Antimicrobial Selective Pressure Research Articles

Structure of Extracellular Vesicles and Their Effect on Bacterial Biofilms Change with the Development of Antibiotic Resistance in the Probiotic Strain Lactiplantibacillus plantarum 8p-a3.

Significant changes in lactobacillus-derived extracellular membrane vesicles, which transfer lipids, polysaccharides, proteins, and nucleic acids, were for the first time observed to accompany the development of resistance to antibiotics (amoxicillin and clarithromycin) in vitro in the probiotic strain Lactiplantibacillus plantarum 8p-a3. The changes occur together with large-scale genome rearrangements, changes in the profile of phenotypic sensitivity to antimicrobials of various groups, and evolution of virulence. Changes in vesicles affected their structure, composition, and activity against biofilms of opportunistic bacteria. The data provide for a better understanding of the molecular mechanisms of microbial survival under selective pressure of antimicrobials, the functional potential of probiotic vesicles, and probiotic safety assessments.

Characterization of a novel multi-resistant Pseudomonas juntendi strain from China with chromosomal blaVIM-2 and a megaplasmid coharboring blaIMP-1-like and blaOXA-1.

Pseudomonas juntendi is a newly identified opportunistic pathogen, of which we have limited understanding. P. juntendi strains are often multidrug resistant, which complicates clinical management of infection. A strain of Pseudomonas juntendi (strain L4326) isolated from feces was characterized by MALDI-TOF-MS and Average Nucleotide Identity BLAST. This strain was further subject to whole-genome sequencing and Maximum Likelihood phylogenetic analysis. The strain was phenotypically characterized by antimicrobial susceptibility testing and conjugation assays. We have isolated the novel P. juntendi strain L4236, which was multidrug resistant, but retained sensitivity to amikacin. L4236 harbored a megaplasmid that encoded blaOXA-1 and a novel blaIMP-1 resistance gene variant. P. juntendi strain L4236 was phylogenetically related to P. juntendi strain SAMN30525517. A rare P. juntendi strain was isolated from human feces in southern China with a megaplasmid coharboring blaIMP-1-like and blaOXA-1. Antimicrobial selection pressures may have driven acquisition of drug-resistance gene mutations and carriage of the megaplasmid.

Evaluation of antimicrobial selective pressure using the multicenter semiautomatic surveillance system Japan surveillance for infection prevention and healthcare epidemiology

BackgroundEvaluating the selective pressure of antimicrobials on bacteria is important for promoting antimicrobial stewardship programs (ASPs). The aim of this study was to assess the selective pressure of antimicrobials by evaluating their use (carbapenem [CBP] and CBP-sparing therapy) over time and the detection status of CBP-resistant organisms using multicenter data. MethodsAmong the facilities whose data were registered in the Japan Surveillance for Infection Prevention and Healthcare Epidemiology from 2017 to 2020, those that had data on the use of CBP and CBP-sparing therapy (fluoroquinolones [FQs], cefmetazole [CMZ], piperacillin–tazobactam [PIP/TAZ], ampicillin–sulbactam [ABPC/SBT], ceftriaxone/cefotaxime [CTRX/CTX], CAZ (ceftazidime), cefepime [CFPM], and aminoglycosides [AGs]) as well as on CBP-resistant Enterobacterales (CRE) and CBP-resistant Pseudomonas aeruginosa (CRPA) detection were included. Alcohol-based hand rubbing (ABHR) usage was also analyzed. Regression analyses, including multivariable regression analysis, were performed to evaluate trends. The association of antimicrobial use density (AUD) with CRE and CRPA detection rates was evaluated. ResultsIn 28 facilities nationwide, CBP, FQ, CAZ, AG, and PIP/TAZ use decreased over the 3-year period, whereas the use of CMZ, ABPC/SBT, CTRX/CTX, CFPM, and ABHR as well as the rates of CRE and CRPA detection increased. The average AUD did not significantly correlate with CRE and CRPA detection rates. The multivariable regression analysis did not reveal any significant correlation between each AUD or ABHR and CRE or CRPA detection. ConclusionCBP and ABHR use showed a decreasing and an increasing trend, respectively, while CRPA and CRE detection rates exhibited a gradual increase. The considerably low CRE and CRPA detection rates suggest that slight differences in numbers may have been observed as excessive trend changes. Further investigation is warranted to evaluate selective pressure while considering the characteristics of ASP and the mechanisms underlying resistance.

Generation and maintenance of the circularized multimeric IS26-associated translocatable unit encoding multidrug resistance

In gram-negative bacteria, IS26 often exists in multidrug resistance (MDR) regions, forming a pseudocompound transposon (PCTn) that can be tandemly amplified. It also generates a circular intermediate called the “translocatable unit (TU)”, but the TU has been detected only by PCR. Here, we demonstrate that in a Klebsiella pneumoniae MDR clone, mono- and multimeric forms of the TU were generated from the PCTn in a preexisting MDR plasmid where the inserted form of the TU was also tandemly amplified. The two modes of amplification were reproduced by culturing the original clone under antimicrobial selection pressure, and the amplified state was maintained in the absence of antibiotics. Mono- and multimeric forms of the circularized TU were generated in a RecA-dependent manner from the tandemly amplified TU, which can be generated in RecA-dependent and independent manners. These findings provide novel insights into the dynamic processes of genome amplification in bacteria.

Whole genome-based antimicrobial resistance, virulence, and phylogenetic characteristics of Trueperella pyogenes clinical isolates from humans and animals

Trueperella pyogenes is an opportunistic zoonotic bacterial pathogen, whose antimicrobial resistance, virulence, and genetic relatedness between strains from animals and humans are barely studied. These characteristics were therefore analyzed for clinical T. pyogenes strains from 31 animals of 11 different species and 8 humans determining their complete circular genome sequence and antimicrobial susceptibility. The MICs of 19 antimicrobials including 3 antiseptics correlated to the resistance genes identified in silico within the genomes revealing a predominance of resistance to streptomycin (aadA9), sulfamethoxazole (sul1), and tetracycline (tet(33), tet(W/N/W)) among strains from humans and cattle. Additional resistance genes (erm(X), erm(56), cmx, drfA1, aadA1, aph(3'')-Ib (strA), aph(6)-Id (strB), aac(3)-IVa, aph(4)-Ia) were found only sporadically. The resistance genes were localized on genetic elements integrated into the chromosome. A cgMLST-based phylogenetic analysis revealed two major clusters each containing genetically diverse strains. The human strains showed the closest relatedness to strains from cattle. Virulence genes coding for fimbriae (fimA, fimC), neuroamidase (nanP, nanH), pyolysin (plo), and collagen binding protein (cbpA) were identified in strains from different hosts, but no correlation was observed between virulence factors and strain origin. The existence of resistance genes typically found in Gram-negative bacteria within the Gram-positive T. pyogenes indicates a wider capacity to adapt to antimicrobial selective pressure. Moreover, the presence of similar antimicrobial resistance profiles found in cattle and human strains as well as their closest relatedness suggests common zoonotic features and cattle as the potential source for human infections.

Correlation between the spread of IMP-producing bacteria and the promoter strength of blaIMP genes.

The first report of transmissible carbapenem resistance encoded by blaIMP-1 was discovered in Pseudomonas aeruginosa GN17203 in 1988, and blaIMP-1 has since been detected in other bacteria, including Enterobacterales. Currently, many variants of blaIMPs exist, and point mutations in the blaIMP promoter have been shown to alter promoter strength. For example, the promoter (Pc) of blaIMP-1, first reported in P. aeruginosa GN17203, was a weak promoter (PcW) with low-level expression intensity. This study investigates whether point mutations in the promoter region have helped to create strong promoters under antimicrobial selection pressure. Using bioinformatic approaches, we retrieved 115 blaIMPs from 14,529 genome data of Pseudomonadota and performed multiple alignment analyses. The results of promoter analysis of the 115 retrieved blaIMPs showed that most of them used the Pc located in class 1 integrons (n = 112, 97.4%). The promoter analysis by year revealed that the blaIMP population with the strong promoter, PcS, was transient. In contrast, the PcW-TG population, which had acquired a TGn-extended -10 motif in PcW and had an intermediate promoter strength, gradually spread throughout the world. An inverse correlation between Pc promoter strength and Intl1 integrase excision efficiency has been reported previously [1]. Because of this trade-off, it is unlikely that blaIMPs with strong promoters will increase rapidly, but the possibility that promoter strength will increase with the use of other integrons cannot be ruled out. Monitoring of the blaIMP genes, including promoter analysis, is necessary for global surveillance of carbapenem-resistant bacteria.

Whole-genome sequencing-based antimicrobial resistance and shedding dynamics of Escherichia coli isolated from calves before and after antimicrobial group treatments.

The continued emergence and spread of antimicrobial resistance (AMR) determinants are serious global concerns. The dynamics of AMR spread and persistence in bacterial and animal host populations are complex and not solely driven by antimicrobial selection pressure. In calf fattening, both antimicrobial use and carriage prevalence of antimicrobial-resistant bacteria are generally recognized as high. This study provides new insights into the short-term, within-farm dynamics and transmission of AMR determinants in Escherichia coli from the dominant fecal flora of calves subjected to antimicrobial group treatments during the rearing period. The diversity of E. coli strains decreased over time, although, in contrast to previous observations in extended-spectrum β-lactamase-producing Enterobacterales, the predominance of a few clones was not observed. The spread of AMR determinants occurred through the dissemination of clonal strains among calves. The median number per strain of AMR determinants conferring resistance to selected antimicrobials decreased toward the end of the rearing period.

Population Analysis of Daptomycin-non-Susceptible Methicillin-Resistant Staphylococcus aureus Reveals the Presence of Variants That Contribute to Daptomycin Resistance.

In this study, population analysis (PA) of methicillin-resistant Staphylococcus aureus (MRSA), before and after long-duration daptomycin (DAP) treatment, was used to detect subpopulations with different susceptibilities to DAP and to verify the changes in the number of resistant cells. Furthermore, we aimed to characterize the bacteriology of the variants present in the non-susceptible cell subpopulation. A DAP non-susceptible (NS) MRSA phenotype (D2) that emerged from a DAP- susceptible MRSA phenotype (D1) during treatment of an open wound, was used for testing. We performed bacteriological and genetic analyses of cryptic DAP-NS MRSA variants detected by PA to study the variants present in the resistant cell subpopulation. PA results suggest that MRSA adapted to survival in the presence of DAP are selected leading to reduced susceptibility. Within the cell population growing in media containing 2.0mg/L of DAP, three variants with different pigment production and colony size were detected. Variant 3 was an orange colony due to enhanced production of staphyloxanthin. Our results revealed that the DAP minimum inhibitory concentration (MIC) value increased two-fold (4mg/L) in variant 3, in which pigment production was most enhanced, compared to the parental strain D2. In conclusion, our results indicate that long-duration DAP treatment can lead to the emergence and increased proportion of DAP-NS subpopulations. Furthermore, slow-growing variants that can be detected only under antimicrobial selective pressure are present among DAP-NS cells, suggesting that these variants may also contribute to the development of DAP resistance.

Emergence of carbapenemase-producing Enterobacterales (CPE) in patients with severe COVID-19 infection and successful control in intensive care.

We aim to highlight therisks of acquiring carbapenemase-producing Enterobacterales (CPE) resistance genes in patients with severe coronavirus disease 2019 (COVID-19) in intensive care. Outbreak analysis to assess for a transmission risk area (TRA) conducted after identification ofpotential CPE outbreak within shared room spaces in intensive care. Analysis conducted within a24-bed single-room model intensive-care department within a level-3 tertiary center public hospital in regional Victoria, Australia. 3 patients, with severe COVID-19 admitted to intensive care over a3-month period with shared room spaces requiring prolonged mechanical ventilation and broad-spectrum antimicrobials, identified and were managed for CPE isolated from sputum. Overlap carbapenemase genes were identified among different organisms raising suspicion of transmitted resistance genes. A subsequent case managed for severe community-acquired pneumonia isolated CPE 3 months beyond these cases. Outbreak analysis via weekly cross-sectional point prevalence screening of fecal samples or rectal swabs for CPE from patients admitted to the intensive-care department over a4-week period. 34 patients were included in the analysis with 51 tests for CPE screening conducted. No further cases of CPE were identified. Statewide Infection Surveillance team and theDepartment of Health and Human Services did not find the cases to derive from a TRA. No further action including environmental screening was indicated. These cases highlight theindependent acquisition of CPE genes in patients with severe COVID-19 and antimicrobial selective pressures resulting in significant morbidity and mortality. Increasing awareness, robust antimicrobial stewardship, and infection prevention measures could reduce pressures driving CPE resistance mutations and therisk of CPE transmission.

Genotypic Evolution of Klebsiella pneumoniae Sequence Type 512 during Ceftazidime/Avibactam, Meropenem/Vaborbactam, and Cefiderocol Treatment, Italy.

In February 2022, a critically ill patient colonized with a carbapenem-resistant K. pneumoniae producing KPC-3 and VIM-1 carbapenemases was hospitalized for SARS-CoV-2 in the intensive care unit of Policlinico Umberto I hospital in Rome, Italy. During 95 days of hospitalization, ceftazidime/avibactam, meropenem/vaborbactam, and cefiderocol were administered consecutively to treat 3 respiratory tract infections sustained by different bacterial agents. Those therapies altered the resistome of K. pneumoniae sequence type 512 colonizing or infecting the patient during the hospitalization period. In vivo evolution of the K. pneumoniae sequence type 512 resistome occurred through plasmid loss, outer membrane porin alteration, and a nonsense mutation in the cirA siderophore gene, resulting in high levels of cefiderocol resistance. Cross-selection can occur between K. pneumoniae and treatments prescribed for other infective agents. K. pneumoniae can stably colonize a patient, and antimicrobial-selective pressure can promote progressive K. pneumoniae resistome evolution, indicating a substantial public health threat.

Coexistence of blaKPC-2 and blaNDM-1 in one IncHI5 plasmid confers transferable carbapenem resistance from a clinical isolate of Klebsiella michiganensis in China

This study firstly identified an IncHI5 plasmid pK254-KPC_NDM co-carrying two different class carbapenemase genes blaKPC-2 and blaNDM-1 in Klebsiella michiganensis K254. The strain K254 was sequenced by high-throughput genome sequencing. A detailed genomic and phenotypic characterization of pK254-KPC_NDM was performed. pK254-KPC_NDM displayed the conserve IncHI5 backbone and carried a resistant accessory region: Tn1696-related transposon Tn7414 containing blaKPC-2 and blaNDM-1. A sequence comparison was applied to a collection of four Tn1696-related transposons (Tn7414-Tn7417) harbouring carbapenemase genes. For all these four transposons, the blaNDM-1 was carried by Tn125 derivatives within three different mobile genetic elements. Tn7414 further acquired another carbapenemase gene, blaKPC-2, because of the integration of the local blaKPC-2 genetic environment from Tn6296, resulting in the high-level carbapenem resistance of K. michiganensis K254. The conjugal transfer and plasmid stability experiments confirmed that pK254-KPC_NDM could be transferred intercellularly and keep the stable vertical inheritance in different bacteria, which would contribute to the further dissemination of multiple carbapenemase genes and enhance the adaption and survival of K. michiganensis under complex and diverse antimicrobial selection pressures. This study was the first to report the K. michiganensis isolate coharbouring blaKPC-2 and blaNDM-1 in the Tn1696-related transposon in IncHI5 plasmid. The emergence of novel transposons simultaneously carrying multiple carbapenemase genes might contribute to the further dissemination of high-level carbapenem resistance in the isolates of the hospital settings and pose new challenges for the treatment of nosocomial infection.

Effectiveness of antimicrobial interventions directed at tackling antimicrobial resistance in animal production: A systematic review and meta-analysis.

In the last decades, a more prudent and rational use of antimicrobials has been progressively directed towards animal production to reduce antimicrobial selective pressure and antimicrobial resistance (AMR) in microorganisms and safeguard the antimicrobial efficacy of treatments in human medicine. This systematic review evaluated the effectiveness of interventions that have been applied to reduce or improve veterinary antimicrobial usage and aimed at decreasing resistant bacteria in chicken broiler and pig production contexts. Original articles were identified by searching PubMed™, Scopus™, The Cochrane Library™, and Web of Science™, and grey literature by searching DANS EASY™, WorldCat™ and RCAAP™. Inclusion criteria included: chicken broiler or pig populations (predestined for meat production), interventions intended to reduce/improve antimicrobial use, comparator with standard or no use of antimicrobials, outcomes related to prevalence of resistant bacteria, farm level studies, original data, and analytical observational studies. Data was extracted from eligible studies and meta-analysis using random or fixed effects models was conducted for combinations including type of intervention, bacterial species, production type and animal populations. Models were selected according to heterogeneity between studies. The effectiveness of interventions was assessed using pooled odds ratio of resistance to antimicrobial substances/classes by bacteria for associations between animal populations with and without intervention. A total of 46 studies were eligible for review. For chicken broilers, most interventions were identified as antimicrobial restrictions on all non-therapeutic use (46%), complete restriction (27%), and prohibition on antimicrobials used for growth promotion (23%). As for pig populations, restrictions were mainly observed on all non-therapeutic use (37%), complete restriction (37%) and group treatments (22%). For meta-analysis, 21 studies were pooled after assessment of existing combinations. These combinations demonstrated a protective effect for most antimicrobial classes in Escherichia coli, Campylobacter and Enterococcus isolates from samples of chicken broilers as well in Escherichia coli and Campylobacter spp. from samples of pigs, compared to animals raised under conventional production or without intervention. Increased odds of resistance were only observed for cephalosporins in E. coli and broilers raised without antimicrobials, and to fluoroquinolones and quinolones in Campylobacter and pigs raised without antimicrobials, compared to conventional production. Our study indicates that organic production, antimicrobial-free farms, and group treatment restrictions are recommended for AMR reduction, providing information that may support decision-making to tackle AMR and better reporting to improve comparability of results between studies.

Genetic Diversity and Antimicrobial Resistance of Extraintestinal E. coli Populations Pre- and Post-Antimicrobial Therapy on Broilers Affected by Colisepticemia.

The aim of the present study was to investigate the genetic diversity and antimicrobial resistance (AMR) of E. coli during enrofloxacin therapy in broilers affected by colisepticemia. Three unrelated farms with ongoing colibacillosis outbreaks were sampled at day 1 before treatment and at days 5, 10 and 24 post-treatment. A total of 179 E. coli isolates were collected from extraintestinal organs and submitted to serotyping, PFGE and the minimum inhibitory concentration (MIC) against enrofloxacin. PFGE clusters shifted from 3-6 at D1 to 10-16 at D5, D10 and D24, suggesting an increased population diversity after the treatment. The majority of strains belonged to NT or O78 and to ST117 or ST23. PFGE results were confirmed with SNP calling: no persistent isolates were identified. An increase in resistance to fluoroquinolones in E. coli isolates was observed along the treatment. Resistome analyses revealed qnrB19 and qnrS1 genes along with mutations in the gyrA, parC and parE genes. Interestingly, despite a fluoroquinolone selective pressure, qnr-carrying plasmids did not persist. On the contrary, two conjugative AMR plasmid clusters (AB233 and AA474) harboring AMR genes other than qnr were persistent since they were identified in both D1 and D10 genomes in two farms. Further studies should be performed in order to confirm plasmid persistence not associated (in vivo) to antimicrobial selective pressure.

Prevalence and antimicrobial susceptibility of Mycoplasma bovis from the upper and lower respiratory tracts of healthy feedlot cattle and those diagnosed with bovine respiratory disease

Mycoplasma bovis is an important respiratory pathogen of cattle. In this study, the prevalence and antimicrobial susceptibility of M. bovis were evaluated from two Cohorts of feedlot cattle spanning an 8-year period. In the first study conducted in 2008–2009, nasopharyngeal swabs from cattle sampled at feedlot entry and after 60 days on feed were collected (Cohort 1). In a second study conducted in 2015–2016, nasopharyngeal and trans-tracheal samples were collected from cattle diagnosed with bovine respiratory disease (BRD) and matching healthy controls (Cohort 2). For Cohort 1, the prevalence of M. bovis was lower in cattle at entry compared to when the same individuals were sampled ≥60 days later (P < 0.05). For Cohort 2, the prevalence of M. bovis was greater in both nasopharyngeal and tracheal samples from cattle diagnosed with BRD, compared to controls (P < 0.05). In both Cohorts, almost all isolates were resistant to tilmicosin. Compared to M. bovis from Cohort 1, isolates of Cohort 2 exhibited increased resistance to clindamycin, enrofloxacin, florfenicol, tylosin, and tulathromycin, with the latter showing resistance levels >90 %. These data suggest that antimicrobials used to prevent and treat BRD selected for resistance in M. bovis over the 8-year period. For macrolides, cross-resistance occurred and M. bovis can retain resistance even when antimicrobial selection pressure is removed. Within 9 years of commercial availability of tulathromycin, the majority of M. bovis displayed resistance. Therefore, longitudinal evaluation of resistance in respiratory pathogens is important to ensure efficacious treatment of BRD.

Antibiotic, Heavy Metal, and Biocide Concentrations in a Wastewater Treatment Plant and Its Receiving Water Body Exceed PNEC Limits: Potential for Antimicrobial Resistance Selective Pressure.

Although the rise in antimicrobial resistance has been attributed mainly to the extensive and indiscriminate use of antimicrobials such as antibiotics and biocides in humans, animals and on plants, studies investigating the impact of this use on water environments in Africa are minimal. This study quantified selected antibiotics, heavy metals, and biocides in an urban wastewater treatment plant (WWTP) and its receiving water body in Kwazulu-Natal, South Africa, in the context of the predicted no-effect concentrations (PNEC) for the selection of antimicrobial resistance (AMR). Water samples were collected from the WWTP effluent discharge point and upstream and downstream from this point. Heavy metals were identified and quantified using the United States Environmental Protection Agency (US EPA) method 200.7. Biocides and antibiotic residues were determined using validated ultra-high-performance liquid chromatography with tandem mass spectrometry-based methods. The overall highest mean antibiotic, metal and biocide concentrations were observed for sulfamethoxazole (286.180 µg/L), neodymium (Nd; 27.734 mg/L), and benzalkonium chloride (BAC 12) (7.805 µg/L), respectively. In decreasing order per sampling site, the pollutant concentrations were effluent > downstream > upstream. This implies that the WWTP significantly contributed to the observed pollution in the receiving water. Furthermore, most of the pollutants measured recorded values exceeding the recommended predicted no-effect concentration (PNEC) values, suggesting that the microbes in such water environments were at risk of developing resistance due to the selection pressure exerted by these antimicrobials. Further studies are required to establish such a relationship.

The fitness cost of horizontally transferred and mutational antimicrobial resistance in Escherichia coli.

Antimicrobial resistance (AMR) in bacteria implies a tradeoff between the benefit of resistance under antimicrobial selection pressure and the incurred fitness cost in the absence of antimicrobials. The fitness cost of a resistance determinant is expected to depend on its genetic support, such as a chromosomal mutation or a plasmid acquisition, and on its impact on cell metabolism, such as an alteration in an essential metabolic pathway or the production of a new enzyme. To provide a global picture of the factors that influence AMR fitness cost, we conducted a systematic review and meta-analysis focused on a single species, Escherichia coli. By combining results from 46 high-quality studies in a multilevel meta-analysis framework, we find that the fitness cost of AMR is smaller when provided by horizontally transferable genes such as those encoding beta-lactamases, compared to mutations in core genes such as those involved in fluoroquinolone and rifampicin resistance. We observe that the accumulation of acquired AMR genes imposes a much smaller burden on the host cell than the accumulation of AMR mutations, and we provide quantitative estimates of the additional cost of a new gene or mutation. These findings highlight that gene acquisition is more efficient than the accumulation of mutations to evolve multidrug resistance, which can contribute to the observed dominance of horizontally transferred genes in the current AMR epidemic.

Prevalence of carbapenem-resistant Enterobacteriaceae in in- and out-of-hospital environments

Introduction: Healthcare is recognized as a condition of high selective pressure (SP) due to the use of antimicrobials. The emergence of community bacterial species resistant to the antimicrobials mostextensively used in hospitals is of concern, as is the case with Gram-negative rods producing extended-spectrum beta-lactamases (ESBL). Hospital colonization by carbapenem-resistant Enterobacteria (CRE) can persist for many months after hospital discharge, and their encoding genes can be transferred among different species. Methodology: Because of the scarcity of options due to resistance to carbapenems, we sought to evaluate the occurrence of CRE in stool samples from outpatients and inpatients of a University Hospital, for whom clinical and epidemiological data were obtained from the analysis of medical records. After parasitological examination, stool samples from outpatients and inpatients were diluted in saline (1:20, 1:1000 and 1:2000) solution and 0.1 mL seeding was performed on MacConkey Agar (MA) containing gentamicin 8 μg/mL and MA containing cephalexin 32 μg/mL. Once isolated, strains were identified by biochemical tests and AST (with imipenem, ertapenem, and meropenem) was performed according to CLSI (2020). Results: We isolated carbapenem-resistant strains of enterobacteria in 13 (43.3%) of the 30 inpatients and in 13 (29%) of the 31 outpatients (p&gt;0.05). In 7 (23.3%) of the 30 inpatients and in 5 (16.1%) of the 31 outpatients, we isolated CRE at 1:1000 or 1:5000 dilutions of stool samples, which correspond to “microbial loads”. Ten bacterial species were isolated in the CRE related to the 13 inpatients and 13 outpatients, among whom we identified two or more species in 9 (69.2%) and 10 (76.9%), respectively. Even taking into account the limitations of the study due to the possibility of bias arising from the absence of data in the medical records, the detection of intestinal colonization with CRE in non-hospitalized individuals is of concern and may jeopardize the implementation of rational empirical therapy in patients of that community. Conclusion: It is possible that, as was the case of ESBL-producing strains decades ago, infections by carbapenem-resistant strains have become endemic in the community.In general, this development is a cause of concern, for plasmid-associated antimicrobial resistance strains, due to the fact that an often-recognized plasmid-associated fitness of bacterial cell favors persistence of the strains, even in the absence of antimicrobial (co-)selective pressures.

Short-term antibiotic therapy for the most common bacterial respiratory infections in infants and children.

Overuse and misuse of antibiotics have strongly accelerated the progressive increase in bacterial antimicrobial resistance (AMR). The evidence that antimicrobial selective pressure was greater the longer the antibiotic therapy was continued has led some experts to reconsider duration of antibiotic therapy testing the use of short-term drug administration. If as effective as long-term therapy, short-term therapy could have been an easy measure to limit AMR emergence. In the present narrative review, whether present knowledge on short-term therapy of acute streptococcal pharyngitis (ASF), acute otitis media (AOM) and mild to moderate community-acquired pneumonia (CAP) allows systematic use of short-term therapy in infants and children with these diseases is discussed. Literature analysis showed that reducing the duration of antibiotic therapy for some of the most common pediatric respiratory infections could be a valid measure to contain the antibiotic abuse and the consequent impact on the emergence of AMR. Several data seem to indicate that this type of intervention is possible, as short-term therapy has been found as effective as the traditionally recommended long-term therapy in some cases of ASF, AOM and mild to moderate CAP. However, further studies are needed to better characterize infants and children who can have benefit with short-term antibiotic therapy in common bacterial respiratory infections.

Characterizations of blaCTX-M-14 and blaCTX-M-64 in a clinical isolate of Escherichia coli from China.

Extended-spectrum beta-lactamase-producing Gram-negative bacteria are common in the community and hospitals. To monitor ESBLs mediated by the CTX-M genotype, we collected clinical ESBL pathogenic strains from a hospital in central China and observed a strain of Escherichia coli, namely Ec15103 carrying blaCTX-M-14, blaCTX-M-64 and blaTEM-1, isolated from the blood of a 7-day-old infant in 2015. Strain Ec15103 contains two drug resistance plasmids: pEc15103A, an IncFI-type plasmid that cannot be conjugatively transferred and carries the drug resistance genes blaTEM-1, aacC2, aadA5, sul1, mph(A), sul2, strAB, and tetA(A); and pEc15103B, an IncK2/Z-type plasmid that carries the conjugation transfer gene and blaCTX-M-14. In addition, blaCTX-M-64 is located on the chromosome of Ec15103, and it is the first report of pathogen with blaCTX-M-64 located on its chromosome (the search terms used "blaCTX-M-64" and "chromosome"). blaCTX-M-14 and blaCTX-M-64 are carried by ISEcp1-mediated transposon Tn6503a and Tn6502, respectively. The conjugation transfer ability of pEc15103B was significantly inhibited by zidovudine (AZT) and linoleic acid (LA) and that expression of blaCTX-M-14, blaCTX-M-64 and blaTEM-1 at the mRNA level did not change based on the concentration of cefotaxime or ampicillin. Co-occurrence of blaCTX-M-14 and blaCTX-M-64 in a single isolate will enhance the drug resistance of bacteria, and the presence of blaCTX-M-64 in the chromosome may make the resistance more maintain. This fact will facilitate its dissemination and persistence under different antimicrobial selection pressures. It is essential to prevent these strains from further spreading in a hospital environment.

Comparison of PCV10, PCV13, PCV15, PCV20 and PPSV23 vaccine coverage of invasive Streptococcus pneumoniae isolate serotypes in Canada: the SAVE study, 2011-20.

As pneumococci evolve under vaccine, antimicrobial and other selective pressures, it is important to track isolates covered by established (PCV10, PCV13 and PPSV23) and new (PCV15 and PCV20) vaccine formulations. To compare invasive pneumococcal disease (IPD) isolates from serotypes covered by PCV10, PCV13, PCV15, PCV20 and PPSV23, collected in Canada from 2011 to 2020, by demographic category and antimicrobial resistance phenotype. IPD isolates from the SAVE study were initially collected by members of the Canadian Public Health Laboratory Network (CPHLN) as part of a collaboration between the Canadian Antimicrobial Resistance Alliance (CARA) and the Public Health Agency of Canada (PHAC). Serotypes were determined by quellung reaction, and antimicrobial susceptibility testing was performed using the CLSI broth microdilution method. A total of 14 138 invasive isolates were collected from 2011 to 2020, with 30.7% of isolates covered by the PCV13 vaccine, 43.6% of isolates covered by the PCV15 vaccine (including 12.9% non-PCV13 serotypes 22F and 33F), and 62.6% of isolates covered by the PCV20 vaccine (including 19.0% non-PCV15 serotypes 8, 10A, 11A, 12F and 15B/C). Non-PCV20 serotypes 2, 9N, 17F and 20, but not 6A (present in PPSV23) represented 8.8% of all IPD isolates. Higher-valency vaccine formulations covered significantly more isolates by age, sex, region and resistance phenotype including MDR isolates. Coverage of XDR isolates did not significantly differ between vaccine formulations. When compared with PCV13 and PCV15, PCV20 covered significantly more IPD isolates stratified by patient age, region, sex, individual antimicrobial resistance phenotypes and MDR phenotype.