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Escherichia coli and Klebsiella pneumonia; along with other Gram-negative bacteria (GNB), are known for their resilience to a broad spectrum of antibiotics, including last-resort options such as colistin, which is regarded as the last line of defense in treating multidrugresistant (MDR) infections. This elevated level of resistance presents a substantial threat to the healthcare system, paving the way for a post-antibiotic era. In the present study, a checkerboard assay was employed to pinpoint the optimal combinations of Origanum majorana (OM) essential oil and colistin (CS) for the purpose of enhancing colistin's antibacterial and antibiofilm activities (FICI ≤ 0.5). The investigation focused on eight colistin-resistant (CS_R) bacterial isolates, with a particular focus on two strains carrying the mobile colistin resistance mcr-1 gene. The CS@OM combination resulted in a 2- to 16-fold improvement of colistin efficacy, resulting in a biofilm eradication rate ranging from ∼ 52 % to 82 %. Furthermore, it evinced a complete eradication of the free-floating cells of the isolates, inclusive of those harboring the mcr-1 gene. The bactericidal activity of the CS@OM combination was achieved through various mechanisms, including bacterial membrane disruption and leakage of internal bacterial constituents, leading to bacterial death. Consequently, the CS@OM combination evinced a high capacity to prevent the development of drug resistance in the examined isolates. To the best of our knowledge, this is the first study to appraise the combination of colistin and Origanum majorana essential oil against colistinresistant isolates, including those carrying mcr-1 gene with the aim of regaining colistin susceptibility.

1Laboratorio de Investigaciones en Mecanismos de Resistencia a Antibio´ ticos, Instituto de Investigaciones en Microbiologı ´a y Parasitologı ´a Me´ dica, Facultad de Medicina, Universidad de Buenos Aires - Consejo Nacional de Investigaciones Cientı ´ficas y Tecnolo´ gicas (IMPaM, UBACONICET), Buenos Aires, Argentina, [ABSTRACT FROM AUTHOR]

ObjectiveTo provide technical support for the molecular surveillance of pathogenic bacteria strains carrying mobile colistin resistance-1 （mcr⁃1） gene isolate from inlet of wastewater treatment plants （WWTP）.MethodsThe Enterobacteriaceae strains carrying mcr⁃1 resistance gene isolate from inlet of WWTP during April 1 to June 30， 2023 in Shanghai were cultured on blood-rich and SS culture medium and were identified using a mass spectrometry analyzer. The mcr⁃1 gene and copy number were detected by real-time fluorescence quantitative PCR. Drug susceptibility test was performed by microbroth dilution method. The copy numbers of Escherichia coli carrying mcr⁃1 gene isolated from wastewater and human fecel were statistically analyzed by SPSS 25.0.ResultsA total of 14 strains carrying the mcr⁃1 gene were isolated from 49 WWTP samples， and the positive isolation rate was 28.6%， including 12 non-diarrheal E. coli strains and 2 Klebsiella pneumoniae strains. The drug susceptibility results showed that all 14 strains were multi-drug resistant bacteria. They were all sensitive to imipenem and tigecycline， but were ampicillin- and cefazolin-resistant. There was no significant difference in the copy number between human-sourced diarrheal E. coli and wastewater-sourced non-diarrheal E. coli （t=0.647， P>0.05）.ConclusionThe isolation and identification of strains carrying the mcr⁃1 gene from inlet of WWTP samples were firstly established in Shanghai. The multi-drug resistance among the isolated strains is severe. To effectively prevent and control the spread of colistin-resistant bacteria， more attention should be paid to the surveillance of mcr⁃1 gene.

Introduction: Hospitals are breeding grounds for multidrug-resistant (MDR) bacteria, posing treatment challenges and increasing the risk of spreading "superbugs." This study investigates the prevalence of colistinresistant bacteria, a last-resort antibiotic, in wastewater from tertiary hospitals in Enugu, Nigeria. Methods: Twenty wastewater samples were collected over three months from two tertiary hospitals in Enugu. A standardized protocol by the American Public Health Association (APHA) was followed. Samples were collected aseptically from key drainage points and transported to the lab within 2 hours. Bacteria were isolated using the pour-plate method and characterized by morphological and biochemical tests, including Catalase, Oxidase, and Glucose Fermentation. Antibiotic susceptibility was assessed using Kirby-Bauer disc diffusion, and colistin resistance was confirmed via broth microdilution. Multiplex PCR detected mcr genes indicating plasmid-mediated resistance. Data were analyzed using SPSS version 23 with Chi-Square and ANOVA tests at a significance level of P < 0.05. Results: Gram-negative bacteria were isolated from 63.1% of samples, with Klebsiella spp. being the most prevalent, accounting for 24.6%. Colistin resistance was phenotypically observed in E. coli (83%), Klebsiella spp. (75%), and Pseudomonas aeruginosa (100%). Genotypically, E. coli harbored mcr-1 (17%) and mcr-3 (83%), while all Klebsiella and Pseudomonas isolates carried multiple mcr genes. Additionally, these bacteria showed resistance to multiple antibiotics, including Septrin, Gentamycin, and Ceftriaxone. Conclusion: The significant presence of colistin-resistant bacteria, especially E. coli and Klebsiella, poses a public health concern, potentially leading to treatment failures and spreading resistance genes. Stricter monitoring of hospital wastewater is necessary to identify emerging resistance trends and improve antibiotic practices in hospitals. [ABSTRACT FROM AUTHOR]

Addressing the emergence of antimicrobial resistance (AMR) poses a significant challenge in veterinary and public health. In this study, we focused on determining the presence, phenotypic background, and genetic epidemiology of plasmid-mediated colistin resistance (mcr) in Escherichia coli bacteria isolated from camels farmed in the United Arab Emirates (UAE). Fecal samples were collected from 50 camels at a Dubai-based farm in the UAE and colistin-resistant Gram-negative bacilli were isolated using selective culture. Subsequently, a multiplex PCR targeting a range of mcr-genes, plasmid profiling, and whole-genome sequencing (WGS) were conducted. Eleven of fifty camel fecal samples (22%) yielded colonies positive for E. coli isolates carrying the mcr-1 gene on mobile genetic elements. No other mcr-gene variants and no chromosomally located colistin resistance genes were detected. Following plasmid profiling and WGS, nine E. coli isolates from eight camels were selected for in-depth analysis. E. coli sequence types (STs) identified included ST7, ST21, ST24, ST399, ST649, ST999, and STdaa2. Seven IncI2(delta) and two IncX4 plasmids were found to be associated with mcr-1 carriage in these isolates. These findings represent the first identification of mcr-1-carrying plasmids associated with camels in the Gulf region. The presence of mcr-1 in camels from this region was previously unreported and serves as a novel finding in the field of AMR surveillance. [ABSTRACT FROM AUTHOR]

Human consumption of antibiotics has increased their concentrations in many parts of the environment, including rivers, sediments, soil, and wastewater. Consequently, resistant bacteria originating from these environments are distributed to humans, resulting in illness. The aim of this study was to identify mobilized colistin-resistant (mcr) genes and quinolone-resistant (qnr) genes in E. coli strains obtained from clinical samples. Additionally, the study explored the impact of different radiation dosages on the expression of antibiotic-resistance genes. In this study, conducted in Beni-Suef, Egypt, samples from 430 community-acquired urinary tract infection (UTI) cases resulted in the isolation of 85 different strains of E. coli. Conventional microbiological procedures were employed to identify these bacterial isolates. Three bacterial isolates with resistance to both quinolones and colistin underwent examination for their corresponding genetic determinants, which subsequently proved the presence of their respective genes. Furthermore, the expression levels of the mcr-1 and qnr-S genes were assessed using real-time PCR after exposure to gamma irradiation. Remarkably, the use of a sublethal dosage of 3 kGy gamma irradiation treatment on bacterial cells increased their susceptibility to colistin and quinolones post-irradiation. Additionally, there was a notable reduction in the expression levels of both mcr-1 and qnr-S genes, which could be helpful for preventing the storage of antibiotic-resistant E. coli in the environment.

Human consumption of antibiotics has increased their concentrations in many parts of the environment, including rivers, sediments, soil, and wastewater. Consequently, resistant bacteria originating from these environments are distributed to humans, resulting in illness. The aim of this study was to identify mobilized colistin-resistant (mcr) genes and quinolone-resistant (qnr) genes in E. coli strains obtained from clinical samples. Additionally, the study explored the impact of different radiation dosages on the expression of antibiotic-resistance genes. In this study, conducted in Beni-Suef, Egypt, samples from 430 community-acquired urinary tract infection (UTI) cases resulted in the isolation of 85 different strains of E. coli. Conventional microbiological procedures were employed to identify these bacterial isolates. Three bacterial isolates with resistance to both quinolones and colistin underwent examination for their corresponding genetic determinants, which subsequently proved the presence of their respective genes. Furthermore, the expression levels of the mcr-1 and qnr-S genes were assessed using real-time PCR after exposure to gamma irradiation. Remarkably, the use of a sublethal dosage of 3 kGy gamma irradiation treatment on bacterial cells increased their susceptibility to colistin and quinolones post-irradiation. Additionally, there was a notable reduction in the expression levels of both mcr-1 and qnr-S genes, which could be helpful for preventing the storage of antibiotic-resistant E. coli in the environment. [ABSTRACT FROM AUTHOR]

There is scarce information concerning the role of sporadic clones in the dissemination of antimicrobial resistance genes (ARGs) within the nosocomial niche. We confirmed that the clinical Escherichia coli M19736 ST615 strain, one of the first isolates of Latin America that harbors a plasmid with an mcr-1 gene, could receive crucial ARG by transformation and conjugation using as donors critical plasmids that harbor blaCTX-M-15, blaKPC-2, blaNDM-5, blaNDM-1, or aadB genes. Escherichia coli M19736 acquired blaCTX-M-15, blaKPC-2, blaNDM-5, blaNDM-1, and aadB genes, being only blaNDM-1 maintained at 100% on the 10th day of subculture. In addition, when the evolved MDR-E. coli M19736 acquired sequentially blaCTX-M-15 and blaNDM-1 genes, the maintenance pattern of the plasmids changed. In addition, when the evolved XDR-E. coli M19736 acquired in an ulterior step the paadB plasmid, a different pattern of the plasmid’s maintenance was found. Interestingly, the evolved E. coli M19736 strains disseminated simultaneously the acquired conjugative plasmids in different combinations though selection was ceftazidime in all cases. Finally, we isolated and characterized the extracellular vesicles (EVs) from the native and evolved XDR-E. coli M19736 strains. Interestingly, EVs from the evolved XDR-E. coli M19736 harbored blaCTX-M-15 though the pDCAG1-CTX-M-15 was previously lost as shown by WGS and experiments, suggesting that EV could be a relevant reservoir of ARG for susceptible bacteria. These results evidenced the genetic plasticity of a sporadic clone of E. coli such as ST615 that could play a relevant transitional link in the clinical dynamics and evolution to multidrug/extensively/pandrug-resistant phenotypes of superbugs within the nosocomial niche by acting simultaneously as a vector and reservoir of multiple ARGs which later could be disseminated.

Background Antimicrobial resistance has become one of the most widespread threats to humans. Acinetobacter baumannii is one of the pathogens responsible for healthcare-associated infections (nosocomial). Colistin is considered the last resort antibiotic against infections with pandrug-resistant (PDR) pathogens. Results: Eleven isolates were detected phenotypically as PDR A. baumannii and were confirmed molecularly using 16S rDNA. The MCR-1 gene was not detected within the chromosomal DNA of the selected isolates. Plasmid bearing the MCR-1 gene was identified in 10 selected isolates of A. baumannii that had not been previously observed to carry the MCR-1 gene. Moreover, the use of colistin in combination with anionic antibiotics or natural compound pterostilbene poses a viable therapeutic alternative for PDR and revives colistin’s bactericidal effects on MCR-1-positive A. baumannii. Finally, the transmission electron microscopy studies proved the synergistic effect of these combinations and revealed the disruption of resistant A. baumannii’s outer membrane and alteration of the permeability properties that allowed overcoming the resistance of the isolates to colistin. Conclusions: Antimicrobial resistance of A. baumannii is related to the presence of the transferable plasmid-bearing MCR-1 gene. This study proved the ability of the combinations of colistin with anionic antibiotics and/or natural compound pterostilbene to restore the bactericidal effect of colistin. Overall, these combinations could be novel promising clinical alternatives against the increasing threat of the widespread multidrug-resistant A. baumannii.

Klebsiella pneumoniae is a nosocomial pathogen with high morbidity and mortality rates in hospitalized patients. The emergence of multidrug-resistant K. pneumoniae has become more challenging to treat, with the prevalence of colistin-resistance. Therefore, reliable methods for detecting colistin resistance are required. Many plants' essential oils have antimicrobial activity and have been used to combat multiple antibiotic resistances. This study aimed to investigate the characterization and prevalence of the colistin resistance gene mcr-1 in K. pneumoniae in Egypt, evaluate rapid polymyxin NP test, determine the transferability of mcr-1 gene, and study the synergistic activity of eugenol combined with colistin against K. pneumoniae isolates. Eighty-two K. pneumonia isolates were collected from different human samples, followed by antibiotic susceptibility testing, rapid polymyxin NP test, and detection of the mcr-1 gene and its transfer frequency. Determination of the MICs of colistin alone and in combination with eugenol was performed, then mcr-1 gene expression was determined in the presence of eugenol. Thirty-two isolates (39%) were colistin-resistant. Rapid polymyxin NP test failed to detect resistant isolates with MICs below 32 µg/mL. Detection of mcr-1 gene was made in 27 (84%) of colistin resistant isolates. The rest isolates possess alteration in the mgrB gene which probably causes colistin resistance. The mcr-1 gene was transferred by conjugation to other sensitive isolates. MIC of eugenol ranged from 416 to 1664 µg/mL, and FICI ranged from 0.265 to 0.75. Results also revealed suppression of mcr-1 gene expression in the presence of sub MIC of eugenol. Our results demonstrated a high prevalence of mcr-1 in Egypt and its ability to transfer to other strains. Difficult determination of colistin-resistant isolates with low values with rapid polymyxin NP test was apparent. Eugenol exerted a synergistic effect with colistin and improved its antimicrobial activity.

Addressing the emergence of antimicrobial resistance (AMR) poses a significant challenge in veterinary and public health. In this study, we focused on determining the presence, phenotypic background, and genetic epidemiology of plasmid-mediated colistin resistance (mcr) in Escherichia coli bacteria isolated from camels farmed in the United Arab Emirates (UAE). Fecal samples were collected from 50 camels at a Dubai-based farm in the UAE and colistin-resistant Gram-negative bacilli were isolated using selective culture. Subsequently, a multiplex PCR targeting a range of mcr-genes, plasmid profiling, and whole-genome sequencing (WGS) were conducted. Eleven of fifty camel fecal samples (22%) yielded colonies positive for E. coli isolates carrying the mcr-1 gene on mobile genetic elements. No other mcr-gene variants and no chromosomally located colistin resistance genes were detected. Following plasmid profiling and WGS, nine E. coli isolates from eight camels were selected for in-depth analysis. E. coli sequence types (STs) identified included ST7, ST21, ST24, ST399, ST649, ST999, and STdaa2. Seven IncI2(delta) and two IncX4 plasmids were found to be associated with mcr-1 carriage in these isolates. These findings represent the first identification of mcr-1-carrying plasmids associated with camels in the Gulf region. The presence of mcr-1 in camels from this region was previously unreported and serves as a novel finding in the field of AMR surveillance.

Purpose: The aim of this study was to investigate the presence of the mcr-1 gene, which is responsible for colistin resistance, in carbapenem-resistant Gram-negative bacteria that cause difficult-to-treat infections in a research hospital in Turkey. Materials and Methods: The mcr-1 gene was examined using PCR in 103 carbapenem-resistant isolates, including 75 Acinetobacter baumannii, 19 Pseudomonas aeruginosa, and 9 Klebsiella pneumoniae. DNA sequencing was performed to confirm the mcr-1 positivity. Other antimicrobial resistance genes were investigated in isolates that were found to be mcr-1-positive by PCR and colistinresistant isolates. Results: Four (3.9% of the 103 carbapenem-resistant isolates and 5.3% of the 75 A. baumannii isolates) A. baumannii isolates, all susceptible to colistin, were found to be mcr-1-positive by PCR, whereas mcr-1 was not detected in four colistin-resistant isolates, one in A. baumannii and three in K. pneumoniae. DNA sequencing analysis determined that none of the amplification products was the targeted fragment, but they matched more than 70% with the chromosomal DNA fragments of A. baumannii strains. Therefore, these results were considered false-positive. Although these false-positive isolates were susceptible to colistin, they were extensively drug-resistant (XDR). Two of them were found to carry blaOXA23-like and blaTEM genes, another blaOXA23- like, blaTEM and blaOXA48-like genes, and the fourth one to have blaOXA23-like and blaCTXM genes. Conclusion: Although the specificity of the primers used to detect the mcr-1 gene by PCR was reported as 100% in most studies, we concluded that PCR tests are insufficient yet to use alone or with antibiotic susceptibility tests in rapid routine diagnosis. Confirming at least PCR-positive samples using DNA sequence analysis would be appropriate for a certain period. [ABSTRACT FROM AUTHOR]

Few studies have addressed drug resistance of Enterobacterales in rural communities in developing countries. This study aimed to determine the coexistence of extended-spectrum β-lactamase (ESBL) and carbapenemase genes in Escherichia coli and Klebsiella pneumoniae strains carrying the mcr-1 gene in rural communities in Ecuador from healthy humans and their backyard animals. Sixty-two strains, thirty E. coli and thirty-two K. pneumoniae strains carrying the mcr-1 gene were selected from a previous study. PCR were performed for the presence of ESBLs and carbapenemase genes. The strains were further characterized, and the genetic relationship was studied with multi-locus sequencing typing (MLST) of seven housekeeping genes. Fifty-nine of the sixty-two mcr-1 isolates (95%) harbored at least on β-lactam resistance gene. The most prevalent ESBL genes were the blaTEM genes (present in in 80% of the E. coli strains) and the blaSHV gene (present in 84% of the K. pneumoniae strains). MSLT analysis revealed 28 different sequence types (ST); 15 for E. coli and 12 for K. pneumoniae, with most ST never described in humans and animals. The coexistence of mcr-1 and β-lactams resistant genes in E. coli and K. pneumoniae strains is alarming and threatens the efficacy of last-resort antibiotics. Our findings highlight backyard animals as a reservoir of mcr-1/β-lactams resistant genes. [ABSTRACT FROM AUTHOR]

Klebsiella pneumoniae is a nosocomial pathogen with high morbidity and mortality rates in hospitalized patients. The emergence of multidrug-resistant K. pneumoniae has become more challenging to treat, with the prevalence of colistin-resistance. Therefore, reliable methods for detecting colistin resistance are required. Many plants' essential oils have antimicrobial activity and have been used to combat multiple antibiotic resistances. This study aimed to investigate the characterization and prevalence of the colistin resistance gene mcr-1 in K. pneumoniae in Egypt, evaluate rapid polymyxin NP test, determine the transferability of mcr-1 gene, and study the synergistic activity of eugenol combined with colistin against K. pneumoniae isolates. Eighty-two K. pneumonia isolates were collected from different human samples, followed by antibiotic susceptibility testing, rapid polymyxin NP test, and detection of the mcr-1 gene and its transfer frequency. Determination of the MICs of colistin alone and in combination with eugenol was performed, then mcr-1 gene expression was determined in the presence of eugenol. Thirty-two isolates (39%) were colistin-resistant. Rapid polymyxin NP test failed to detect resistant isolates with MICs below 32 µg/mL. Detection of mcr-1 gene was made in 27 (84%) of colistin resistant isolates. The rest isolates possess alteration in the mgrB gene which probably causes colistin resistance. The mcr-1 gene was transferred by conjugation to other sensitive isolates. MIC of eugenol ranged from 416 to 1664 µg/mL, and FICI ranged from 0.265 to 0.75. Results also revealed suppression of mcr-1 gene expression in the presence of sub MIC of eugenol. Our results demonstrated a high prevalence of mcr-1 in Egypt and its ability to transfer to other strains. Difficult determination of colistin-resistant isolates with low values with rapid polymyxin NP test was apparent. Eugenol exerted a synergistic effect with colistin and improved its antimicrobial activity. [ABSTRACT FROM AUTHOR]

Resistance to colistin generated by the mcr-1 gene in Enterobacteriaceae is of great concern due to its efficient worldwide spread. Despite the fact that the Lima region has a third of the Peruvian population and more than half of the national pig and poultry production, there are no reports of the occurrence of the mcr-1 gene in Escherichia coli isolated from livestock. In the present work, we studied the occurrence of E. coli carrying the mcr-1 gene in chicken and pig farms in Lima between 2019 and 2020 and described the genomic context of the mcr-1 gene. We collected fecal samples from 15 farms in 4 provinces of Lima including the capital Lima Metropolitana and recovered 341 E. coli isolates. We found that 21.3% (42/197) and 12.5% (18/144) of the chicken and pig strains were mcr-1-positive by PCR, respectively. The whole genome sequencing of 14 mcr-1-positive isolates revealed diverse sequence types (e.g., ST48 and ST602) and the presence of other 38 genes that confer resistance to 10 different classes of antibiotics, including beta-lactamase blaCTX-M-55. The mcr-1 gene was located on diverse plasmids belonging to the IncI2 and IncHI1A:IncHI1B replicon types. A comparative analysis of the plasmids showed that they contained the mcr-1 gene within varied structures (mikB–mcr1–pap2, ISApl1–mcr1–pap2, and Tn6330). To the best of our knowledge, this is the first attempt to study the prevalence of the mcr-1 gene in livestock in Peru, revealing its high occurrence in pig and chicken farms. The genetic diversity of mcr-1-positive strains suggests a complex local epidemiology calling for a coordinated surveillance under the One-Health approach that includes animals, retail meat, farmers, hospitals and the environment to effectively detect and limit the spread of colistin-resistant bacteria. [ABSTRACT FROM AUTHOR]

Abstract Objectives This study aimed to explore mechanism of colistin resistance amongst Klebsiella pneumoniae isolates through plasmid mediated mcr-1 gene in Pakistan. Carbapenem and Colistin resistant K. pneumoniae isolates (n = 34) stored at − 80 °C as part of the Aga Khan University Clinical Laboratory strain bank were randomly selected and subjected to mcr-1 gene PCR. To investigate mechanisms of resistance, other than plasmid mediated mcr-1 gene, whole genome sequencing was performed on 8 clinical isolates, including 6 with colistin resistance (MIC > 4 μg/ml) and 2 with intermediate resistance to colistin (MIC > 2 μg/ml). Results RT-PCR conducted revealed absence of mcr-1 gene in all isolates tested. Whole genome sequencing results revealed modifications in Lipid A-Ara4N pathway. Modifications in Lipid A-Ara4N pathway were detected in ArnA_ DH/FT, UgdH, ArnC and ArnT genes. Mutation in ArnA_ DH/FT gene were detected in S3, S5, S6 and S7 isolates. UgdH gene modifications were found in all isolates except S3, mutations in ArnC were present in all except S1, S2 and S8 and ArnT were detected in all except S4 and S7. In the absence of known mutations linked with colistin resistance, lipid pathway modifications may possibly explain the phenotype resistance to colistin, but this needs further exploration.

Colistin resistance (Col-R) has been rising worldwide with high rate of morbidity and mortality. The emergence of mobile colistin resistance (mcr) harboring microorganisms become of a health concern. Hence, screening for fecal carriage of Col-R Enterobacteriaceae could aid in the prevention and control efforts of Col-R. This study aimed to screen for Col-R Enterobacteriaceae in the stool specimens of the hospitalized patients; explore for colistin minimal inhibitory concentrations (MIC) and the genetic determinants of these isolates, and to predict the risk factors among the studied patients' groups. Stool specimens from 290 hospitalized adult patients were screened for the presence of Col-R bacterial isolates using CHROMagar COL-APSE medium. Colistin MIC was estimated for Col-R Enterobacteriaceae using the broth microdilution (BMD) assay. Bacterial isolates were screened through the Polymerase chain reaction (PCR) for the existence of mcr-1 and mcr-2 genes. The fecal carriage of Col-R among the studied patients was 16.8 %. About 72 Col-R bacterial isolates were recovered. Col-R Enterobacteriaceae were predominant and were detected in 89.7 % of the bacterial isolates. Using the BMD, Col-R was confirmed and most of the isolates showed low resistance MIC titer (4 μg/ ml; 55.7 %). In addition, mcr-1 gene was the most frequent Col-R gene detected (69.2 %), while mcr-2 gene was less prevalent (11.5 %). The current study reported high prevalence of the Col-R and mcr-1 gene harbored by the fecal flora; with the risk to be easily transmitted inside the hospitals and within the different communities. This highlights the need for active surveillance in addition to the infection control programs

Colistin (polymyxin E) is a bactericidal antibiotic used to treat severe infections caused by multidrug-resistant Gram-negative bacteria. The product of the mcr1 gene generates transferable plasmid-mediated colistin resistance, which has arisen as a worldwide health-care problem. This study aimed to isolate and identify colistin-resistant bacteria, and evaluate the ability of essential oils in its fights. Twenty-seven bacterial isolates were collected from patients who were admitted to the National Cancer Institute, Cairo, Egypt, and processed using standard microbiological methods. Essential oils were purchased from AB Chem Company, Egypt, screened for antibacterial, cytotoxic activity, and (GC-MS) analysis. A total of 5 bacterial isolates were resistant to colistin with minimum inhibitory concentration (MIC) ranging from 6.25->200 µg/ml. Cinnamon oil exhibited the highest activity against colistin-resistant strains followed by thyme and eucalyptus oil. The (MIC) of cinnamon oils against resistant strains ranged from 4.88 to 312.5 µg/ml. Moreover, mcr-1 gene expression was extremely down-regulated after the treatment of bacterial strains with cinnamon oil and decreased to 20–35-fold. Examination of treated bacterial cells with sub-inhibitory concentrations under transmission electron microscopy showed various abnormalities occurred in most of these cells. Cinnamon oil exhibits antibacterial activity against colistin-resistant strains, showing it as a promising natural alternative in clinical therapy. [ABSTRACT FROM AUTHOR]

Background: Actually, no data on the prevalence of plasmid colistin resistance in Tunisia are available among clinical bacteria. Objectives: This study aimed to investigate the current epidemiology of colistin resistance and the spread of the mcr gene in clinical Gram-negative bacteria (GNB) isolated from six Tunisian university hospitals. Methods: A total of 836 GNB strains were inoculated on COL-R agar plates with selective screening agar for the isolation of GNB resistant to colistin. For the selected isolates, mcr genes, beta-lactamases associated-resistance genes and molecular characterisation were screened by PCRs and sequencing. Results: Colistin-resistance was detected in 5.02% (42/836) of the isolates and colistin-resistant isolates harboured an ESBL (blaCTX-M-15) and/or a carbapenemase (blaOXA-48, blaVIM) encoding gene in 45.2% of the cases. The mcr-1 gene was detected in four E. coli isolates (0.59%) causing urinary tract infections and all these isolates also contained the blaTEM-1 gene. The blaCTX-M-15 gene was detected in three isolates that also carried the IncY and IncFIB replicons. The genetic environment surrounding the mcr-carrying plasmid indicated the presence of pap-2 gene upstream mcr-1 resistance marker with unusual missing of ISApl1 insertion sequence. The Conclusions: This study reports the first description of the mcr-1 gene among clinical E. coli isolates in Tunisia and provides an incentive to conduct routine colistin susceptibility testing in GNB clinical isolates. [ABSTRACT FROM AUTHOR]

Few studies have addressed drug resistance of Enterobacterales in rural communities in developing countries. This study aimed to determine the coexistence of extended-spectrum β-lactamase (ESBL) and carbapenemase genes in Escherichia coli and Klebsiella pneumoniae strains carrying the mcr-1 gene in rural communities in Ecuador from healthy humans and their backyard animals. Sixty-two strains, thirty E. coli and thirty-two K. pneumoniae strains carrying the mcr-1 gene were selected from a previous study. PCR were performed for the presence of ESBLs and carbapenemase genes. The strains were further characterized, and the genetic relationship was studied with multi-locus sequencing typing (MLST) of seven housekeeping genes. Fifty-nine of the sixty-two mcr-1 isolates (95%) harbored at least on β-lactam resistance gene. The most prevalent ESBL genes were the blaTEM genes (present in in 80% of the E. coli strains) and the blaSHV gene (present in 84% of the K. pneumoniae strains). MSLT analysis revealed 28 different sequence types (ST); 15 for E. coli and 12 for K. pneumoniae, with most ST never described in humans and animals. The coexistence of mcr-1 and β-lactams resistant genes in E. coli and K. pneumoniae strains is alarming and threatens the efficacy of last-resort antibiotics. Our findings highlight backyard animals as a reservoir of mcr-1/β-lactams resistant genes.

ABSTRACT: Plasmid-mediated polymyxin resistance was first described in 2015, in China, in Escherichia coli carrying the mcr-1 (Mobile Colistin Resistance-1) gene. Since then, it has become a major public health challenge worldwide, representing a major threat to human and animal health. In addition, there are still few reports on the prevalence of mcr-1 in Enterobacteriaceae isolated from humans, animals and food. Therefore, the purpose of the study was to investigate the occurrence of the mcr-1 gene in bacterial isolates with phenotypic resistance to polymyxin B obtained from clinical specimens of companion animals. Phenotypic resistance to polymyxin B were determined by broth microdilution and the susceptibility profile to other antimicrobials (amikacin, amoxicillin/clavulanate, ampicillin, ampicillin/sulbactam, aztreonam, cefazolin, cefepime, cefotaxime, cefoxitin, ceftazidime, ceftriaxone, chloramphenicol, ciprofloxacin, doxycycline, ertapenem, gentamicin, imipenem, marbofloxacin, meropenem, phosphomycin, piperacillin/tazobactam, tetracycline, ticarcillin/clavulanate, tobramycin and trimethoprim/sulfamethoxazole) by disc-diffusion agar method. The extraction of bacterial DNA was performed via heat shock followed by spectrophotometric evaluation. To verify the presence of mcr-1, the Polymerase Chain Reaction was employed using specific primers, followed by agarose gel electrophoresis. The positive isolates had the corresponding amplicons sequenced. In this study, there were identified the first isolates of Escherichia coli, Klebsiella spp. and Enterobacter spp. carrying the mcr-1 gene derived from specimens of companion animals in Brazil. Our results suggest the dissemination of resistance to polymyxins in the community and the environment, highlighting the need for surveillance and optimized treatment guidelines.

Objectives: This study aimed to explore mechanism of colistin resistance amongst Klebsiella pneumoniae isolates through plasmid mediated mcr-1 gene in Pakistan. Carbapenem and Colistin resistant K. pneumoniae isolates (n = 34) stored at − 80 °C as part of the Aga Khan University Clinical Laboratory strain bank were randomly selected and subjected to mcr-1 gene PCR. To investigate mechanisms of resistance, other than plasmid mediated mcr-1 gene, whole genome sequencing was performed on 8 clinical isolates, including 6 with colistin resistance (MIC > 4 μg/ml) and 2 with intermediate resistance to colistin (MIC > 2 μg/ml). Results: RT-PCR conducted revealed absence of mcr-1 gene in all isolates tested. Whole genome sequencing results revealed modifications in Lipid A-Ara4N pathway. Modifications in Lipid A-Ara4N pathway were detected in ArnA_ DH/FT, UgdH, ArnC and ArnT genes. Mutation in ArnA_ DH/FT gene were detected in S3, S5, S6 and S7 isolates. UgdH gene modifications were found in all isolates except S3, mutations in ArnC were present in all except S1, S2 and S8 and ArnT were detected in all except S4 and S7. In the absence of known mutations linked with colistin resistance, lipid pathway modifications may possibly explain the phenotype resistance to colistin, but this needs further exploration. [ABSTRACT FROM AUTHOR]

Aims: Antimicrobial resistance (AMR) spreads not only by pathogenic but also by commensal bacteria, and the latter can become a reservoir for resistance genes. This study was aimed to investigate the AMR patterns along with the presence of mobilized colistin resistance (mcr) genes in commensal Escherichia coli circulating in chickens, farm environments, street foods, and human patients. Materials and Methods: By a cross-sectional survey, isolates obtained from 530 samples were tested for their AMR profiles against 9 antimicrobials. Minimum inhibitory concentration (MIC) of the phenotypically colistin-resistant isolates was determined and screened for a set of mcr genes followed by sequencing of mcr-1 gene in the multidrug-resistant (MDR) isolates. Results: A total of 313 E. coli strains were isolated and confirmed by polymerase chain reaction. Antimicrobial susceptibility testing revealed that about 98% (confidence interval [95% CI] 95–99) of the isolates were MDR, and 58% (95% CI 52–63) isolates exhibited resistance to colistin. MIC values of colistin against the isolates ranged from 4 to 64 mg/L. Except for human patients, 20.4% colistin-resistant isolates from other sources of isolation had mcr-1 gene. Conclusions: There is abundance of commensal MDR E. coli strains with the acquisition of mcr-1 gene circulating in chickens and farm environments in Bangladesh. [ABSTRACT FROM AUTHOR]

Colistin is considered as the last resort for treatment of bacterial infections caused by carbapenem-resistant Gram-negative bacilli (GNB). Colistin resistance can increase due to the spread of plasmid-borne mcr-1 gene. This study aimed to determine colistin susceptibility and to detect the presence of mcr-1 gene in the clinical isolates of GNB recovered from different clinical samples collected from Ain Shams University Hospital, Cairo, Egypt. About thirty-five GNB isolates were recovered from the different clinical samples that were collected during the period from February-April, 2019. These isolates were subjected to antibiotic susceptibility testing using disc diffusion assay, and Colistin susceptibility through the E-test. In addition, conventional Polymerase chain reaction (PCR) was carried out for detection of mcr-1 gene among the colistin GNB resistant isolates. Most of the GNB isolates (60 %) were recovered from blood samples. Klebsiella pneumoniae (K. pneumoniae) was the most common isolated bacterium; that was represented by 24 isolates (68%). Out of the 35 GNB, only five isolates (14.3 %) were resistant to colistin by E-test, with MIC >256 𝜇g/ ml. The mcr-1 gene was detected only in one Pseudomonas aeruginosa (P. aeruginosa) isolate. This study concluded the low frequency of mcr-1 gene among the current GNB isolates. However, a large scale study is recommended to detect colistin resistance among GNB.

Colistin is considered the last option for treatment of infections caused by Multidrug Resistant (MDR) Gram negative pathogens. The mcr-1 gene could transfer the resistance to colistin between different species. Therefore, screening of this resistance mechanism will help greatly in the control of further spread of colistin resistance and enhancing the outcomes of patients. The current work aimed to study the frequency of colistin resistance among MDR Gram negative pathogens isolated from clinical samples at Ain Shams University Hospitals and to explore if the mcr-1 gene was the responsible mechanism for this resistance. These pathogens were isolated from various samples including; blood, pleural fluid, sputum, urine, swabs from surgical and burn wounds, that were submitted from wards and intensive care units to the central microbiology laboratory at Ain Shams University Hospital, Cairo, Egypt, during the period from June to December 2019. Culture and bacterial identification were done by conventional microbiologic techniques. Eighty Gram negative bacterial pathogens were assayed for antimicrobial susceptibility by disc diffusion test. Sixty MDR Gram negative isolates were identified and further studied for colistin susceptibility by E- test, as well as real- time PCR to detect mcr-1 gene. Totally four isolates (6.7%) were phenotypically resistant to colistin. We found mcr-1 gene in three of the obtained isolates (5%); 1 Pseudomonas aeruginosa (5.3%) and 2 Acinetobacter baumannii (14.3%). In conclusion, although we detected a low prevalence of mcr-1 positive isolates from human clinical samples, continuous monitoring and implementation of infection control precautions are greatly required, to interfere with the further occurrence and transfer of bacterial species carrying the mcr-1 gene.

Resistance to colistin generated by the mcr-1 gene in Enterobacteriaceae is of great concern due to its efficient worldwide spread. Despite the fact that the Lima region has a third of the Peruvian population and more than half of the national pig and poultry production, there are no reports of the occurrence of the mcr-1 gene in Escherichia coli isolated from livestock. In the present work, we studied the occurrence of E. coli carrying the mcr-1 gene in chicken and pig farms in Lima between 2019 and 2020 and described the genomic context of the mcr-1 gene. We collected fecal samples from 15 farms in 4 provinces of Lima including the capital Lima Metropolitana and recovered 341 E. coli isolates. We found that 21.3% (42/197) and 12.5% (18/144) of the chicken and pig strains were mcr-1-positive by PCR, respectively. The whole genome sequencing of 14 mcr-1-positive isolates revealed diverse sequence types (e.g., ST48 and ST602) and the presence of other 38 genes that confer resistance to 10 different classes of antibiotics, including beta-lactamase blaCTX-M-55. The mcr-1 gene was located on diverse plasmids belonging to the IncI2 and IncHI1A:IncHI1B replicon types. A comparative analysis of the plasmids showed that they contained the mcr-1 gene within varied structures (mikB–mcr1–pap2, ISApl1–mcr1–pap2, and Tn6330). To the best of our knowledge, this is the first attempt to study the prevalence of the mcr-1 gene in livestock in Peru, revealing its high occurrence in pig and chicken farms. The genetic diversity of mcr-1-positive strains suggests a complex local epidemiology calling for a coordinated surveillance under the One-Health approach that includes animals, retail meat, farmers, hospitals and the environment to effectively detect and limit the spread of colistin-resistant bacteria.

Background: Actually, no data on the prevalence of plasmid colistin resistance in Tunisia are available among clinical bacteria. Objectives: This study aimed to investigate the current epidemiology of colistin resistance and the spread of the mcr gene in clinical Gram-negative bacteria (GNB) isolated from six Tunisian university hospitals. Methods: A total of 836 GNB strains were inoculated on COL-R agar plates with selective screening agar for the isolation of GNB resistant to colistin. For the selected isolates, mcr genes, beta-lactamases associated-resistance genes and molecular characterisation were screened by PCRs and sequencing. Results: Colistin-resistance was detected in 5.02% (42/836) of the isolates and colistin-resistant isolates harboured an ESBL (blaCTX-M-15) and/or a carbapenemase (blaOXA-48, blaVIM) encoding gene in 45.2% of the cases. The mcr-1 gene was detected in four E. coli isolates (0.59%) causing urinary tract infections and all these isolates also contained the blaTEM-1 gene. The blaCTX-M-15 gene was detected in three isolates that also carried the IncY and IncFIB replicons. The genetic environment surrounding the mcr-carrying plasmid indicated the presence of pap-2 gene upstream mcr-1 resistance marker with unusual missing of ISApl1 insertion sequence. The Conclusions: This study reports the first description of the mcr-1 gene among clinical E. coli isolates in Tunisia and provides an incentive to conduct routine colistin susceptibility testing in GNB clinical isolates.

The emergence of the plasmid-mediated colistin resistance gene mcr-1 is threatening the last-line role of colistin in human medicine. With mcr-1-positive Escherichia coli (E. coli) isolated from food animal being frequently reported in China, the prevalence of mcr-1 in food animal has attracted public attention. In the present study, a total of 105 colistin-resistant E. coli strains were isolated from 200 fecal samples collected from six swine farms in northeastern China. mcr-PCR revealed that the prevalence of mcr-1 in colistin-resistant E. coli was 53.33% (56/105). mcr-1-positive E. coli showed extensive antimicrobial resistance profiles with the presence of additional resistance genes, increased expression of multidrug efflux pump-associated genes, and increased biofilm formation ability. MLST differentiated all the mcr-1-positive E. coli into 25 sequence types (STs) and five unknown ST, and the most common ST was ST10 (n = 11). By phylogenetic group classification, the distribution of all mcr-1-positive E. coli belonging to groups A, B1, B2, and D was 46.43, 35.71, 5.36, and 5.36%, respectively. Conjugation experiment demonstrated that most of the mcr-1 were transferable at frequencies of 2.68 × 10–6–3.73 × 10–3 among 30 representative mcr-1-positive E. coli. The plasmid replicon types IncI2 (n = 9), IncX4 (n = 5), IncHI2 (n = 3), IncN (n = 3), and IncP (n = 1) were detected in the transconjugants. The results of growth assay, competition experiment, and plasmid stability testing showed that acquisition of mcr-1-harboring plasmids could reduce the fitness of bacterial hosts, but mcr-1 remained stable in the recipient strain. Due to the potential possibility of these mcr-1-positive E. coli being transmitted to humans through the food chain or through horizontal transmission, therefore, it is necessary to continuously monitor the prevalence and dissemination of mcr-1 in food animal, particularly in swine.

The emergence of the plasmid-mediated colistin resistance gene mcr-1 is threatening the last-line role of colistin in human medicine. With mcr-1 -positive Escherichia coli (E. coli) isolated from food animal being frequently reported in China, the prevalence of mcr-1 in food animal has attracted public attention. In the present study, a total of 105 colistin-resistant E. coli strains were isolated from 200 fecal samples collected from six swine farms in northeastern China. mcr -PCR revealed that the prevalence of mcr-1 in colistin-resistant E. coli was 53.33% (56/105). mcr-1 -positive E. coli showed extensive antimicrobial resistance profiles with the presence of additional resistance genes, increased expression of multidrug efflux pump-associated genes, and increased biofilm formation ability. MLST differentiated all the mcr-1- positive E. coli into 25 sequence types (STs) and five unknown ST, and the most common ST was ST10 (n = 11). By phylogenetic group classification, the distribution of all mcr-1 -positive E. coli belonging to groups A, B1, B2, and D was 46.43, 35.71, 5.36, and 5.36%, respectively. Conjugation experiment demonstrated that most of the mcr-1 were transferable at frequencies of 2.68 × 10–6–3.73 × 10–3 among 30 representative mcr - 1- positive E. coli. The plasmid replicon types IncI2 (n = 9), IncX4 (n = 5), IncHI2 (n = 3), IncN (n = 3), and IncP (n = 1) were detected in the transconjugants. The results of growth assay, competition experiment, and plasmid stability testing showed that acquisition of mcr-1- harboring plasmids could reduce the fitness of bacterial hosts, but mcr-1 remained stable in the recipient strain. Due to the potential possibility of these mcr-1 -positive E. coli being transmitted to humans through the food chain or through horizontal transmission, therefore, it is necessary to continuously monitor the prevalence and dissemination of mcr-1 in food animal, particularly in swine. [ABSTRACT FROM AUTHOR]

The occurrence and spreading of multi-resistant, as well as pan-resistant bacterial isolates, presents a global problem of modern medicine. Narrowed therapeutic options actualized the use of "old" antibiotics such as polymyxins. The use of polymyxins in human medicine has been reduced since the 1960s due to the discovery of safer antibiotics. Modern researches provided a better understanding of their pharmacokinetics and pharmacodynamics, as well as dosing regimens with minimum side effects. However, the increased usage in therapy consequently led to the occurrence of resistance to these "last-line" antibiotics. Most of polymyxin resistant bacterial isolates carried chromosomally mediated resistance, but the discovery of the plasmid mcr-1 gene in 2015 in China changed the paradigm of the origin and spreading of polymyxin resistance. Animals are the main reservoirs of bacteria carrying plasmid with the mcr-1 gene, because of widespread polymyxins application in veterinary medicine and in food industry, as well. Many studies confirmed the transfer of polymyxins resistance genes from animal bacterial isolates to human isolates, as well as between different bacterial species in vivo or in vitro. These findings indicated the need for more detailed epidemiological research and surveillance, as the European Centre for Disease Prevention and Control recommended.

Colistin is one of the most effective antibiotics against multidrug resistant Gram-negative bacteria. However, the recent emergence of plasmid-borne mobilized colistin resistance (mcr) genes is considered a serious antimicrobial resistance challenge worldwide. In this study, we report detection of an mcr-1 carrying Escherichia coli isolate (named ATAVET mcr-1 Turkey) from retail raw chicken meat in Turkey. Of the 11 (from 500 total tested) phenotypically colistin-resistant isolates, 1 was shown to carry the mcr-1 gene by PCR. Whole-genome sequencing indicated that mcr-1 was located on a ∼13 kb-long contig that was almost identical to the corresponding part in pZJ1635, an IncI2 plasmid encoding mcr-1 in the same genetic context in another E. coli strain. In addition, ATAVET mcr-1 Turkey harbored blaCTX-M-8, qnrB19, mdf(A), tet(A), sul2, aph(3″)-Ib, aph(6)-Id, and floR resistance genes. Phylogenetic analysis based on whole genome and multilocus sequence typing indicated that ATAVET mcr-1 Turkey was more closely related to mcr-1 carrying E. coli isolates from food and human clinical samples previously reported from different parts of the world than to those from Turkey. These findings further emphasize the worldwide emergence and spread of mcr meditated colistin resistance in bacteria with zoonotic potential within animals and the food chain. [ABSTRACT FROM AUTHOR]

Introduction: Gram-negative isolates harboring mobilized colistin resistance (mcr-1) gene are a great threat to human health. They have been reported worldwide among various bacterial isolates. This work aimed to study the prevalence of colistin resistance among Gram-negative bacteria and the incidence of mcr-1 gene among these isolates. Methods: A descriptive cross-sectional study was done at a tertiary care center from June 2016 to February 2017. An ethical approval was taken from review board of the Nepal Health Research Council (Reg. no: 274/2016). Convenience sampling was used. The data was collected and analyzed using Microsoft Excel 2010 and Statistical Package for Social Sciences (SPSS) Version 16 . Point estimate at 95% Confidence Interval was calculated along with frequency and proportion for binary data. Results: Among 485 gram-negative isolates, only 13 (2.68%) (1.26-6.62 at 95% Confidence Interval) isolates were colistin-resistant and mcr-1 was present in two isolates. Predominant colistin-resistant isolates were E. coli 6 (4.1%), Enterobacter spp 2 (2.81%), and Acinetobacter spp 2 (2.81%). A high level of colistin-resistance was noted in 4 (30.7%) isolates as indicated by the very high value of colistin MIC (>256 µg/ml). ICU was the major site of isolation of colistin-resistant and mcr-1 positive pathogens. The majority of colistin-resistant isolates were highly drug-resistant and were sensitive only to polymyxin B. Antibiotics like imipenem, amikacin, gentamicin, aztreonam, ciprofloxacin, and piperacillin-tazobactam were effective for few of these isolates. Conclusions: Though the prevalence of mcr-1 gene was low among colistin-resistant gram-negative isolates, the resistant pattern was quite alarming as these isolates were highly drug-resistant.

Background: Colistin is the last choice for serious infections caused by multidrug-resistant Gram negative bacteria and one of the prominent causes for spreading the resistance is Plasmid-borne Mobile Colistin Resistance (mcr). Broth microdilution method (BMD) is the reference tool for colistin minimum inhibitory concentrations (MIC) determination, but it has many obstacles, so commercial BMD methods had been developed that are more userfriendly than the reference method and (Liofilchem ® ComASPTM) is one of them, which we used to determine colistin MIC in this study. Objective: To detect colistin resistant Gram negative bacilli (GNB) by ComASPTM colistin (formerly Sensi Test™Colistin) among Intensive Care Units (ICUs) patients admitted to Ain Shams university hospitals and to screen the presence of mcr-1 gene by Polymerase Chain Reaction (PCR) in Colistin resistant isolates. Method: This Observational cross-sectional study was performed in the Medical Microbiology and Immunology Department, Faculty of Medicine, Ain Shams University between June 2019 to November 2019. One hundred isolates of Gram negative bacilli were obtained from patients admitted at different ICUs of Ain Shams University Hospitals. Full identification was done by conventional microbiological methods, Then MIC was measured for all isolated organisms by using commercial BMD ComASPTM colistin, PCR was done for colistin resistant isolates to detect mcr-1 gene. Results: 60% of the GNB isolates were K.pneumoniea. Colistin resistance was 14% among 100 GNB, 35.7% of these colistin resistant were K.pneumoniea obtained from urine samples. Prevalence of mcr-1 gene was 7.1%. Conclusion: Commercial BMD ComASPTM colistin is simple and uncomplicated method for detection colistin susceptibility. [ABSTRACT FROM AUTHOR]

There is scarce information concerning the role of sporadic clones in the dissemination of antimicrobial resistance genes (ARGs) within the nosocomial niche. We confirmed that the clinical Escherichia coli M19736 ST615 strain, one of the first isolates of Latin America that harbors a plasmid with an mcr-1 gene, could receive crucial ARG by transformation and conjugation using as donors critical plasmids that harbor bla CTX-M-15 , bla KPC-2 , bla NDM-5 , bla NDM-1 , or aadB genes. Escherichia coli M19736 acquired bla CTX-M-15 , bla KPC-2 , bla NDM-5 , bla NDM-1 , and aadB genes, being only blaNDM-1 maintained at 100% on the 10th day of subculture. In addition, when the evolved MDR- E. coli M19736 acquired sequentially bla CTX-M-15 and bla NDM-1 genes, the maintenance pattern of the plasmids changed. In addition, when the evolved XDR- E. coli M19736 acquired in an ulterior step the paadB plasmid, a different pattern of the plasmid's maintenance was found. Interestingly, the evolved E. coli M19736 strains disseminated simultaneously the acquired conjugative plasmids in different combinations though selection was ceftazidime in all cases. Finally, we isolated and characterized the extracellular vesicles (EVs) from the native and evolved XDR- E. coli M19736 strains. Interestingly, EVs from the evolved XDR- E. coli M19736 harbored bla CTX-M-15 though the pDCAG1-CTX-M-15 was previously lost as shown by WGS and experiments, suggesting that EV could be a relevant reservoir of ARG for susceptible bacteria. These results evidenced the genetic plasticity of a sporadic clone of E. coli such as ST615 that could play a relevant transitional link in the clinical dynamics and evolution to multidrug/extensively/pandrug-resistant phenotypes of superbugs within the nosocomial niche by acting simultaneously as a vector and reservoir of multiple ARGs which later could be disseminated., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The author(s) declared that they were an editorial board member of Frontiers, at the time of submission. This had no impact on the peer review process and the final decision., (Copyright © 2024 Carrera Páez, Olivier, Gambino, Poklepovich, Aguilar, Quiroga and Centrón.)

Zoonotic and antimicrobial-resistant Escherichia coli (hereafter, E. coli) is a global public health threat which can lead to detrimental effects on human health. Here, we aim to investigate the antimicrobial resistance and the presence of mcr-1 gene in E. coli isolated from chicken feces. Ninety-four E. coli isolates were obtained from samples collected from different locations in Bangladesh, and the isolates were identified using conventional microbiological tests. Phenotypic disk diffusion tests using 20 antimicrobial agents were performed according to CLSI-EUCAST guidelines, and minimum inhibitory concentrations (MICs) were determined for a subset of samples. E. coli isolates showed high resistance to colistin (88.30%), ciprofloxacin (77.66%), trimethoprim/sulfamethoxazole (76.60%), tigecycline (75.53%), and enrofloxacin (71.28%). Additionally, the pathotype eaeA gene was confirmed in ten randomly selected E. coli isolates using primer-specific polymerase chain reaction (PCR). The presence of mcr-1 gene was confirmed using PCR and sequencing analysis in six out of ten E. coli isolates. Furthermore, sequencing and phylogenetic analyses revealed a similarity between the catalytic domain of Neisseria meningitidis lipooligosaccharide phosphoethanolamine transferase A (LptA) and MCR proteins, indicating that the six tested isolates were colistin resistant. Finally, the findings of the present study showed that E. coli isolated from chicken harbored mcr-1 gene, and multidrug and colistin resistance. These findings accentuate the need to implement strict measures to limit the imprudent use of antibiotics, particularly colistin, in agriculture and poultry farms.

Abstract Background Bacterial infections have become more challenging to treat due to the emergence of multidrug-resistant pathogenic bacteria. Combined antibiotics prove to be a relatively effective method to control such resistant strains. This study aim to investigate synergistic activity of eugenol combined with colistin against a collection of clinical isolated Escherichia coli (E.coli) strains, and to evaluate potential interaction. Methods Antimicrobial susceptibility, minimum inhibitory concentration (MIC) and fractional inhibitory concentration index (FICI) of the bacteria were determined by disk diffusion assay, broth microdilution method and checkerboard assay, respectively. The mcr-1 mRNA expression was measured by Real-time PCR. To predict possible interactions between eugenol and MCR-1, molecular docking assay was taken. Results For total fourteen strains including eight colistin-resistant strains, eugenol was determined with MIC values of 4 to 8 μg/mL. Checkerboard dilution test suggested that eugenol exhibited synergistic activity when combined with colistin (FICI ranging from 0.375 to 0.625). Comparison analysis of Real-time PCR showed that synergy could significantly down-regulate expression of mcr-1 gene. A metal ion coordination bond with catalytic zinc atom and a hydrogen bond with crucial amino acid residue Ser284 of MCR-1 were observed after molecular docking, indicating antibacterial activity and direct molecular interactions of eugenol with MCR-1 protein. Conclusions Our results demonstrated that eugenol exhibited synergistic effect with colistin and enhanced its antimicrobial activity. This might further contribute to the antibacterial actions against colistin-resistant E.coli strains. Graphical abstract Synergistic effect of eugenol with colistin against colistin-resistant Escherichia coli isolates.

Purpose: The recurrence of multi-drug resistant (MDR) pathogens to the latest antibiotics and the limited development of new antibacterial agents have reduced the options for the treatment of severe infections. The reintroduction of old antibiotics, such as colistin, represents an effective strategy, since the latest antibiotics are over-consumed and ineffective against MDR pathogens. In 2015, Liu (Lancet Infect Dis 16:161–168, 2016) reported Escherichia coli (E. coli) isolates carrying plasmid-mediated colistin resistance gene mcr-1. The first of mcr-1 positive colistin-resistant (col-R) E. coli from a human blood culture was observed in 2012 in Latin America, while in Italy was reported for the first time by our center in 2016. The present study aimed to describe the prevalence of mcr-1 positive col-R strains in E. coli-related bloodstream infection among patients hospitalized in Fondazione IRCCS Policlinico San Matteo in Pavia, Italy, from 2012 to 2018, including the three cases already published. Methods: All col-R E. coli strains isolated from blood cultures collected during the study period were analyzed. The minimal inhibitory concentration of colistin was determined using broth microdilution and detection of mcr-1 and mcr-2 genes was performed by PCR. The sequence type of E. coli mcr-1 positive was determined according to Multilocus sequence typing. Results: Out of 1557 samples, 14 strains (0.90%) were col-R. and positive for the presence of the mcr-1 gene, with no mcr-2 detected. The most common ST was ST10 (n = 3), followed by ST410 (n = 2). The remaining strains exhibited different MLST profiles, indicating that they were genetically unrelated. Conclusions: Proper reporting of the presence of mcr-1 genes is an essential component to anticipate the spread of colistin resistance. This public health issue is particularly alarming in Italy due to the consistent circulation of MDR bacteria. [ABSTRACT FROM AUTHOR]

In recent years, colistin has been the drug of choice for treatment of nosocomial infections, especially in bloodstream infections, lower respiratory tract infections, or urinary tract infections. In this study, 65 multidrug-resistant Klebsiella pneumoniae isolated from different clinical samples were included. Minimum inhibitory concentration (MIC) of colistin was detected by broth microdilution method in three different ways. For selected K. pneumoniae strains, eazyplex SuperBug mcr-1 test was performed. This test detects mcr-1 gene, which encodes a colistin-resistance determinant. Most of the analyzed K. pneumoniae strains were resistant to colistin in all applied methods. The exception was two strains, where MIC of colistin was 2 mg/L in SensiTest Colistin and MIC-Strip Colistin tests. In MIC COL test, MIC for these strains was 4 mg/L. All analyzed strains produced extended-spectrum beta-lactamases and 11 (16.9%) metallo-beta-lactamases. Eleven (16.9%) K. pneumoniae strains were resistant to all antibiotics, whereas 17 (26.1%) were susceptible to only one drug. Colistin MIC values varied from 2 to >64 mg/L in MIC-Strip Colistin test; from 2 to >16 mg/L in SensiTest Colistin and from 4 to >16 mg/L in MIC COL test. None of the analyzed K. pneumoniae strains carried mcr-1 gene. Data of this work suggest that resistance to colistin emerged among multidrug-resistant K. pneumoniae strains. The tests allowed for reliable estimation of susceptibility to colistin and could be used in microbiological diagnostics. [ABSTRACT FROM AUTHOR]

An antibiotic susceptibility monitoring programme was conducted from 2004 to 2010, resulting in a collection of 143 Escherichia coli cultured from bovine faecal samples (diarrhoea) and milk-aliquots (mastitis). The isolates were subjected to whole-genome sequencing and were distributed in phylogroups A, B1, B2, C, D, E, and G with no correlation for particular genotypes with pathotypes. In fact, the population structure showed that the strains belonging to the different phylogroups matched broadly to ST complexes; however, the isolates are randomly associated with the diseases, highlighting the necessity to investigate the virulence factors more accurately in order to identify the mechanisms by which they cause disease. The antimicrobial resistance was assessed phenotypically, confirming the genomic prediction on three isolates that were resistant to colistin, although one isolate was positive for the presence of the gene mcr-1 but susceptible to colistin. To further characterise the genomic context, the four strains were sequenced by using a single-molecule long read approach. Genetic analyses indicated that these four isolates harboured complex and diverse plasmids encoding not only antibiotic resistant genes (including mcr-1 and bla) but also virulence genes (siderophore, ColV, T4SS). A detailed description of the plasmids of these four E. coli strains, which are linked to bovine mastitis and diarrhoea, is presented for the first time along with the characterisation of the predicted antibiotic resistance genes. The study highlighted the diversity of incompatibility types encoding complex antibiotic resistance elements such as Tn6330, ISEcp1, Tn6029, and IS5075. The mcr-1 resistance determinant was identified in IncHI2 plasmids pCFS3273-1 and pCFS3292-1, thus providing some of the earliest examples of mcr-1 reported in Europe, and these sequences may be a representative of the early mcr-1 plasmidome characterisation in the EU/EEA.

We identified discrete importation events of the mcr-1 gene on incompatibility group IncI2 plasmids in Escherichia coli isolated from patients in New South Wales, Australia, in 2011 and 2013. mcr-1 is present in a small minority of colistin-resistant Enterobacteriaceae and appears not to be established locally.

We detected the mcr-1 gene in 21 (14.8%) Salmonella isolates from pigs at slaughter; 19 were serovar Typhimurium sequence type 34. The gene was located on IncHI2-like plasmids that also harbored IncF replicons and lacked a conjugative transfer region. These findings highlight the need to prevent further spread of colistin resistance in animals and humans.

Colistin is a last resort antibiotic used for the treatment of human infections associated with carbapenemase-producing Enterobacteriales. Here, we evaluated the occurrence of mcr-1 and -2 plasmid-mediated colistin resistance in colistin and/or carbapenem resistant human clinical Enterobacteriales and other gram-negative bacteria (n = 543) as well as third generation cephalosporin-resistant (3GCR) Escherichia coli isolates from poultry abattoir workers (n = 15) and poultry fecal samples (n = 92) collected from two geographically separate abattoirs in Romania. which revealed that mcr-1 was present within four sequence types (STs): ST744 (n = 7), ST57 (n = 7), ST156 (n = 2), and ST10 (n = 1). Within STs, serotypes were conserved and, notably, all except one of the mcr-1-positive isolates were found to exhibit fluoroquinolone-resistance (FQR) associated SNPs in both gyrA and parC. While there were variations in genotypes, all isolates belonging to ST744, ST57, and ST156 were rich in resistance determinants, carrying aminoglycoside-modifying enzymes genes, sulfonamide resistance gene blaTEM–1 as well as blaCMY–2 AmpC β-lactamase resistance genes. They also exhibited high similarity in carriage of virulence genes; ST10, however, only carried the mcr-1 gene. Whole genome sequencing (WGS) analysis also revealed that although the mcr-1 gene was identified in a diverse population of E. coli, two STs (ST57 and ST744) predominated and interestingly, were found in isolates across both abattoirs providing evidence for clonal transmission. Also, two main genomic contexts of mcr-1 isolates were revealed with all ST57 isolates harboring the mcr-1 gene between two copies of ISApl1 (or the Tn6330 transposon) whilst a common mcr-1 containing scaffold, highly similar to IncX type mcr-1-bearing plasmids (pWI2-mcr, Accession number: LT838201), was present among mcr-1 isolates of varying phylogenetic backgrounds (ST10, ST744 and ST156). The high prevalence of the mcr-1 gene in poultry E. coli isolates with co-resistance to cephalosporins and quinolones, in a country where antimicrobial use in food production species is poorly regulated, is concerning and the findings from this study should lead to better surveillance of antimicrobial resistance (AMR) in food-production animals in Romania.

Present study characterized five Escherichia coli co-expressing ESBL and MCR-1 recovered from food, food-producing animals, and companion animals in China. Antimicrobial susceptibility tests, conjugation experiments, and plasmid typing were performed. Whole genome sequencing (WGS) was undertaken for all five isolates using either PacBio RS II or Illumina HiSeq 2500 platforms. The cefotaxime and colistin resistance encoded by blaCTX–M and mcr-1 genes, respectively, was transferable by conjugation either together or separately for all five strains. Interestingly, the ESBL and mcr-1 genes could be co-selected by cefotaxime, while the colistin only selected the mcr-1-carrying plasmids during the conjugation experiments. Five E. coli sequence types (ST88, ST93, ST602, ST162, and ST457) were detected. Although diverse plasmid profiles were identified, IncI2, IncFIB, and IncFII plasmid types were predominant. These five clonally unrelated isolates harbored the mcr-1 gene located on similar plasmid backbones, which showed high nucleotide similarity to plasmid pHNSHP45. The mcr-1 gene can be co-transmitted with blaCTX–M genes through IncI2 plasmids with or without ISApl1 in our study. Characterization of these co-existence ESBL and mcr-1 isolates extends our understanding on the dissemination of these resistance markers among bacteria of diverse origins.

Pseudomonas aeruginosa has caused high rates of mortality due to the appearance of strains with multidrug resistance (MDR) profiles. This study aimed to characterize the molecular profile of virulence and resistance genes in 99 isolates of P. aeruginosa recovered from different clinical specimens. The isolates were identified by the automated method Vitek2, and the antibiotic susceptibility profile was determined using different classes of antimicrobials. The genomic DNA was extracted and amplified by multiplex polymerase chain reaction (mPCR) to detect different virulence and antimicrobial resistance genes. Molecular typing was performed using the enterobacterial repetitive intergenic consensus (ERIC-PCR) technique to determine the clonal relationship among P. aeruginosa isolates. The drug susceptibility profiles of P. aeruginosa for all strains showed high levels of drug resistance, particularly, 27 (27.3%) isolates that exhibited extensively drug-resistant (XDR) profiles, and the other isolates showed MDR profiles. We detected the polymyxin E (mcr-1) gene in one strain that showed resistance against colistin. The genes that confer resistance to oxacillin (blaOXA-23 and blaOXA-51) were present in three isolates. One of these isolates carried both genes. As far as we know from the literature, this is the first report of the presence of blaOXA-23 and blaOXA-51 genes in P. aeruginosa.

Colistin is a last resort antibiotic used for the treatment of human infections associated with carbapenemase-producing Enterobacteriales. Here, we evaluated the occurrence of mcr-1 and -2 plasmid-mediated colistin resistance in colistin and/or carbapenem resistant human clinical Enterobacteriales and other gram-negative bacteria (n = 543) as well as third generation cephalosporin-resistant (3GCR) Escherichia coli isolates from poultry abattoir workers (n = 15) and poultry fecal samples (n = 92) collected from two geographically separate abattoirs in Romania. which revealed that mcr-1 was present within four sequence types (STs): ST744 (n = 7), ST57 (n = 7), ST156 (n = 2), and ST10 (n = 1). Within STs, serotypes were conserved and, notably, all except one of the mcr-1- positive isolates were found to exhibit fluoroquinolone-resistance (FQR) associated SNPs in both gyrA and parC. While there were variations in genotypes, all isolates belonging to ST744, ST57, and ST156 were rich in resistance determinants, carrying aminoglycoside-modifying enzymes genes, sulfonamide resistance gene bla TEM–1 as well as bla CMY–2 AmpC β-lactamase resistance genes. They also exhibited high similarity in carriage of virulence genes; ST10, however, only carried the mcr-1 gene. Whole genome sequencing (WGS) analysis also revealed that although the mcr-1 gene was identified in a diverse population of E. coli , two STs (ST57 and ST744) predominated and interestingly, were found in isolates across both abattoirs providing evidence for clonal transmission. Also, two main genomic contexts of mcr-1 isolates were revealed with all ST57 isolates harboring the mcr-1 gene between two copies of IS Apl1 (or the Tn 6330 transposon) whilst a common mcr-1 containing scaffold, highly similar to IncX type mcr-1 -bearing plasmids (pWI2-mcr, Accession number: LT838201), was present among mcr-1 isolates of varying phylogenetic backgrounds (ST10, ST744 and ST156). The high prevalence of the mcr-1 gene in poultry E. coli isolates with co-resistance to cephalosporins and quinolones, in a country where antimicrobial use in food production species is poorly regulated, is concerning and the findings from this study should lead to better surveillance of antimicrobial resistance (AMR) in food-production animals in Romania. [ABSTRACT FROM AUTHOR]

There is emerging evidence that food of animal origin may be responsible for the spread of multidrug resistant extraintestinal pathogenic Escherichia coli in the community. Here, we describe the emergence of colistin resistance gene, mcr-1 , in a strain belonging to the dominant uropathogenic E. coli ST69 lineage. E. coli strain CFSAN061770 was isolated during monitoring of the popular Egyptian raw milk cheese, karish cheese, for the presence of colistin resistance. The complete genome of E. coli strain CFSAN061770 comprises a chromosome of 5,292,297 bp with a G + C content of 50.6%. Further, three plasmids named pEGY1-MCR-1, pEGY2 and pEGY3 of 228,947 bp, 103,234 bp and 87,012 bp were detected, respectively. Plasmid pEGY1-MCR-1 belongs to the IncHI2 incompatibility group and carries the colistin resistance mcr-1 gene flanked by two I SApl1 elements and forms a composite transposon. It mediates resistance to aminoglycosides (aadA1 and aadA2), phenicol (cmlA1 and floR), sulfonamides (sul3), and tetracycline (tet (A)), and these loci were found clustered in a multidrug resistant region. Plasmid pEGY3 carries a complex multiple resistance locus (CMR) (aph(3′)-Ia, strA, strB, sul2 , and bla TEM-1) encoding resistance to different classes of antibiotics. Interestingly, the closest plasmids to plasmid pEGY1-MCR-1 detected from the NCBI Blast search belonged to the incompatibility group IncHI2 and were from the Kingdom of Saudi Arabia and Qatar which suggests a dissemination of pEGY1-MCR-1-like plasmids in the Middle East. Most striking, and of great public health concern is that strain CFSAN061770 carries five virulence genes (iss, fimH, iutA, kpsMIII and kpsTIII) which were identified in clinical extraintestinal pathogenic E. coli. Besides that, it carries the astA gene, which codes for the enteroaggregative E. coli heat-stable toxin 1 (EAST1). • Uropathogenic E. coli ST69 lineage strain CFSAN061770 was isolated from raw milk cheese. • Strain CFSAN061770 carries multiple antimicrobial resistance genes located on plasmids. • It carries virulence genes which were identified in extraintestinal pathogenic E. coli. [ABSTRACT FROM AUTHOR]

Present study characterized five Escherichia coli co-expressing ESBL and MCR-1 recovered from food, food-producing animals, and companion animals in China. Antimicrobial susceptibility tests, conjugation experiments, and plasmid typing were performed. Whole genome sequencing (WGS) was undertaken for all five isolates using either PacBio RS II or Illumina HiSeq 2500 platforms. The cefotaxime and colistin resistance encoded by bla CTX–M and mcr-1 genes, respectively, was transferable by conjugation either together or separately for all five strains. Interestingly, the ESBL and mcr-1 genes could be co-selected by cefotaxime, while the colistin only selected the mcr-1 -carrying plasmids during the conjugation experiments. Five E. coli sequence types (ST88, ST93, ST602, ST162, and ST457) were detected. Although diverse plasmid profiles were identified, IncI2, IncFIB, and IncFII plasmid types were predominant. These five clonally unrelated isolates harbored the mcr-1 gene located on similar plasmid backbones, which showed high nucleotide similarity to plasmid pHNSHP45. The mcr-1 gene can be co-transmitted with bla CTX–M genes through IncI2 plasmids with or without IS Apl1 in our study. Characterization of these co-existence ESBL and mcr-1 isolates extends our understanding on the dissemination of these resistance markers among bacteria of diverse origins. [ABSTRACT FROM AUTHOR]

Bacterial infections in immunocompromised patients are associated with a high mortality and morbidity rate. In this high-risk group, the presence of multidrug-resistant (MDR) bacteria, particularly bacteria that harbor a transferable antibiotic resistance gene, complicates the management of bacterial infections. In this study, we investigated the presence of the transferable colistin resistance mcr genes in patients with leukemia in Spain. 217 fecal samples collected in 2013–2015 from 56 patients with acute leukemia and colonized with MDR Enterobacteriaceae strains, were screened on September 2017 for the presence of the colistin resistance mcr genes (mcr-1 to -5) by multiplex PCR. mcr positive strains selected on LBJMR and MacConkey supplemented with colistin (2 μg/ml) media were phenotypically and molecularly characterized by antimicrobial susceptibility testing, minimum inhibitory concentration, multilocus sequence typing and plasmid characterization. Among 217 fecal samples, 5 samples collected from 3 patients were positive for the presence of the mcr-1 colistin-resistance gene. Four Escherichia coli strains were isolated and exhibited resistance to colistin with MIC = 4 μg/ml. Other genes conferring the resistance to β-lactam antibiotics have also been identified in mcr-1 positive strains, including bla TEM-206 and bla TEM-98. Three different sequence types were identified, including ST1196, ST140 and ST10. Plasmid characterization allowed us to detect the mcr-1 colistin resistance gene on conjugative IncP plasmid type. To the best of our knowledge, we have identified the mcr-1 gene for the first time in leukemia patients in Spain. In light of these results, strict measures have been implemented to prevent its dissemination. [ABSTRACT FROM AUTHOR]

The antimicrobial susceptibility profiles of 13 Salmonella enterica isolated from chicken eggs from various areas of Metro Manila, Philippines were determined. Results showed that Salmonella isolates had the highest resistance to erythromycin (92.31%), doxycycline (53.85%), sulphamethoxazole--trimethoprim (53.85%), tetracycline (46.15%), streptomycin (38.46%) and spectinomycin (38.46%); and highest antibiotic susceptibilities to amikacin (92.31%), ciprofloxacin (84.62%), gentamicin (84.62%), kanamycin (84.62%), norfloxacin (84.62%), amoxicillin clavulanic acid (76.92%), neomycin (69.23%) and ofloxacin (61.54%) using the standard disk diffusion method. Multidrug resistance was observed in eight (61.54%) isolates, in which two (15.38%) isolates were resistant to at least 10 different antimicrobials and one (7.69%) isolate was resistant to at least eight different classes of antimicrobials tested in the study. Based on WHO classification system, high antimicrobial susceptibilities were observed for most of the Highest Priority Critically Important Antimicrobials (61.54%-84.62%) and High Priority Critically Important Antimicrobials (69.23%- 92.31%), however, low antimicrobial susceptibilities were observed for Highly Important Antimicrobials (0 to 46.15%) and Important Antimicrobials (38.46%). PCR analysis showed that all isolates were negative for the mcr-1 resistance gene. Data obtained in this study may serve as a guide for poultry clinicians and farm owners in their selection of effective antimicrobials against Salmonella and formulation of more effective control programs against Salmonella infection in layer chickens. [ABSTRACT FROM AUTHOR]