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This study focuses on the genomic characterization of a multidrug-resistant Escherichia coli strain responsible for a severe gastrointestinal infection in a 33-year-old male. The patient initially received sulfamethoxazole/trimethoprim treatment, which proved ineffective. Fecal culture confirmed the presence of E. coli displaying a MDR profile to ampicillin, nalidixic acid, ciprofloxacin, sulfamethoxazole, trimethoprim, and tetracycline. Serotyping identified the strain as ONT:H19. Virulence analysis indicated a highly virulent profile with numerous virulence markers. Plasmid analysis uncovered various plasmids carrying both antimicrobial resistance and virulence genes. MLST assigned the strain to ST10955. Phylogenomic analysis revealed similarity to an older Brazilian isolate, suggesting the persistence of a common lineage with evolving antimicrobial resistance. This report highlights the first identification of a multidrug-resistant ST10955 E. coli strain with a wide resistome and virulence potential, emphasizing the importance of ongoing surveillance of E. coli strains due to their potential for severe infections, resistance development, and virulence., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

The emergence of highly successful genetic lineages of methicillin-resistant Staphylococcus aureus (MRSA) poses a challenge in human healthcare due to increased morbidity and mortality rates. The RdJ clone (CC5-ST105-SCCmecII-t002 lineage), previously identified in Rio de Janeiro, Brazil, was linked to bloodstream infections and features a mutation in the aur gene (encoding aureolysin). Additionally, clinical isolates derived from this clone were more effective at evading monocytic immune responses. This study aimed to detect the RdJ clone among clinical MRSA isolated in Santa Catarina (SC) and examine its antimicrobial resistance and phagocytosis evasion capabilities. Our findings revealed the RdJ clone in 20 % of MRSA isolates, all exhibiting multiresistance. RdJ clone isolates from SC did not demonstrate a decreased rate of phagocytosis compared to CC5 non-RdJ isolates. Structural analysis suggests that the aur mutation is unlikely to significantly impact aureolysin activity. Genomic analysis of one isolate unveiled a genetic variant of the RdJ clone, sharing lineage and gene distribution but lacking the aur mutation. This study enhances the understanding of the clinical and epidemiologic risks associated with the RdJ clone and the biological mechanisms underlying its spreading in SC., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Klebsiella pneumoniae strains that produce Klebsiella pneumoniae Carbapenemase (KPC) variants displaying resistance to ceftazidime-avibactam (CZA) often remain susceptible to meropenem (MEM), suggesting a potential therapeutic use of this carbapenem antibiotic. However, in vitro studies indicate that these sorts of strains can mutate becoming MEM-resistant, raising concerns about the effectiveness of carbapenems as treatment option. We have studied mutation rates occurring from the reversion of MEM-susceptible KPC-114 to MEM-resistant KPC-2, in CZA-resistant K. pneumoniae belonging to ST11. Two-step fluctuation assays (FAs) were conducted. In brief, initial cultures of KPC-114-producing K. pneumoniae showing 1 µg/mL MEM MIC were spread on Mueller-Hinton agar plates containing 2-8 µg/mL MEM. A second step of FA, at 4-16 µg/mL MEM was performed from a mutant colony obtained at 2 µg/mL MEM. Mutation rates were calculated using maximum likelihood estimation. Parental and mutant strains were sequenced by Illumina NextSeq, and mutations were predicted by variant-calling analysis. At 8 µg/mL MEM, mutants derived from parental CZA-resistant (MIC ≥ 64 µg/mL)/MEM-susceptible (MIC = 1 µg/mL) KPC-114-positive K. pneumoniae exhibited an accumulative mutation rate of 3.05 × 10 -19 mutations/cell/generation, whereas at 16 µg/mL MEM an accumulative mutation rate of 1.33 × 10 -19 mutations/cell/generation resulted in the reversion of KPC-114 (S181_P182 deletion) to KPC-2. These findings highlight that the reversion of MEM-susceptible KPC-114 to MEM-resistant KPC-2, in CZA-resistant K. pneumoniae ST11 is related to low mutation rates suggesting a low risk of therapeutic failure. In vivo investigations are necessary to confirm the clinical potential of MEM against CZA-resistant KPC variants.IMPORTANCEThe emergence of ceftazidime-avibactam (CZA) resistance among carbapenem-resistant Klebsiella pneumoniae is a major concern due to the limited therapeutic options. Strikingly, KPC mutations mediating CZA resistance are generally associated with meropenem susceptibility, suggesting a potential therapeutic use of this carbapenem antibiotic. However, the reversion of meropenem-susceptible to meropenem-resistant could be expected. Therefore, knowing the mutation rate related to this genetic event is essential to estimate the potential use of meropenem against CZA-resistant KPC-producing K. pneumoniae . In this study, we demonstrate, in vitro , that under high concentrations of meropenem, reversion of KPC-114 to KPC-2 in CZA-resistant/meropenem-susceptible K. pneumoniae belonging to the global high-risk ST11 is related to low mutation rates.

Objectives: Klebsiella spp. are leading causes of nosocomial infections. Their ability to harbour antimicrobial resistance genes makes them an important public health threat. This study aimed to report the genomic background of carbapenemase-producing Klebsiella quasipneumoniae (HV55B) and Klebsiella michiganensis (HV55D) strains isolated from fresh vegetables destined for hospitalized inpatients., Methods: Microbiological and molecular methods were used to isolate and identify the strains, which were submitted to the antimicrobial susceptibility test and pH tolerance assays. Whole genome sequencing was performed on MiSeq and NextSeq platforms, and online available tools were applied to bioinformatic analysis of clinically relevant information., Results: Both isolates were considered multidrug-resistant and tolerated pH ≥ 4 for 24 h. HV55B belonged to sequence type (ST) ST668, and presented a broad resistome and plasmids from four incompatibility groups. HV55D belonged to ST40. Both strains HV55B and HV55D were genetically close to isolates responsible for human infections around the world, which stands for the plausibility of such bacteria to cause disease in patients of the studied institution., Conclusions: Our results confirm the presence of carbapenemase-producing Klebsiella spp. in fresh foodstuffs intended for hospitalized inpatients' consumption. The genomes characterized here also provide clinically and genomically relevant information to forthcoming epidemiological surveillance studies., (Copyright © 2024 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

The overuse of antimicrobials in livestock has contributed to the emergence and selection of clinically relevant multidrug-resistant bacteria. In Brazil, there is no conclusive information on the occurrence of Escherichia coli producing extended-spectrum β-lactamase (ESβL) in cattle breeding, which is an important sector of agribusiness in this country. Herein, we investigated the presence of ESβL-positive E. coli strains in dairy cattle from a commercial farm with routine practice of therapeutic cephalosporins. Ninety-five rectal swab samples were collected from healthy dairy calves and cows under treatment with ceftiofur. Samples were screened for the presence of ESβL producers, and positive isolates were identified by MALDI-TOF, with subsequent screening for genes encoding ESβL variants by PCR and sequencing. The presence of ESβL (CTX-M-15)-producing E. coli was confirmed in calves, and lactating and dry cows. Most ESβL strains with genetic homologies ≥ 90% were grouped into two major PFGE clusters, confirming the suscessful expansion of clonally related lineages in animals from different lactating cycles, on the same property. Four representatives CTX-M-15-positive E. coli strains had their genomes sequenced, belonging to the clonal complex (CC) 23 and sequence type (ST) 90. A phylogeographical landscape of ST90 was performed revealing a global One Health linkage. Our results highlight the intestinal microbiota of dairy cattle as a hotspot for the spread of critical priority ESβL-producing E. coli and demonstrate that ST90 is an international clone genomically adapted to human and animal hosts, which deserve additional investigation to determine its zoonotic potential and impact in food chain., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

In this study, rapid diagnostic of multidrug-resistant (MDR) sepsis pathogens, directly from positive blood culture (BC) bottles, was evaluated by combining MALDI-TOF and the EUCAST Rapid Antimicrobial Susceptibility Testing (RAST). Carbapenemase production in Escherichia coli and Klebsiella pneumoniae isolates was also evaluated by RAST. From 171 positive BC bottles analyzed, 79 (46 %) MDR species, including E. coli (4/34, 12 %), K. pneumoniae (33/48, 69 %), Pseudomonas aeruginosa (12/12, 100 %), Acinetobacter baumannii (15/15, 100 %), and Staphylococcus aureus (14/37, 38 %) displaying resistance to beta-lactams, fluoroquinolones, aminoglycosides, and/or trimethoprim/sulphamethoxazole, were identified. In this regard, turnaround time of direct MALDI-TOF identification and RAST was < 7 h, which was significantly (p< 0.05) lower than our routine method. Carbapenemase detection by RAST displayed 100% sensitivity and 88.7 % specificity at 8 h. This protocol could offer advantages for the treatment and clinical outcomes of septic patients, improving the rapid diagnostic of sepsis by MDR pathogens., Competing Interests: Declaration of competing interest The authors declare that there is no conflict of interest., (Copyright © 2024. Published by Elsevier Inc.)

Background: The rapid and global spread of Escherichia coli carrying mcr-type genes at the human-animal-environmental interface has become a serious global public health problem., Objective: To perform a genomic investigation of a colistin-resistant E. coli strain (14005RM) causing urinary tract infection, using a hybrid de novo assembly of Illumina/Nanopore sequence data, presenting phylogenomic insights into the relationship with mcr-1-positive strains circulating at the human-animal-environmental interface, in Brazil., Methods: Genomic DNA was sequenced using both the Illumina NexSeq and Nanopore MinION platforms. De novo hybrid assembly was performed by Unicycler. Genomic data were assessed by in silico prediction and bioinformatic tools., Results: The genome assembly size was 5 333 039 bp. The mcr-1.5-positive E. coli strain 14005RM belongs to the sequence type ST354 and presented a broad resistome (antibiotics, heavy metals, disinfectants, and glyphosate) and virulome. The mcr-1.5 gene was carried by an IncI2 plasmid (p14005RM, sizing 65,458 kb). Full genome SNP-based phylogenetic analysis reveals that mcr-1.5-producing E. coli strain 14005RM is highly related (> 98% identity) to colistin-resistant mcr-1.1-positive ST354 lineages associated with urinary tract infections in Brazil since 2015., Conclusion: Mobile colistin resistance within the Brazilian One Health microbiosphere is mediated by mcr gene variants propagated by IncX4, IncHI2, and IncI2 plasmids, circulating among global clones of E. coli., (Copyright © 2024 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

The spread of antimicrobial resistance (AMR) through multiple reservoirs is a global concern. Wastewater is a critical AMR dissemination source, so this study aimed to assess the persistence of resistance genetic markers in wastewater using a culture-independent approach. Raw and treated wastewater samples (n = 121) from a wastewater treatment plant (WWTP), a human hospital, a veterinary hospital, and a pig farm were monthly collected and concentrated by filtration. DNA was extracted directly from filter membranes, and PCR was used in the qualitative search of 32 antimicrobial resistance genes (ARGs). Selected genes (bla CTX-M , bla KPC , qnrB, and mcr-1) were enumerated by quantitative real-time PCR (qPCR). Twenty-six ARGs were detected in the qualitative ARGs search, while quantitative data showed a low variation of the ARG's relative abundance (RA) throughout the months, especially at the human hospital and the WWTP. At the WWTP, despite significantly reducing the absolute number of gene copies/L after each treatment stage (p < 0.05), slight increases (p > 0.05) in the RAs of genes bla CTX-M , qnrB, and mcr-1 were observed in reused water (tertiary treatment) when compared with secondary effluent. Although the increase is not statistically significant, it is worth noting that there was some level of ARGs concentration after the disinfection process. No significant absolute or relative after-treatment quantification reductions were observed for any ARGs at the veterinary hospital or the pig farm. The spread of ARGs through sewage needs to be continuously addressed, because their release into natural environments may pose potential risks of exposure to resistant bacteria and impact local ecosystems., (© 2024 John Wiley & Sons Ltd.)

Competing Interests: We declare no competing interests. This work was funded by the National Council for Scientific and Technological Development (CNPq; 304905/2022-4, 314336/2021-4, and 150712/2022-7) and the Coordination for the Improvement of Higher Education Personnel (CAPES; 88887.463868/2019-00 and Finance Code 001).

The use, misuse, and overuse of antimicrobials is one of the main public health threats of the 21st century. We investigated the risk factor of the presence of extended-spectrum, cephalosporin-resistant Enterobacterales in feces of non-domestic and domestic birds and other domestic animals in Piauí State, northeast Brazil. We collected a total of 387 cloacal and rectal swab samples of free-living birds, domestic birds, and domestic mammals in five municipalities: Amarante, Água Branca, Lagoa Alegre, Parnaíba, and Teresina. A total of 59/387 (15.2%) of these samples harbored extended spectrum beta-lactamase (ESBL)-producing Enterobacterales. Using the MALDI-TOF technique, we identified fifty-seven samples as Escherichia coli and two samples as Klebsiella pneumoniae . Teresina and Parnaíba had the highest prevalence of animals with resistant bacteria (32.1% and 27.1%, respectively) and highest exposure risk factor (OR of 16.06 and 8.58, respectively, and p < 0.001 for all). Multidrug-resistant, ESBL-producing Enterobacterales were observed in 72.8% of the samples (43/59). For the free-living birds, the positive samples belonged to a great kiskadee ( Pitangus sulphuratus ) and a semipalmated sandpiper ( Calidris pusilla ) in migratory and resident species, respectively. For domestic animals, the swine samples showed the highest prevalence of antimicrobial resistance. The lack of access to veterinary care and information regarding antimicrobial therapy, along with the easy access to antimicrobials without medical prescription, favors the inadequate use of antimicrobials in Piauí.

Competing Interests: None declared.

Background: Carbapenemase-producing Citrobacter freundii has been reported as a leading cause of healthcare-associated infections. Particularly, C. freundii belonging to the sequence type (ST) 18 is considered to be an emerging nosocomial clone., Objectives: To report the genomic background and phylogenomic analysis of a multidrug-resistant NDM-1-producing C. freundii ST18 (strain CF135931) isolated from an endangered green sea turtle affected by plastic pollution in Brazil., Methods: Genomic DNA was extracted and sequenced using the Illumina NextSeq platform. De novo assembly was performed by CLC Workbench, and in silico analysis accomplished by bioinformatics tools. For phylogenomic analysis, publicly available C. freundii (txid:546) genome assemblies were retrieved from the NCBI database., Results: The genome size was calculated at 5 290 351 bp, comprising 5263 total genes, 4 rRNAs, 77 tRNAs, 11ncRNAs, and 176 pseudogenes. The strain belonged to C. freundii ST18, whereas resistome analysis predicted genes encoding resistance to β-lactams (bla NDM-1 , bla OXA-1 , bla CMY-117 , and bla TEM-1C ), aminoglycosides (aph(3'')-Ib, aadA16, aph(3')-VI, aac(6')-Ib-cr, and aph(6)-Id), quinolones (aac(6')-Ib-cr), macrolides (mph(A) and erm(B)), sulphonamides (sul1 and sul2), tetracyclines (tetA and tetD), and trimethoprim (dfrA27). The phylogenomic analysis revealed that CF135931 strain is closely related to international human-associated ST18 clones producing NDM-1., Conclusion: Genomic surveillance efforts are necessary for robust monitoring of the emergence of drug-resistant strains and WHO critical priority pathogens within a One Health framework. In this regard, this draft genome and associated data can improve understanding of dissemination dynamics of nosocomial clones of carbapenemase-producing C. freundii beyond hospital walls. In fact, the emergence of NDM-1-producing C. freundii of global ST18 in wildlife deserves considerable attention., (Copyright © 2024 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Background: The global spread of extended-spectrum β-lactamase (ESβL)-producing Escherichia coli has been considered a One Health issue that demands continuous genomic epidemiology surveillance in humans and non-human hosts., Objectives: To report the occurrence and genomic data of ESβL-producing E. coli strains isolated from South American llamas inhabiting a protected area with public access in the Andean Highlands of Peru., Methods: Two ESβL-producing E. coli strains (E. coli L1LB and L2BHI) were identified by MALDI-TOF. Genomic DNAs were extracted and sequenced using the Illumina NextSeq platform. De novo assembly was performed by CLC Genomic Workbench and in silico prediction was accomplished by curated bioinformatics tools. SNP-based phylogenomic analysis was performed using publicly available genomes of global E. coli ST10., Results: Escherichia coli L1LB generated a total of 4 000 11 and L2BHI a total of 4 002 54 paired-end reads of ca.164 × and ca. 157 ×, respectively. Both E. coli strains were assigned to serotype O8:H4, fimH41, and ST10. The bla CTX-M-65 ESβL gene, along with other medically important antimicrobial resistance genes, was predicted. Broad virulomes, including the presence of the astA gene, were confirmed. The phylogenomic analysis revealed that E. coli L1LB and L2BHI strains are closely related to isolates from companion animals and human hosts, as well as environmental strains, previously reported in North America, South America, Africa, and Asia., Conclusion: Presence of ESβL-producing E. coli ST10 in South American camelids with historical and cultural importance supports successful expansion of international clones of priority pathogens in natural areas with public access., (Copyright © 2023. Published by Elsevier Ltd.)

Migratory birds have contributed to the dissemination of multidrug-resistant (MDR) bacteria across the continents. A CTX-M-2-producing Escherichia coli was isolated from a black skimmer ( Rynchops niger ) in Southeast Brazil. The whole genome was sequenced using the Illumina NextSeq platform and de novo assembled by CLC. Bioinformatic analyses were carried out using tools from the Center for Genomic Epidemiology. The genome size was estimated at 4.9 Mb, with 4790 coding sequences. A wide resistome was detected, with genes encoding resistance to several clinically significant antimicrobials, heavy metals, and biocides. The bla CTX-M-2 gene was inserted in an In 229 class 1 integron inside a ∆Tn As3 transposon located in an IncHI2/ST2 plasmid. The strain was assigned to ST5506, CH type fumC19/fimH32 , serotype O8:K87, and phylogroup B1. Virulence genes associated with survival in acid conditions, increased serum survival, and adherence were also identified. These data highlight the role of migratory seabirds as reservoirs and carriers of antimicrobial resistance determinants and can help to elucidate the antimicrobial resistance dynamics under a One Health perspective.

Mangrove ecosystems are hotspots of biodiversity, but have been threatened by anthropogenic activities. Vancomycin-resistant enterococci (VRE) are nosocomial bacteria classified as high priority by the World Health Organization (WHO). Herein, we describe the identification and genomic characteristics of a vancomycin-resistant Enterococcus faecalis strain isolated from a highly impacted mangrove ecosystem of the northeastern Brazilian, in 2021. Genomic analysis confirmed the existence of the transposon Tn1546-vanA and clinically relevant antimicrobial resistance genes, such as streptogramins, tetracycline, phenicols, and fluoroquinolones. Virulome analysis identified several genes associated to adherence, immune modulation, biofilm, and exoenzymes production. The UFSEfl strain was assigned to sequence type (ST9), whereas phylogenomic analysis with publicly available genomes from a worldwide confirmed clonal relatedness with a hospital-associated Brazilian clone. Our findings highlight the successful expansion of hospital-associated VRE in a mangrove area and shed light on the need for strengthening genomic surveillance of WHO priority pathogens in these vital ecosystems., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Stenotrophomonas maltophilia is an opportunistic human pathogen associated with nosocomial and community-acquired infections. We have conducted a microbiological and genomic surveillance study of broad-spectrum cephalosporin- and carbapenem-resistant Gram-negative bacteria colonizing wild birds inhabiting the Brazilian Amazonia. Strikingly, two S. maltophilia strains (SM79 and SM115) were identified in Plain-throated antwren (Isleria hauxwelli) passerines affected by Amazonian fragmentation and degradation. Noteworthy, SM79 and SM115 strains belonged to new sequence types (STs) ST474 and ST473, respectively, displaying resistance to broad-spectrum β-lactams, aminoglycosides and/or fluoroquinolones. In this regard, resistome analysis confirmed efflux pumps (smeABC, smeDEF, emrAB-tolC and macB), bla L1 and bla L2 , aph(3')-IIc and aac(6')-Iak, and Smqnr resistance genes. Comparative phylogenomic analysis with publicly available S. maltophilia genomes clustered ST473 and ST474 with human strains, whereas the ST474 was also grouped with S. maltophilia strains isolated from water and poultry samples. In summary, we report two novel sequence types of S. maltophilia colonizing wild Amazonian birds. The presence of opportunistic multidrug-resistant pathogens in wild birds, from remotes areas, could represent an ecological problem since these animals could easily promote long-distance dispersal of medically important antimicrobial-resistant bacteria. Therefore, while our results could provide a baseline for future epidemiological genomic studies, considering the limited information regarding S. maltophilia circulating among wild animals, additional studies are necessary to evaluate the clinical impact and degree of pathogenicity of this human opportunistic pathogen in wild birds., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Two novel variants of Klebsiella pneumoniae carbapenemase (KPC) associated with resistance to ceftazidime-avibactam (CZA) and designated as KPC-113 and KPC-114 by NCBI were identified in 2020, in clinical isolates of Klebsiella pneumoniae in Brazil. While K. pneumoniae of ST16 harbored the bla KPC-113 variant on an IncFII-IncFIB plasmid, K. pneumoniae of ST11 carried the bla KPC-114 variant on an IncN plasmid. Both isolates displayed resistance to broad-spectrum cephalosporins, β-lactam inhibitors, and ertapenem and doripenem, whereas K. pneumoniae producing KPC-114 showed susceptibility to imipenem and meropenem. Whole-genome sequencing and in silico analysis revealed that KPC-113 presented a Gly insertion between Ambler positions 264 and 265 (R264&#95;A265insG), whereas KPC-114 displayed two amino acid insertions (Ser-Ser) between Ambler positions 181 and 182 (S181&#95;P182insSS) in KPC-2, responsible for CZA resistance profiles. Our results confirm the emergence of novel KPC variants associated with resistance to CZA in international clones of K. pneumoniae circulating in South America. IMPORTANCE KPC-2 carbapenemases are endemic in Latin America. In this regard, in 2018, ceftazidime-avibactam (CZA) was authorized for clinical use in Brazil due to its significant activity against KPC-2 producers. In recent years, reports of resistance to CZA have increased in this country, limiting its clinical application. In this study, we report the emergence of two novel KPC-2 variants, named KPC-113 and KPC-114, associated with CZA resistance in Klebsiella pneumoniae strains belonging to high-risk clones ST11 and ST16. Our finding suggests that novel mutations in KPC-2 are increasing in South America, which is a critical issue deserving active surveillance.

The global dissemination of extended-spectrum-β-lactamase (ESBL)-producing Escherichia coli has been considered a critical issue within a One Health framework. The aim of this study was to perform a genomic investigation of an ESBL-producing E. coli strain belonging to the globally spread sequence type/clonal complex ST90/CC23, isolated from gastrointestinal tract of a dog, in Brazil. Besides CTX-M-15 ESBL, this E. coli isolate carried mutations conferring resistance to human and veterinary fluoroquinolones (GyrA [Ser83Leu, Asp87Asn], ParC [Ser80Ile] and ParE [Ser458Ala]), and resistance determinants to disinfectants and pesticides. Noteworthy, phylogenomic analysis revealed that this multidrug E. coli strain clustered with ST90 lineages isolated from human, dog, and livestock in Brazil. The phylogenetic tree also revealed that this E. coli strain shares a common ancestor with isolates from the United States, Russia, Germany, and China, highlighting the potential global spreading of this clone. In summary, we report genomic data of CTX-M-15-positive E.coli ST90 colonizing a pet. Colonization of companion animals by critical resistant pathogens highlights the need for close monitoring to better understand the epidemiology and genetic factors contributing for successful adaptation of global clones at the human-animal interface.

The spread of carbapenemase-producing Klebsiella pneumoniae beyond hospital settings is a global critical issue within a public health and One Health perspective. Another worrisome concern is the convergence of virulence and resistance in healthcare-associated lineages of K. pneumoniae leading to unfavorable clinical outcomes. During a surveillance study of WHO critical priority pathogens circulating in an impacted urban river in São Paulo, Brazil, we isolate two hypermucoviscous and multidrug-resistant K. pneumoniae strains (PINH-4250 and PINH-4900) from two different locations near to medical centers. Genomic investigation revealed that both strains belonged to the global high-risk sequence type (ST) ST11, carrying the bla KPC-2 carbapenemase gene, besides other medically important antimicrobial resistance determinants. A broad virulome was predicted and associated with hypervirulent behavior in the Galleria mellonella infection model. Comparative phylogenomic analysis of PINH-4250 and PINH-4900 along to an international collection of publicly available genomes of K. pneumoniae ST11 revealed that both environmental strains were closely related to hospital-associated K. pneumoniae strains recovered from clinical samples between 2006 and 2018, in São Paulo city. Our findings support that healthcare-associated KPC-2-positive K. pneumoniae of ST11 clone has successfully expanded beyond hospital settings. In summary, aquatic environments can become potential sources of international clones of K. pneumoniae displaying carbapenem resistance and hypervirulent behaviors, which is a critical issue within a One Health perspective., Competing Interests: All authors declare no conflicts of interest., (© 2023 The Authors. Published by Elsevier B.V.)

Olive ridley ( Lepidochelys olivacea ) turtles migrate across tropical regions of the Atlantic, Pacific, and Indian Oceans. Worryingly, olive ridley populations have been declining substantially and is now considered a threatened species. In this regard, habitat degradation, anthropogenic pollution, and infectious diseases have been the most notorious threats for this species. We isolated a metallo-β-lactamase (NDM-1)-producing Citrobacter portucalensis from the blood sample of an infected migratory olive ridley turtle found stranded sick in the coast of Brazil. Genomic analysis of C. portucalensis confirmed a novel sequence type (ST), named ST264, and a wide resistome to broad-spectrum antibiotics. The production of NDM-1 by the strain contributed to treatment failure and death of the animal. Phylogenomic relationship with environmental and human strains from African, European and Asian countries confirmed that critical priority clones of C. portucalensis are spreading beyond hospital settings, representing an emerging ecological threat to marine ecosystems., Competing Interests: All authors declare no conflicts of interest., (© 2023 The Authors.)

Wild birds have emerged as novel reservoirs and potential spreaders of antibiotic-resistant priority pathogens, being proposed as sentinels of anthropogenic activities related to the use of antimicrobial compounds. The aim of this study was to investigate the occurrence and genomic features of extended-spectrum β-lactamase (ESBL)-producing bacteria in wild birds in South America. In this regard, we have identified two ESBL (CTX-M-55 and CTX-M-65)-positive Escherichia coli (UNB7 and GP188 strains) colonizing Creamy-bellied Thrush ( Turdus amaurochalinus ) and Variable Hawk ( Geranoaetus polyosoma ) inhabiting synanthropic and wildlife environments from Brazil and Chile, respectively. Whole-genome sequence (WGS) analysis revealed that E. coli UNB7 and GP188 belonged to the globally disseminated clone ST602, carrying a wide resistome against antibiotics (β-lactams), heavy metals (arsenic, copper, mercury), disinfectants (quaternary ammonium compounds), and pesticides (glyphosate). Additionally, E. coli UNB7 and GP188 strains harbored virulence genes encoding hemolysin E, type II and III secretion systems, increased serum survival, adhesins and siderophores. SNP-based phylogenomic analysis, using an international genome database, revealed genomic relatedness (19-363 SNP differences) of GP188 with livestock and poultry strains, and genomic relatedness (61-318 differences) of UNB7 with environmental, human and livestock strains (Table S1), whereas phylogeographical analysis confirmed successful expansion of ST602 as a global clone of One Health concern. In summary, our results support that ESBL-producing E. coli ST602 harboring a wide resistome and virulome have begun colonizing wild birds in South America, highlighting a potential new reservoir of critical priority pathogens., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (© 2023 The Authors. Published by Elsevier B.V.)

Objectives: The aim of this study was to characterise a broad-spectrum cephalosporin-resistant AmpC-positive Enterobacter huaxiensis colonising the skin of a Neotropical frog (Phyllomedusa distincta) inhabiting the Brazilian Atlantic Forest., Methods: During a genomic surveillance study of antimicrobial resistance, we screened skin samples from P. distincta. Gram-negative bacteria growing on MacConkey agar plates supplemented with 2 µg/mL ceftriaxone were identified by matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry. A cephalosporin-resistant E. huaxiensis was sequenced using the Illumina NextSeq platform. Genomic data were analysed using bioinformatics tools, whereas AmpC β-lactamase was characterised in depth by comparative analysis of amino acids, in silico modelling, and analysis of susceptibility to β-lactam antibiotics and combinations of β-lactamase inhibitors., Results: Whole-genome sequencing analysis revealed a novel variant of AmpC β-lactamase belonging to the ACT family, designated ACT-107 by NCBI. This variant contains 12 novel amino acid mutations within the ACT family, 5 in the signal peptide sequence (Ile2, Met14, Tyr16, Gly18 and Thr20), and 7 in the mature protein (Gln22, His43, Cys60, Thr157, Glu225, Ala252 and Asn310). In silico modelling showed that substitutions occurring in the mature chain are localised in the solvent-accessible surface of the protein, where they are not expected to affect the β-lactamase activity, as observed in the resistance profile. Strikingly, 'not designated' ACT variants from E. huaxiensis were clustered (> 96% identity) with ACT-107., Conclusion: Since E. huaxiensis has been isolated from human infection, ACT-107 requires surveillance and the attention of clinicians., Competing Interests: Competing interests None declared., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Objectives: The aim of this study was to perform a genomic investigation of a multiple fluoroquinolone-resistant Leclercia adecarboxylata strain isolated from a synanthropic pigeon in São Paulo, Brazil., Methods: Whole-genome sequencing was performed using an Illumina platform, and in silico deep analyses of the resistome were performed. Comparative phylogenomics was conducted using a global collection of publicly available genomes of L. adecarboxylata strains isolated from human and animal hosts., Results: L. adecarboxylata strain P62P1 displayed resistance to human (norfloxacin, ofloxacin, ciprofloxacin, and levofloxacin) and veterinary (enrofloxacin) fluoroquinolones. This multiple quinolone-resistant profile was associated with mutations in the gyrA (S83I) and parC (S80I) genes and the presence of the qnrS gene within an ISKpn19-orf-qnrS1-ΔIS3-bla LAP-2 module, previously identified in L. adecarboxylata strains isolated from pig feed and faeces in China. Genes associated with arsenic, silver, copper, and mercury resistance were also predicted. Phylogenomic analysis revealed clustering (378-496 single nucleotide polymorphism differences) with two L. adecarboxylata strains isolated from human and fish sources in China and Portugal, respectively., Conclusions: L. adecarboxylata is a Gram-negative bacterium of the Enterobacterales order and is considered an emergent opportunistic pathogen. Since L. adecarboxylata has adapted to human and animal hosts, genomic surveillance is highly recommended, in order to identify the emergence and spread of resistant lineages and high-risk clones. In this regard, this study provides genomic data that can help clarify the role of synanthropic animals in the dissemination of clinically relevant L. adecarboxylata within a One Health context., (Copyright © 2023 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Extended spectrum β-lactamase (ESBL)-producing Enterobacterales has spread rapidly around the world, reaching remote areas. In this regard, wild birds that acquire ESBL producers from anthropogenically impacted areas can become reservoirs, contributing to further dissemination of antimicrobial-resistant bacteria categorized as critical priority pathogens to remote environments, during migration seasons. We have conducted a microbiological and genomic investigation on the occurrence and features of ESBL-producing Enterobacterales in wild birds from the remote Acuy Island, in the Gulf of Corcovado, at Chilean Patagonia. Strikingly, five ESBL-producing Escherichia coli were isolated from migratory and resident gulls. Whole-genome sequencing (WGS) analysis revealed the presence of two E. coli clones belonging to international sequence types (STs) ST295 and ST388, producing CTX-M-55 and CTX-M-1 ESBLs, respectively. Moreover, E. coli carried a wide resistome and virulome associated with human and animal infections. Phylogenomic analysis of global and publicly genomes of E. coli ST388 (n = 51) and ST295 (n = 85) clustered gulls isolates along to E. coli strains isolated from the environment, companion animal and livestock in the United States of America, within or close to the migratory route of Franklin's gull, suggesting a possible trans hemispheric movement of international clones of WHO critical priority ESBL producing pathogens., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Staphylococcus aureus remains one of the leading causes of infections worldwide and a common cause of bacteraemia. However, studies documenting the epidemiology of S. aureus in South America using genomics are scarce. We hereby report on the largest genomic epidemiology study to date of both methicillin-resistant S. aureus (MRSA) and methicillin-susceptible S. aureus (MSSA) in South America, conducted by the StaphNET-SA network. We characterised 404 genomes recovered from a prospective observational study of S. aureus bacteraemia in 58 hospitals from Argentina, Bolivia, Brazil, Paraguay and Uruguay between April and October 2019. We show that a minority of S. aureus isolates are phenotypically multi-drug resistant (5.2%), but more than a quarter are resistant to macrolide-lincosamide-streptogramin B (MLSb). MSSA were more genetically diverse than MRSA. Lower rates of associated antimicrobial resistance in community-associated(CA)-MRSA versus hospital-associated (HA)-MRSA were found in association with three S. aureus genotypes dominating the MRSA population: CC30-MRSA-IVc- t019-lukS/F-PV+ , CC5-MRSA-IV- t002-lukS/F-PV- and CC8-MRSA-IVc- t008-lukS/F-PV+ -COMER+. These are historically from a CA origin, carry on average fewer antimicrobial resistance determinants, and often lack key virulence genes. Surprisingly, CC398-MSSA- t1451-lukS/F-PV- related to the CC398 human-associated lineage is widely disseminated throughout the region, and is described here for the first time as the most prevalent MSSA lineage in South America. Moreover, CC398 strains carrying ermT (largely responsible for the MLSb resistance rates of MSSA strains: inducible iMLSb phenotype) and sh&#95;fabI (related to triclosan resistance) were recovered from both CA and HA origin. The frequency of MRSA and MSSA lineages differed between countries but the most prevalent S. aureus genotypes are high-risk clones widely distributed in the South American region without a clear country-specific phylogeographical structure. Therefore, our findings underline the need for continuous genomic surveillance by regional networks such as StaphNET-SA. This article contains data hosted by Microreact.

Salmonella serovars Heidelberg and Minnesota encoding antimicrobial resistance to third-generation cephalosporins and fluoroquinolones are often detected in poultry/poultry meat. We analysed the genomes of 10 Salmonella Heidelberg (SH) and 4 Salmonella Minnesota (SM) from faecal isolates of Brazilian poultry. These featured virulent and multidrug-resistant characteristics, with AmpC beta-lactamase (bla CMY-2 ) predominance (9/14), for all SM (4/4) and some SH (3/10) located on IncC plasmid replicons. IncC carrying bla CTX-M-2 was only detected among SH (3/10). Mutation in the gyrA/parC genes was present in all SH, whereas SM harboured parC mutation plus qnrB19 on ColRNAI plasmids (3/4). In silico resistance overall corroborated with phenotypic results. Core genome phylogenies showed close clustering and high similarities between the Brazilian and poultry meat/food isolates from Europe, and to human isolates from European countries with documented import of Brazilian poultry meat. Conjugation assays with SM successfully transferred bla CMY-2 , and qnrB19 to an Escherichia coli recipient. The findings reinforce the ongoing antimicrobial resistance acquisition of SH and Minnesota and the risks for disseminating resistant strains and/or mobile elements which may increasingly affect importing countries and the need for controlling AMR in major poultry-exporting countries like Brazil., (© 2022 The Authors. Environmental Microbiology Reports published by Applied Microbiology International and John Wiley & Sons Ltd.)

The unbridled dissemination of multidrug-resistant pathogens is a major threat to global health and urgently demands novel therapeutic alternatives. Antimicrobial photodynamic therapy (aPDT) has been developed as a promising approach to treat localized infections regardless of drug resistance profile or taxonomy. Even though this technique has been known for more than a century, discussions and speculations regarding the biochemical mechanisms of microbial inactivation have never reached a consensus on what is the primary cause of cell death. Since photochemically generated oxidants promote ubiquitous reactions with various biomolecules, researchers simply assumed that all cellular structures are equally damaged. In this study, biochemical, molecular, biological and advanced microscopy techniques were employed to investigate whether protein, membrane or DNA damage correlates better with dose-dependent microbial inactivation kinetics. We showed that although mild membrane permeabilization and late DNA damage occur, no correlation with inactivation kinetics was found. On the other hand, protein degradation was analyzed by three different methods and showed a dose-dependent trend that matches microbial inactivation kinetics. Our results provide a deeper mechanistic understanding of aPDT that can guide the scientific community toward the development of optimized photosensitizing drugs and also rationally propose synergistic combinations with antimicrobial chemotherapy., (© 2022 American Society for Photobiology.)

Antimicrobial blue light (aBL) offers efficacy and safety in treating infections. However, the bacterial targets for aBL are still poorly understood and may be dependent on bacterial species. Here, we investigated the biological targets of bacterial killing by aBL (λ = 410 nm) on three pathogens: Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa. Initially, we evaluated the killing kinetics of bacteria exposed to aBL and used this information to calculate the lethal doses (LD) responsible for killing 90 and 99.9% of bacteria. We also quantified endogenous porphyrins and assessed their spatial distribution. We then quantified and suppressed reactive oxygen species (ROS) production in bacteria to investigate their role in bacterial killing by aBL. We also assessed aBL-induced DNA damage, protein carbonylation, lipid peroxidation, and membrane permeability in bacteria. Our data showed that P. aeruginosa was more susceptible to aBL (LD 99.9 = 54.7 J/cm 2 ) relative to S. aureus (LD 99.9 = 158.9 J/cm 2 ) and E. coli (LD 99.9 = 195 J/cm 2 ). P. aeruginosa exhibited the highest concentration of endogenous porphyrins and level of ROS production relative to the other species. However, unlike other species, DNA degradation was not observed in P. aeruginosa. Sublethal doses of blue light (90 ) could damage the cell membrane in Gram-negative species but not in S. aureus. In all bacteria, oxidative damage to bacterial DNA (except P. aeruginosa), proteins, and lipids occurred after high aBL exposures (>LD 99.9 ). We conclude that the primary targets of aBL depend on the species, which are probably driven by variable antioxidant and DNA-repair mechanisms. IMPORTANCE Antimicrobial-drug development is facing increased scrutiny following the worldwide antibiotic crisis. Scientists across the world have recognized the urgent need for new antimicrobial therapies. In this sense, antimicrobial blue light (aBL) is a promising option due to its antimicrobial properties. Although aBL can damage different cell structures, the targets responsible for bacterial inactivation have still not been completely established and require further exploration. In our study, we conducted a thorough investigation to identify the possible aBL targets and gain insights into the bactericidal effects of aBL on three relevant pathogens: Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa. This research not only adds new content to blue light studies but opens new perspectives to antimicrobial applications.

WHO priority pathogens have disseminated beyond hospital settings and are now being detected in urban and wild animals worldwide. In this regard, synanthropic animals such as urban pigeons ( Columba livia ) and rodents ( Rattus rattus , Rattus norvegicus and Mus musculus ) are of interest to public health due to their role as reservoirs of pathogens that can cause severe diseases. These animals usually live in highly contaminated environments and have frequent interactions with humans, domestic animals, and food chain, becoming sentinels of anthropogenic activities. In this study, we report genomic data of Escherichia coli strains selected for ceftriaxone and ciprofloxacin resistance, isolated from pigeons and black rats. Genomic analysis revealed the occurrence of international clones belonging to ST10, ST155, ST224 and ST457, carrying a broad resistome to beta-lactams, aminoglycosides, trimethoprim/sulfamethoxazole, fluoroquinolones, tetracyclines and/or phenicols. SNP-based phylogenomic investigation confirmed clonal relatedness with high-risk lineages circulating at the human-animal-environmental interface globally. Our results confirm the dissemination of WHO priority CTX-M-positive E. coli in urban rodents and pigeons in Brazil, highlighting potential of these animals as infection sources and hotspot for dissemination of clinically relevant pathogens and their resistance genes, which is a critical issue within a One Health perspective., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (© 2022 The Authors.)

Resistance to colistin generated by the mcr-1 gene in Enterobacteriaceae is of great concern due to its efficient worldwide spread. Despite the fact that the Lima region has a third of the Peruvian population and more than half of the national pig and poultry production, there are no reports of the occurrence of the mcr-1 gene in Escherichia coli isolated from livestock. In the present work, we studied the occurrence of E. coli carrying the mcr-1 gene in chicken and pig farms in Lima between 2019 and 2020 and described the genomic context of the mcr-1 gene. We collected fecal samples from 15 farms in 4 provinces of Lima including the capital Lima Metropolitana and recovered 341 E. coli isolates. We found that 21.3% (42/197) and 12.5% (18/144) of the chicken and pig strains were mcr-1 -positive by PCR, respectively. The whole genome sequencing of 14 mcr-1 -positive isolates revealed diverse sequence types (e.g., ST48 and ST602) and the presence of other 38 genes that confer resistance to 10 different classes of antibiotics, including beta-lactamase bla CTX-M-55 . The mcr-1 gene was located on diverse plasmids belonging to the IncI2 and IncHI1A:IncHI1B replicon types. A comparative analysis of the plasmids showed that they contained the mcr-1 gene within varied structures ( mikB - mcr1 - pap2, IS Apl1 - mcr1 - pap2, and Tn6330). To the best of our knowledge, this is the first attempt to study the prevalence of the mcr-1 gene in livestock in Peru, revealing its high occurrence in pig and chicken farms. The genetic diversity of mcr-1 -positive strains suggests a complex local epidemiology calling for a coordinated surveillance under the One-Health approach that includes animals, retail meat, farmers, hospitals and the environment to effectively detect and limit the spread of colistin-resistant bacteria.

Escherichia coli are gut commensal bacteria and opportunistic pathogens, and the emergence of antimicrobial resistance threatens the safety of the food chain. To know the E. coli strains circulating in the Brazilian poultry sector is important since the country corresponds to a significant chicken meat production. Thus, we analyzed 90 publicly genomes available in a database using web-based tools. Genomic analysis revealed that sul alleles were the most detected resistance genes, followed by aadA, bla CTX-M , and dfrA. Plasmids of the IncF family were important, followed by IncI1-Iα, Col-like, and p0111. Genes of specific metabolic pathways that contribute to virulence (terC and gad) were predominant, followed by sitA, traT, and iss. Additionally, pap, usp, vat, sfa/foc, ibeA, cnf1, eae, and sat were also predicted. In this regard, 11 E. coli were characterized as avian pathogenic E. coli and one as atypical enteropathogenic E. coli. Phylogenetic analysis confirmed the predominant occurrence of B1 but also A, D, B2, F, E, G, C, and Clade I phylogroups, whereas international clones ST38, ST73, ST117, ST155, and ST224 were predicted among 53 different sequence types identified. Serotypes O6:H1 and:H25 were prevalent, and fimH31 and fimH32 were the most representatives among the 36 FimH types detected. Finally, single nucleotide polymorphisms-based phylogenetic analysis confirmed high genomic diversity among E. coli strains. While international E. coli clones have adapted to the Brazilian poultry sector, the virulome background of these strains support a pathogenic potential to humans and animals, with lineages carrying resistance genes that can lead to hard-to-treat infections., (© 2022. The Author(s) under exclusive licence to Sociedade Brasileira de Microbiologia.)

Acquired antibiotic resistance in bacteria has become an important worldwide challenge. Currently, several bacteria, including Escherichia coli, have multidrug resistance profiles. Genes such as bla CTX-M-24 and bla KPC-2 (carbapenemase) are widespread. This research letter reports about a genomic surveillance study where multidrug-resistant E. coli containing CTX-M-24(IncF [F-:A1:B32]) and KPC-2(IncX3/IncU) plasmids were obtained from community- acquired urinary tract infection in Brazil., Competing Interests: Conflicts of interest The authors declare no conflicts of interest., (Copyright © 2022 Sociedade Brasileira de Infectologia. Published by Elsevier España, S.L.U. All rights reserved.)

Reports of Gram-negative bacteria harboring multiple carbapenemase genes have increased in South America, leading to an urgent need for appropriate microbiological diagnosis. We evaluated phenotypic methods for detecting Klebsiella pneumoniae carbapenemase 2 (KPC-2) and New Delhi metallo-β-lactamase-1 (NDM-1) coexpression in members of the K. pneumoniae complex (i.e., K. pneumoniae, K. quasipneumoniae, and K. variicola) isolated from human and animal hosts, based on inhibition of ceftazidime-avibactam (CZA) and aztreonam (ATM) by dipicolinic acid (DPA), EDTA, or avibactam (AVI). While the presence of bla KPC-2 and bla NDM-1 genes was confirmed by whole-genome sequencing, PCR, and/or GeneXpert, coexpression was successfully detected based on the following: (i) a ≥5-mm increase in the zone diameter of ATM (30 µg) disks plus AVI (4 or 20 µg) and ≥4-mm and ≥10-mm increases in the zone diameters for "CZA 50" (30 µg ceftazidime [CAZ] and 20 µg AVI) and "CZA 14" (10 µg CAZ and 4 µg AVI) disks, respectively, when we added DPA (1 mg/disk) or EDTA (5 mM) in a combined disk test (CDT); (ii) a positive ghost zone (synergism) between ATM (30 µg) and CZA 50 disks and between CZA 50 and DPA (1 mg) disks, using the double-disk synergy test (DDST) at a disk-disk distance of 2.5 cm; (iii) ≥3-fold MIC reductions of ATM and CZA in the presence of AVI (4 µg/mL), DPA (500 µg/mL), or EDTA (320 µg/mL); and (iv) immunochromatography. Although our results demonstrated that inhibition by AVI, DPA, and EDTA may provide simple and inexpensive methods for the presumptive detection of coexpression of KPC-2 and NDM-1 in members of the K. pneumoniae complex, additional studies are necessary to confirm the accuracy of these methodologies by testing other Gram-negative bacterial species and other KPC and NDM variants coexpressed by WHO critical priority pathogens detected worldwide. IMPORTANCE Alerts regarding the emergence and increase of combinations of carbapenemases in Enterobacterales in Latin America and the Caribbean have recently been issued by PAHO and WHO, emphasizing the importance of appropriate microbiological diagnosis and the effective and articulated implementation of infection prevention and control programs. In this study, we evaluated methods based on inhibition of ceftazidime (CAZ), ceftazidime-avibactam (CZA), and aztreonam (ATM) by dipicolinic acid (DPA), EDTA, and avibactam (AVI) inhibitors for the identification of KPC-2- and NDM-1-coexpression in members of the K. pneumoniae complex recovered from human and animal hosts. Our results demonstrate that inhibition by AVI, DPA, and EDTA may provide simple and inexpensive methods for the presumptive detection of coexpression of KPC-2 and NDM-1 in members of the K. pneumoniae complex.

Salmonella Heidelberg is a clinically-important serovar linked to food-borne illness, and commonly isolated from poultry products. Since 1962, Salmonella Heidelberg has been widely reported from poultry production systems in several countries, including Brazil. The emergence of multidrug-resistant (MDR) Salmonella Heidelberg strains in food animals underscores a significant food safety hazard. In our study, we performed antimicrobial susceptibility testing (AST) and Whole-genome sequencing (WGS) to identify the antimicrobial resistance (AMR) genes, pathogenicity mechanisms and virulence factors (VF) in Salmonella Heidelberg E2 strain recovered from a chicken carcass in Southern Brazil. Salmonella Heidelberg strain belonged to ST15 and showed to be susceptible to colistin (MIC ≤2 μg/mL) and multidrug-resistant to amoxicillin-clavulanic acid, gentamicin, ampicillin, cefaclor, cefazolin, ceftiofur, nalidixic acid, azithromycin, erythromycin, doxycycline, tetracycline and sulfonamide. We identified AMR genes mediating resistance to aminoglycosides (aac(6')-Iaa, aac(3)-VIa, aph(3')-Ia, aadA, 16S rrsD), β-lactams (blaCTX-M-2), quinolones (parC), macrolides (acrB), tetracyclines (tet(A)), fosfomycin (fosA7) and sulfonamide (sul1). Interestingly, the mutation in parC T255S has never been reported among Salmonella Heidelberg strains. To our knowledge, this is the first report of a Salmonella enterica strain harbouring 16S rrsD 471G > A, acrB F28L and acrB L40P chromosomal point mutations. Three plasmid replicon types, ST2-IncHI2, ST2-IncHI2A and IncX1 were identified. Nine Salmonella Pathogenicity Islands and 98 virulence genes encoding virulence factors were identified associated with cell adhesion, invasion, intracellular survival and resistance to antimicrobial peptides. Although Salmonella Heidelberg E2 strain likely originated from poultry, cross-contamination during meat processing cannot be excluded. This study adds to our understanding of Salmonella Heidelberg transmission along the food-chain and informs ongoing regulatory discussions on Salmonella Heidelberg in poultry., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Methicillin-resistant staphylococci have become leading cause of infectious diseases in humans and animals, being categorized as high priority pathogens by the World Health Organization. Although methicillin-resistant Staphylococcus sciuri (recently moved to Mammaliicoccus sciuri) has been widely reported in companion animals, there is scarce information regarding their clinical impact and genomic features. Herein, we reported the occurrence and genomic characteristics of methicillin-resistant M. sciuri recovered from fatal infections in pets admitted to an intensive care unit of a veterinary hospital, in Brazil. Two M. sciuri strains were isolated from bronchoalveolar lavage samples collected from dog (strain SS01) and cat (strain SS02) presenting with sepsis and acute respiratory distress syndrome. Both isolates displayed a multidrug-resistant profile, whereas whole-genome sequencing analysis confirmed the presence of the mecA gene, along to genetic determinant conferring resistance to macrolides, streptogramins, aminoglycosides, and trimethoprim. For both strains, the mec and crr gene complex shared high identity (≥97%) with analogue sequences from a M. sciuri isolated from a human wound infection, in the Czech Republic. Strains were assigned to the sequence type ST52 and the novel ST74. Phylogenomic analysis revealed a broad host range association of these strains with several hosts and sources, including humans, animals, food, and the environment through different years and geographic locations. Our findings demonstrate that infections caused by mecA-positive M. sciuri strains can be a serious threat for veterinary intensive care patients and the medical staff, with additional implications for One Health approaches., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Nilton Lincopan reports financial support was provided by Fundação de Amparo à Pesquisa do Estado de São Paulo., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

An unusual biotype of KPC-2-producing Klebsiella pneumoniae (KPC-Kpn) isolates was detected in Corrientes, Argentina, which, to their isolation date, had been free of KPC-Kpn outbreaks. Our aim was to describe the clinical epidemiology focused on genomic characterization of atypical urease-negative KPC-Kpn clinical isolates belonging to the high-risk hospital-associated clonal lineage ST340/CC258. Thirteen isolates were recovered, all of them from inpatients with KPC-Kpn infection (August 2015 to January 2016). These isolates displayed identical enterobacterial repetitive intergenic consensus-PCR electropherotype belonging to a single clonal sequence type ST340. Whole genome sequencing was performed on two KPC-Kpn and the resistome analyses revealed the following acquired resistance genes: bla KPC-2 , bla CTX-M-15, bla OXA-1, bla SHV-11, aac(3)-IId , aph(3')-Ia , aac(6')-Ib-cr, sul1 , dfrA14, catB3, fosA , and arr-3. Mutations in GyrA (S83I) and ParC (S80I) were also identified. Among the virulence determinants, yersiniabactin was detected in both strains, specifically the ybt9 locus located in ICE Kp3 . Five plasmid incompatibility groups were observed in this clone and an unusual IncP6 plasmid bearing bla KPC-2 gene (named pKpn3KP) was fully characterized. In this study, we present the first draft genome sequences of two clinical isolates of KPC-2/CTX-M-15-producing K. pneumoniae belonging to the high-risk clonal lineage ST340/CC258 associated with nosocomial outbreaks in Argentina.

Carbapenem-resistant Enterobacterales (CRE) is a critical public health problem in South America, where the prevalence of NDM metallo-betalactamases has increased substantially in recent years. In this study, we used whole genome sequencing to characterize a multidrug-resistant (MDR) Klebsiella pneumoniae (UCO-361 strain) clinical isolate from a teaching hospital in Chile. Using long-read (Nanopore) and short-read (Illumina) sequence data, we identified a novel un-typeable megaplasmid (314,976 kb, pNDM-1&#95;UCO-361) carrying the bla NDM-1 carbapenem resistance gene within a Tn 3000 transposon. Strikingly, conjugal transfer of pNDM-1&#95;UCO-361 plasmid only occurs at low temperatures with a high frequency of 4.3 × 10 -6 transconjugants/receptors at 27 °C. UCO-361 belonged to the ST1588 clone, previously identified in Latin America, and harbored aminoglycoside, extended-spectrum β-lactamases (ESBLs), carbapenem, and quinolone-resistance determinants. These findings suggest that bla NDM-1 -bearing megaplasmids can be adapted to carriage by some K. pneumoniae lineages, whereas its conjugation at low temperatures could contribute to rapid dissemination at the human-environmental interface.

Antimicrobial resistance is an ancient natural phenomenon increasingly pressured by anthropogenic activities. Escherichia coli has been used as markers of environmental contamination and human-related activity. Seabirds may be bioindicators of clinically relevant bacterial pathogens and their antimicrobial resistance genes, including extended-spectrum-beta-lactamase (ESBL) and/or plasmid-encoded AmpC (pAmpC), in anthropized and remote areas. We evaluated cloacal swabs of 20 wild magnificent frigatebirds ( Fregata magnificens ) of the Alcatrazes Archipelago, the biggest breeding colony of magnificent frigatebirds in the southern Atlantic and a natural protected area with no history of human occupation, located in the anthropized southeastern Brazilian coast. We characterized a highly virulent multidrug-resistant ST648 (O153:H9) pandemic clone, harboring bla CTX - M -2 , bla CMY -2 , qnrB, tetB, sul1, sul2, aadA1, aac(3)-VIa and mdfA , and virulence genes characteristic of avian pathogenic (APEC) ( hlyF, iroN, iss, iutA , and ompT ) and other extraintestinal E. coli (ExPEC) ( chuA, kpsMII , and papC ). To our knowledge, this is the first report of ST648 E. coli co-producing ESBL and pAmpC in wild birds inhabiting insular environments. We suggest this potentially zoonotic and pathogenic lineage was likely acquired through indirect anthropogenic contamination of the marine environment, ingestion of contaminated seafood, or by intra and/or interspecific contact. Our findings reinforce the role of wild birds as anthropization sentinels in insular environments and the importance of wildlife surveillance studies on pathogens of critical priority classified by the World Health Organization., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Ewbank, Fuentes-Castillo, Sacristán, Esposito, Fuga, Cardoso, Godoy, Zamana, Gattamorta, Catão-Dias and Lincopan.)

Psittacine birds are commonly kept as companion birds and the maintenance of these birds in captivity may represent a zoonotic risk and contribute to the propagation of multidrug-resistant and β-lactamase extended-spectrum (ESBLs)-producing pathogens. This study aimed to identify and characterize strains of the Klebsiella pneumoniae complex isolated from diseased psittacine birds, determining virulence and resistance profiles. K. pneumoniae strains were isolated from 16 birds (16/46). All strains carried more than three virulence genes, with a high frequency of fimH and kpn (93.75%), uge (87.52%), and irp-2 (81.25%) genes. The antimicrobial susceptibility revealed that 3/16 strains were ESBL producers. Genomic analysis revealed that CTX-M-15-positive strains belonged to sequence types (STs) ST15, ST147, and ST307, characterized as international clones associated with outbreaks of healthcare-associated infections (HAIs) worldwide.

Extended spectrum β-lactamase (ESBL)-producing Klebsiella pneumoniae is a medically important pathogen that commonly causes human nosocomial infections. Since veterinary emergency and critical care services have also significantly progressed over the last decades, there are increasing reports of ESBL-producing K. pneumoniae causing hospital-associated infections in companion animals. We present microbiological and genomic analysis of a multidrug-resistant ESBL-positive K. pneumoniae (LCKp01) isolated from a fatal infection in a dog admitted to a veterinary intensive care unit. LCKp01 strain belonged to the sequence type ST392 and displays a KL27 (wzi-187) and O-locus 4 (O4). A broad resistome and presence of the bla CTX-M-15 ESBL gene were predicted. SNP-based phylogenomic analysis, using an international genome database, clustered LCKp01 (60-80 SNPs differences) with K. pneumoniae ST392 from human and animal infections, isolated at 4-year interval, whereas phylogeographical analysis confirmed successful expansion of ST392 as a global clone of One Health concern., Competing Interests: All authors declare no conflicts of interest., (© 2022 The Authors.)

Salmonella enterica subsp. enterica serovar Heidelberg has been associated with a broad host range, such as poultry, dairy calves, swine, wild birds, environment, and humans. The continuous evolution of S . Heidelberg raises a public health concern since there is a global dispersal of lineages harboring a wide resistome and virulome on a global scale. Here, we characterized the resistome, phylogenetic structure and clustered regularly interspaced short palindromic repeats (CRISPR) array composition of 81 S . Heidelberg strains isolated from broiler farms ( n = 16), transport and lairage ( n = 5), slaughterhouse ( n = 22), and retail market ( n = 38) of the poultry production chain in Brazil, between 2015 and 2016 using high-resolution approaches including whole-genome sequencing (WGS) and WGS-derived CRISPR genotyping. More than 91% of the S . Heidelberg strains were multidrug-resistant. The total antimicrobial resistance (AMR) gene abundances did not vary significantly across regions and sources suggesting the widespread distribution of antibiotic-resistant strains from farm to market. The highest AMR gene abundance was observed for fosA7 , aac(6')-Iaa , sul2 , tet(A) , gyrA , and parC for 100% of the isolates, followed by 88.8% for bla CMY -2 . The β-lactam resistance was essentially driven by the presence of the plasmid-mediated AmpC (pAmpC) bla CMY -2 gene, given the isolates which did not carry this gene were susceptible to cefoxitin (FOX). Most S . Heidelberg strains were classified within international lineages, which were phylogenetically nested with Salmonella strains from European countries; while CRISPR genotyping analysis revealed that the spacer content was overall highly conserved, but distributed into 13 distinct groups. In summary, our findings underscore the potential role of S . Heidelberg as a key pathogen disseminated from farm to fork in Brazil and reinforce the importance of CRISPR-based genotyping for salmonellae. Hence, we emphasized the need for continuous mitigation programs to monitor the dissemination of this high-priority pathogen., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Monte, Nethery, Berman, Keelara, Lincopan, Fedorka-Cray, Barrangou and Landgraf.)

Carbapenemase-producing Enterobacterales are rapidly spreading and adapting to different environments beyond hospital settings. During COVID-19 lockdown, a carbapenem-resistant NDM-1-positive Escherichia coli isolate (BA01 strain) was recovered from a pygmy sperm whale ( Kogia breviceps ), which was found stranded on the southern coast of Brazil. BA01 strain belonged to the global sequence type (ST) 162 and carried the bla NDM-1 , besides other medically important antimicrobial resistance genes. Additionally, genes associated with resistance to heavy metals, biocides, and glyphosate were also detected. Halophilic behavior (tolerance to > 10% NaCl) of BA01 strain was confirmed by tolerance tests of NaCl minimal inhibitory concentration, whereas halotolerance associated genes katE and nhaA , which encodes for catalase and Na + /H + antiporter cytoplasmic membrane, respectively, were in silico confirmed. Phylogenomics clustered BA01 with poultry- and human-associated ST162 lineages circulating in European and Asian countries. Important virulence genes, including the astA (a gene encoding an enterotoxin associated with human and animal infections) were detected, whereas in vivo experiments using the Galleria mellonella infection model confirmed the virulent behavior of the BA01 strain. WHO critical priority carbapenemase-producing pathogens in coastal water are an emerging threat that deserves the urgent need to assess the role of the aquatic environment in its global epidemiology., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Sellera, Cardoso, Fuentes-Castillo, Esposito, Sano, Fontana, Fuga, Goldberg, Seabra, Antonelli, Sandri, Kolesnikovas and Lincopan.)