Antimicrobial Resistance Profiles and Molecular Characteristics of Extended-Spectrum ß-Lactamase-Producing Salmonella enterica Serovar Typhimurium Isolates from Food Animals During 2010-2021 in South Korea.

Extended-spectrum ß-lactamase (ESBL)-producing Salmonella is emerging as a worldwide public health concern. In this study, we aimed to investigate the antimicrobial resistance profiles and molecular characteristics of ESBL-producing Salmonella enterica serovar Typhimurium (S. Typhimurium). We obtained a total of 995 S. Typhimurium isolates from the feces and carcasses of pigs (n = 678), chickens (n = 202), and cattle (n = 115) during 2010-2021 in Korea. We found that 35 S. Typhimurium isolates (3.5%) showed resistance to ceftiofur: pigs (51.4%, 18/35) and cattle (42.9%, 15/35). All of the ceftiofur-resistant S. Typhimurium isolates demonstrated multidrug resistance. Moreover, ceftiofur-resistant S. Typhimurium isolates displayed significantly higher rates of resistance to chloramphenicol and trimethoprim/sulfamethoxazole than ceftiofur-susceptible S. Typhimurium isolates (p < 0.05). The ceftiofur-resistant S. Typhimurium isolates produced four different CTX-M-type ß-lactamase, comprising blaCTX-M-55 in the majority (51.4%, 18/35), followed by blaCTX-M-65 (28.6%, 10/35), blaCTX-M-14 (17.1%, 6/35), and blaCTX-M-1 (2.9%, 1/35). Among the 35 ceftiofur-resistant S. Typhimurium isolates, 16 blaCTX-M-55-positive isolates and one blaCTX-M-1-positive isolate were transferred to recipient Escherichia coli RG488 by conjugation. The predominantly found transposable units were blaCTX-M-55-orf477 (45.7%, 16/35), followed by blaCTX-M-65-IS903 (28.6%, 10/35) and blaCTX-M-14-IS903 (17.1%, 6/35). Ceftiofur-resistant S. Typhimurium represented 19 types, with types P1-19 (22.9%, 8/35) and P12-34 (22.9%, 8/35) making up the majority and being found in most farms nationwide. Sequence types (STs) were different by animal species: ST19 (48.6%, 17/35) and ST34 (42.9%, 15/35) were mostly found STs in pigs and cattle, respectively. These findings showed that food animals, especially pigs and cattle, act as reservoirs of blaCTX-M-harboring S. Typhimurium that can potentially be spread to humans.

Nationwide surveillance and characterization of the third-generation cephalosporin-resistant Salmonella enterica serovar infantis isolated from chickens in South Korea between 2010 and 2022.

The occurrence of extended-spectrum ß-lactamase (ESBL)/AmpC ß-lactamase-producing Salmonella conferring resistance to third-generation cephalosporin has emerged as a global public health concern. In this study, we aimed to investigate the prevalence and molecular characterization of third-generation cephalosporin-resistant Salmonella enterica serovar Infantis. In total, 409 S. Infatis isolates were collected from the feces and carcasses of healthy and diseased food animals, including chickens (n = 348), pigs (n = 48), cattle (n = 8), and ducks (n = 5) between 2010 and 2022 nationwide in South Korea. Among them, 61.9 % (253/409) of S. Infantis strains displayed resistance to ceftiofur, with the most resistant isolates obtained from chickens (98.4 %, 249/253). Moreover, S. Infantis isolates showed high resistance (47.7-67.2 %) to streptomycin, ampicillin, nalidixic acid, sulfisoxazole, chloramphenicol, tetracycline, and trimethoprim/sulfamethoxazole. Additionally, the multidrug resistance (MDR) was significantly greater in the ceftiofur-resistant isolates compared to the ceftiofur-susceptible isolates (p < 0.05). All the ceftiofur-resistant S. Infantis strains produced CTX-M/CMY-2 ß-lactamase enzymes, with bla CTX-M-65 comprising the most (98.4 %, 249/253), followed by bla CTX-M-15 (1.2 %, 3/253), and bla CMY-2 (0.4 %, 1/253). The ceftiofur-resistant S. Infantis belonged to 37 different pulsotypes, with X1A1 (26.1 %, 66/253), X1A2 (20.9 %, 53/253), and X5A3 (9.1 %) being the most prevalent, representing a total of 56.1 % (142/253). Furthermore, the S. Infantis sequence type (ST)32 was the most common, accounting for 91.9 % (34/37) of the three distinct STs (ST32, ST16, and ST11) detected across farms located in various provinces nationwide. Most of the bla CMX-M-65 genes (77.5 %, 193/249), all of the bla CTX-M-15 genes (100 %, 3/3), and the bla CMY-2 gene (100 %, 1/1) were transferred to the recipient E. coli RG488 by conjugation. In addition, the majority of the transconjugants (98.9 %, 191/193) containing bla CTX-M-65 genes belong to the IncFIB replicon type, playing an important role in the quick and widespread dissemination of S. Infantis. Thus, ceftiofur-resistant S. Infantis carrying the ß-lactamase genes in chickens has the potential to be transmitted to humans.

Emergence of a clinical Klebsiella pneumoniae harboring an acrAB-tolC in chromosome and carrying the two repetitive tandem core structures for bla KPC-2 and bla CTX-M-65 in a plasmid.

Objective: The emergence of clinical Klebsiella pneumoniae strains harboring acrAB-tolC genes in the chromosome, along with the presence of two repetitive tandem core structures for bla KPC-2 and bla CTX-M-65 genes on a plasmid, has presented a significant clinical challenge. Methods: In order to study the detailed genetic features of K. pneumoniae strain SC35, both the bacterial chromosome and plasmids were sequenced using Illumina and nanopore platforms. Furthermore, bioinformatics methods were employed to analyze the mobile genetic elements associated with antibiotic resistance genes. Results: K. pneumoniae strain SC35 was found to possess a class A beta-lactamase and demonstrated resistance to all tested antibiotics. This resistance was attributed to the presence of efflux pump genes, specifically acrAB-tolC, on the SC35 chromosome. Additionally, the SC35 plasmid p1 carried the two repetitive tandem core structures for bla KPC-2 and bla CTX-M-65, as well as bla TEM-1 with rmtB, which shared overlapping structures with mobile genetic elements as In413, Tn3, and TnAs3. Through plasmid transfer assays, it was determined that the SC35 plasmid p1 could be successfully transferred with an average conjugation frequency of 6.85 × 10-4. Conclusion: The structure of the SC35 plasmid p1 appears to have evolved in correlation with other plasmids such as pKPC2_130119, pDD01754-2, and F4_plasmid pA. The infectious strain SC35 exhibits no susceptibility to tested antibioticst, thus effective measures should be taken to prevent the spread and epidemic of this strain.

Genetic analysis of resistance and virulence characteristics of clinical multidrug-resistant Proteus mirabilis isolates.

Objective: Proteus mirabilis is the one of most important pathogens of catheter-associated urinary tract infections. The emergence of multidrug-resistant (MDR) P. mirabilis severely limits antibiotic treatments, which poses a public health risk. This study aims to investigate the resistance characteristics and virulence potential for a collection of P. mirabilis clinical isolates. Methods and results: Antibiotic susceptibility testing revealed fourteen MDR strains, which showed high resistance to most ß-lactams and trimethoprim/sulfamethoxazole, and a lesser extent to quinolones. All the MDR strains were sensitive to carbapenems (except imipenem), ceftazidime, and amikacin, and most of them were also sensitive to aminoglycosides. The obtained MDR isolates were sequenced using an Illumina HiSeq. The core genome-based phylogenetic tree reveals the high genetic diversity of these MDR P. mirabilis isolates and highlights the possibility of clonal spread of them across China. Mobile genetic elements SXT/R391 ICEs were commonly (10/14) detected in these MDR P. mirabilis strains, whereas the presence of resistance island PmGRI1 and plasmid was sporadic. All ICEs except for ICEPmiChn31006 carried abundant antimicrobial resistance genes (ARGs) in the HS4 region, including the extended-spectrum ß-lactamase (ESBL) gene blaCTX-M-65. ICEPmiChn31006 contained the sole ARG blaCMY-2 and was nearly identical to the global epidemic ICEPmiJpn1. The findings highlight the important roles of ICEs in mediating the spread of ARGs in P. mirabilis strains. Additionally, these MDR P. mirabilis strains have great virulence potential as they exhibited significant virulence-related phenotypes including strong crystalline biofilm, hemolysis, urease production, and robust swarming motility, and harbored abundant virulence genes. Conclusion: In conclusion, the prevalence of MDR P. mirabilis with high virulence potential poses an urgent threat to public health. Intensive monitoring is needed to reduce the incidence of infections by MDR P. mirabilis.

Epidemiological characteristics of SHV, cmlv, and FosA6-producing carbapenem-resistant Klebsiella pneumoniae based on whole genome sequences in Jiangsu, China.

Carbapenem-resistant Klebsiella pneumoniae (CRKP), particularly those with high virulence, cause invasive disease in clinical settings. An epidemiological investigation was conducted on the evolution, virulence, and antimicrobial resistance of CRKP isolates in two tertiary teaching hospitals in Jiangsu, China from November 2020 to December 2021. There were 31 different CRKP strains discovered. We performed whole genome sequencing (WGS) on 13 SHV, cmlv, and FosA6-producing CRKP to reveal molecular characteristics. Five ST15/ST11 isolates had CRISPR-Cas systems. By conjugation tests, KPC-2 can be transmitted horizontally to E. coil. A conjugative pHN7A8-related multi-resistance plasmid (KPC-2, blaCTX-M-65, blaTEM-1, fosA3, catII, and rmtB) was first discovered in CRKP clinical isolates. Using bacteriological testing, a serum killing assay, and an infection model with Galleria mellonella, three ST11-K64 KPC-2 generating carbapenem-resistant hypervirulent Klebsiella pneumoniae (CR-hvKP) were identified. These strains harbored a virulent plasmid and an IncFII-family pKPC/pHN7A8 conjugative plasmid, which led to hypervirulence and resistance. One of these CR-hvKPs, which co-harbored KPC-2, NDM-6, SHV-182, SHV-64, and blaCTX-M-122 genes, was first discovered. Importantly, this CR-hvKP strain also produced biofilm and had non-inferior fitness. The widespread use of ceftazidime/avibactam might provide this CR-hvKP with a selective advantage; hence, immediate action is required to stop its dissemination. Another important finding is the novel ST6136 in K. pneumoniae. Finally, the sterilization efficiency rates of Fe2C nanoparticles in CRKP were more than 98%. Furthermore, our novel antibacterial Fe2C nanoparticles may also provide a therapeutic strategy for infections.

Co-Existence of KPC-2, LAP-2, and CTX-M-65 in an ST1469 Multidrug-Resistant Klebsiella pneumoniae Strain in China.

Purpose: Beta-lactamase-producing Klebsiella pneumoniae is common in the clinic, but research associated with the co-existence of KPC-2, LAP-2, and CTX-M-65 in K. pneumoniae is still rare. In this study, the phenotypic and genetic characteristics of a multidrug-resistant K. pneumoniae strain SJ25 co-harboring bla KPC-2, bla LAP-2, and bla CTX-M-65 with rare ST1469 were investigated. Methods and Results: Antimicrobial susceptibility testing revealed that strain SJ25 was resistant to various common antibiotics, except ciprofloxacin, fosfomycin, colistin, and tigecycline. Whole-genome analysis revealed that strain SJ25 carries a variety of antimicrobial resistance genes and virulence determinants. Plasmid analysis confirmed that the bla KPC-2 and bla CTX-M-65 genes were located on an ~136 kb transferrable IncFII/IncR plasmid and that bla LAP-2 was located on an untypeable plasmid. Conclusion: Our findings emphasized the need for continuous surveillance of ß-lactamase-bearing K. pneumoniae in the clinic to control potential dissemination and outbreak.

Molecular Characterization of Salmonella Detected along the Broiler Production Chain in Trinidad and Tobago.

This cross-sectional study determined the serovars, antimicrobial resistance genes, and virulence factors of Salmonella isolated from hatcheries, broiler farms, processing plants, and retail outlets in Trinidad and Tobago. Salmonella in silico serotyping detected 23 different serovars where Kentucky 20.5% (30/146), Javiana 19.2% (28/146), Infantis 13.7% (20/146), and Albany 8.9% (13/146) were the predominant serovars. There was a 76.0% (111/146) agreement between serotyping results using traditional conventional methods and whole-genome sequencing (WGS) in in silico analysis. In silico identification of antimicrobial resistance genes conferring resistance to aminoglycosides, cephalosporins, peptides, sulfonamides, and antiseptics were detected. Multidrug resistance (MDR) was detected in 6.8% (10/146) of the isolates of which 100% originated from broiler farms. Overall, virulence factors associated with secretion systems and fimbrial adherence determinants accounted for 69.3% (3091/4463), and 29.2% (1302/4463) counts, respectively. Ten of 20 isolates of serovar Infantis (50.0%) showed MDR and contained the blaCTX-M-65 gene. This is the first molecular characterization of Salmonella isolates detected along the entire broiler production continuum in the Caribbean region using WGS. The availability of these genomes will help future source tracking during epidemiological investigations associated with Salmonella foodborne outbreaks in the region and worldwide.

Genomic Analysis of Two MDR Isolates of Salmonella enterica Serovar Infantis from a Spanish Hospital Bearing the blaCTX-M-65 Gene with or without fosA3 in pESI-like Plasmids.

Salmonella enterica serovar Infantis (S. Infantis) is a broiler-associated pathogen which ranks in the fourth position as a cause of human salmonellosis in the European Union. Here, we report a comparative genomic analysis of two clinical S. Infantis isolates recovered in Spain from children who just returned from Peru. The isolates were selected on the basis of resistance to cefotaxime, one of the antibiotics of choice for treatment of S. enterica infections. Antimicrobial susceptibility testing demonstrated that they were resistant to eight classes of antimicrobial agents: penicillins, cephalosporins, phenicols, aminoglycosides, tetracyclines, inhibitors of folate synthesis, (fluoro)quinolones and nitrofurans, and one of them was also resistant to fosfomycin. As shown by whole-genome sequence analysis, each isolate carried a pESI-like megaplasmid of ca. 300 kb harboring multiple resistance genes [blaCTX-M-65, aph(4)-Ia, aac(3)-IVa, aph(3')-Ia, floR, dfrA14, sul1, tet(A), aadA1 ± fosA3], as well as genes for resistance to heavy metals and disinfectants (mer, ars and qacEΔ1). These genes were distributed in two complex regions, separated by DNA belonging to the plasmid backbone, and associated with a wealth of transposable elements. The two isolates had a D87Y amino acid substitution in the GyrA protein, and truncated variants of the nitroreductase genes nfsA and nsfB, accounting for chromosomally encoded resistances to nalidixic acid and nitrofurantoin, respectively. The two S. Infantis isolates were assigned to sequence type ST32 by in silico multilocus sequence typing (MLST). Phylogenetic analysis revealed that they were closely related, differing only by 12 SNPs, although they were recovered from different children two years apart. They were also genetically similar to blaCTX-M-65-positive ± fosA3 isolates obtained from humans and along the poultry production chain in the USA, South America, as well as from humans in several European countries, usually associated with a travel history to America. However, this is the first time that the S. Infantis blaCTX-M-65 ± fosA3 MDR clone has been reported in Spain.

Successful Treatment of Severe Post-craniotomy Meningitis Caused by an Escherichia coli Sequence Type 410 Strain Coharboring bla NDM - 5 and bla CTX - M - 65.

Intracranial infections caused by multidrug-resistant Gram-negative bacterium have led to considerable mortality due to extremely limited treatment options. Herein, we firstly reported a clinical carbapenem-resistant Escherichia coli isolate coharboring bla NDM - 5 and bla CTX - M - 65 from a patient with post-craniotomy meningitis. The carbapenem-resistant Escherichia coli strain CNEC001 belonging to Sequence Type 410 was only susceptible to amikacin and tigecycline, both of which have poor penetration through the blood-brain barrier (BBB). The bla CTX - M - 65 gene was expressed on a 135,794 bp IncY plasmid. The bla NDM - 5 gene was located on a genomic island region of an IncX3-type plasmid pNDM5-CNEC001. Based on the characteristics of the strain, we presented the successful treatment protocol of intravenous (IV) tigecycline and amikacin combined with intrathecal (ITH) amikacin in this study. Intracranial infection caused by Escherichia coli coharboring bla NDM - 5 and bla CTX - M - 65 is rare and fatal. Continuous surveillance and infection control measures for such strain need critical attention in clinical settings.

Incidence and Characterization of Salmonella Isolates From Raw Meat Products Sold at Small Markets in Hubei Province, China.

Salmonella is a leading cause of foodborne disease and is often associated with the consumption of foods of animal origin. In this study, sixty-six Salmonella isolates were obtained from 631 raw meat samples purchased at small retail suppliers in Hubei Province, China. The most prevalent Salmonella serotypes were Thompson (18.2%) and Agona (13.6%). Frequent antimicrobial resistance was observed for the sulfonamides (43.9%), tetracycline (43.9%), and the ß-lactams amoxicillin and ampicillin (36.4% for each). Interestingly, a high incidence of resistance to cephazolin was observed in strains of the most common serotype, S. Thompson. Class I integrons were found in 27.3% (18/66) of the isolates and five of these integrons contained different gene cassettes (aacA4C-arr-3-dfr2, dfrA12-aadA21, aadA2, dfrA12-aadA2, dfr17-aadA5). Additional antimicrobial resistance genes, including bla TEM-1, bla CTX-M-65, bla CTX-M-15, qnrB, and qnrS, were also identified among these Salmonella isolates. Results of replicon typing and conjugation experiments revealed that an integron with qnrB and bla CTX-M-15 genes was present on incH12 mobile plasmid in S. Thompson strain. Multilocus sequence typing (MLST) analysis revealed 32 sequence types, indicating that these isolates were phenotypically and genetically diverse, among which ST26 (18.2%) and ST541 (12.1%) were the predominant sequence types. The integrons, along with multiple antimicrobial resistance genes on mobile plasmids, are likely contributors to the dissemination of multidrug resistance in Salmonella.

Salmonella enterica serovar Infantis from Food and Human Infections, Switzerland, 2010-2015: Poultry-Related Multidrug Resistant Clones and an Emerging ESBL Producing Clonal Lineage.

Objectives: The aim of this study was to characterize a collection of 520 Salmonella enterica serovar Infantis strains isolated from food (poultry meat), human infections and environmental sources from the years 2010, 2013 and 2015 in Switzerland. Methods: We performed antimicrobial susceptibility testing and pulsed-field gel electrophoresis (PFGE) analysis on all 520 S. Infantis isolates, and whole genome sequencing (WGS) on 32 selected isolates. Results: The majority (74.8%) of the isolates was multidrug resistant (MDR). PFGE analysis revealed that 270 (51.9%) isolates shared an identity of 90%. All isolates subjected to WGS belonged to sequence type (ST) 32 or a double-locus variant thereof (one isolate). Seven (21.9%) of the sequenced isolates were phylogenetically related to the broiler-associated clone B that emerged in Hungary and subsequently spread within and outside of Europe. In addition, three isolates harboring blaCTX-M-65 on a predicted large (∼320 kb) plasmid grouped in a distinct cluster. Conclusion: This study documents the presence of the Hungarian clone B and related clones in food and human isolates between 2010 and 2015, and the emergence of a blaCTX-M-65 harboring MDR S. serovar Infantis lineage.