Ceftazidime/avibactam resistance is associated with PER-3-producing ST309 lineage in Chilean clinical isolates of non-carbapenemase producing Pseudomonas aeruginosa.

Introduction: Ceftazidime/avibactam (CZA) is indicated against multidrug-resistant Pseudomonas aeruginosa, particularly those that are carbapenem resistant. CZA resistance in P. aeruginosa producing PER, a class A extended-spectrum ß-lactamase, has been well documented in vitro. However, data regarding clinical isolates are scarce. Our aim was to analyze the contribution of PER to CZA resistance in non-carbapenemase-producing P. aeruginosa clinical isolates that were ceftazidime and/or carbapenem non-susceptible. Methods: Antimicrobial susceptibility was determined through agar dilution and broth microdilution, while bla PER gene was screened through PCR. All PER-positive isolates and five PER-negative isolates were analyzed through Whole Genome Sequencing. The mutational resistome associated to CZA resistance was determined through sequence analysis of genes coding for PBPs 1b, 3 and 4, MexAB-OprM regulators MexZ, MexR, NalC and NalD, AmpC regulators AmpD and AmpR, and OprD porin. Loss of bla PER-3 gene was induced in a PER-positive isolate by successive passages at 43°C without antibiotics. Results: Twenty-six of 287 isolates studied (9.1%) were CZA-resistant. Thirteen of 26 CZA-resistant isolates (50%) carried bla PER. One isolate carried bla PER but was CZA-susceptible. PER-producing isolates had significantly higher MICs for CZA, amikacin, gentamicin, ceftazidime, meropenem and ciprofloxacin than non-PER-producing isolates. All PER-producing isolates were ST309 and their bla PER-3 gene was associated to ISCR1, an insertion sequence known to mobilize adjacent DNA. PER-negative isolates were classified as ST41, ST235 (two isolates), ST395 and ST253. PER-negative isolates carried genes for narrow-spectrum ß-lactamases and the mutational resistome showed that all isolates had one major alteration in at least one of the genes analyzed. Loss of bla PER-3 gene restored susceptibility to CZA, ceftolozane/tazobactam and other ß-lactamsin the in vitro evolved isolate. Discussion: PER-3-producing ST309 P. aeruginosa is a successful multidrug-resistant clone with blaPER-3 gene implicated in resistance to CZA and other ß-lactams.

Genomic features of an extensively drug-resistant and NDM-1-positive Klebsiella pneumoniae ST340 isolated from river water.

The environmental contamination plays a significant role in the emergence of antimicrobial resistance. In this study, we report a genomic analysis of an extensively drug-resistant and blaNDM-1-producing Klebsiella pneumoniae (EW807) strain recovered from a surface water sample. Strain EW807 belonged to sequence type (ST) 340 and serotype O4:KL15, a high-risk clone of the clonal group 258. This strain carried a broad resistome, including blaNDM-1 and blaCTX-M-15. The core genome multilocus sequence typing phylogenetic analysis revealed that the EW807 strain was most related to strains from Brazil and the USA. An IncX3 plasmid was identified harboring the blaNDM-1 gene, while an IncFIB(K) plasmid was detected carrying the blaCTX-M-15 in addition to multidrug resistance and multimetal tolerance regions. IncX3 and IncFIB(K) plasmids shared high similarity with plasmids from a human in China and a dog in Brazil, respectively. The regions harboring the blaNDM-1 and blaCTX-M-15 genes contained sequences from the Tn3 family. These findings suggest that IncX3 plasmid could play a role in the spread of NDM-1 in a post-pandemic scenario. To the best of our knowledge, this is the first report of blaNDM-1-producing K. pneumoniae ST340 O4:KL15 strain in the environment. Therefore, the presence of high-risk clones of K. pneumoniae carrying carbapenemases in the environment requires strict surveillance.

Empirical treatment and mortality in bacteremia due to extended spectrum ß-lactamase producing Enterobacterales (ESßL-E), a retrospective cross-sectional study in a tertiary referral hospital from Colombia.

BACKGROUND: Infections caused by extended spectrum ß-lactamase (ESßL) producing bacteria are common and problematic. When they cause bloodstream infections, they are associated with significant morbidity and mortality. METHODS: A retrospective cross-sectional observational study was conducted in a single center in Pereira, Colombia. It included people hospitalized with bacteremia due to gram-negative bacilli with the extended-spectrum ß-lactamase producing phenotype. A logistic regression analysis was constructed. Clinical characteristics and risk factors for death from sepsis were established. RESULTS: The prevalence of bacteremia due to Enterobacterales with extended-spectrum ß-lactamase producing phenotype was 17%. 110 patients were analyzed. Most patients were men (62%) with a median age of 58 years, hospital mortality was 38%. Admission to intensive care was 45%. The following risk factors for mortality were established: shock requiring vasoactive support, Pitt score > 3 points, and not having an infectious disease consultation (IDC). CONCLUSIONS: bacteremia due to Enterobacterales with extended-spectrum ß-lactamase producing phenotype have a high mortality. Early recognition of sepsis, identification of risk factors for antimicrobial resistance, and prompt initiation of appropriate empiric antibiotic treatment are important. An infectious disease consultation may help improve outcomes.

Removal of Extended-Spectrum Beta-Lactamase-Producing Escherichia coli, ST98, in Water for Human Consumption by Black Ceramic Water Filters in Low-Income Ecuadorian Highlands.

Fecal contamination in natural water sources is a common problem in low-income countries. Several health risks are associated with unprotected water sources, such as gastrointestinal infections caused by parasites, viruses, and bacteria. Moreover, antibiotic-resistant bacteria in water sources have become an increasing problem worldwide. This study aimed to evaluate the bacterial pathogens present in water within a rural context in Ecuador, along with the efficiency of black ceramic water filters (BCWFs) as a sustainable household water treatment. We monitored five natural water sources that were used for human consumption in the highlands of Ecuador and analyzed the total coliforms and E. coli before and after BCWF installation. The results indicated a variable bacterial contamination (29-300 colony-forming units/100mL) in all unfiltered samples, and they were considered as high risk for human consumption, but after filtration, no bacteria were present. Moreover, extended-spectrum beta-lactamase-producing E. coli with blaTEM, blaCTX-M9, and blaCTX-M1 genes, and two E. coli classified in the clonal complex ST10 (ST98) were detected in two of the locations sampled; these strains can severely impact public health. The clonal complex ST10, found in the E. coli isolates, possesses the potential to spread bacteria-resistant genes to humans and animals. The results of the use of BCWFs, however, argue for the filters' potential impact within those contexts, as the BCWFs completely removed even antibiotic-resistant contaminants from the water.

Genomic features of a multidrug-resistant and mercury-tolerant environmental Escherichia coli recovered after a mining dam disaster in South America.

Mining dam disasters contribute to the contamination of aquatic environments, impacting associated ecosystems and wildlife. A multidrug-resistant Escherichia coli strain (B2C) was isolated from a river water sample in Brazil after the Mariana mining dam disaster. The genome was sequenced using the Illumina MiSeq platform, and de novo assembled using Unicycler. Resistome, virulome, and plasmidome were predicted using bioinformatics tools. Data analysis revealed that E. coli B2C belonged to sequence type ST219 and phylogroup E. Strikingly, a broad resistome (antibiotics, hazardous heavy metals, and biocides) was predicted, including the presence of the clinically relevant blaCTX-M-2 extended-spectrum ß-lactamase (ESBL) gene, qacE∆1 efflux pump gene, and the mer (mercury resistance) operon. SNP-based analysis revealed that environmental E. coli B2C was clustered along to ESBL-negative E. coli strains of ST219 isolated between 1980 and 2021 from livestock in the United States of America. Acquisition of clinically relevant genes by ST219 seems to be a recent genetic event related to anthropogenic activities, where polluted water environments may contribute to its dissemination at the human-animal-environment interface. In addition, the presence of genes conferring resistance to heavy metals could be related to environmental pollution from mining activities. Antimicrobial resistance genes could be essential biomarkers of environmental exposure to human and mining pollution.

CTX-M-15-producing Klebsiella pneumoniae ST273 associated with nasal infection in a domestic cat.

OBJECTIVES: The aim of this study was to investigate the genetic context of expanded-spectrum ß-lactam resistance in a Klebsiella pneumoniae strain causing a hard-to-treat nasal infection in a domestic cat. METHODS: A K. pneumoniae isolate was recovered from a 4-year-old male cat hospitalised in a veterinary hospital in Paraíba, Northeastern Brazil. Following phenotypic confirmation of multidrug resistance by the disk diffusion method, the genome was sequenced using an Illumina MiSeq system. Multilocus sequence typing (MLST) and structural features related to antimicrobial resistance were determined by downstream bioinformatics analyses. RESULTS: The strain was confirmed as sequence type 273 (ST273) K. pneumoniae harbouring a variety of genes conferring antimicrobial resistance to phenicols tetracyclines, aminoglycosides, ß-lactams, fosfomycin, sulfonamides and quinolones. Two plasmids were identified. Plasmid p114PB_I co-harboured a set of plasmid-borne resistance genes [blaCTX-M-15, blaTEM-1, qnrS1, tetD, tetR, sul2, aph(6)-Id, aph(3'') and cat2]. Notably, the multiresistance region was characterised as a chimeric plasmid structure sharing high sequence homology with several plasmids from Enterobacteriaceae. The second plasmid (p114PB_II) was characterised as a plasmid present in many genomes belonging to K. pneumoniae. CONCLUSION: The genetic context of the plasmid sequences harboured by a veterinary pathogenic K. pneumoniae isolate reveals the high complexity of horizontal gene transfer mechanisms in the acquisition of antimicrobial resistance genes. The emergence, dissemination and evolution of antimicrobial resistance must be investigated from a One Health perspective.

Rapid Detection of Carbapenemase and Extended-Spectrum ß-Lactamase Producing Gram-Negative Bacteria Directly from Positive Blood Cultures Using a Novel Protocol.

BACKGROUND: Early and adequate antibiotic treatment is the cornerstone of improving clinical outcomes in patients with bloodstream infections (BSI). Delays in appropriate antimicrobial therapy have catastrophic consequences for patients with BSI. Microbiological characterization of multi-drug resistant pathogens (MDRP) allows clinicians to provide appropriate treatments. Current microbiologic techniques may take up to 96 h to identify causative pathogens and their resistant patterns. Therefore, there is an important need to develop rapid diagnostic strategies for MDRP. We tested a modified protocol to detect carbapenemase and extended-spectrum ß-lactamase (ESBL) producing Gram-negative bacteria (GNB) from positive blood cultures. METHODS: This is a prospective cohort study of consecutive patients with bacteremia. We developed a modified protocol using the HB&L® system to detect MDRP. The operational characteristics were analyzed for each test (HB&L-ESBL/AmpC® and HB&L-Carbapenemase® kits). The kappa coefficient, sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), likelihood ratios (LR) with 95% confidence intervals (CI), and reduction in identification time of this novel method were calculated. RESULTS: Ninety-six patients with BSI were included in the study. A total of 161 positive blood cultures were analyzed. Escherichia coli (50%, 81/161) was the most frequently identified pathogen, followed by Klebsiella pneumoniae (15%, 24/161) and Pseudomonas aeruginosa (8%, 13/161). Thirty-three percent of isolations had usual resistance patterns. However, 34/161 (21%) of identified pathogens were producers of carbapenemases and 21/161 (13%) of extended-spectrum ß-lactamases. Concordance between our HB&L® modified protocol and the traditional method was 99% (159/161). Finally, identification times were significantly shorter using our HB&L®-modified protocol than traditional methods: median (IQR) 19 h (18, 22) vs. 61 h (60, 64), p < 0.001. CONCLUSIONS: Here, we provide novel evidence that using our HB&L®-modified protocol is an effective strategy to reduce the time to detect MDRP producers of carbapenemases or extended-spectrum ß-lactamases, with an excellent concordance rate when compared to the gold standard. Further studies are needed to confirm these findings and to determine whether this method may improve clinical outcomes.

Genetic Diversity and New Sequence Types of Escherichia coli Coharboring ß-Lactamases and PMQR Genes Isolated from Domestic Dogs in Central Panama.

Background: ß-lactamase-producing Escherichia coli are a widely distributed source of antimicrobial resistance for animals and humans. Little is known about the susceptibility profile and genetic characteristics of E. coli strains isolated from domestic dogs in Latin America. Methods: We report on a cross-sectional study that evaluated E. coli strains isolated from fecal samples of domestic dogs in central Panama. The extended-spectrum ß-lactamase (ESBL), AmpC genes, and plasmid-mediated quinolone resistance were investigated. Molecular typing using Pasteur's multilocus sequence typing (MLST) was conducted. Results: A total of 40 E. coli isolates were obtained, of which 80% (32/40) were resistant to at least one of the antibiotics tested, while 20% (8/40) were sensitive to all antibiotics analyzed in this study (p < 0.001). Forty percent of the strains were resistant to three or more antibiotics. The most common resistance was to tetracycline (45%) and ampicillin (30%) while 2.5% showed an ESBL phenotype. Antibiotic resistance genes were detected for one ß-lactamase (blaTEM-1) and two plasmid-mediated quinolone resistance (PMQR) enzymes (qnrS and qnrB). In addition, mutations in the chromosomal AmpC gene were observed at positions −35, −28, −18, −1, and +58. Fourteen different sequence types (STs) were identified; the most frequent were ST399 and ST425 (12% each). ST3 and ST88, which have been previously identified in human clinical isolates, were also evidenced. Three new STs were found for the first time: ST1015, ST1016 (carrier of the blaTEM-1 gene), and ST1017 (carrier of the blaTEM-1, qnrS, and qnrB genes). Conclusions: In the intestinal strains of E. coli isolated from domestic dogs, there was a high frequency of resistance to antibiotics. The presence of genes from plasmids and chromosomal mutations that conferred antibiotic resistance, the identification of isolates previously reported in humans, and the genetic diversity of STs (including three that were newly identified) confirmed the determinants of resistance to antibiotics in the domestic dogs from central Panama.

Imported One-Day-Old Chicks as Trojan Horses for Multidrug-Resistant Priority Pathogens Harboring mcr-9, rmtG, and Extended-Spectrum ß-Lactamase Genes.

Antimicrobial resistance is a critical issue that is no longer restricted to hospital settings but also represents a growing problem involving intensive animal production systems. In this study, we performed a microbiological and molecular investigation of priority pathogens carrying transferable resistance genes to critical antimicrobials in 1-day-old chickens imported from Brazil to Uruguay. Bacterial identification was performed by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry, and antibiotic susceptibility was determined by Sensititre. Antimicrobial resistance genes were sought by PCR, and clonality was assessed by pulsed-field gel electrophoresis (PFGE). Four multidrug-resistant (MDR) representative strains were sequenced by an Illumina and/or Oxford Nanopore Technologies device. Twenty-eight MDR isolates were identified as Escherichia coli (n = 14), Enterobacter cloacae (n = 11), or Klebsiella pneumoniae (n = 3). While resistance to oxyiminocephalosporins was due to blaCTX-M-2, blaCTX-M-8, blaCTX-M-15, blaCTX-M-55, and blaCMY-2, plasmid-mediated quinolone resistance was associated with the qnrB19, qnrE1, and qnrB2 genes. Finally, resistance to aminoglycosides and fosfomycin was due to the presence of 16S rRNA methyltransferase rmtG and fosA-type genes, respectively. Short- and long-read genome sequencing of E. cloacae strain ODC_Eclo3 revealed the presence of IncQ/rmtG (pUR-EC3.1; 7,400 bp), IncHI2A/mcr-9.1/blaCTX-M-2 (pUR-EC3.2, ST16 [pMLST; 408,436 bp), and IncN2/qnrB19/aacC3/aph(3″)-Ib (pUR-EC3.3) resistance plasmids. Strikingly, the blaCTX-M-2 gene was carried by a novel Tn1696-like composite transposon designated Tn7337. In summary, we report that imported 1-day-old chicks can act as Trojan horses for the hidden spread of WHO critical-priority MDR pathogens harboring mcr-9, rmtG, and extended-spectrum ß-lactamase genes in poultry farms, which is a critical issue from a One Health perspective. IMPORTANCE Antimicrobial resistance is considered a significant problem for global health, including within the concept of One Health; therefore, the food chain connects human health and animal health directly. In this work, we searched for microorganisms resistant to antibiotics considered critical for human health in intestinal microbiota of 1-day-old baby chicks imported to Uruguay from Brazil. We describe genes for resistance to antibiotics whose use the WHO has indicated to "watch" or "reserve" (AWaRe classification), such as rmtG and mcr9.1, which confer resistance to all the aminoglycosides and colistin, respectively, among other genes, and their presence in new mobile genetic elements that favor its dissemination. The sustained entry of these microorganisms evades the sanitary measures implemented by the countries and production establishments to reduce the selection of resistant microorganisms. These silently imported resistant microorganisms could explain a considerable part of the antimicrobial resistance problems found in the production stages of the system.

Colistin-resistant mcr-1-positive Escherichia coli ST1775-H137 co-harboring blaCTX-M-2 and blaCMY-2 recovered from an urban stream.

The rapid dissemination of colistin resistance mcr-type genes and extended-spectrum ß-lactamase-encoding genes at the human-animal-environment interface has raised concerns worldwide. In this study, we performed a genomic investigation of a multidrug (MDR)- and colistin-resistant Escherichia coli strain recovered from an urban stream strongly affected by pollution and used for recreational purposes in Brazil. E. coli strain EW827 was resistant to clinically significant antimicrobials, including polymyxins, extended-spectrum cephalosporins, and fluoroquinolones. Whole-genome sequencing analysis revealed that EW827 strain belonged to ST1775 and carried the fimH137 allele, clinically relevant antimicrobial resistance genes (e.g., mcr-1.1, blaCTX-M-2, and blaCMY-2), tolerance genes to metals, and biocide resistance genes. Moreover, IncX4 and IncI1-ST12 replicon types were identified carrying mcr-1.1 and blaCMY-2, respectively. A novel genetic environment of the mcr-1.1 gene, in which a 258-bp ∆IS5-like was inserted in the opposite orientation upstream of the mcr-1.1-pap2 element, was also detected. Additionally, the blaCTX-M-2 gene was harbored by a Tn21-like element on the chromosome. The occurrence of MDR E. coli co-harboring mcr-1.1, blaCTX-M-2, and blaCMY-2 in urban water represents a potential risk to humans, animals, and environmental safety. Therefore, epidemiological studies are required to monitoring multidrug-resistant bacteria and their antimicrobial resistance genes in aquatic ecosystems to determine possible routes and fates of these genes.

Sickle cell disease children's gut colonization by extended-spectrum ß-lactamase (ESBL)-producing Enterobacterales: an antibiotic prophylaxis effect?

Introduction. Sickle cell disease (SCD) children have a high susceptibility to pneumococcal infection. For this reason, they are routinely immunized with pneumococcal vaccines and use antibiotic prophylaxis (AP).Hypothesis/Gap Statement. Yet, little is known about SCD children's gut microbiota. If antibiotic-resistant Enterobacterales may colonize people on AP, we hypothesized that SCD children on AP are colonized by resistant enterobacteria species.Objective. To evaluate the effect of continuous AP on Enterobacterales gut colonization from children with SCD.Methodology. We analysed 30 faecal swabs from SCD children on AP and 21 swabs from children without the same condition. Enterobacterales was isolated on MacConkey agar plates and identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) (bioMérieux, Marcy l'Etoile, France). We performed the antibiogram by Vitek 2 system (bioMérieux, Marcy l'Etoile, France), and the resistance genes were identified by multiplex PCR.Results. We found four different species with resistance to one or more different antibiotic types in the AP-SCD children's group: Escherichia coli, Klebsiella pneumoniae, Citrobacter freundii, and Citrobacter farmeri. Colonization by resistant E. coli was associated with AP (prevalence ratio 2.69, 95 % confidence interval [CI], 1.98-3.67, P<0.001). Strains producing extended-spectrum ß-lactamases (ESBL) were identified only in SCD children, E. coli, 4/30 (13 %), and K. pneumoniae, 2/30 (7 %). The ESBL-producing Enterobacterales were associated with penicillin G benzathine use (95 % CI, 22.91-86.71, P<0.001). CTX-M-1 was the most prevalent among ESBL-producers (3/6, 50 %), followed by CTX-M-9 (2/6, 33 %), and CTX-M-2 (1/6, 17 %).Conclusion. Resistant enterobacteria colonize SCD children on AP, and this therapy raises the chance of ESBL-producing Enterobacterales colonization. Future studies should focus on prophylactic vaccines as exclusive therapy against pneumococcal infections.

Colistin-Resistant mcr-1-Positive Escherichia coli ST131-H22 Carrying bla CTX-M-15 and qnrB19 in Agricultural Soil.

The pandemic Escherichia coli sequence type 131 (ST131) carrying plasmid-mediated colistin resistance mcr genes has emerged worldwide causing extraintestinal infections, with lineages belonging to three major clades (A, B, and C). Clade B is the most prevalent in animals, contaminating associated meat products, and can be transmitted zoonotically. However, the bla CTX-M-15 gene has only been associated with C2 subclade so far. In this study, we performed a genomic investigation of an E. coli (strain S802) isolated from a kale crop in Brazil, which exhibited a multidrug-resistant (MDR) profile to clinically significant antimicrobials (i.e., polymyxin, broad-spectrum cephalosporins, aminoglycosides, and fluoroquinolones). Whole-genome sequencing analysis revealed that the S802 strain belonged to serotype O25:H4, ST131/CC131, phylogenetic group B2, and virotype D5. Furthermore, S802 carried the clade B-associated fimH22 allele, genes encoding resistance to clinically important antimicrobials, metals, and biocides, and was phylogenetically related to human, avian, and swine ST131-H22 strains. Additionally, IncHI2-IncQ1, IncF [F2:A-:B1], and ColE1-like plasmids were identified harboring mcr-1.1, bla CTX-M-15, and qnrB19, respectively. The emergence of the E. coli ST131-H22 sublineage carrying mcr-1.1, bla CTX-M-15, and qnrB19 in agricultural soil represents a threat to food and environmental safety. Therefore, a One Health approach to genomic surveillance studies is required to effectively detect and limit the spread of antimicrobial-resistant bacteria and their resistance genes.

Antimicrobial Resistance Patterns and Dynamics of Extended-Spectrum ß-Lactamase-Producing Uropathogenic Escherichia coli in Cusco, Peru.

Urinary tract infections (UTIs) are a common human infection. Antibiotic resistance in extended-spectrum ß-lactamase (ESBL)-producing uropathogenic E. coli (UPEC) is a major therapeutic challenge due to limited treatment alternatives. The aim was to characterize the antimicrobial resistance (AMR) and dynamics of ESBL-producing UPEC isolates from UTI cases seen at a local hospital in Cusco, Peru. Ninety-nine isolates from respective patients were characterized against 18 different antibiotics. Latent class analysis (LCA) was used to evaluate the dynamics across the study time according to resistance patterns. The median age of patients was 51 years old, and nearly half were women. ESBL-producing UPEC isolates were slightly more frequent in outpatient services than emergency rooms, and there were higher resistance rates in males compared to females. Half of the ESBL producers were resistant to aminoglycosides and nitrofurantoin. Cefoxitin and fosfomycin resistance was 29.3% and 14.1%, respectively. Resistance to carbapenems was not observed. All isolates were multidrug-resistant bacteria, and 16.2% (16/99) were also classified as extensively drug-resistant bacteria. The resistance patterns varied across the study time and differed regarding sex and healthcare service. The study revealed high levels of AMR to commonly used antimicrobials and a dynamic circulation of ESBL-producing UPEC isolates with varying resistance patterns.

Molecular characterization of antimicrobial resistance in Klebsiella pneumoniae isolated from Brazilian dairy herds.

In this observational study, phenotypic and genotypic patterns of antimicrobial resistance (AMR) in Klebsiella pneumoniae isolated from intramammary infections, clinical mastitis, fresh feces, rectal swabs, animal hindlimbs, and bulk tank milk samples from Brazilian dairy herds were investigated. In addition, we identified specific genetic variants present among extended-spectrum ß-lactamase (ESBL) producers. We obtained 169 isolates of K. pneumoniae from 2009 to 2011 on 24 Brazilian dairy farms located in 4 Brazilian states. The AMR profile of all isolates was determined using disk-diffusion assays. The antimicrobial panel included drugs commonly used as mastitis treatment in Brazilian dairy herds (gentamicin, cephalosporins, sulfamethoxazole-trimethoprim, tetracycline) as well as antimicrobials of critical importance for human health (meropenem, ceftazidime, fluoroquinolones). The K. pneumoniae isolates resistant to tetracycline, fluoroquinolones, sulfamethoxazole-trimethoprim, or chloramphenicol were screened for presence of drug-specific AMR genes [tet, qnr, aac(6')-Ib, floR, catA2, cm1A, dfr, sul] using PCR. In addition, we identified ESBL genes present among ESBL-producers by using whole genome sequencing. Genomes were assembled and annotated, and patterns of AMR genes were investigated. Resistance was commonly detected against tetracycline (22.5% of all isolates), streptomycin (20.7%), and sulfamethoxazole-trimethoprim (9.5%). Antimicrobial resistance rates were higher in K. pneumoniae isolated from intramammary infections in comparison with isolates from feces (19.2 and 0% of multidrug resistance in intramammary and fecal isolates, respectively). In contrast, no difference in AMR rates was observed when contrasting hind limbs and isolates from intramammary infections. The genes tetA, sul2, and floR were the most frequently observed AMR genes in K. pneumoniae resistant to tetracycline, sulfamethoxazole-trimethoprim, and chloramphenicol, respectively. The tetA gene was present exclusively in isolates from milk. The genes blaCTX-M8 and blaSHV-108 were present in 3 ESBL-producing K. pneumoniae, including an isolate from bulk tank milk. The 3 isolates were of sequence type 281 and had similar mobile genetic elements and virulence genes. Our study reinforced the epidemiological importance and dissemination of blaCTX-M-8 pST114 plasmid in food-producing animals in Brazil.

Multiresistant and blaCTX-M-14-Carrying Salmonella ser. Typhimurium Isolated During a Salmonellosis Outbreak in an Equine Hospital in Argentina.

Salmonella spp. causes digestive clinical signs in horses. Foals and hospitalized animals are more susceptible to the disease. Nowadays, the report of multidrug-resistant Salmonella spp. producer of extended-spectrum ß-lactamases, is more frequent. The aim of this work was to study the clonal relationship and antimicrobial susceptibility profiles among Salmonella ser. Typhimurium isolates, obtained during a salmonellosis outbreak in an Argentinian equine hospital. Thus, in 2017, we studied the genotypic profiles and the susceptibility to antimicrobials of the strains isolated from three animals with diarrhea in an equine hospital of Argentina. The pulsotype identified by pulsed-field gel electrophoresis was the same among the isolates. Also, this pulsotype had been previously detected in human and porcine isolates, suggesting the circulation of the same strains in different species. Multidrug-resistant isolates with different ß-lactam susceptibility profiles were identified and blaCTX-M-14 was detected for the first time from an isolate of equine-origin in Argentina. Salmonella ser. Typhimurium is an important pathogen in public and veterinary health, so our results emphasize the relevance of appropriate measures to prevent and control this disease. Furthermore, routine antibiotic susceptibility tests of local strains are needed to improve the empiric treatment of equine salmonellosis.

Pseudomonas aeruginosa epidemic high-risk clones and their association with horizontally-acquired ß-lactamases: 2020 update.

Pseudomonas aeruginosa global clones associated with multidrug-resistant (MDR) and extensively drug-resistant (XDR) phenotypes, denominated high-risk clones, are a growing threat in hospitals worldwide. Here we provide a 2020 update on nosocomial MDR/XDR high-risk P. aeruginosa clones. According to their prevalence, global spread and association with MDR/XDR profiles and regarding extended-spectrum ß-lactamases (ESBLs) and carbapenemases, the worldwide top 10 P. aeruginosa high-risk clones includes ST235, ST111, ST233, ST244, ST357, ST308, ST175, ST277, ST654 and ST298. ST235 is certainly the most relevant high-risk clone, showing a worldwide dissemination associated with over 60 different ß-lactamase variants, including multiple carbapenemases from classes A and B. Moreover, ST235 shows a highly virulent phenotype associated with a high mortality rate, likely due to the production of the ExoU cytotoxin. ST111 and ST233 are also worldwide disseminated MDR/XDR clones, particularly linked to VIM-2 metallo-ß-lactamase (MBL), whereas ST244 is a very prevalent clone not always associated with MDR/XDR profiles. ST357, ST308 and ST298 are also exoU+ and are therefore potentially associated with higher virulence. In contrast, ST175, prevalent in some European countries, shows a MDR/XDR phenotype frequently caused by specific chromosomal mutations and is associated with lower virulence. Finally, ST277 is highly prevalent in Brazil and is specifically associated with the SPM MBL. A deeper understanding of the underlying factors driving the success of high-risk clones, including the reported increased capacity for acquiring exogenous determinants, increased spontaneous mutation rates or greater ability to develop biofilms, is required to develop global strategies to combat them.

Development and validation of a scoring system for predicting cancer patients at risk of extended-spectrum b-lactamase-producing Enterobacteriaceae infections.

BACKGROUND: Extended-spectrum beta-lactamase producing Enterobacteriaceae (ESBL-PE) infections are frequent and highly impact cancer patients. We developed and validated a scoring system to identify cancer patients harboring ESBL-PE at the National Institute of Cancer of Colombia. METHODS: We retrospectively analyzed medical records of 1695 cancer patients. Derivation phase included 710 patients admitted between 2013 to 2015, ESBL-PE positive culture (n = 265) paired by month and hospitalization ward with Non-ESBL-PE (n = 445). A crude and weighted score was developed by conditional logistic regression. The model was evaluated in a Validation cohort (n = 985) with the same eligibility criteria between 2016 to 2017. RESULTS: The score was based on eight variables (reported with Odds Ratio and 95% confidence interval): Hospitalization ≥7 days (5.39 [2.46-11.80]), Hospitalization during the previous year (4, 87 [2.99-7.93]), immunosuppressive therapy during the previous 3 months (2.97 [1.44-6.08]), Neutropenia (1.90 [1.12-3.24]), Exposure to Betalactams during previous month (1.61 [1.06-2.42]), Invasive devices (1.51 [1.012-2.25]), Neoplasia in remission (2.78 [1.25-1.17]), No chemotherapy during the previous 3 months (1.90 [1.22-2.97]). The model demonstrated an acceptable discriminatory capacity in the Derivation phase, but poor in the Validation phase (Recipient Operating Characteristic Curve: 0.68 and 0.55 respectively). CONCLUSIONS: Cancer patients have a high prevalence of risk factors for ESBL-PE infection. The scoring system did not adequately discriminate patients with ESBL-PE. In a high-risk population, other strategies should be sought to identify patients at risk of resistant ESBL-PE infection.

Geographic mapping of Enterobacteriaceae with extended-spectrum ß-lactamase (ESBL) phenotype in Pereira, Colombia.

BACKGROUND: Antimicrobial resistance is an ecological and multicausal problem. Infections caused by extended-spectrum ß-lactamase producing Enterobacteriaceae (ESBL-E) can be acquired and transmitted in the community. Data on community-associated ESBL-E infections/colonizations in Colombia are scarce. Georeferencing tools can be used to study the dynamics of antimicrobial resistance at the community level. METHODS: We conducted a study of geographic mapping using modern tools based on geographic information systems (GIS). Two study centers from the city of Pereira, Colombia were involved. The records of patients who had ESBL-producing Enterobacteriaceae were reviewed. Antimicrobial susceptibility testing and phenotypic detection of ESBL was done according to CLSI standards. RESULTS: A population of 415 patients with community-acquired infections/colonizations and 77 hospital discharges were obtained. Geographic distribution was established and heat maps were created. Several hotspots were evidenced in some geographical areas of the south-west and north-east of the city. Many of the affected areas were near tertiary hospitals, rivers, and poultry industry areas. CONCLUSIONS: There are foci of antimicrobial resistance at the community level. This was demonstrated in the case of antimicrobial resistance caused by ESBL in a city in Colombia. Causality with tertiary hospitals in the city, some rivers and the poultry industry is proposed as an explanation of the evidenced phenomenon. Geographic mapping tools are useful for monitoring antimicrobial resistance in the community.

Widespread high-risk clones of multidrug-resistant extended-spectrum ß-lactamase-producing Escherichia coli B2-ST131 and F-ST648 in public aquatic environments.

Aquatic environments are considered a reservoir for the dissemination of multidrug-resistant (MDR) bacteria, principally Escherichia coli, with the consequent spread of acquired antimicrobial resistance genes (ARGs). Widespread high-risk clones of MDR E. coli are responsible for human infections worldwide. This study aimed to characterise, through whole-genome sequencing (WGS), isolates of MDR E. coli harbouring ARGs obtained from public aquatic environments in Brazil. MDR E. coli isolates were obtained from rivers, streams and lakes that presented different Water Quality Index records and were submitted to WGS. The resistome, mobilome and virulome showed a great diversity of ARGs, plasmids and virulence genes, respectively. In addition, mutations in the quinolone resistance-determining regions of GyrA, ParC and ParE as well as several metal resistance genes (MRGs) and antibacterial biocide resistance genes (ABGs) were detected. Typing and subtyping of MDR E. coli revealed different lineages, with two belonging to widespread high-risk clones (i.e. B2-ST131-fimH30 and F-ST648-fimH27), which are grouped by core genome multilocus sequence typing (cgMLST) in clusters with E. coli lineages obtained from different sources distributed worldwide. MDR bacteria carrying MRGs and ABGs have emerged as a global human and environmental health problem. Detection of widespread high-risk clones calls for attention to the dissemination of fluoroquinolone-resistant QnrS1- and CTX-M-producing E. coli lineages associated with human infections in public aquatic environments.

Multidrug-resistant CTX-M-15-positive Klebsiella pneumoniae ST307 causing urinary tract infection in a dog in Brazil.

OBJECTIVES: Klebsiella pneumoniae sequence type 307 (ST307) has emerged as a new high-risk clone worldwide. The aim of this study was to report the draft genome sequence of a multidrug-resistant CTX-M-15-positive K. pneumoniae strain causing urinary tract infection in a companion animal in Brazil. METHODS: Whole-genome sequencing was performed on an Illumina NextSeq platform and the genome was de novo assembled using SPAdes. Data analysis was performed using online tools from the Center for Genomic Epidemiology. RESULTS: The genome size of K. pneumoniae strain PRETA was calculated at 5 450 575 bp, containing a total of 5355 protein-coding sequences. The strain was assigned to the high-risk ST307 and carried the clinically relevant blaCTX-M-15 gene, besides other genes conferring resistance to aminoglycosides, tetracyclines, quinolones, trimethoprim and fosfomycin. CONCLUSION: These data offer novel information for comparative genomic analyses in order to track the transmission dynamics and epidemiology of the newly emerging high-risk clone ST307.