Colistin Resistance Gene Mcr-1 Research Articles

Prevalence and genetic diversity of multidrug-resistant Salmonella Typhimurium monophasic variant in a swine farm from China.

Salmonella 4,[5],12:i:-, a monophasic variant of S. Typhimurium, has become a global serovar causing animal and human infections since its first emergence in the late 1980's. Several previous studies showed the increasing prevalence of S. 4,[5],12:i:- in China, most of which were from swine with multidrug resistance (MDR) profiles. However, the molecular characteristic and evolution of S. 4,[5],12:i:- in the same swine farm are still unknown. In this study, a total of 54 S. enterica strains were isolated from different fattening pigs aged 1, 3, and 6 months, most of which belonged to S. 4,[5],12:i:-. Whole-genome sequencing revealed that all 45 S. 4,[5],12:i:- strains belonged to ST34 and were further divided into two different ribosomal STs and nine different core-genome STs. Phylogenetic analysis of 286 S. 4,[5],12:i:- strains in China, including 241 from the EnteroBase Salmonella database, revealed the genetic diversity of S. 4,[5],12:i:- and indicated that S. 4,[5],12:i:- in this swine farm might have multiple origins. Three different IncHI2 plasmids carrying various resistance genes were characterized by nanopore sequencing and could be conjugated to Escherichia coli. The colistin resistance gene mcr-1 and ESBLs gene blaCTX - M-14 were co-located on the chromosome of one strain. The dynamic changes in antimicrobial resistance regions and transferability of IncHI2 plasmids, as well as the chromosomal location of resistance genes, facilitated the diversity of the antimicrobial resistance characteristics in S. 4,[5],12:i:-. Since the swine farm is regarded as the important reservoir of MDR S. 4,[5],12:i:-, the prevalence and evolution of S. 4,[5],12:i:- from swine farms to pig products and humans should be continually monitored.

First report of mobile colistin resistance gene mcr-1 in avian pathogenic Escherichia coli isolated from turkeys in the Gaza Strip, Palestine.

Colistin is used to treat avian pathogenic Escherichia coli (APEC), a microorganism that affects turkey meat production in the Gaza Strip and worldwide. However, the recent emergence of plasmid-borne mobile colistin resistance (mcr) genes in pathogenic E. coli strains is a serious antimicrobial resistance (AMR) challenge for both human and animal health. In December 2018, colistin was banned as a veterinary antimicrobial in the Gaza Strip. This study aimed to detect and track the prevalence of colistin-resistant APEC isolated from turkey flocks in the Gaza Strip. This study investigated 239 APEC isolates from turkey flocks in the Gaza Strip between October 2018 and December 2021 (at 6-month intervals). The colistin-resistant APEC strains were detected using the broth microdilution method. The mcr-1 gene was identified using a polymerase chain reaction. The overall colistin resistance among the isolated APECs was 32.2% during the study period. The average resistance in the first interval was 37.5%, which significantly decreased to 9.3% in the last interval. Among the 77 phenotypically resistant isolates, 32.4% were positive for mcr-1. The average abundance of mcr-1 in the first interval was 66.6%, which decreased to 25% in the last interval. To the best of our knowledge, this is the first study reporting the presence of the mcr-1 gene among the APEC isolates from turkeys in the Gaza Strip. Banned veterinary use of colistin significantly decreased the percentage of resistant APEC isolates from turkeys in Gaza Strip. Further studies are needed to investigate other colistin resistance genes and track the emergence of AMR.

Gigantol restores the sensitivity of mcr carrying multidrug-resistant bacteria to colistin

BackgroundThe emergence and wide spread of plasmid-mediated colistin resistance gene (mcr-1) and its mutants have immensely limited the efficacy of colistin in treating multidrug-resistant (MDR) Gram-negative bacterial infections. The development of synergistic combinations of antibiotics with a natural product that coped with the resistance of MDR bacteria was an economic strategy to restore antibiotics activity. Herein, we investigated gigantol, a bibenzyl phytocompound, for restoring in vitro and in vivo, the sensitivity of mcr-positive bacteria to colistin. MethodsThe synergistic activity of gigantol and colistin against multidrug-resistant Enterobacterales was studied via checkerboard assay and time-killing curve. Subsequently, the transcription and protein expression levels of mcr-1 gene were determined by RT-PCR and Western blots. The interaction of gigantol and MCR-1 was simulated via molecular docking and verified via site-directed mutagenesis of MCR-1. Hemolytic activity and cytotoxicity assay were used to evaluate the safety of gigantol. Finally, the in vivo synergistic effect was evaluated via two animal infection models. ResultsGigantol restored the activity of colistin against mcr-positive bacteria E.coli B2 (MIC from 4 μg/ml to 0.25 μg/ml), Salmonella 15E343 (MIC from 8 μg/ml to 1 μg/ml), K. pneumoniae 19-2-1 (MIC from 32 μg/ml to 2 μg/ml) carrying mcr-1, mcr-3, mcr-8, respectively. Mechanistic studies revealed that gigantol down-regulated the expression of genes involved in LPS-modification, reduced the MCR-1 products and inhibited the activity of MCR-1 by binding to amino acid residues Tyr287 and Pro481 in its D-glucose-binding pocket. Safety evaluation showed that the addition of gigantol relieves the hemolysis caused by colistin. Compared with monotherapy, the combination of gigantol and colistin significantly improved the survival rate of Gallgallella mellonella larvae and mice infected by E.coli B2. Moreover, there was a considerable decrease in the bacterial load present in the viscera of mice. ConclusionOur results confirmed that gigantol was a potential colistin adjuvant, and could be used to tackle multi-drug resistant Gram-negative pathogen infections combined with colistin.

Low prevalence of mobilized resistance genes blaNDM, mcr-1, and tet(X4) in Escherichia coli from a hospital in China.

The emergence and spread of carbapenemase genes, colistin resistance genes mcr-1, and tigecycline resistance gene tet(X) represent a significant threat to clinical therapy and public health. In this study, we investigated the presence of carbapenemase genes, mcr-1, and tet(X) in 298 Escherichia coli strains obtained from a teaching hospital in China. In total, eight (2.68%), six (2.01%), and one (0.34%) E. coli isolates carried blaNDM, mcr-1, and tet(X4), respectively. The blaNDM gene was located on IncX3 (n = 4), F2:A-:B- (n = 3), and F2:A1:B1 (n = 1) plasmids, with high similarity to multiple plasmids belonging to the same incompatibility type from Enterobacteriaceae. Six MCR-producing strains contained mcr-1-carrying IncI2 plasmids, organized similarly to other mcr-1-bearing IncI2 plasmids from animals in China. The blaCTX-M-55/64/132/199 gene located within a typical transposition unit (ISEcp1-blaCTX-M-orf477Δ) was inserted near dnaJ to generate 5-bp direct repeats in four mcr-1-positive plasmids. The tet(X) and another four resistance genes [aadA2, tet(A), floR, and Δlnu(F)] were co-located on an IncX1 plasmid, highly similar to other tet(X4)-carrying IncX1 plasmids from Escherichia and Klebsiella of animal or food origin, except that the conjugative transfer region of IncX1 plasmids was absent in our plasmid. Although a low prevalence of blaNDM, mcr-1, and tet(X) was observed in E. coli from patients in this study, their dissemination associated with some successful pandemic plasmids is of great concern. The continued surveillance of these crucial resistance genes in patients should be strengthened.

Characterization of two novel colistin resistance gene mcr-1 variants originated from Moraxella spp.

This study aimed to characterize two novel mcr-1 variants, mcr-1.35 and mcr-1.36, which originated from Moraxella spp. that were isolated from diseased pigs in China. The Moraxella spp. carrying novel mcr-1 variants were subjected to whole-genome sequencing (WGS) and phylogenetic analysis based on the 16S rRNA gene. The mcr-1 variants mcr-1.35 and mcr-1.36 were characterized using phylogenetic analysis, a comparison of genetic environments, and protein structure prediction. The WGS indicated that two novel mcr-1 variants were located in the chromosomes of three Moraxella spp. with a genetic environment of mcr-1-pap2. In addition to the novel colistin resistance genes mcr-1.35 and mcr-1.36, the three Moraxella spp. contained other antimicrobial resistance genes, including aac(3)-IId, tet(O), sul2, floR, and blaROB-3. A functional cloning assay indicated that either the mcr-1.35 or mcr-1.36 gene could confer resistance to colistin in Escherichia coli DH5α and JM109. The nucleotide sequences of mcr-1.35 and mcr-1.36 presented 95.33 and 95.33% identities, respectively, to mcr-1.1. The phylogenetic analysis showed that mcr-1.35 and mcr-1.36 were derived from Moraxella spp. that belonged to subclades that were different from those of the mcr-1 variants (mcr-1.1 to mcr-1.34 except mcr-1.10) originating from Enterobacteriaceae. The deduced amino acid sequences of MCR-1.35 (MCR-1.36) showed 96.67% (96.49%), 82.59% (82.04%), 84.07% (83.52%), 55.52% (55.17%), 59.75% (59.57%), and 61.88% (61.69%) identity to MCR-1.10, MCR-2.2, MCR-6.1, MCR-LIN, MCR-OSL, and MCR-POR, respectively, that originated from Moraxella sp. Notably, protein structure alignment showed only a few changes in amino acid residues between MCR-1.1 and MCR-1.35, as well as between MCR-1.1 and MCR-1.36. In conclusion, this study identified Moraxella spp. carrying two novel mcr-1 variants, mcr-1.35 and mcr-1.36, conferring resistance to colistin, which were isolated from pig farms in China. In addition, mcr-like variants were observed to be located in the chromosomes of some species of Moraxella isolated from pig samples.

High prevalence of Escherichia coli co-harboring conjugative plasmids with colistin- and carbapenem resistance genes in a wastewater treatment plant in China.

Emergence and dissemination of resistance to last-resort antibiotics such as carbapenem and colistin is a growing, global health concern. Wastewater treatment plants (WWTPs) link human activities and the environment, can act as reservoirs and sources for emerging antibiotic resistance, and likely play a large role in antibiotic resistance transmission. The aim of this study was to investigate occurrence and characteristics of colistin- and carbapenem-resistant Escherichia coli (CCREC) in wastewater and sludge samples collected over a one-year period from different functional areas of an urban WWTP in Jinan city, Shandong, China. A total of 8 CCREC were isolated from 168 samples with selective agar and PCR, corresponding to high prevalence of 4.8%, co-harboring carbapenem resistance genes (blaNDM) and colistin resistance gene (mcr-1) and subsequently whole-genome sequenced. Additionally, all isolates were multidrug-resistant by antimicrobial susceptibility testing and carried a variety of antibiotic resistance genes. Two isolates carrying virulence genes associated with avian pathogenic E. coli were identified, one belonging to the high-risk clone O101:H9-ST167. Southern blotting was used to characterize CCREC isolates and plasmids carrying blaNDM-genes or mcr-1 could be transferred to a recipient strain E. coli J53 by in vitro conjugation assays. Resistance to other antibiotic classes were sporadically co-transferred to the transconjugant. Transposition of and mcr-1-carrying element from a transferable IncHI2-plasmid was observed among two CCREC clones isolated within 4 days of each other. The occurrence of multidrug-resistant CCREC capable of transferring their antibiotic resistance genotypes via conjugative plasmids is alarming. WWTPs bring bacteria from different sources together, providing opportunities for horizontal exchange of DNA among compatible hosts. Further dissemination of the colistin-, carbapenem-, or both colistin- and carbapenem resistant E. coli could lead to a serious threat to public health.

MC-PRPA-HLFIA Cascade Detection System for Point-of-Care Testing Pan-Drug-Resistant Genes in Urinary Tract Infection Samples.

Recently, urinary tract infection (UTI) triggered by bacteria carrying pan-drug-resistant genes, including carbapenem resistance gene blaNDM and blaKPC, colistin resistance gene mcr-1, and tet(X) for tigecycline resistance, have been reported, posing a serious challenge to the treatment of clinical UTI. Therefore, point-of-care (POC) detection of these genes in UTI samples without the need for pre-culturing is urgently needed. Based on PEG 200-enhanced recombinase polymerase amplification (RPA) and a refined Chelex-100 lysis method with HRP-catalyzed lateral flow immunoassay (LFIA), we developed an MCL-PRPA-HLFIA cascade assay system for detecting these genes in UTI samples. The refined Chelex-100 lysis method extracts target DNA from UTI samples in 20 min without high-speed centrifugation or pre-incubation of urine samples. Following optimization, the cascade detection system achieved an LOD of 102 CFU/mL with satisfactory specificity and could detect these genes in both simulated and actual UTI samples. It takes less than an hour to complete the process without the use of high-speed centrifuges or other specialized equipment, such as PCR amplifiers. The MCL-PRPA-HLFIA cascade assay system provides new ideas for the construction of rapid detection methods for pan-drug-resistant genes in clinical UTI samples and provides the necessary medication guidance for UTI treatment.

Cajanin stilbene acid: A direct inhibitor of colistin resistance protein MCR-1 that restores the efficacy of polymyxin B against resistant Gram-negative bacteria

BackgroundThe resistance of Gram-negative bacteria to polymyxin B, caused by the plasmid-mediated colistin resistance gene mcr-1, which encodes a phosphoethanolamine transferase (MCR-1), is a serious threat to global public health. Therefore, it is urgent to find new drugs that can effectively alleviate polymyxin B resistance. Through the screening of 78 natural compounds, we found that cajanin stilbene acid (CSA) can significantly restore the susceptibility of polymyxin B to mcr-1 positive Escherichia coli (E. coli). PurposeIn this study, we tried to evaluate the ability of CSA to restore the susceptibility of polymyxin B towards the E. coli, and explore the mechanism of sensitivity recovery. Study design and methodsCheckerboard MICs, time-killing curves, scanning electron microscope, lethal and semi-lethal models of infection in mice were used to assess the ability of CSA to restore the susceptibility of polymyxyn to E. coli. The interaction between CSA and MCR-1 was evaluated using surface plasmon resonance (SPR), and molecular docking experiments. ResultsHere, we find that CSA, a potential direct inhibitor of MCR-1, effectively restores the sensitivity of E. coli to polymyxin B. CSA can restore the sensitivity of polymyxin B to drug-resistant E. coli, and the MIC value can be reduced to 1 μg/ml. The time killing curve and scanning electron microscopy results also showed that CSA can effectively restore polymyxin B sensitivity. In vivo experiments showed that the simultaneous use of CSA and polymyxin B can effectively reduce the infection of drug-resistant E. coli in mice. SPR and molecular docking experiments confirmed that CSA strongly bound to MCR-1. The 17-carbonyl oxygen and 12- and 18‑hydroxyl oxygens of CSA were the key sites binding to MCR-1. ConclusionCSA is able to significantly restore the sensitivity of polymyxin B to E. coli in vivo and in vitro. CSA inhibits the enzymatic activity of the MCR-1 protein by binding to key amino acids at the active center of the MCR-1 protein.

Gram-negative ESKAPE bacteria bloodstream infections in patients during the COVID-19 pandemic.

Bloodstream infections due to bacteria are a highly consequential nosocomial occurrences and the organisms responsible for them are usually multidrug-resistant. The aims of this study were to describe the incidence of bacteremia caused by Gram-negative ESKAPE bacilli during the COVID-19 pandemic and characterize the clinical and microbiological findings including antimicrobial resistance. A total of 115 Gram-negative ESKAPE isolates were collected from patients with nosocomial bacteremia (18% of the total bacteremias) in a tertiary care center in Mexico City from February 2020 to January 2021. These isolates were more frequently derived from the Respiratory Diseases Ward (27), followed by the Neurosurgery (12), Intensive Care Unit (11), Internal Medicine (11), and Infectious Diseases Unit (7). The most frequently isolated bacteria were Acinetobacter baumannii (34%), followed by Klebsiella pneumoniae (28%), Pseudomonas aeruginosa (23%) and Enterobacter spp (16%). A. baumannii showed the highest levels of multidrug-resistance (100%), followed by K. pneumoniae (87%), Enterobacter spp (34%) and P. aeruginosa (20%). The bla CTX-M-15 and bla TEM-1 genes were identified in all beta-lactam-resistant K. pneumoniae (27), while bla TEM-1 was found in 84.6% (33/39) of A. baumannii isolates. The carbapenemase gene bla OXA-398 was predominant among carbapenem-resistant A. baumannii (74%, 29/39) and bla OXA-24was detected in four isolates. One P. aeruginosa isolate was bla VIM-2 gene carrier, while two K. pneumoniae and one Enterobacter spp were bla NDM gene carriers. Among colistin-resistant isolates mcr-1 gene was not detected. Clonal diversity was observed in K. pneumoniae, P. aeruginosa and Enterobacter spp. Two outbreaks caused by A. baumannii ST208 and ST369 were detected, both belonging to the clonal complex CC92 and IC2. A. baumannii was associated with a death rate of 72% (28/32), most of them (86%, 24/28) extensively drug-resistant or pandrug-resistant isolates, mainly in patients with COVID-19 (86%, 24/28) in the Respiratory Diseases Ward. A. baumannii isolates had a higher mortality rate (72%), which was higher in patients with COVID-19. There was no statistically significant association between the multidrug-resistant profile in Gram-negative ESKAPE bacilli and COVID-19 disease. The results point to the important role of multidrug-resistant Gram-negative ESKAPE bacteria causing bacteremia in nosocomial settings before and during the COVID-19 epidemic. Additionally, we were unable to identify a local impact of the COVID-19 pandemic on antimicrobial resistance rates, at least in the short term.

Epidemic characteristics of the SXT/R391 integrated conjugative elements in multidrug-resistant Proteus mirabilis isolated from chicken farm

This study was designed to depict prevalence and antimicrobial resistance characteristics of Proteus mirabilis (P. mirabilis) strains in 4 chicken farms and to probe the transfer mechanism of resistance genes. A total of 187 P. mirabilis isolates were isolated from 4 chicken farms. The susceptibility testing of these isolates to 14 antimicrobials showed that the multidrug resistance (MDR) rate was as high as 100%. The β-lactamase resistance genes blaOXA-1, blaCTX-M-1G, blaCTX-M-9G and colistin resistance gene mcr-1 were highly carried in the P. mirabilis isolates. An MDR strain W47 was selected for whole genome sequencing (WGS) and conjugation experiment. The results showed that W47 carried 23 resistance genes and 64 virulence genes, and an SXT/R391 integrated conjugative elements (ICEs) named ICEPmiChn5 carrying 17 genes was identified in chromosome. ICEPmiChn5 was able to be excised from the chromosome of W47 forming a circular intermediate, but repeated conjugation experiments were unsuccessful. Among 187 P. mirabilis isolates, 144 (77.01%, 144/187) isolates carried ICEPmiChn5-like ICEs, suggesting that ICEs may be the major vector for the transmission of resistance genes among MDR chicken P. mirabilis strains in this study. The findings were conducive to insight into the resistance mechanism of chicken P. mirabilis strains and provide a theoretical basis for the use of antibiotics for the treatment of MDR P. mirabilis infections in veterinary clinic.

Identification of a novel adjuvant loperamide that enhances the antibacterial activity of colistin against MCR-1-positive pathogens in vitro/vivo.

Severe infection with multidrug-resistant Enterobacterales caused by the plasmid-induced colistin resistance gene MCR-1 is a serious public health challenge. In this case, it is necessary and pressing to find a treatment to overcome antibiotic resistance. Here, we investigated the synergistic effect and mechanism of loperamide combined with colistin against MCR-1-positive pathogens. We evaluated the combined effect of loperamide and colistin using the checkerboard method and the time-kill experiment. The results showed that loperamide could enhance the bactericidal ability of colistin, and this combination regimen could completely kill the tested bacteria within 4 h. Subsequently, spectrofluorimetric methods were used to explore the mechanism of loperamide combined with colistin. The results indicated that the mode of action of loperamide combined with colistin was found to involve mechanical disruption of the membrane. Furthermore, molecular simulation and microscale thermophoresis results revealed that loperamide reduced the impact of MCR-1 protein by directly binding to its active site. In addition, the combined regimen of loperamide and colistin effectively reduced the bacterial load in the thighs of mice while increasing the protection rate by 70%. In short, as a potential lead compound, loperamide can enhance the killing effect of colistin on pathogenic Enterobacterales carrying MCR-1 by causing membrane damage and inhibiting MCR-1 protein activity.

Enterobacteriaceae genome-wide analysis reveals roles for P1-like phage-plasmids in transmission of mcr-1, tetX4 and other antibiotic resistance genes

P1 -like phage-plasmids (PPs) are important gene vehicles in isolated pathogens. In this study, we conducted genome-wide and cross-species analysis of antimicrobial resistance genes (ARGs) from 35 ARG-positive P1-like PPs. LS-BSR analysis reveal that P1-like PPs had in common 7 highly variable regions and carried 48 different ARG subtypes. The most prevalent gene groups were the colistin resistance gene mcr-1 and a class 1 integron. Analysis of the flanking sequences of mcr-1 indicated an “IS30-mcr-1-ORF-IS30” as the core cluster. In particular, we found an mcr-1- and blaCTX-M-55-coharboring large fusion P1-like PP. Also, tet(X4) was detected and flanking sequences indicated tet(X4)-bearing cluster can formed a larger size fusion plasmid mediated a wider spread via IS26 hotspots. Overall, this study demonstrated that P1-like PPs can not only mobilize a large number of ARGs in variable regions but also form larger hybrid P1-like PPs that would increase their ability to spread antimicrobial resistance.

Outbreak of colistin resistant, carbapenemase (bla NDM, bla OXA-232) producing Klebsiella pneumoniae causing blood stream infection among neonates at a tertiary care hospital in India.

Infections caused by multi-drug resistant Klebsiella pneumoniae are a leading cause of mortality and morbidity among hospitalized patients. In neonatal intensive care units (NICU), blood stream infections by K. pneumoniae are one of the most common nosocomial infections leading to poor clinical outcomes and prolonged hospital stays. Here, we describe an outbreak of multi-drug resistant K. pneumoniae among neonates admitted at the NICU of a large tertiary care hospital in India. The outbreak involved 5 out of 7 neonates admitted in the NICU. The antibiotic sensitivity profiles revealed that all K. pneumoniae isolates were multi-drug resistant including carbapenems and colistin. The isolates belonged to three different sequence types namely, ST-11, ST-16 and ST-101. The isolates harboured carbapenemase genes, mainly bla NDM-1, bla NDM-5 and bla OXA-232 besides extended-spectrum β-lactamases however the colistin resistance gene mcr-1, mcr-2 and mcr-3 could not be detected. Extensive environmental screening of the ward and healthcare personnel led to the isolation of K. pneumoniae ST101 from filtered incubator water, harboring bla NDM-5, bla OXA-232 and ESBL genes (bla CTX-M) but was negative for the mcr genes. Strict infection control measures were applied and the outbreak was contained. This study emphasizes that early detection of such high-risk clones of multi-drug resistant isolates, surveillance and proper infection control practices are crucial to prevent outbreaks and further spread into the community.

Discovery, structure and function of plasmid mediated shufflon

Antimicrobial resistance has become a major public health issue of global concern. Conjugation is an important way for fast spreading drug-resistant plasmids, during which the type Ⅳ pili plays an important role. Type Ⅳ pili can adhere on the surfaces of host cell and other medium, facilitating formation of bacterial biofilms, bacterial aggregations and microcolonies, and is also a critical factor in liquid conjugation. PilV is an adhesin-type protein found on the tip of type Ⅳ pili encoded by plasmid R64, and can recognize the lipopolysaccharid (LPS) molecules that locate on bacterial membrane. The shufflon is a clustered inversion region that diversifies the PilV protein, which consequently affects the recipient recognition and conjugation frequency in liquid mating. The shufflon was firstly discovered on an IncI1 plasmid R64 and has been identified subsequently in plasmids IncI2, IncK and IncZ, as well as the pathogenicity island of Salmonella typhi. The shufflon consists of four segments including A, B, C, and D, and a specific recombination site named sfx. The shufflon is regulated by its downstream-located recombinase-encoding gene rci, and different rearrangements of the shufflon region in different plasmids were observed. Mobile colistin resistance gene mcr-1, which has attracted substantial attentions recently, is mainly located in IncI2 plasmid. The shufflon may be one of the contributors to fast spread of mcr-1. Herein, we reviewed the discovery, structure, function and prevalence of plasmid mediated shufflon, aiming to provide a theoretical basis on transmission mechanism and control strategy of drug-resistant plasmids.

Gold drugs as colistin adjuvants in the fight against MCR-1 producing bacteria.

The emergence and rapid spread of the mobile colistin resistance gene mcr-1 among bacterial species and hosts significantly challenge the efficacy of "last-line" antibiotic colistin. Previously, we reported silver nitrate and auranofin serve as colistin adjuvants for combating mcr-1-positive bacteria. Herein, we uncovered more gold-based drugs and nanoparticles, and found that they exhibited varying degree of synergisms with colistin on killing mcr-1-positive bacteria. However, pre-activation of the drugs by either glutathione or N-acetyl cysteine, thus releasing and accumulating gold ions, is perquisite for their abilities to substitute zinc cofactor from MCR-1 enzyme. X-ray crystallography and biophysical studies further supported the proposed mechanism. This study not only provides basis for combining gold-based drugs and colistin for combating mcr-1-positive bacterial infections, but also undoubtedly opens a new horizon for metabolism details of gold-based drugs in overcoming antimicrobial resistance.

Detection and Dissemination of MCR-1 Colistin Resistance Gene in a Hospital Setting in Abakaliki, Nigeria

Detection and Dissemination of MCR-1 Colistin Resistance Gene in a Hospital Setting in Abakaliki, Nigeria

Carriage and Transmission of mcr-1 in Salmonella Typhimurium and Its Monophasic 1,4,[5],12:i:- Variants from Diarrheal Outpatients: a 10-Year Genomic Epidemiology in Guangdong, Southern China.

The banning of colistin as a feed additive for food-producing animals in mainland China in 2017 caused the decline in the prevalence of Escherichia coli-mobilized colistin resistance (mcr-1) in China. Salmonella Typhimurium and its monophasic 1,4,[5],12:i:- variants are also the main species associated with the spread of mcr-1; however, the evidence of the prevalence and transmission of mcr-1 among Salmonella is lacking. Herein, the 5,354 Salmonella isolates recovered from fecal samples of diarrheal patients in Guangdong, Southern China, from 2009 to 2019 were screened for colistin resistance and mcr-1, and mcr-1-positive isolates were characterized based on whole-genome sequencing (WGS) data. Relatively high prevalence rates of colistin resistance and mcr-1 (4.05%/4.50%) were identified, and more importantly, the prevalence trends of colistin-resistant and mcr-1-positive Salmonella isolates had a similar dynamic profile, i.e., both were first detected in 2012 and rapidly increased during 2013 to 2016, followed by a sharp decrease since 2017. WGS and phylogenetic analysis indicate that, whether before or after the ban, the persistence and cross-hospital transmission of mcr-1 are primarily determined by IncHI2 plasmids with similar backbones and sequence type 34 (ST34) Salmonella in specific clades that are associated with a high prevalence of IncHI2 plasmids and clinically important antimicrobial resistance genes, including blaCTX-M-14-fosA3-oqxAB-floR genotypes. Our work reveals the difference in the prevalence rate of mcr-1 in clinical Salmonella before and after the Chinese colistin ban, whereas mcr-1 transmission was closely linked to multidrug-resistant IncHI2 plasmid and ST34 Salmonella across diverse hospitals over 10 years. Continued surveillance is required to explore the factors related to a sharp decrease in mcr-1 after the recent ban and determine whether the ban has affected the carriage of mcr-1 in Salmonella circulating in the health care system. IMPORTANCE Colistin is one of the last-line antibiotics for the clinical treatment of Enterobacteriaceae. However, the emergence of the mobilized colistin resistance (mcr-1) gene has spread throughout the entire human health system and largely threatens the usage of colistin in the clinical setting. In this study, we investigated the existence of mcr-1 in clinical Salmonella from a 10-year continuous surveillance and genomic study. Overall, the colistin resistance rate and mcr-1 carriage of Salmonella in tertiary hospitals in Guangdong (2009 to 2019) were relatively high and, importantly, rapidly increased from 2013 to 2016 and significantly decreased after the Chinese colistin withdrawal. However, before or after the ban, the MDR IncHI2 plasmid with a similar backbone and ST34 Salmonella were the main vectors involved in the spread of mcr-1. Interestingly, these Chinese mcr-1-carrying Salmonella obtain phylogenetically and phylogeographically distinct patterns compared with those from other continents and are frequently associated with clinically important ARGs including the extended-spectrum β-lactamases. Our data confirmed that the national stewardship intervention seems to be successful in blocking antibiotic resistance determinants and that continued surveillance of colistin resistance in clinical settings, farm animals, and related products is necessary.

Spread of the mcr-1 colistin-resistance gene in Escherichia coli through plasmid transmission and chromosomal transposition in French goats.

Colistin-resistance widely disseminated in food-producing animals due to decades of colistin use to treat diarrhea. The plasmid-borne mcr-1 gene has been extensively reported from bovine, swine and chicken worldwide, but smaller productions such as the goat farming sector were much less surveyed. We looked for colistin-resistant isolates presenting plasmid-borne genes of the mcr family in both breeding (n=80) and fattening farms (n=5). Localization of the mcr-1 gene was performed using Southern blot analysis coupled to short-read and long-read sequencing. Only the mcr-1 gene was identified in 10% (8/80) of the breeding farms and four over the five fattening farms. In total, 4.2% (65/1561) of the animals tested in breeding farms and 60.0% (84/140) of those tested in fattening farms presented a mcr-1-positive E. coli. The mcr-1 gene was located either on the chromosome (32.2%) or on IncX4 (38.9%) and IncHI2 (26.8%) plasmids. As expected, both clonal expansion and plasmidic transfers were observed in farms where the mcr-1 gene was carried by plasmids. Tn6330 transposition was observed in the chromosome of diverse E. coli sequence types within the same farm. Our results show that the mcr-1 gene is circulating in goat production and is located either on plasmids or on the chromosome. Evidence of Tn6330 transposition highlighted the fact that chromosomal insertion does not impair the transmission capability of the mcr-1 gene. Only strict hygiene and biosecurity procedures in breeding farms, as well as a prudent use of antibiotics in fattening farms, can avoid such complex contamination pathways.

Dihydroartemisinin inhibits plasmid transfer in drug-resistant Escherichia coli via limiting energy supply.

Conjugative transfer of antibiotic resistance genes (ARGs) by plasmids is an important route for ARG dissemination. An increasing number of antibiotic and nonantibiotic compounds have been reported to aid the spread of ARGs, highlighting potential challenges for controlling this type of horizontal transfer. Development of conjugation inhibitors that block or delay the transfer of ARG-bearing plasmids is a promising strategy to control the propagation of antibiotic resistance. Although such inhibitors are rare, they typically exhibit relatively high toxicity and low efficacy in vivo and their mechanisms of action are inadequately understood. Here, we studied the effects of dihydroartemisinin (DHA), an artemisinin derivative used to treat malaria, on conjugation. DHA inhibited the conjugation of the IncI2 and IncX4 plasmids carrying the mobile colistin resistance gene ( mcr-1) by more than 160-fold in vitro in Escherichia coli, and more than two-fold (IncI2 plasmid) in vivo in a mouse model. It also suppressed the transfer of the IncX3 plasmid carrying the carbapenem resistance gene bla NDM-5 by more than two-fold in vitro. Detection of intracellular adenosine triphosphate (ATP) and proton motive force (PMF), in combination with transcriptomic and metabolomic analyses, revealed that DHA impaired the function of the electron transport chain (ETC) by inhibiting the tricarboxylic acid (TCA) cycle pathway, thereby disrupting PMF and limiting the availability of intracellular ATP for plasmid conjugative transfer. Furthermore, expression levels of genes related to conjugation and pilus generation were significantly down-regulated during DHA exposure, indicating that the transfer apparatus for conjugation may be inhibited. Our findings provide new insights into the control of antibiotic resistance and the potential use of DHA.

Carbapenem and Colistin Resistance, Integrons and Plasmid Replicon Types in Multi-Drug Resistant Klebsiella Strains Isolated in Turkey.

Multidrug-resistant (MDR) Klebsiella is a globally important nosocomial pathogen. In the present study, 101 multidrug-resistant Klebsiella strains isolated from various clinical specimens obtained from two different Medical Faculties' hospitals were involved. We aimed to find out the prevalence of carbapenemase, mobile colistin resistance genes, and integrons in MDR Klebsiella strains. The antibiotic susceptibilities of strains were determined by Kirby Bauer disc-diffusion method and resistance to colistin was confirmed by detection of minimum inhibitory concentrations. The prevalence of carba-penemase genes (blaOXA-48, blaNDM, blaIMP, blaVIM, blaKPC), mobile colistin-resistance genes (mcr-1 and mcr-2), and integrons (class I, II and III) were examined in Klebsiella strains by polymerase chain reaction. All strains were resistant to β-lactam antibiotics, carbapenems, and quinolones. On the other hand, only nine (8.9%) strains were resistant to colistin. The most common carbapenemase genes were blaNDM (64.3%) and blaOXA-48 (53.5%). Besides, 28 (27.7%) strains were found to harbor both blaNDM and blaOXA-48. These 28 strains be-longed to the IncA/C (18.7%), IncL/M (7.7%), and IncFIIs (1.1%) plasmid replicon types. No strain was positive for blaIMP, mcr-1, and mcr-2. Class I and Class II integrons were shown to be harbored in 83.2% and 63.3% of strains, respectively. In total, 63 (63.6%) of strains harbored both classes I and II integrons. Class III integron was not detected. There was a statistically significant relationship between the presence of integrons and antibiotic resistance for cefotaxime (p = 0.024), ciprofloxacin (p < 0.001) trimethoprim/sulfamethoxazole (p < 0.001) and levofloxacin (p = 0.002). To our knowledge, this study represents the first report of a human isolate for the co-presence of blaNDM, blaOXA-48 and both Class I and Class II integrons, from Turkey. Our findings also highlight the dissemination of integrons and carbapenemases and the importance of surveillance on emerging antibiotic resistance.