) Generates Cell Debris that Increases Extracellular Antibiotic Resistance Gene Transfer via Proximal Adsorption to Recipients and Upregulated Transformation Genes.

To advance the understanding of antibiotic resistance propagation from wastewater treatment plants, it is important to elucidate how different effluent disinfection processes affect the dissemination of predominantly extracellular antibiotic resistance genes (eARGs). Here, we show that, by facilitating proximal adsorption to recipient cells, bacterial debris generated by chlorination (but not by UV irradiation) increases the natural transformation frequency of their adsorbed eARG by 2.9 to 7.2-fold relative to free eARGs. This is because chlorination increases the bacterial surface roughness by 1.1 to 6.7-fold and the affinity toward eARGs by 1.6 to 5.8-fold, and 98% of the total eARGs released after chlorination were adsorbed to cell debris. In contrast, UV irradiation released predominantly free eARGs with 18% to 56% lower transformation frequency. The collision theory indicates that the ARG donor-recipient collision frequency increased by 35.1-fold for eARGs adsorbed onto chlorination-generated bacterial debris, and the xDLVO model infers a 29% lower donor-recipient contact energy barrier for these ARGs. Exposure to chlorination-generated bacterial debris also upregulated genes associated with natural transformation in Vibrio vulnificus (e.g., tfoX encoding the major activator of natural transformation) by 2.6 to 5.2-fold, likely due to the generation of chlorinated molecules (5.1-fold higher Cl content after chlorination) and persistent reactive species (e.g., carbon-centered radicals) on bacterial debris. Increased proximal eARG adsorption to bacterial debris was also observed in the secondary effluent after chlorination; this decreased eARG decay by 64% and increased the relative abundance of ARGs by 7.2-fold. Overall, this study highlights that different disinfection approaches can result in different physical states of eARGs that affect their resulting dissemination potential via transformation.

Triclosan Promotes Conjugative Transfer of Antibiotic Resistance Genes to Opportunistic Pathogens in Environmental Microbiome.

Although triclosan, as a widely used antiseptic chemical, is known to promote the transmission of antibiotic resistance to diverse hosts in pure culture, it is still unclear whether and how triclosan could affect the transmission of broad-host-range plasmids among complex microbial communities. Here, bacterial culturing, fluorescence-based cell sorting, and high-throughput 16S rRNA gene amplicon sequencing were combined to investigate contributions of triclosan on the transfer rate and range of an IncP-type plasmid from a proteobacterial donor to an activated sludge microbiome. Our results demonstrate that triclosan significantly enhances the conjugative transfer of the RP4 plasmid among activated sludge communities at environmentally relevant concentrations. High-throughput 16S rRNA gene sequencing on sorted transconjugants demonstrates that triclosan not only promoted the intergenera transfer but also the intragenera transfer of the RP4 plasmid among activated sludge communities. Moreover, triclosan mediated the transfer of the RP4 plasmid to opportunistic human pathogens, for example, Legionella spp. The mechanism of triclosan-mediated conjugative transfer is primarily associated with excessive oxidative stress, followed by increased membrane permeability and provoked SOS response. Our findings offer insights into the impacts of triclosan on the dissemination of antibiotic resistance in the aquatic environmental microbiome.

Removal behavior and key drivers of antibiotic resistance genes in two full-scale leachate treatment plants

Leachate is a critical reservoir of antibiotic resistance genes (ARGs) and its proper treatment is closely related to human health and ecosystem safety. Here, we used high-throughput qPCR to explore the removal behavior of ARGs in two full-scale leachate treatment plants (LTPs) where biological treatment and membrane filtration processes were integrated. A total of 286 ARGs and 55 mobile genetic elements (MGEs) were detected, with aminoglycoside, multidrug and MLSB resistance genes being the most prevalent and abundant. Anaerobic digestion was found to be an important pretreatment process for leachate, while anoxic/aerobic tanks in membrane bioreactor (MBR) acted as incubators for ARGs due to their significant proliferation effect on ARGs. Integrated membrane filtration (UF-NF-RO) excelled in ARGs removal with absolute abundances reduced by 3 to 6 orders of magnitude, from about 109 copies/mL in raw leachate to 103-105 copies/mL in effluents. Our results also showed that leachate treatment processes significantly altered the composition of ARGs and bacterial communities. Procrustes analysis and network analysis revealed strong associations between microbes and ARGs, with several hub genes and bacterial genera identified. Structural equation models (SEMs) indicated that bacterial composition, MGEs and basic water properties were the key drivers shaping ARGs dynamics in the raw leachate, biological system and filtration system, respectively. Notably, several pathogens (e.g., Klebsiella, Vibrio, Aeromonas) were closely correlated with ARGs in raw leachate and may amplify the dissemination risks of ARGs. Moreover, insertion sequences in biological systems would accelerate the horizontal gene transfer of ARGs. In short, this study provides new insights into the mechanisms of ARGs removal and dissemination behavior in industrial-scale LTPs.

Characterization and Transferability of erm and tet Antibiotic Resistance Genes in Lactobacillus spp. Isolated from Traditional Fermented Milk.

Lactobacillus is a widely used bacteria and consumed through various fermented foods and beverages. Strains have been shown to carry resistance genes and mobile genetic elements with their ability to transfer the resistance to sensitive pathogenic strains. To study this, 4 cultures of Lactobacillus were isolated from traditional fermented milk. The isolates were able to grow up to 4% (w/v) NaCl concentration and 45°C temperature, and showed > 97% 16S rRNA gene similarities with Lactobacillus fermentum. All the isolates were phenotypically screened for the presence of antibiotic resistance. Minimum inhibitory concentration (MIC) as microbiological breakpoints were observed against a varied class of antibiotics. Isolates AKO 94.6, DVM 95.7, and NIFTEM 95.8 were explicitly resistant to ampicillin, ciprofloxacin and vancomycin with MIC well beyond the maximum range of 256µg/ml in the E-strip test. While isolate SKL1 was sensitive to ampicillin and showed MIC at 0.25µg/ml but resistant to streptomycin and trimethoprim (MIC > 256µg/ml). Molecular characterization showed the presence of tet(M) gene in three isolates SKL1, DVM 95.7, and NIFTEM 95.8 which was chromosomally associated resistance determinants while erm(B) resistance gene was detected in isolates DVM 95.7 and NIFTEM 95.8 only which was a plasmid associated gene and could be transferrable conjugally. Gene for Tn916 family (xis) was also observed in isolates DVM 95.7 and NIFTEM 95.8. Transferability of antibiotic resistance to pathogenic recipient strains was examined in isolates DVM 95.7 and NIFTEM 95.8 in different food matrices. The highest conjugation frequency with ~ 10-1 was obtained in alfalfa seed sprouts. This study reports the presence of acquired gene resistance in Lactobacillus species and dissemination to susceptible strains of bacteria in different food matrices. 16S rRNA gene sequences of isolates were uploaded to the NCBI GenBank database to retrieve the accession number.

Effects and mechanisms of phthalates on the transformation of antibiotic resistance genes into &amp;lt;italic&amp;gt;Escherichia coli&amp;lt;/italic&amp;gt;

<p indent="0mm">The horizontal transfer of antibiotic resistance genes (ARGs) is the fundamental cause of the spread of antibiotic resistance, which seriously threatens ecological security and public health. Transformation is an important way of ARGs horizontal transfer. Phthalates (PAEs) and ARGs can form compound pollution. However, the effects of PAEs on the transformation process of ARGs remain poorly understood. Thus, transformation experiments were carried out to explore the influence and mechanism of typical PAEs on the transformation of ARGs into <italic>Escherichia coli </italic>(<italic>E</italic>.<italic> coli</italic>). It was found that dimethyl phthalate (DMP), diethyl phthalate (DEP), dibutyl phthalate (DBP), di(2-ethylhexyl) phthalate (DEHP), and benzyl butyl phthalate (BBP) at lower than its minimum inhibitory concentration (MIC) respectively reduced the transformation efficiency of ARGs by 27.55%, 32.71%, 46.40%, 38.36%, and 59.87%, and the transformation frequency of ARGs also decreased obviously with the increase of PAEs concentrations. The results indicated that PAEs inhibited the transformation of ARGs into <italic>E</italic>.<italic> coli</italic>. PAEs induced plasmids carrying ARGs to form larger aggregates, reduced cell membrane permeability of recipient cells, caused an increase of oxidative stress in recipient cells, down-regulated the expression of genes involved in adenosine triphosphate (ATP) conversion, finally resulted in the inhibitory effect of ARGs transformation. This work revealed the mechanism of PAEs inhibiting the transformation of ARGs at cellular and molecular levels. The results will have scientific significance for understanding the molecular ecological effects of PAEs pollution in the environment.

Evaluation of the Microbiological Quality of Water in the Rice-Growing System of the Lis Valley, Portugal

Society’s demand for the preservation of water resources has stimulated technological advances and improved agricultural management, to limit its negative impacts, favor socio-economic development and promote the conservation of natural resources. This study assessed the impact of a rice farming system in the Mediterranean area on the microbiological quality of water, representing the irrigation and cultivation conditions of the Lis Valley Irrigation District (LVID). Indicators used include the counting of total coliforms (TC), fecal coliforms (FC), fecal enterococci (FE) and ampicillin-resistant Enterobacteriaceae (Ampr), and a phenotypic characterization of antibiotic (AB) resistances. The results show that the water at the LVID inlets contained high microbiological contamination due to sources outside the perimeter. The rice paddy agroecosystem had a beneficial impact on the load of TC, FC and FE of the respective drainage water, with a decrease of two orders of magnitude between the values of these counts at the inlets and at the outlets of the rice fields. However, the relative frequency of Enterobacteriaceae Ampr increased in the rice paddy soil, possibly as a result of the favorable conditions provided by this wetland ecosystem for the transfer of AB resistance genes. The analysis of AB resistance patterns revealed high relative resistance percentages for both β-lactams and non-β-lactams in all samples. This study made it possible to assess the risks of microbiological contamination of irrigation water and the beneficial effect of the rice-growing system in controlling the applied indicators.

Regulatory strategies for inhibiting horizontal gene transfer of ARGs in paddy and dryland soil through computer-based methods

Antibiotic resistance genes (ARGs) have been regarded as emerging pollutants due to their potential risk of resistance. Horizontal gene transfer (HGT) is the main pathway for ARGs to lead to environmental threats. Therefore, the inhabitation of ARGs' HGT can effectively inhibit ARGs' potential drug resistance risk within a single strain. In this paper, the characteristics of ARGs' HGT in paddy and dryland soils were identified and regulated by a combination of ARGs' HGT feature identification, transfer mechanism analysis and transfer process regulation. The homology modeling algorithm was used to simulate the construction of the Tn5 plasmid transposase of Escherichia coli (E. coli) for identifying ARGs' HGT characteristics. The GCG (212.617 Å) was thus determined as the target codon. Through integrated computer-based methods, results showed that the most important environmental disturbance factors for the HGT of ARGs in the paddy and dryland soils were rough farmyard manure/sewage irrigation and mining pollution, respectively. Under the disturbance of key environmental factors, the inhibitory effect of HGT of ARGs in paddy and dryland soil was reduced by 35.01 % and 34.74 %, respectively. Results demonstrated that the proposed theoretical mechanism and control strategies could effectively inhibit the HGT of E. coli ARGs in the soil environment.

Photophysical and in vitro photoinactivation of Escherichia coli using cationic 5,10,15,20-tetra(pyridin-3-yl) porphyrin and Zn(II) derivative conjugated to graphene quantum dots

Pathogenic microorganisms may continue causing infection through the transfer of antibiotic resistance genes. As a result, the efficacy of pharmaceuticals in microbial inactivation is deteriorating. The present study was conducted to investigate the antimicrobial activity of neutral and quaternized free base and Zn 5,10,15,20-tetra(pyridin-3-yl) porphyrins on Escherichia coli (E. coli), a gram-negative bacterium that causes cholecystitis, pneumonia and urinary tract infections. Conjugation of the porphyrin to graphene quantum dots (GQDs) was implemented to enhance photocatalysis and reactive oxygen species generation. Density functional theory (DFT) geometry optimizations for free base and Zn porphyrin based on the B3LYP (Becke 3-Parameter (Exchange), Lee, Yang and Parr) functional of the Gaussian09 program package and Time-dependent density-functional theory (TD-DFT) calculations of the associated UV-visible absorption spectra are reported to analyse the electronic structure and optical properties of the porphyrins. The TD-DFT calculations showed that for both porphyrins the value of highest occupied molecular orbital (ΔHOMO) is greater than that of lowest unoccupied molecular orbital (ΔLUMO) which tells that there is no unusual splitting of (LUMO) orbitals which may be caused by systematic error in TD-DFT calculations. Due to the red shift in the spectrum of ZnT(3-Py)P and the ΔLUMO being higher, the HOMO-LUMO gap was expected to be lower than that of H2T(3-Py)P. The photophysical properties and Photodynamic antimicrobial chemotherapy activities of these nanoconjugates were investigated. The highest ΦΔ was that of Q-ZnT(3-Py)P- GDQs at 0.69 with the log reduction of 9.42.

Early life dynamics of ARG and MGE associated with intestinal virome in neonatal piglets

The pre- and post-weaning stages for piglets are critical periods for the maturation of intestinal functions and contamination with antibiotic resistant bacterial pathogens will threaten their intestinal health. The presence of bacteriophage can also alter bacterial populations in the intestine but whether transmission of antibiotic resistance genes (ARG) is affected by phage during maturation of the neonatal piglet intestine is not known. We therefore identified the intestinal virome along with ARGs and mobile genetic elements (MGE) from piglet fecal samples collected from 3 to 28 days representing the different growth stages. We found wide fluctuations for the intestinal virome of weaning piglets and most virus - related antibiotic resistance was derived from temperate phage suggesting a reservoir of multidrug resistance was present in the neonatal porcine gut. Our results provide a comprehensive understanding of ARGs associated with the intestinal virome that therefore represents a potential risk for horizontal ARG transfer to pathogenic bacteria.

Dynamic distribution and potential transmission of antibiotic resistance genes in activated sludge.

Public concerns are increasing regarding the prevalence and transmission of antibiotic resistance genes (ARGs) in wastewater treatment plants (WWTPs), especially ARG persistence and dissemination in activated sludge (AS). However, the temporal dynamics of ARGs in the AS of WWTPs over a long period of time and their transfer potential after the treatment process upgrade (e.g., total nitrogen reduction from 20 to 15mg/L in effluent) remain poorly explored. Here, metagenomic sequencing was performed to quantify the ARGs in AS samples from two WWTPs with different treatment processes over a 2-year period. A total of 368 and 426 ARG subtypes affiliated with 20 ARG types were identified separately in the two WWTPs and the similar core ARGs were shared by all 54 samples. There were significant differences in ARG composition in different treatment processes, yet the abundance and diversity of ARGs in the AS samples demonstrated no distinct seasonal patterns. Notably, after the treatment process upgrade, the relative abundance of sulfonamide, beta-lactam, and aminoglycoside resistance genes was reduced by more than 10%, and the transfer potential of ARGs in bacterial pathogens decreased greatly, which suggested that an upgrade could limit the prevalence and transmission of ARGs. Variation partitioning analysis showed that metal resistance genes rather than bacterial community represented the significantly influential factor in shaping ARGs, and some key genera correlated with ARGs were identified through network analysis. These results will deepen our understanding of the dynamic changes in ARG profiles in AS systems and guide wastewater treatment plant upgrades. KEY POINTS: • The potential transfer of ARGs decreased after the treatment process upgrade • Metal resistance genes were the most influential factor in shaping ARG composition • Co-occurrence networks displayed potential hosts of beta-lactam resistance genes.

Pangasius Hypophthalmus Viscera as a Potential Vector of Bacterial Cross-Contamination and Resistance of Escherichia coli to Antibiotics

The viscera of Pangasius fish was studied to provide baseline information about the presence of antibiotic-resistant E. coli on it. This aimed to assess the possible resistance of bacterial pathogens to antibiotics and cross-contamination into the fish's muscles during processing, as well as to evaluate the effect of starvation on the microbial loads of Pangasius fish viscera. The resistance of E. coli was tested against 15 antimicrobial agents using the disk diffusion method. The findings revealed that starvation reduced microbial loads on the viscera compared to non-starvation Pangasius. LAB, coliforms, and E. coli count on viscera of non-starved Pangasius were 7.0±0.5, 5.5±0.9 and 5.4±1.0 log CFU g-1, whereas those of the starved fish were 2.6±0.8, 3.8±0.4 and 3.1±0.3 log CFU g-1, respectively. A total of 55 E. coli isolated from Pangasius viscera were tested for antimicrobial susceptibility as stated above. Surprisingly, 69.09% of E. coli isolates were multi-antibiotic resistant from three to fifteen antibiotics tested. A high level of resistance to ampicillin (63.64%), ceftazidime (69.09%), nalidixic acid (78.18%) was observed. More importantly, 9.09% of the E. coli isolates were resistant to all kinds of antibiotics tested. As E. coli is a potential vector for transfer of antibiotic resistance gene, causing cross-resistance with human enteric pathogens, there is a need for both the prudent use of these antimicrobial agents in aquaculture and stringent appropriate infection control in the processing chain in Vietnam.

Whole-Genome Analysis of Acinetobacter baumannii Strain AB43 Containing a Type I-Fb CRISPR-Cas System: Insights into the Relationship with Drug Resistance.

The CRISPR-Cas system is a bacterial and archaea adaptive immune system and is a newly recognized mechanism for controlling antibiotic resistance gene transfer. Acinetobacter baumannii (A. baumannii) is an important organism responsible for a variety of nosocomial infections. A. baumannii infections have become problematic worldwide because of the resistance of A. baumannii to multiple antibiotics. Thus, it is clinically significant to explore the relationship between the CRISPR-Cas system and drug resistance in A. baumannii. This study aimed to analyze the genomic characteristics of the A. baumannii strain AB3 containing the type I-Fb CRISPR-Cas system, which was isolated from a tertiary care hospital in China, and to investigate the relationship between the CRISPR-Cas system and antibiotic resistance in this strain. The whole-genome sequencing (WGS) of the AB43 strain was performed using Illumina and PacBio sequencing. The complete genome of AB43 consisted of a 3,854,806 bp chromosome and a 104,309 bp plasmid. The specific characteristics of the CRISPR-Cas system in AB43 are described as follows: (1) The strain AB43 carries a complete type I-Fb CRISPR-Cas system; (2) Homology analysis confirmed that the cas genes in AB43 share high sequence similarity with the same subtype cas genes; (3) A total of 28 of 105 A. baumannii AB43 CRISPR spacers matched genes in the bacteriophage genome database and the plasmid database, implying that the CRISPR-Cas system in AB43 provides immunity against invasive bacteriophage and plasmids; (4) None of the CRISPR spacers in A. baumannii AB43 were matched with antimicrobial resistance genes in the NCBI database. In addition, we analyzed the presence of antibiotic resistance genes and insertion sequences in the AB43 strain and found that the number of antibiotic resistance genes was not lower than in the “no CRISPR-Cas system” strain. This study supports the idea that the CRISPR-Cas system may inhibit drug-resistance gene expression via endogenous gene regulation, except to the published mechanism that the CRISPR-Cas system efficiently limits the acquisition of antibiotic resistance genes that make bacteria sensitive to antibiotics.