Klebsiella pneumoniae is an important multidrug-resistant (MDR) pathogen affecting humans and a major source for hospital infections associated with high morbidity and mortality due to limited treatment options. We summarize the wide resistome of this pathogen, which encompasses plentiful chromosomal and plasmid-encoded antibiotic resistance genes (ARGs). Under antibiotic selective pressure, K. pneumoniae continuously accumulates ARGs, by de novo mutations, and via acquisition of plasmids and transferable genetic elements, leading to extremely drug resistant (XDR) strains harboring a 'super resistome'. In the last two decades, numerous high-risk (HiR) MDR and XDR K. pneumoniae sequence types have emerged showing superior ability to cause multicontinent outbreaks, and continuous global dissemination. The data highlight the complex evolution of MDR and XDR K. pneumoniae, involving transfer and spread of ARGs, and epidemic plasmids in highly disseminating successful clones. With the worldwide catastrophe of antibiotic resistance and the urgent need to identify the main pathogens that pose a threat on the future of infectious diseases, further studies are warranted to determine the epidemic traits and plasmid acquisition in K. pneumoniae. There is a need for future genomic and translational studies to decipher specific targets in HiR clones to design targeted prevention and treatment.

extraintestinal multidrug resistant Escherichia coli sequence type (ST) 131 is a worldwide pandemic pathogen and a major cause of urinary tract and bloodstream infections. the role of this pandemic lineage in multidrug resistance plasmid dissemination is still scarce. We herein performed a metaanalysis on E. coli ST131 whole-genome sequence (WGS) databases to unravel ST131 plasmidome and specifically to decipher CTX-M encoding plasmids-clade associations. We mined 880 ST131 WGS data and proved that CTX-M-27-encoding IncF[F1:A2:B20] (Group1) plasmids are strictly found in clade C1, whereas CTX-M-15-encoding IncF[F2:A1:B-] (Group2) plasmids exist only in clade C2 suggesting strong plasmid-clade adaptations. Specific Col-like replicons (Col156, Col(MG828), and Col8282) were also found to be clade C1-associated. BLAST-based search revealed that Group1 and Group2 plasmids are narrow-host-range and restricted to E.coli. Among a collection of 20 newly sequenced Israeli ST131 CTX-M-encoding plasmids (2003-2016), Group1 and Group2 plasmids were dominant and associated with the expected clades. We found, for the first time in ST131, a CTX-M-15-encoding phage-like plasmid group (Group3) and followed its spread in the WGS data. This study offers a comprehensive way to decipher plasmid-bacterium associations and demonstrates that the CTX-M-encoding ST131 Group1 and Group2 plasmids are clade-restricted and presumably less transmissible, potentially contributing to ST131 clonal superiority. Multidrug resistant ESBL-producing Escherichia coli ST131 is a high-risk extraintestinal pathogenic E. coli (ExPEC) lineage that has spread explosively throughout the world 1-4 , mostly by patient-to-patient spread and contaminated food 5. This lineage may colonize healthy humans and animals 6 and is the causative pathogen of urinary tract and bloodstream infections 7-9. E. coli ST131 nowadays is recognized as the major E. coli lineage responsible for the spread of multidrug resistance (MDR) 3 and in specific CTX-M ESBL genes 10. The global expansion of this lineage is driven by spread of two major fluoroquinolone-resistance clades which possess fimH30 allele-H30R/C1 and H30Rx/C2, where the later harbors bla CTX-M-15 ESBL gene 4,11-13. Plasmid-bacterium associations in ST131 clades connected CTX-M-15-encoding IncF[F2:A1:B-] (FAB formula) plasmids to clade C2 11,12,14,15 , and non-CTX-M-15-encoding IncF[F1:A2:B20] plasmids to clade C1 4,11. The major CTX-M genes described in clade С1 were CTX-M-14 and −27 alleles of the CTX-M-9 group 16 , with this later allele-C1-M27 16 , recognized recently as a new subclade with global occurrence 17,18. MDR often disseminates in bacteria through horizontal gene transfer of mobile genetic elements, especially plasmid transmission 19,20. However, if there are highly adaptive plasmid-bacterium associations within ST131 lineage, the main driving force for MDR spread related to this pandemic clone is due to clonal expansion rather than to horizontal plasmid spread 11,12,15. The existence of successful plasmid-S...

Simple SummaryMultidrug-resistant (MDR) Enterobacteriaceae are becoming a major worldwide concern in human and veterinary medicine, mainly due to the production of extended-spectrum β-lactamases (ESBLs). These bacteria have been investigated in adult horses, but not in neonatal foals. In this study, we investigated extended-spectrum β-lactamase Enterobacteriaceae (ESBL-E) shedding and infection in hospitalized mares and their neonatal foals. Overall, we sampled rectal swabs from 55 pairs of mares and their foals on admission, and 33 of them were re-sampled on the 3rd day of hospitalization. We also collected clinical samples, when available. We found that shedding rates and bacterial species diversity increased significantly during hospitalization, both in mares and foals. On admission to hospital, foals’ shedding was associated with umbilical infection. During hospitalization, it was associated with ampicillin treatment. Foals’ shedding was independent of their mares’ shedding. Four foals were infected with ESBL-E strains, including umbilical infections and wounds. We suggest further investigation and surveillance of ESBL-E in neonatal foals, in order to reduce resistance rates and infections.AbstractExtended-spectrum β-lactamase Enterobacteriaceae (ESBL-E) have been investigated in adult horses, but not in foals. We aimed to determine shedding and infection in neonatal foals and mares. Rectal swabs were sampled from mare and foal pairs on admission and on the 3rd day of hospitalization; enriched, plated, and bacteria were verified for ESBL production. Identification and antibiotic susceptibility profiles were determined (Vitek2). Genotyping was performed by multilocus sequence typing (MLST). Genes were identified by PCR and Sanger sequencing. Medical data were analyzed for risk factors (SPSS). On admission, 55 pairs were sampled, of which 33 pairs were re-sampled. Shedding rates on admission in foals and mares were 33% (95% CI 21–47%) and 16% (95% CI 8–29%), respectively, and during hospitalization, these increased significantly to 85% (95% CI 70–94%) and 58% (95% CI 40–73%), respectively. Foal shedding was associated with umbilical infection on admission (P = 0.016) and with ampicillin treatment during hospitalization (p = 0.011), and was independent of the mare’s shedding. The most common ESBL-E was Escherichia coli. During hospitalization, species diversity increased. Four foals were infected with ESBL-E strains, including umbilical infections and wounds. This study substantiates an alarming prevalence of shedding in neonatal foals, which should be further investigated in order to reduce resistance rates.

The emergence of extended-spectrum β-lactamase (ESBL)-producing multidrug resistant Klebsiella pneumoniae causing community urinary tract infections (CA-UTI) in healthy women undermines effective treatment and poses a public health concern. We performed a comprehensive genomic analysis (Illumina and MinION) and virulence studies using Caenorhabditis elegans nematodes to evaluate KpnU95, a blaCTX-M-15-producing CA-UTI K. pneumoniae strain. Whole genome sequencing identified KpnU95 as sequence type 1412 and revealed the chromosomal and plasmid-encoding resistome, virulome and persistence features. KpnU95 possess a wide virulome and caused complete C. elegans killing. The strain harbored a single novel 180.3Kb IncFIB(K) plasmid (pKpnU95), which encodes ten antibiotic resistance genes, including blaCTX-M-15 and qnrS1 alongside a wide persistome encoding heavy metal and UV resistance. Plasmid curing and reconstitution were used for loss and gain studies to evaluate its role on bacterial resistance, fitness and virulence. Plasmid curing abolished the ESBL phenotype, decreased ciprofloxacin MIC and improved bacterial fitness in artificial urine accompanied with enhanced copper tolerance, without affecting bacterial virulence. Meta-analysis supported the uniqueness of pKpnU95 and revealed plasmid-ST1412 lineage adaptation. Overall, our findings provide translational data on a CA-UTI K. pneumoniae ST1412 strain and demonstrates that ESBL-encoding plasmids play key roles in multidrug resistance and in bacterial fitness and persistence.

Salmonella enterica is a major causative pathogen of human and animal gastroenteritis. Antibiotic resistant strains have emerged due to the production of extended-spectrum β-lactamases (ESBLs) posing a major health concern. With the increasing reports on ESBL-producing Enterobacterales that colonize companion animals, we aimed to investigate ESBL dissemination among ESBL-producing Salmonella enterica (ESBL-S) in hospitalized horses. We prospectively collected ESBL-S isolates from hospitalized horses in a Veterinary-Teaching Hospital during Dec 2015–Dec 2017. Selection criteria for ESBL-S were white colonies on CHROMagarESBL plates and an ESBL phenotypic confirmation. Salmonella enterica serovars were determined using the Kaufmann-White-Le-Minor serological scheme. ESBL-encoding plasmids were purified, transformed and compared using restriction fragment length polymorphism (RFLP). Whole genome sequencing (Illumina and MinION platforms) were performed for detailed phylogenetic and plasmid analyses. Twelve ESBL-S were included in this study. Molecular investigation and Sequence Read Archive (SRA) meta-analysis revealed the presence of three unique Salmonella enterica serovars, Cerro, Havana and Liverpool, all reported for the first time in horses. PFGE revealed the clonal spread of S. Cerro between seven horses. All twelve isolates carried blaCTX–M–3 and showed an identical multidrug resistance profile with co-resistance to trimethoprim/sulfamethoxazole and to aminoglycosides. Plasmid RFLP proved the inter-serovar horizontal spread of a single blaCTX–M–3-encoding plasmid. Complete sequence of a representative plasmid (S. Havana strain 373.3.1), designated pSEIL-3 was a -86.4 Kb IncM2 plasmid, that encoded nine antibiotic resistance genes. pSEIL-3 was virtually identical to pCTX-M3 from Citrobacter freundii, and showed high identity (>95%) to six other blaCTX–M–3 or blaNDM–1 IncM2 broad host range plasmids from various Enterobacterales of human origin. Using a specific six gene-based multiplex PCR, we detected pSEIL-3 in various Enterobacterales species that co-colonized the horses’ gut. Together, our findings show the alarming emergence of ESBL-S in hospitalized horses associated with gut shedding and foal morbidity and mortality. We demonstrated the dissemination of CTX-M-3 ESBL among different Salmonella enterica serovars due to transmission of a broad host range plasmid. This report highlights horses as a zoonotic reservoir for ESBL-S, including highly transmissible plasmids that may represent a ‘One-Health’ hazard. This risk calls for the implementation of infection control measures to monitor and control the spread of ESBL-S in hospitalized horses.