Antimicrobial Resistance Mechanisms Research Articles

Effects of removing in-feed antibiotics and zinc oxide on the taxonomy and functionality of the microbiota in post weaning pigs

Abstract Background Post weaning diarrhoea (PWD) causes piglet morbidity and mortality at weaning and is a major driver for antimicrobial use worldwide. New regulations in the EU limit the use of in-feed antibiotics (Ab) and therapeutic zinc oxide (ZnO) to prevent PWD. New approaches to control PWD are needed, and understanding the role of the microbiota in this context is key. In this study, shotgun metagenome sequencing was used to describe the taxonomic and functional evolution of the faecal microbiota of the piglet during the first two weeks post weaning within three experimental groups, Ab, ZnO and no medication, on commercial farms using antimicrobials regularly in the post weaning period. Results Diversity was affected by day post weaning (dpw), treatment used and diarrhoea but not by the farm. Microbiota composition evolved towards the dominance of groups of species such as Prevotella spp. at day 14dpw. ZnO inhibited E. coli overgrowth, promoted higher abundance of the family Bacteroidaceae and decreased Megasphaera spp. Animals treated with Ab exhibited inconsistent taxonomic changes across time points, with an overall increase of Limosilactobacillus reuteri and Megasphaera elsdenii. Samples from non-medicated pigs showed virulence-related functions at 7dpw, and specific ETEC-related virulence factors were detected in all samples presenting diarrhoea. Differential microbiota functions of pigs treated with ZnO were related to sulphur and DNA metabolism, as well as mechanisms of antimicrobial and heavy metal resistance, whereas Ab treated animals exhibited functions related to antimicrobial resistance and virulence. Conclusion Ab and particularly ZnO maintained a stable microbiota composition and functionality during the two weeks post weaning, by limiting E. coli overgrowth, and ultimately preventing microbiota dysbiosis. Future approaches to support piglet health should be able to reproduce this stable gut microbiota transition during the post weaning period, in order to maintain optimal gut physiological and productive conditions.

Biofilm inhibition/eradication: exploring strategies and confronting challenges in combatting biofilm.

Biofilms are complex communities of microorganisms enclosed in a self-produced extracellular matrix, posing a significant threat to different sectors, including healthcare and industry. This review provides an overview of the challenges faced due to biofilm formation and different novel strategies that can combat biofilm formation. Bacteria inside the biofilm exhibit increased resistance against different antimicrobial agents, including conventional antibiotics, which can lead to severe problems in livestock and animals, including humans. In addition, biofilm formation also imposes heavy economic pressure on industries. Hence it becomes necessary to explore newer alternatives to eradicate biofilms effectively without applying selection pressure on the bacteria. Excessive usage of antibiotics may also lead to an increase in the number of resistant strains as bacteria employ an advanced antimicrobial resistance mechanism. This review provides insight into multifaceted technologies like quorum sensing inhibition, enzymes, antimicrobial peptides, bacteriophage, phytocompounds, and nanotechnology to neutralize biofilms without developing antimicrobial resistance (AMR). Furthermore, it will pave the way for developing newer therapeutic agents to deal with biofilms more efficiently.

AMR mechanisms in L. interrogans serovars: a comprehensive study

Antimicrobial resistance (AMR) is one of the global health challenges of the 21st century. Data regarding AMR mechanisms in Leptospira interrogans, the causative agents of leptospirosis, have been relatively limited. Therefore, our study aimed to identify resistance genes and explore potential resistance mechanisms specific to particular serovars. We conducted a comprehensive analysis of 98 Leptospira strains, representing 10 common serovars, using whole-genome sequencing (WGS) FASTA files. Employing the PATRIC tool from the Bacterial and Viral Bioinformatics Resource Center (BV-BRC), we scrutinized the genomes for AMR genes. Our investigation revealed 32 genes associated with AMR, with 20 key genes consistently prevalent across most strains. Notably, we identified unique efflux pump systems in serovar Pomona, indicating distinctive resistance mechanisms in this serovar. In summary, our findings shed light on the genetic landscape of AMR in Leptospira, uncovering both common and serovar-specific resistance elements. The presence of unique efflux pump systems in serovar Pomona introduces a novel dimension to our understanding of resistance mechanisms. These insights underscore the importance of tailored intervention strategies and collaborative efforts between human and veterinary healthcare professionals, as well as environmental scientists, to address the complex dynamics of leptospirosis and its implications for antibiotic resistance.

A hypothesis-based hop microbiology laboratory module testing the plausibility of the mythical origin of the India Pale Ale.

As one of the most famous fermented drinks in the world, beer is an especially relatable topic for microbiology courses. Here, we describe a short and easily adaptable module based on the antibacterial properties of hops used in brewing. By the 15th century, beer recipes included hops (the flower of the Humulus lupulus plant) as a bittering agent and antimicrobial. By the 19th century, the highly hopped Indian Pale Ale (IPA) became popular, and a modern myth has emerged that IPAs were invented to survive long ocean voyages such as from Britain to India. With that myth in mind, we designed a hypothesis-driven microbiology lab module that tests the plausibility of this brewing myth-namely that highly hopped beers possess enough antibacterial activity to prevent spoilage, while lowly hopped beers do not. The overall design of the module is to test the antimicrobial properties of hops using petri plates containing varying concentrations of hop extract. The module includes hypothesis generation and testing related to bacterial physiology and cell envelope morphology (hops are not equally effective against Gram-positive and Gram-negative bacteria) and to mechanisms of antimicrobial resistance (as beer spoilage bacteria have repeatedly evolved hop resistance). Pre- and post-assessment showed that students made significant gains in the learning objectives for the module, which encourages critical thinking and hypothesis testing by linking microbial physiology and antimicrobial resistance to an important and topical real-world application.

Antimicrobial Resistance Mechanisms of Gram-Positive and Gram-Negative Bacteria

The discovery of antibiotics has long helped humans in the battle against bacteria. However, the misuse of antibiotics in industries and medical systems has unintentionally provided an ideal environment for bacteria to develop resistance mechanisms through mutations and gene transfer, resulting in the emergence of antimicrobial-resistant bacteria. This has proven to be an urgent and pressing issue in the global healthcare system, resulting in increased mortality. Therefore, increasing resources are invested to study their different resistance mechanisms and develop corresponding novel drugs and treatment methods. This review briefly introduces several key resistance mechanisms with examples from both Gram-negative and Gram-positive bacteria, the current and novel methods for treating multiple drug-resistant bacteria as well as the potential actions that could be done to improve the situation.

Structural and kinetic analysis of the monofunctional Staphylococcus aureus PBP1

Staphylococcus aureus, an ESKAPE pathogen, is a major clinical concern due to its pathogenicity and manifold antimicrobial resistance mechanisms. The commonly used β-lactam antibiotics target bacterial penicillin-binding proteins (PBPs) and inhibit crosslinking of peptidoglycan strands that comprise the bacterial cell wall mesh, initiating a cascade of effects leading to bacterial cell death. S. aureus PBP1 is involved in synthesis of the bacterial cell wall during division and its presence is essential for survival of both antibiotic susceptible and resistant S. aureus strains. Here, we present X-ray crystallographic data for S. aureus PBP1 in its apo form as well as acyl-enzyme structures with distinct classes of β-lactam antibiotics representing the penicillins, carbapenems, and cephalosporins, respectively: oxacillin, ertapenem and cephalexin. Our structural data suggest that the PBP1 active site is readily accessible for substrate, with little conformational change in key structural elements required for its covalent acylation of β-lactam inhibitors. Stopped-flow kinetic analysis and gel-based competition assays support the structural observations, with even the weakest performing β-lactams still having comparatively high acylation rates and affinities for PBP1. Our structural and kinetic analysis sheds insight into the ligand–PBP interactions that drive antibiotic efficacy against these historically useful antimicrobial targets and expands on current knowledge for future drug design and treatment of S. aureus infections.

Comparative analysis of antimicrobial resistance phenotype and genotype of Riemerella anatipestifer

Riemerella anatipestifer is one of the important bacterial pathogens that threaten the waterfowl farming industry. In this study, 157 suspected R. anatipestifer strains were isolated from diseased ducks and geese from seven regions of China during 2019–2020, and identified using multiple polymerase chain reaction (PCR). Antimicrobial susceptibility tests and whole-genome sequence (WGS) analysis were then performed for comparative analysis of antimicrobial resistance phenotypes and genotypes. The results showed that these strains were susceptible to florfenicol, ceftriaxone, spectinomycin, sulfafurazole and cefepime, but resistant to kanamycin, amikacin, gentamicin, and streptomycin, exhibiting multiple antimicrobial resistance phenotypes. WGS analysis revealed a wide distribution of genotypes among the 157 strains with no apparent regional pattern. Through next-generation sequencing analysis of antimicrobial resistance genes, a total of 88 resistance genes were identified. Of them, 19 tetracycline resistance genes were most commonly found, followed by 15 efflux pump resistance genes, 11 glycopeptide resistance genes and seven macrolide resistance genes. The 157 R. anatipestifer strains contained 42–55 resistance genes each, with the strains carrying 47 different resistance genes being the most abundant. By comparing the antimicrobial resistance phenotype and genotype, it was observed that a high correlation between them for most antimicrobial resistance properties was detected, except for a difference in aminoglycoside resistance phenotype and genotype. In conclusion, 157 R. anatipestifer strains exhibited severe multiple antimicrobial resistance phenotypes and genotypes, emphasizing the need for improved antimicrobial usage guidelines. The wide distribution and diverse types of resistance genes among these strains provide a foundation for studying novel mechanisms of antimicrobial resistance.

Upregulation of outer membrane porin gene ompC contributed to enhancement of azithromycin susceptibility in multidrug-resistant Escherichia coli.

The outer membrane (OM) in gram-negative bacteria contains proteins that regulate the passive or active uptake of small molecules for growth and cell function, as well as mediate the emergence of antibiotic resistance. This study aims to explore the potential mechanisms for restoring bacteria to azithromycin susceptibility based on transcriptome analysis of bacterial membrane-related genes. Transcriptome sequencing was performed by treating multidrug-resistant Escherichia coli T28R with azithromycin or in combination with colistin and confirmed by reverse transcription-quantitative PCR (RT-qPCR). Azithromycin enzyme-linked immunosorbent assay (ELISA) test, ompC gene overexpression, and molecular docking were utilized to conduct the confirmatory research of the potential mechanisms. We found that colistin combined with azithromycin led to 48 differentially expressed genes, compared to azithromycin alone, such as downregulation of tolA, eptB, lpxP, and opgE and upregulation of ompC gene. Interestingly, the addition of colistin to azithromycin differentially downregulated the mph(A) gene mediating azithromycin resistance, facilitating the intracellular accumulation of azithromycin. Also, overexpression of the ompC elevated azithromycin susceptibility, and colistin contributed to further suppression of the Mph(A) activity in the presence of azithromycin. These findings suggested that colistin firstly enhanced the permeability of bacterial OM, causing intracellular drug accumulation, and then had a repressive effect on the Mph(A) activity along with azithromycin. Our study provides a novel perspective that the improvement of azithromycin susceptibility is related not only to the downregulation of the mph(A) gene and conformational remodeling of the Mph(A) protein but also the upregulation of the membrane porin gene ompC.IMPORTANCEUsually, active efflux via efflux pumps is an important mechanism of antimicrobial resistance, such as the AcrAB-TolC complex and MdtEF. Also, bacterial porins exhibited a substantial fraction of the total number of outer membrane proteins in Enterobacteriaceae, which are involved in mediating the development of the resistance. We found that the upregulation or overexpression of the ompC gene contributed to the enhancement of resistant bacteria to azithromycin susceptibility, probably due to the augment of drug uptakes caused and the opportunity of Mph(A) function suppressed by azithromycin with colistin. Under the combination of colistin and azithromycin treatment, OmpC exhibited an increased selectivity for cationic molecules and played a key role in the restoral of the antibiotic susceptibility. Investigations on the regulation of porin expression that mediated drug resistance would be important in clinical isolates treated with antibiotics.

Molecular epidemiological and antimicrobial-resistant mechanisms analysis of prolonged Neisseria gonorrhoeae collection between 1971 and 2005 in Japan.

As antimicrobial-resistant (AMR) Neisseria gonorrhoeae strains have emerged, humans have adjusted the antimicrobials used to treat infections. We identified shifts in the N. gonorrhoeae population and the determinants of AMR strains isolated during the recurring emergence of resistant strains and changes in antimicrobial therapies. We examined 243 N. gonorrhoeae strains corrected at the Kanagawa Prefectural Institute of Public Health, Kanagawa, Japan, these isolated in 1971-2005. We performed multilocus sequence typing and AMR determinants (penA, mtrR, porB, ponA, 23S rRNA, gyrA and parC) mainly using high-throughput genotyping methods together with draft whole-genome sequencing on the MiSeq (Illumina) platform. All 243 strains were divided into 83 STs. ST1901 (n = 17) was predominant and first identified after 2001. Forty-two STs were isolated in the 1970s, 34 in the 1980s, 22 in the 1990s and 13 in the 2000s, indicating a decline in ST diversity over these decades. Among the 29 strains isolated after 2001, 28 were highly resistant to ciprofloxacin (MIC ≥ 8 mg/L) with two or more amino-acid substitutions in quinolone-resistance-determining regions. Seven strains belonging to ST7363 (n = 3), ST1596 (n = 3) and ST1901 (n = 1) were not susceptible to cefixime, and six strains carried penA alleles with mosaic-like penicillin-binding protein 2 (PBP2; penA 10.001 and 10.016) or PBP2 substitutions A501V and A517G. We observed a significant reduction in the diversity of N. gonorrhoeae over 35 years in Japan. Since 2001, ST1901, which is resistant to ciprofloxacin, has superseded previous strains, becoming the predominant ST population.

Antimicrobial Resistance: Techniques to Fight AMR in Bacteria – A Review

Antimicrobials or antibiotics were the important revelations of the last century, however, it came along with a silent curse that people care less to talk about. Antimicrobial resistance (AMR) which emerged alongside antibiotics in the last century has been a significant concern for scientists and policymakers. Since their discovery, it has been noted that the widespread use of antibiotics is the primary cause of bacteria developing antimicrobial drug resistance. Despite the recognition of this issue, it is challenging to curtail the widespread use of antibiotics because they are essential for treating various infections. Paradoxically, the necessity of using these drugs becomes an inadvertent advantage for bacteria to evolve resistance mechanisms. This dilemma creates a seeming stalemate in our battle against these tiny microorganisms. Delaying action could have dire consequences, potentially leading to the emergence of stronger superbugs that pose a serious threat to the entire human population. The recent COVID-19 pandemic serves as a stark reminder of the devastating impact a small microbe can have on global health. This paper delves into the mechanisms of antimicrobial resistance in bacteria, the evolution of superbugs and the innovative techniques employed by scientists to combat these challenges. Taking proactive steps is crucial to avoid a future where we are at the mercy of increasingly resilient microbes.

New Approaches to Overcoming Antimicrobial Resistance in Endophthalmitis.

Endophthalmitis is a rare but vision-threatening infection characterized by marked inflammation of intraocular fluids and tissues, uncommonly seen following surgery and intravitreal injection. Antimicrobials are used worldwide in the prophylaxis and treatment of bacterial and fungal infections of the eye and are standard treatment in the preoperative and postoperative care of surgical patients. However, antimicrobials are reported to be overprescribed in many parts of the world, which contributes to antimicrobial resistance (AMR). AMR complicates the prophylaxis and treatment of endophthalmitis. This article examines the prevalence and mechanisms of AMR in ocular microorganisms, emphasizing the importance of understanding AMR patterns for tailored treatments. It also explores prophylaxis and management strategies for endophthalmitis, with a discussion on the use of intracameral antibiotic administration. The use of prophylactic intracameral antibiotics during cataract surgery is common in many parts of the world but is still controversial in some locations, especially in the US. Finally, it highlights the role of stewardship in ophthalmology and its benefits in the treatment of endophthalmitis.

Molecular Aspects of the Functioning of Pathogenic Bacteria Biofilm Based on Quorum Sensing (QS) Signal-Response System and Innovative Non-Antibiotic Strategies for Their Elimination.

One of the key mechanisms enabling bacterial cells to create biofilms and regulate crucial life functions in a global and highly synchronized way is a bacterial communication system called quorum sensing (QS). QS is a bacterial cell-to-cell communication process that depends on the bacterial population density and is mediated by small signalling molecules called autoinducers (AIs). In bacteria, QS controls the biofilm formation through the global regulation of gene expression involved in the extracellular polymeric matrix (EPS) synthesis, virulence factor production, stress tolerance and metabolic adaptation. Forming biofilm is one of the crucial mechanisms of bacterial antimicrobial resistance (AMR). A common feature of human pathogens is the ability to form biofilm, which poses a serious medical issue due to their high susceptibility to traditional antibiotics. Because QS is associated with virulence and biofilm formation, there is a belief that inhibition of QS activity called quorum quenching (QQ) may provide alternative therapeutic methods for treating microbial infections. This review summarises recent progress in biofilm research, focusing on the mechanisms by which biofilms, especially those formed by pathogenic bacteria, become resistant to antibiotic treatment. Subsequently, a potential alternative approach to QS inhibition highlighting innovative non-antibiotic strategies to control AMR and biofilm formation of pathogenic bacteria has been discussed.

Genomic insights into an extensively drug-resistant and hypervirulent Burkholderia dolosa N149 isolate of a novel sequence type (ST2237) from a Vietnamese patient hospitalised for stroke

Burkholderia dolosa is a clinically important opportunistic pathogen in inpatients. Here we characterised an extensively drug-resistant and hypervirulent B. dolosa isolate from a patient hospitalised for stroke. Resistance to 41 antibiotics was tested with the agar disc diffusion, minimum inhibitory concentration, or broth microdilution method. The complete genome was assembled using short-reads and long-reads and the hybrid de novo assembly method. Allelic profiles obtained by multilocus sequence typing were analysed using the PubMLST database. Antibiotic-resistance and virulence genes were predicted in silico using public databases and the 'baargin' workflow. B. dolosa N149 phylogenetic relationships with all available B. dolosa strains and Burkholderia cepacia complex strains were analysed using the pangenome obtained with Roary. B. dolosa N149 displayed extensive resistance to 31 antibiotics and intermediate resistance to 4 antibiotics. The complete genome included three circular chromosomes (6 338 630 bp in total) and one plasmid (167 591 bp). Genotypic analysis revealed various gene clusters (acr, amr, amp, emr, ade, bla and tet) associated with resistance to 35 antibiotic classes. The major intrinsic resistance mechanisms were multidrug efflux pump alterations, inactivation and reduced permeability of targeted antibiotics. Moreover, 91 virulence genes (encoding proteins involved in adherence, formation of capsule, biofilm and colony, motility, phagocytosis inhibition, secretion systems, protease secretion, transmission and quorum sensing) were identified. B. dolosa N149 was assigned to a novel sequence type (ST2237) and formed a mono-phylogenetic clade separated from other B. dolosa strains. This study provided insights into the antimicrobial resistance and virulence mechanisms of B. dolosa.

Reimagining Chitosan‐Based Antimicrobial Biomaterials to Mitigate Antibiotic Resistance and Alleviate Antibiotic Overuse: A Review

AbstractThis paper reports recent advancements in chitosan‐based antimicrobial biomaterials that aim to address the issue of antimicrobial resistance (AMR). Today, AMR is one of the most significant public health challenges the world is facing. To combat the overuse of antibiotics, a range of materials, including advanced nanosized polymers, have been explored as potential antimicrobial agents. In this direction, chitosan scores highly because of its unique combination of properties such as biodegradability, biocompatibility, bioactivity, non‐toxic, and, most importantly, its intrinsic antibacterial and antifungal activity. Herein, an overview of chitosan and its derivatives as potential alternatives to conventional antimicrobial therapeutics is reported. This review starts with understanding the AMR mechanism, which is critical for developing suitable materials for treating infectious diseases. Then, the inherent characteristics, modes of antimicrobial action, and factors that impact the antimicrobial effectiveness of chitosan and its derivatives are discussed. The authors further summarize various clinical trial results of chitosan‐based materials as antimicrobial agents. Finally, various antimicrobial applications of chitosan‐based materials are reported. In summary, this review offers new insights into the design of chitosan‐based sustainable materials for various antimicrobial applications.

Novel functional insights into the microbiome inhabiting marine plastic debris: critical considerations to counteract the challenges of thin biofilms using multi-omics and comparative metaproteomics

BackgroundMicrobial functioning on marine plastic surfaces has been poorly documented, especially within cold climates where temperature likely impacts microbial activity and the presence of hydrocarbonoclastic microorganisms. To date, only two studies have used metaproteomics to unravel microbial genotype–phenotype linkages in the marine ‘plastisphere’, and these have revealed the dominance of photosynthetic microorganisms within warm climates. Advancing the functional representation of the marine plastisphere is vital for the development of specific databases cataloging the functional diversity of the associated microorganisms and their peptide and protein sequences, to fuel biotechnological discoveries. Here, we provide a comprehensive assessment for plastisphere metaproteomics, using multi-omics and data mining on thin plastic biofilms to provide unique insights into plastisphere metabolism. Our robust experimental design assessed DNA/protein co-extraction and cell lysis strategies, proteomics workflows, and diverse protein search databases, to resolve the active plastisphere taxa and their expressed functions from an understudied cold environment.ResultsFor the first time, we demonstrate the predominance and activity of hydrocarbonoclastic genera (Psychrobacter, Flavobacterium, Pseudomonas) within a primarily heterotrophic plastisphere. Correspondingly, oxidative phosphorylation, the citrate cycle, and carbohydrate metabolism were the dominant pathways expressed. Quorum sensing and toxin-associated proteins of Streptomyces were indicative of inter-community interactions. Stress response proteins expressed by Psychrobacter, Planococcus, and Pseudoalteromonas and proteins mediating xenobiotics degradation in Psychrobacter and Pseudoalteromonas suggested phenotypic adaptations to the toxic chemical microenvironment of the plastisphere. Interestingly, a targeted search strategy identified plastic biodegradation enzymes, including polyamidase, hydrolase, and depolymerase, expressed by rare taxa. The expression of virulence factors and mechanisms of antimicrobial resistance suggested pathogenic genera were active, despite representing a minor component of the plastisphere community.ConclusionOur study addresses a critical gap in understanding the functioning of the marine plastisphere, contributing new insights into the function and ecology of an emerging and important microbial niche. Our comprehensive multi-omics and comparative metaproteomics experimental design enhances biological interpretations to provide new perspectives on microorganisms of potential biotechnological significance beyond biodegradation and to improve the assessment of the risks associated with microorganisms colonizing marine plastic pollution.EMsdtEfK7hXyizwjh4GBBGVideo

Whole genome sequence-based molecular characterization of blood isolates of carbapenem-resistant Enterobacter cloacae complex from ICU patients in Kolkata, India, during 2017-2022: emergence of phylogenetically heterogeneous Enterobacter hormaechei subsp. xiangfangensis.

Blood-borne infections caused by the carbapenem-resistant Enterobacter cloacae complex (CR-ECC) are major public threats with respect to the challenges encountered during treatment. This study describes the whole genome sequencing-based molecular characteristics of blood isolates (n = 70) of CR-ECC from patients admitted to the intensive care unit of tertiary care hospitals in Kolkata, India, during 2017-2022 with respect to species identification, antimicrobial resistance (AMR) profiling, mechanism of drug resistance, and molecular subtypes. Vitek2 MALDI and species-specific PCR identified Enterobacter hormaechei subsp. xiangfangensis (47.14%) as the emerging CR-ECC subspecies in Kolkata. The predominating carbapenemase and extended-spectrum β-lactamase genes found were blaNDM-1 (51.42%) and blaCTX-M-15 (27%), respectively. Besides, blaNDM-4, blaNDM-5, blaNDM-7, blaCMH-3, blaSFO-1, blaOXA-181, blaOXA-232, blaKPC-3, and blaDHA-7 genes were also detected, which were not previously reported from India. A multitude of Class 1 integrons (including In180, In4874, In4887, and In4888, which were novel) and plasmid replicon types (IncFIB, IncFII, IncX3, IncHI1-HI2, IncC, and IncR) involved in AMR dissemination were identified. Reverse transcription-PCR and western blot revealed that carbapenem resistance in non-carbapenemase-producing CR-ECC isolates was contributed by elevated levels of ampC, overexpression of acrAB, and loss of ompF. A total of 30 distinct sequence types (STs) were ascertained by multi-locus sequence typing; of which, ST2011, ST2018, ST2055, ST2721, and ST2722 were novel STs. Pulsed-field gel electrophoresis analysis showed heterogeneity (69 pulsotypes with a similarity coefficient of 48.40%) among the circulating isolates, suggesting multiple reservoirs of infections in humans. Phylogenetically and genetically diverse CR-ECC with multiple AMR mechanisms mandates close monitoring of nosocomial infections caused by these isolates to forestall the transmission and dissemination of AMR.IMPORTANCEThe emergence and extensive dissemination of the carbapenem-resistant Enterobacter cloacae complex (CR-ECC) have positioned it as a critical nosocomial global pathogen. The dearth of a comprehensive molecular study pertaining to CR-ECC necessitated this study, which is the first of its kind from India. Characterization of blood isolates of CR-ECC over the last 6 years revealed Enterobacter hormaechei subsp. xiangfangensis as the most prevalent subsp., exhibiting resistance to almost all antibiotics currently in use and harboring diverse transmissible carbapenemase genes. Besides the predominating blaNDM-1 and blaCTX-M-15, we document diverse carbapenemase and AmpC genes, such as blaNDM-4, blaNDM-7, blaOXA-181, blaOXA-232, blaKPC-3, blaCMH-3, blaSFO-1, and blaDHA-7, in CR-ECC, which were not previously reported from India. Furthermore, novel integrons and sequence types were identified. Our findings emphasize the need for strengthened vigilance for molecular epidemiological surveillance of CR-ECC due to the presence of epidemic clones with a phylogenetically diverse and wide array of antimicrobial resistance genes in vulnerable populations.

Genetic epidemiology and plasmid-mediated transmission of mcr-1 by Escherichia coli ST155 from wastewater of long-term care facilities.

Long-term care facilities (LTCFs) for older people play an important and unique role in multidrug-resistant organism transmission. Herein, we investigated the genetic characteristics of mobile colistin resistance gene (mcr-1)-carrying Escherichia coli strains isolated from wastewater of LTCFs in Shanghai. Antimicrobial susceptibility test was carried out by agar dilution methods. Whole-genome sequencing and plasmid sequencing were conducted, and resistance genes and sequence types of colistin in E. coli isolates were analyzed. Core genome multilocus sequence typing (cgMLST) analysis was performed by the Ridom SeqSphere+ software. Phylogenetic tree through the maximum likelihood method was constructed by MEGA X. Out of 306 isolates, only 1 E. coli named ECSJ33 was found, and the plasmid pECSJ33 from ECSJ33 harbored the mcr-1 gene that was located with 59,080 bp belonging to IncI2 type. The plasmid pECSJ33 was capable of conjugation with an efficiency of 2.9 × 10-2. Bioinformatic analysis indicated pECSJ33 shared backbone with the previously reported mcr-1-harboring pHNGDF93 isolated from fish source. Moreover, the cgMLST analysis revealed that ECSJ33 belongs to different lineages from those reported from previous E. coli strains but shared high similarity to NCTC11129 in cluster 11. The phylogenetic tree revealed MCR-1 of ECSJ33 in this study was mostly of animal food origin and that they were closely related. Our study firstly reports detection of genome sequence of a multidrug-resistant mcr-1-harboring E. coli ST155 from wastewater of LTCF source in China. The data may prove that the plasmid pECSJ33 belongs to food origin and help to understand the antimicrobial resistance mechanisms and genomic features of colistin resistance under One Health approach.IMPORTANCEOne Escherichia coli named ECSJ33 was found from wastewater of a long-term care facility (LTCF) and the plasmid pECSJ33 from ECSJ33 harbored the mobile colistin resistance gene (mcr-1) that was located with 59,080 bp belonging to IncI2 type, which was capable of conjugation with an efficiency of 2.9 × 10-2. This paper firstly reports an mcr-1-carrying E. coli strain ST155 isolated from LTCF in China. Comparative genomics analysis indicated pECSJ33 shared backbone with the previously reported mcr-1-harboring pHNGDF93 isolated from fish source. The phylogenetic tree revealed MCR-1 protein of ECSJ33 in this study was mostly of animal food origin and that they were closely related. Therefore, the pECSJ33 could be considered as food-origin transmission mcr-1-harboring plasmid.

Genomic Sequencing of Clinical Cupriavidus gilardii Isolates Revealed Their Diverse Antimicrobial Resistance Mechanisms.

Cupriavidus gilardii is an emerging multidrug-resistant pathogen found in many environments and few clinical samples. The clinical infectiousness, pathogenicity, and resistance mechanisms of C. gilardii are still unclear due to the lack of clinical and sequencing data. We need to obtain insight into the clinical characteristics, virulence, and resistance mechanisms of C. gilardii. We isolated five C. gilardii isolates from hospitalized patients and carried out assay, culture and genome sequencing. We analyzed the genomic features of clinical C. gilardii isolates and took insight into their clinical characteristics, virulence, and resistance mechanisms. These isolates were resistant to meropenem, gentamicin, and other antimicrobials due to intrinsic resistance genes. Furthermore, the sequencing results revealed the widespread presence of the MCR-5.1 gene in C. gilardii. The virulence magnitude of C. gilardii is closely correlated with the number of virulence factors they carry. Some C. gilardii strains can acquire resistance to levofloxacin through gyrA gene mutation during treatment. The diverse antimicrobial resistance mechanisms challenge the treatment of C. gilardii infections. We present the genomic characteristics of clinically isolated C. gilardii to improve (i) our understanding of this pathogen and (ii) treatment options.

Hybrid Molecules of Azithromycin with Chloramphenicol and Metronidazole: Synthesis and Study of Antibacterial Properties.

The sustained rise of antimicrobial resistance (AMR) causes a strong need to develop new antibacterial agents. One of the methods for addressing the problem of antibiotic resistance is through the design of hybrid antibiotics. In this work, we proposed a synthetic route for the conjugation of an azithromycin derivative with chloramphenicol and metronidazole hemisuccinates and synthesized two series of new hybrid molecules 4a-g and 5a-g. While a conjugation did not result in tangible synergy for wild-type bacterial strains, new compounds were able to overcome AMR associated with the inducible expression of the ermC gene on a model E. coli strain resistant to macrolide antibiotics. The newly developed hybrids demonstrated a tendency to induce premature ribosome stalling, which might be crucial since they will not induce a macrolide-resistant phenotype in a number of pathogenic bacterial strains. In summary, the designed structures are considered as a promising direction for the further development of hybrid molecules that can effectively circumvent AMR mechanisms to macrolide antibiotics.

Clostridioides difficile Infection: Diagnosis and Treatment Challenges.

Clostridioides difficile is the most important cause of healthcare-associated diarrhea in the United States. The high incidence and recurrence rates of C. difficile infection (CDI), associated with high morbidity and mortality, pose a public health challenge. Although antibiotics targeting C. difficile bacteria are the first treatment choice, antibiotics also disrupt the indigenous gut flora and, therefore, create an environment that is favorable for recurrent CDI. The challenge of treating CDI is further exacerbated by the rise of antibiotic-resistant strains of C. difficile, placing it among the top five most urgent antibiotic resistance threats in the USA. The evolution of antibiotic resistance in C. difficile involves the acquisition of new resistance mechanisms, which can be shared among various bacterial species and different C. difficile strains within clinical and community settings. This review provides a summary of commonly used diagnostic tests and antibiotic treatment strategies for CDI. In addition, it discusses antibiotic treatment and its resistance mechanisms. This review aims to enhance our current understanding and pinpoint knowledge gaps in antimicrobial resistance mechanisms in C. difficile, with an emphasis on CDI therapies.