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515500.9974The Widespread Multidrug-Resistant Serotype O12 Pseudomonas aeruginosa Clone Emerged through Concomitant Horizontal Transfer of Serotype Antigen and Antibiotic Resistance Gene Clusters. The O-specific antigen (OSA) in Pseudomonas aeruginosa lipopolysaccharide is highly varied by sugar identity, side chains, and bond between O-repeats. These differences classified P. aeruginosa into 20 distinct serotypes. In the past few decades, O12 has emerged as the predominant serotype in clinical settings and outbreaks. These serotype O12 isolates exhibit high levels of resistance to various classes of antibiotics. Here, we explore how the P. aeruginosa OSA biosynthesis gene clusters evolve in the population by investigating the association between the phylogenetic relationships among 83 P. aeruginosa strains and their serotypes. While most serotypes were closely linked to the core genome phylogeny, we observed horizontal exchange of OSA biosynthesis genes among phylogenetically distinct P. aeruginosa strains. Specifically, we identified a "serotype island" ranging from 62 kb to 185 kb containing the P. aeruginosa O12 OSA gene cluster, an antibiotic resistance determinant (gyrA(C248T)), and other genes that have been transferred between P. aeruginosa strains with distinct core genome architectures. We showed that these genes were likely acquired from an O12 serotype strain that is closely related to P. aeruginosa PA7. Acquisition and recombination of the "serotype island" resulted in displacement of the native OSA gene cluster and expression of the O12 serotype in the recipients. Serotype switching by recombination has apparently occurred multiple times involving bacteria of various genomic backgrounds. In conclusion, serotype switching in combination with acquisition of an antibiotic resistance determinant most likely contributed to the dissemination of the O12 serotype in clinical settings. IMPORTANCE: Infection rates in hospital settings by multidrug-resistant (MDR) Pseudomonas aeruginosa clones have increased during the past decades, and serotype O12 is predominant among these epidemic strains. It is not known why the MDR phenotype is associated with serotype O12 and how this clone type has emerged. This study shows that evolution of MDR O12 strains involved a switch from an ancestral O4 serotype to O12. Serotype switching was the result of horizontal transfer and genetic recombination of lipopolysaccharide (LPS) biosynthesis genes originating from an MDR taxonomic outlier P. aeruginosa strain. Moreover, the recombination event also resulted in acquisition of antibiotic resistance genes. These results impact on our understanding of MDR outbreak strain and serotype evolution and can potentially assist in better monitoring and prevention.201526396243
548610.9974Genomic Landscape of Multidrug Resistance and Virulence in Enterococcus faecalis IRMC827A from a Long-Term Patient. We report on a highly virulent, multidrug-resistant strain of Enterococcus faecalis IRMC827A that was found colonizing a long-term male patient at a tertiary hospital in Khobar, Saudi Arabia. The E. faecalis IRMC827A strain carries several antimicrobial drug resistance genes and harbours mobile genetic elements such as Tn6009, which is an integrative conjugative element that can transfer resistance genes between bacteria and ISS1N via an insertion sequence. Whole-genome-sequencing-based antimicrobial susceptibility testing on strains from faecal samples revealed that the isolate E. faecalis IRMC827A is highly resistant to a variety of antibiotics, including tetracycline, doxycycline, minocycline, dalfopristin, virginiamycin, pristinamycin, chloramphenicol, streptomycin, clindamycin, lincomycin, trimethoprim, nalidixic acid and ciprofloxacin. The isolate IRMC827A carries several virulence factors that are significantly associated with adherence, biofilm formation, sortase-assembled pili, manganese uptake, antiphagocytosis, and spreading factor of multidrug resistance. The isolate also encompasses two mutations (G2576T and G2505A) in the 23S rRNA gene associated with linezolid resistance and three more mutations (gyrA p.S83Y, gyrA p.D759N and parC p.S80I) of the antimicrobial resistance phenotype. The findings through next-generation sequencing on the resistome, mobilome and virulome of the isolate in the study highlight the significance of monitoring multidrug-resistant E. faecalis colonization and infection in hospitalized patients. As multidrug-resistant E. faecalis is a serious pathogen, it is particularly difficult to treat and can cause fatal infections. It is important to have quick and accurate diagnostic tests for multidrug-resistant E. faecalis, to track the spread of multidrug-resistant E. faecalis in healthcare settings, and to improve targeted interventions to stop its spread. Further research is necessary to develop novel antibiotics and treatment strategies for multidrug-resistant E. faecalis infections.202337887006
496320.9973Comprehensive Genomic Analysis of Uropathogenic E. coli: Virulence Factors, Antimicrobial Resistance, and Mobile Genetic Elements. Our whole-genome sequencing analysis of sixteen uropathogenic E. coli isolates revealed a concerning picture of multidrug resistance and potentially virulent bacteria. All isolates belonged to four distinct clonal groups, with the highly prevalent ST131 lineage being associated with extensive antibiotic resistance and virulence factors. Notably, all isolates exhibited multidrug resistance, with some resistant to as many as 12 antibiotics. Fluoroquinolone resistance stemmed primarily from efflux pumps and mutations in gyrase and topoisomerase genes. Additionally, we identified genes encoding resistance to extended-spectrum cephalosporins, trimethoprim/sulfamethoxazole, and various heavy metals. The presence of diverse plasmids and phages suggests the potential for horizontal gene transfer and the dissemination of virulence factors. All isolates harbored genomic islands containing virulence factors associated with adhesion, biofilm formation, and invasion. Genes essential for iron acquisition, flagella biosynthesis, secretion systems, and toxin production were also prevalent. Adding further complexity to understanding the isolates' genetic makeup, we identified CRISPR-Cas systems. This study underscores the need for continued genomic surveillance in understanding the pathogenic mechanisms and resistance profiles of uropathogenic E. coli to aid in developing targeted therapeutic strategies.202439338985
549630.9973Genome-Wide Analysis of Escherichia coli Isolated from Dairy Animals Identifies Virulence Factors and Genes Enriched in Multidrug-Resistant Strains. The gastrointestinal tracts of dairy calves and cows are reservoirs of antimicrobial-resistant bacteria (ARB), which are present regardless of previous antimicrobial therapy. Young calves harbor a greater abundance of resistant bacteria than older cows, but the factors driving this high abundance are unknown. Here, we aimed to fully characterize the genomes of multidrug-resistant (MDR) and antimicrobial-susceptible Escherichia coli strains isolated from pre-weaned calves, post-weaned calves, dry cows, and lactating cows and to identify the accessory genes that are associated with the MDR genotype to discover genetic targets that can be exploited to mitigate antimicrobial resistance in dairy farms. Results indicated that both susceptible and resistant E. coli isolates recovered from animals on commercial dairy operations were highly diverse and encoded a large pool of virulence factors. In total, 838 transferrable antimicrobial resistance genes (ARGs) were detected, with genes conferring resistance to aminoglycosides being the most common. Multiple sequence types (STs) associated with mild to severe human gastrointestinal and extraintestinal infections were identified. A Fisher's Exact Test identified 619 genes (ARGs and non-ARGs) that were significantly enriched in MDR isolates and 147 genes that were significantly enriched in susceptible isolates. Significantly enriched genes in MDR isolates included the iron scavenging aerobactin synthesis and receptor genes (iucABCD-iutA) and the sitABCD system, as well as the P fimbriae pap genes, myo-inositol catabolism (iolABCDEG-iatA), and ascorbate transport genes (ulaABC). The results of this study demonstrate a highly diverse population of E. coli in commercial dairy operations, some of which encode virulence genes responsible for severe human infections and resistance to antibiotics of human health significance. Further, the enriched accessory genes in MDR isolates (aerobactin, sit, P fimbriae, and myo-inositol catabolism and ascorbate transport genes) represent potential targets for reducing colonization of antimicrobial-resistant bacteria in the calf gut.202337887260
515940.9972Microevolution of Monophasic Salmonella Typhimurium during Epidemic, United Kingdom, 2005-2010. Microevolution associated with emergence and expansion of new epidemic clones of bacterial pathogens holds the key to epidemiologic success. To determine microevolution associated with monophasic Salmonella Typhimurium during an epidemic, we performed comparative whole-genome sequencing and phylogenomic analysis of isolates from the United Kingdom and Italy during 2005-2012. These isolates formed a single clade distinct from recent monophasic epidemic clones previously described from North America and Spain. The UK monophasic epidemic clones showed a novel genomic island encoding resistance to heavy metals and a composite transposon encoding antimicrobial drug resistance genes not present in other Salmonella Typhimurium isolates, which may have contributed to epidemiologic success. A remarkable amount of genotypic variation accumulated during clonal expansion that occurred during the epidemic, including multiple independent acquisitions of a novel prophage carrying the sopE gene and multiple deletion events affecting the phase II flagellin locus. This high level of microevolution may affect antigenicity, pathogenicity, and transmission.201626982594
515650.9972Pseudomonas aeruginosa strains isolated from animal with high virulence genes content and highly sensitive to antimicrobials. OBJECTIVES: P. aeruginosa is one of the most metabolically versatile bacteria having the ability to survive in multiple environments through its accessory genome. An important hallmark of P. aeruginosa is the high level of antibiotic resistance, which often makes eradication difficult and sometimes impossible. Evolutionary forces have led to this bacterium to develop high antimicrobial resistance with a variety of elements contributing to both intrinsic and acquired resistance. The objectives were to genetically and phenotypically characterizer P. aeruginosa strains isolated from companion animals of different species. METHODS: We characterized a collection of 39 P. aeruginosa strains isolated from infected animals. The genetic characterization was in relation to chromosomal profile by PFGE; content of virulence gene; presence of genomic islands (GIs); genes of the cytotoxins exported by T3SS: exoU, exoS, exoT and exoY; and type IV pili allele. The phenotypic characterization was based on patterns of susceptibility to different antimicrobials. RESULTS: Each strain had a PFGE profile, a high virulence genes content, and a large accessory genome. However, most of the strains presented high sensitivity to almost all antimicrobials tested, showing no acquired resistance (no β-lactamases). The exception to this lack of resistance was seen with penicillin. CONCLUSIONS: P. aeruginosa could be a naturally sensitive bacterium to standard antimicrobials but could rapidly develop intrinsic and acquired resistance when the bacterium is exposed to pressure exerted by antibiotics, as observed in hospital settings.202438452900
504560.9972Emergence of colistin-resistance in extremely drug-resistant Acinetobacter baumannii containing a novel pmrCAB operon during colistin therapy of wound infections. BACKGROUND: Colistin resistance is of concern since it is increasingly needed to treat infections caused by bacteria resistant to all other antibiotics and has been associated with poorer outcomes. Longitudinal data from in vivo series are sparse. METHODS: Under a quality-improvement directive to intensify infection-control measures, extremely drug-resistant (XDR) bacteria undergo phenotypic and molecular analysis. RESULTS: Twenty-eight XDR Acinetobacter baumannii isolates were longitudinally recovered during colistin therapy. Fourteen were susceptible to colistin, and 14 were resistant to colistin. Acquisition of colistin resistance did not alter resistance to other antibiotics. Isolates had low minimum inhibitory concentrations of an investigational aminoglycoside, belonged to multi-locus sequence type 94, were indistinguishable by pulsed-field gel electrophoresis and optical mapping, and harbored a novel pmrC1A1B allele. Colistin resistance was associated with point mutations in the pmrA1 and/or pmrB genes. Additional pmrC homologs, designated eptA-1 and eptA-2, were at distant locations from the operon. Compared with colistin-susceptible isolates, colistin-resistant isolates displayed significantly enhanced expression of pmrC1A1B, eptA-1, and eptA-2; lower growth rates; and lowered fitness. Phylogenetic analysis suggested that colistin resistance emerged from a single progenitor colistin-susceptible isolate. CONCLUSIONS: We provide insights into the in vivo evolution of colistin resistance in a series of XDR A. baumannii isolates recovered during therapy of infections and emphasize the importance of antibiotic stewardship and surveillance.201323812239
187270.9972Escherichia coli ST2797 Is Abundant in Wastewater and Might Be a Novel Emerging Extended-Spectrum Beta-Lactamase E. coli. The increasing prevalence of antibiotic-resistant bacteria is an emerging threat to global health. The analysis of antibiotic-resistant enterobacteria in wastewater can indicate the prevalence and spread of certain clonal groups of multiresistant bacteria. In a previous study of Escherichia coli that were isolated from a pump station in Norway over 15 months, we found a recurring E. coli clone that was resistant to trimethoprim, ampicillin, and tetracycline in 201 of 3,123 analyzed isolates (6.1%). 11 representative isolates were subjected to whole-genome sequencing and were found to belong to the MLST ST2797 E. coli clone with plasmids carrying resistance genes, including bla(TEM-1B), sul2, dfrA7, and tetB. A phenotypic comparison of the ST2797 isolates with the uropathogenic ST131 and ST648 that were repeatedly identified in the same wastewater samples revealed that the ST2797 isolates exhibited a comparable capacity for temporal survival in wastewater, greater biofilm formation, and similar potential for the colonization of mammalian epithelial cells. ST2797 has been isolated from humans and has been found to carry extended spectrum β-lactamase (ESBL) genes in other studies, suggesting that this clonal type is an emerging ESBL E. coli. Collectively, these findings show that ST2797 was more ubiquitous in the studied wastewater than were the infamous ST131 and ST648 and that ST2797 may have similar abilities to survive in the environment and cause infections in humans. IMPORTANCE The incidence of drug-resistant bacteria found in the environment is increasing together with the levels of antibiotic-resistant bacteria that cause infections. The COVID-19 pandemic has shed new light on the importance of monitoring emerging threats and finding early warning systems. Therefore, to mitigate the antimicrobial resistance burden, the monitoring and early identification of antibiotic-resistant bacteria in hot spots, such as wastewater treatment plants, are required to combat the occurrence and spread of antibiotic-resistant bacteria. Here, we applied a PhenePlate system as a phenotypic screening method for genomic surveillance and discovered a dominant and persistent E. coli clone ST2797 with a multidrug resistance pattern and equivalent phenotypic characteristics to those of the major pandemic lineages, namely, ST131 and ST648, which frequently carry ESBL genes. This study highlights the continuous surveillance and report of multidrug resistant bacteria with the potential to spread in One Health settings.202337260395
548480.9972A set of antibiotic-resistance mechanisms and virulence factors in GES-16-producing Klebsiella quasipneumoniae subsp. similipneumoniae from hospital wastewater revealed by whole-genome sequencing. Klebsiella quasipneumoniae subsp. similipneumoniae has emerged as a human pathogen and sporadic isolates from non-clinical sources were reported. Here, we described the phenotypic- and genomic-characteristics of a multidrug-resistant (MDR) and potentially hypervirulent (MDR-hv) Klebsiella quasipneumoniae subsp. similipneumoniae (KqA1) isolated from hospital wastewater. The antibiotic susceptibility profile of KqA1 was investigated using disk-diffusion method, broth microdilution method, and agar dilution method, and the genetic characteristics of antimicrobial resistance, mobile genetics elements, and virulence were evaluated by genomic DNA sequencing on the Illumina® NovaSeq6000 platform as well as by bioinformatic analysis. Resistome analyses revealed the presence of genes related to resistance to β-lactams, aminoglycosides, quinolones, tetracyclines, sulfonamides, trimethoprim, chloramphenicol, macrolides, and fosfomycin. New genetic contexts to bla(GES-16) (carbapenemase gene) and to fosA (fosfomycin resistance gene) were described. A set of mechanisms that can contribute to antibiotic resistance, commonly detected in Klebsiella spp., was also found including chromosomal mutations, efflux systems, proteins, and regulators. Moreover, KqA1 presented genes related to tolerance to metals (arsenic, copper, nickel, cobalt, magnesium, cadmium, zinc, tellurium, selenium) and to biocides (quaternary-ammonium compounds). The isolate was classified as potentially hypervirulent due to a wide range of virulence factors found associated to regulation, motility, biofilm, effector delivery systems, immune modulation, nutritional/metabolic factors, adherence, invasion, and competitive advantage. The occurrence of MDR-hv KqA1 in hospital wastewater points out how this environment matrix plays a crucial role in the maintenance and selection of critical bacterial pathogens. Regarding One Health perspective, it is evident the need for multidisciplinary implementation of control measures for antibiotic-resistant bacteria, not only in hospital settings but also in a general environmental context to mitigate the dissemination of MDR and hv bacteria.202336375580
189890.9972Multiple-Replicon Resistance Plasmids of Klebsiella Mediate Extensive Dissemination of Antimicrobial Genes. Multiple-replicon resistance plasmids have become important carriers of resistance genes in Gram-negative bacteria, and the evolution of multiple-replicon plasmids is still not clear. Here, 56 isolates of Klebsiella isolated from different wild animals and environments between 2018 and 2020 were identified by phenotyping via the micro-broth dilution method and were sequenced and analyzed for bacterial genome-wide association study. Our results revealed that the isolates from non-human sources showed more extensive drug resistance and especially strong resistance to ampicillin (up to 80.36%). The isolates from Malayan pangolin were particularly highly resistant to cephalosporins, chloramphenicol, levofloxacin, and sulfamethoxazole. Genomic analysis showed that the resistance plasmids in these isolates carried many antibiotic resistance genes. Further analysis of 69 plasmids demonstrated that 28 plasmids were multiple-replicon plasmids, mainly carrying beta-lactamase genes such as bla (CTX-M-) (15), bla (CTX-M-) (14), bla (CTX-M-) (55), bla (OXA-) (1), and bla (TEM-) (1). The analysis of plasmids carried by different isolates showed that Klebsiella pneumoniae might be an important multiple-replicon plasmid host. Plasmid skeleton and structure analyses showed that a multiple-replicon plasmid was formed by the fusion of two or more single plasmids, conferring strong adaptability to the antibiotic environment and continuously increasing the ability of drug-resistant isolates to spread around the world. In conclusion, multiple-replicon plasmids are better able to carry resistance genes than non-multiple-replicon plasmids, which may be an important mechanism underlying bacterial responses to environments with high-antibiotic pressure. This phenomenon will be highly significant for exploring bacterial resistance gene transmission and diffusion mechanisms in the future.202134777312
4929100.9972Comparative genomics analysis of Acinetobacter baumannii multi-drug resistant and drug sensitive strains in China. The incidence of multidrug-resistant Acinetobacter baumannii has posed a major challenge for clinical treatment. There is still a significant gap in understanding the mechanism causing multi-drug resistance (MDR). In this study, the genomes of 10 drug sensitive and 10 multi-drug resistant A.baumannii strains isolated from a hospital in China were sequenced and compared. The antibiotic resistance genes, virulence factors were determined and CRIPSR-Cas system along with prophages were detected. The results showed that MDR strains are significantly different from the drug sensitive strains in the CARD entries, patterns of sequences matching up to plasmids, VFDB entries and CRISPR-Cas system. MDR strains contain unique CARD items related to antibiotic resistance which are absent in sensitive strains. Furthermore, sequences from genomes of MDR strains can match up with plasmids from more diversified bacteria genera compared to drug sensitive strains. MDR strains also contain a lower level of CRISPR genes and larger amount of prophages, along with higher levels of spacer sequences. These findings provide new experimental evidences for the study of the antibiotic resistance mechanism of A. baumannii.202235307599
1769110.9972DNA sequence and comparative genomics of pAPEC-O2-R, an avian pathogenic Escherichia coli transmissible R plasmid. In this study, a 101-kb IncF plasmid from an avian pathogenic Escherichia coli (APEC) strain (APEC O2) was sequenced and analyzed, providing the first completed APEC plasmid sequence. This plasmid, pAPEC-O2-R, has functional transfer and antimicrobial resistance-encoding regions. The resistance-encoding region encodes resistance to eight groups of antimicrobial agents, including silver and other heavy metals, quaternary ammonium compounds, tetracycline, sulfonamides, aminoglycosides, trimethoprim, and beta-lactam antimicrobial agents. This region of the plasmid is unique among previously described IncF plasmids in that it possesses a class 1 integron that harbors three gene cassettes and a heavy metal resistance operon. This region spans 33 kb and is flanked by the RepFII plasmid replicon and an assortment of plasmid maintenance genes. pAPEC-O2-R also contains a 32-kb transfer region that is nearly identical to that found in the E. coli F plasmid, rendering it transferable by conjugation to plasmid-less strains of bacteria, including an APEC strain, a fecal E. coli strain from an apparently healthy bird, a Salmonella enterica serovar Typhimurium strain, and a uropathogenic E. coli strain from humans. Differences in the G+C contents of individual open reading frames suggest that various regions of pAPEC-O2-R had dissimilar origins. The presence of pAPEC-O2-R-like plasmids that encode resistance to multiple antimicrobial agents and that are readily transmissible from APEC to other bacteria suggests the possibility that such plasmids may serve as a reservoir of resistance genes for other bacteria of animal and human health significance.200516251312
4926120.9972Complete Assembly of Escherichia coli Sequence Type 131 Genomes Using Long Reads Demonstrates Antibiotic Resistance Gene Variation within Diverse Plasmid and Chromosomal Contexts. The incidence of infections caused by extraintestinal Escherichia coli (ExPEC) is rising globally, which is a major public health concern. ExPEC strains that are resistant to antimicrobials have been associated with excess mortality, prolonged hospital stays, and higher health care costs. E. coli sequence type 131 (ST131) is a major ExPEC clonal group worldwide, with variable plasmid composition, and has an array of genes enabling antimicrobial resistance (AMR). ST131 isolates frequently encode the AMR genes bla(CTX-M-14), bla(CTX-M-15), and bla(CTX-M-27), which are often rearranged, amplified, and translocated by mobile genetic elements (MGEs). Short DNA reads do not fully resolve the architecture of repetitive elements on plasmids to allow MGE structures encoding bla(CTX-M) genes to be fully determined. Here, we performed long-read sequencing to decipher the genome structures of six E. coli ST131 isolates from six patients. Most long-read assemblies generated entire chromosomes and plasmids as single contigs, in contrast to more fragmented assemblies created with short reads alone. The long-read assemblies highlighted diverse accessory genomes with bla(CTX-M-15), bla(CTX-M-14), and bla(CTX-M-27) genes identified in three, one, and one isolates, respectively. One sample had no bla(CTX-M) gene. Two samples had chromosomal bla(CTX-M-14) and bla(CTX-M-15) genes, and the latter was at three distinct locations, likely transposed by the adjacent MGEs: ISEcp1, IS903B, and Tn2 This study showed that AMR genes exist in multiple different chromosomal and plasmid contexts, even between closely related isolates within a clonal group such as E. coli ST131.IMPORTANCE Drug-resistant bacteria are a major cause of illness worldwide, and a specific subtype called Escherichia coli ST131 causes a significant number of these infections. ST131 bacteria become resistant to treatments by modifying their DNA and by transferring genes among one another via large packages of genes called plasmids, like a game of pass-the-parcel. Tackling infections more effectively requires a better understanding of what plasmids are being exchanged and their exact contents. To achieve this, we applied new high-resolution DNA sequencing technology to six ST131 samples from infected patients and compared the output to that of an existing approach. A combination of methods shows that drug resistance genes on plasmids are highly mobile because they can jump into ST131's chromosomes. We found that the plasmids are very elastic and undergo extensive rearrangements even in closely related samples. This application of DNA sequencing technologies illustrates at a new level the highly dynamic nature of ST131 genomes.201931068432
2470130.9972Whole-genome sequencing of Klebsiella pneumoniae MDR circulating in a pediatric hospital setting: a comprehensive genome analysis of isolates from Guayaquil, Ecuador. BACKGROUND: Klebsiella pneumoniae is the major cause of nosocomial infections worldwide and is related to a worsening increase in Multidrug-Resistant Bacteria (MDR) and virulence genes that seriously affect immunosuppressed patients, long-stay intensive care patients, elderly individuals, and children. Whole-Genome Sequencing (WGS) has resulted in a useful strategy for characterizing the genomic components of clinically important bacteria, such as K. pneumoniae, enabling them to monitor genetic changes and understand transmission, highlighting the risk of dissemination of resistance and virulence associated genes in hospitals. In this study, we report on WGS 14 clinical isolates of K. pneumoniae from a pediatric hospital biobank of Guayaquil, Ecuador. RESULTS: The main findings revealed pronounced genetic heterogeneity among the isolates. Multilocus sequencing type ST45 was the predominant lineage among non-KPC isolates, whereas ST629 was found more frequently among KPC isolates. Phylogenetic analysis suggested local transmission dynamics. Comparative genomic analysis revealed a core set of 3511 conserved genes and an open pangenome in neonatal isolates. The diversity of MLSTs and capsular types, and the high genetic diversity among these isolates indicate high intraspecific variability. In terms of virulence factors, we identified genes associated with adherence, biofilm formation, immune evasion, secretion systems, multidrug efflux pump transporters, and a notably high number of genes related to iron uptake. A large number of these genes were detected in the ST45 isolate, whereas iron uptake yersiniabactin genes were found exclusively in the non-KPC isolates. We observed high resistance to commonly used antibiotics and determined that these isolates exhibited multidrug resistance including β-lactams, aminoglycosides, fluoroquinolones, quinolones, trimetropins, fosfomycin and macrolides; additionally, resistance-associated point mutations and cross-resistance genes were identified in all the isolates. We also report the first K. pneumoniae KPC-3 gene producers in Ecuador. CONCLUSIONS: Our WGS results for clinical isolates highlight the importance of MDR in neonatal K. pneumoniae infections and their genetic diversity. WGS will be an imperative strategy for the surveillance of K. pneumoniae in Ecuador, and will contribute to identifying effective treatment strategies for K. pneumoniae infections in critical units in patients at stratified risk.202439367302
2538140.9971Passenger pathogens on physicians. BACKGROUND: Hospital acquired infections pose a significant risk for patients undergoing hematopoietic stem cell transplantation. Horizontal transfer of antimicrobial resistance genes contributes to prevalence of multidrug-resistant infections in this patient population. METHODS: At an academic bone marrow transplantation center, we performed whole genome DNA sequencing (WGS) on commonly used physician items, including badges, stethoscopes, soles of shoes, and smart phones from 6 physicians. Data were analyzed to determine antimicrobial resistance and virulence factor genes. RESULTS: A total of 1,126 unique bacterial species, 495 distinct bacteriophages, 91 unique DNA viruses, and 175 fungal species were observed. Every item contained bacteria with antibiotic and/or antiseptic resistance genes. Stethoscopes contained greatest frequency of antibiotic resistance and more plasmid-carriage of antibiotic resistance. DISCUSSION AND CONCLUSIONS: These data indicate that physician examination tools and personal items possess potentially pathogenic microbes. Infection prevention policies must consider availability of resources to clean physical examination tools as well as provider awareness when enacting hospital policies. Additionally, the prevalence of antimicrobial resistance genes (eg, encoding resistance to aminoglycosides, β-lactams, and quinolones) reinforces need for antimicrobial stewardship, including for immunocompromised patients. Further research is needed to assess whether minute quantities of microbes on physician objects detectable by WGS represents clinically significant inoculums for immunocompromised patients.202336306861
9968150.9971Antibiotic Resistance, Core-Genome and Protein Expression in IncHI1 Plasmids in Salmonella Typhimurium. Conjugative plasmids from the IncHI1 incompatibility group play an important role in transferring antibiotic resistance in Salmonella Typhimurium. However, knowledge of their genome structure or gene expression is limited. In this study, we determined the complete nucleotide sequences of four IncHI1 plasmids transferring resistance to antibiotics by two different next generation sequencing protocols and protein expression by mass spectrometry. Sequence data including additional 11 IncHI1 plasmids from GenBank were used for the definition of the IncHI1 plasmid core-genome and pan-genome. The core-genome consisted of approximately 123 kbp and 122 genes while the total pan-genome represented approximately 600 kbp. When the core-genome sequences were used for multiple alignments, the 15 tested IncHI1 plasmids were separated into two main lineages. GC content in core-genome genes was around 46% and 50% in accessory genome genes. A multidrug resistance region present in all 4 sequenced plasmids extended over 20 kbp and, except for tet(B), the genes responsible for antibiotic resistance were those with the highest GC content. IncHI1 plasmids therefore represent replicons that evolved in low GC content bacteria. From their original host, they spread to Salmonella and during this spread these plasmids acquired multiple accessory genes including those coding for antibiotic resistance. Antibiotic-resistance genes belonged to genes with the highest level of expression and were constitutively expressed even in the absence of antibiotics. This is the likely mechanism that facilitates host cell survival when antibiotics suddenly emerge in the environment.201627189997
1836160.9971Comparative genomics with a multidrug-resistant Klebsiella pneumoniae isolate reveals the panorama of unexplored diversity in Northeast Brazil. The emergence of community acquired infections increases the public health concern on K. pneumoniae and closely related bacteria among which antimicrobial resistance spreads. We report a multidrug-resistant K. pneumoniae isolate, B31, of a patient infected in the community and admitted to an intensive care unit in Northeast Brazil. Antimicrobial susceptibility and genome information were thoroughly investigated to characterize B31 in front of 172 sequenced strains of different countries. Assigned to the Sequence Type 15, which is globally spread, B31 presented extended spectrum beta-lactamase, tigecycline and ciprofloxacin resistance. Genome sequencing revealed most resistance genes being carried by plasmids with high dissemination potential. The absence of main virulence factors, like yersiniabactin and colibactin, apparently suggests a mild pathogenic strain which, on the contrary, persisted and caused severe infection in a previously healthy patient. The present study contributes to unveil the unclear genomic scenario of virulent and multidrug-resistant K. pneumoniae in Brazil.202133373662
1896170.9971Difference analysis and characteristics of incompatibility group plasmid replicons in gram-negative bacteria with different antimicrobial phenotypes in Henan, China. BACKGROUND: Multi-drug-resistant organisms (MDROs) in gram-negative bacteria have caused a global epidemic, especially the bacterial resistance to carbapenem agents. Plasmid is the common vehicle for carrying antimicrobial resistance genes (ARGs), and the transmission of plasmids is also one of the important reasons for the emergence of MDROs. Different incompatibility group plasmid replicons are highly correlated with the acquisition, dissemination, and evolution of resistance genes. Based on this, the study aims to identify relevant characteristics of various plasmids and provide a theoretical foundation for clinical anti-infection treatment. METHODS: 330 gram-negative strains with different antimicrobial phenotypes from a tertiary hospital in Henan Province were included in this study to clarify the difference in incompatibility group plasmid replicons. Additionally, we combined the information from the PLSDB database to elaborate on the potential association between different plasmid replicons and ARGs. The VITEK mass spectrometer was used for species identification, and the VITEK-compact 2 automatic microbial system was used for the antimicrobial susceptibility test (AST). PCR-based replicon typing (PBRT) detected the plasmid profiles, and thirty-three different plasmid replicons were determined. All the carbapenem-resistant organisms (CROs) were tested for the carbapenemase genes. RESULTS: 21 plasmid replicon types were detected in this experiment, with the highest prevalence of IncFII, IncFIB, IncR, and IncFIA. Notably, the detection rate of IncX3 plasmids in CROs is higher, which is different in strains with other antimicrobial phenotypes. The number of plasmid replicons they carried increased with the strain resistance increase. Enterobacterales took a higher number of plasmid replicons than other gram-negative bacteria. The same strain tends to have more than one plasmid replicon type. IncF-type plasmids tend to be associated with MDROs. Combined with PLSDB database analysis, IncFII and IncX3 are critical platforms for taking bla(KPC-2) and bla(NDM). CONCLUSIONS: MDROs tend to carry more complex plasmid replicons compared with non-MDROs. The plasmid replicons that are predominantly prevalent and associated with ARGs differ in various species. The wide distribution of IncF-type plasmids and their close association with MDROs should deserve our attention. Further investigation into the critical role of plasmids in the carriage, evolution, and transmission of ARGs is needed.202438373913
5506180.9971Genomic and phenotypic insight into antimicrobial resistance of Pseudomonas fluorescens from King George Island, Antarctica. The genus Pseudomonas includes metabolically versatile microorganisms occupying diverse niches, from environmental habitats to plant pathogens, and has clinically significant strains. For this reason, Pseudomonas spp. might act as a reservoir of antimicrobial resistance genes, which have been detected even in isolated environments. The aim of this study was to report the antimicrobial susceptibility profile of 25 Pseudomonas fluorescens isolates from soil samples collected on King George Island (Antarctic Peninsula), and to select non-clonal isolates with unusual phenotypes for whole genome sequencing (WGS). Six classes of antimicrobials were assessed with disk diffusion and colistin with minimum inhibitory concentration (MIC) by broth microdilution. In order to confirm the discrepant phenotypes, MIC by agar dilution was performed for the beta-lactams aztreonam, ceftazidime, cefepime and the aminoglycoside neomycin. The genus Pseudomonas was confirmed by matrix-assisted laser desorption/ionization - time of flight (MALDI-TOF) and the clonal relationships were examined using repetitive extragenic palindromic polymerase chain reaction (BOX-PCR), from which 14 strains were selected for WGS. Antimicrobial susceptibility testing revealed that all strains were susceptible to neomycin and exhibited varying degrees of intermediate or full resistance to aztreonam and colistin. Additionally, 11 strains demonstrated intermediate resistance to ceftazidime, and six were resistant to cefepime. The genomic analysis identified various efflux pumps, predominantly from the ABC transporter and resistance-nodulation-division families. Resistance genes were detected against eight classes of antimicrobials, listed by prevalence: beta-lactams, tetracyclines, polymyxins, aminoglycosides, fosmidomycin, fosfomycin, quinolones, and chloramphenicol. Genes associated with heavy-metal resistance, prophages, and adaptations to extreme environments were also investigated. One notable isolate exhibited not only the highest number of pathogenicity and resistance islands, but also presented a carbapenemase-encoding gene (bla (PFM-2)) in its genome. Overall, one plasmid was identified in a distinct isolate, which did not exhibit antimicrobial resistance determinants. The genotypic and phenotypic findings are consistent, suggesting that efflux pumps play a critical role in antimicrobial extrusion. This study offers valuable insight into the evolution of antimicrobial resistance in P. fluorescens, particularly in extreme environments, such as Antarctica. By exploring the antimicrobial resistance mechanisms in P. fluorescens, the study sheds light on how isolated ecosystems drive the natural evolution of resistance genes.202540099188
5691190.9971Rapid and Accurate Antibiotic Susceptibility Determination of tet(X)-Positive E. coli Using RNA Biomarkers. The emergence and prevalence of novel plasmid-mediated tigecycline resistance genes, namely, tet(X) and their variants, pose a serious threat to public health worldwide. Rapid and accurate antibiotic susceptibility testing (AST) that can simultaneously detect the genotype and phenotype of tet(X)-positive bacteria may contribute to the deployment of an effective antibiotic arsenal, mortality reduction, and a decrease in the use of broad-spectrum antimicrobial agents. However, current bacterial growth-based AST methods, such as broth microdilution, are time consuming and delay the prompt treatment of infectious diseases. Here, we developed a rapid RNA-based AST (RBAST) assay to effectively distinguish tet(X)-positive and -negative strains. RBAST works by detecting specific mRNA expression signatures in bacteria after short-term tigecycline exposure. As a proof of concept, a panel of clinical isolates was characterized successfully by using the RBAST method, with a 3-h assay time and 87.9% accuracy (95% confidence interval [CI], 71.8% to 96.6%). Altogether, our findings suggest that RNA signatures upon antibiotic exposure are promising biomarkers for the development of rapid AST, which could inform early antibiotic choices. IMPORTANCE Infections caused by multidrug-resistant (MDR) Gram-negative pathogens are an increasing threat to global health. Tigecycline is one of the last-resort antibiotics for the treatment of these complicated infections; however, the emergence of plasmid-encoded tigecycline resistance genes, namely, tet(X), severely diminishes its clinical efficacy. Currently, there is a lack of rapid and accurate antibiotic susceptibility testing (AST) for the detection of tet(X)-positive bacteria. In this study, we developed a rapid and robust RNA-based antibiotic susceptibility determination (RBAST) assay to effectively distinguish tet(X)-negative and -positive strains using specific RNA biomarkers in bacteria after tigecycline exposure. Using this RBAST method, we successfully characterized a set of clinical strains in 3 h. Our data indicate that the RBAST assay is useful for identifying tet(X)-positive Escherichia coli.202134704829