Impact of the COVID-19 pandemic on extended-spectrum ß-lactamase producing Escherichia coli in urinary tract and blood stream infections: results from a nationwide surveillance network, Finland, 2018 to 2022.

BACKGROUND: Before the COVID-19 pandemic there has been a constant increase in antimicrobial resistance (AMR) of Escherichia coli, the most common cause of urinary tract infections and bloodstream infections. The aim of this study was to investigate the impact of the COVID-19 pandemic on extended-spectrum ß-lactamase (ESBL) production in urine and blood E. coli isolates in Finland to improve our understanding on the source attribution of this major multidrug-resistant pathogen. METHODS: Susceptibility test results of 564,233 urine (88.3% from females) and 23,860 blood E. coli isolates (58.8% from females) were obtained from the nationwide surveillance database of Finnish clinical microbiology laboratories. Susceptibility testing was performed according to EUCAST guidelines. We compared ESBL-producing E. coli proportions and incidence before (2018-2019), during (2020-2021), and after (2022) the pandemic and stratified these by age groups and sex. RESULTS: The annual number of urine E. coli isolates tested for antimicrobial susceptibility decreased 23.3% during 2018-2022 whereas the number of blood E. coli isolates increased 1.1%. The annual proportion of ESBL-producing E. coli in urine E. coli isolates decreased 28.7% among males, from 6.9% (average during 2018-2019) to 4.9% in 2022, and 28.7% among females, from 3.0 to 2.1%. In blood E. coli isolates, the proportion decreased 32.9% among males, from 9.3 to 6.2%, and 26.6% among females, from 6.2 to 4.6%. A significant decreasing trend was also observed in most age groups, but risk remained highest among persons aged ≥ 60 years. CONCLUSIONS: The reduction in the proportions of ESBL-producing E. coli was comprehensive, covering both specimen types, both sexes, and all age groups, showing that the continuously increasing trends could be reversed. Decrease in international travel and antimicrobial use were likely behind this reduction, suggesting that informing travellers about the risk of multidrug-resistant bacteria, hygiene measures, and appropriate antimicrobial use is crucial in prevention. Evaluation of infection control measures in healthcare settings could be beneficial, especially in long-term care.

Screening of antimicrobial activity of Ilex paraguariensis St. Hil. leaf extracts against carbapenemase-producing bacteria.

I. paraguariensis St. Hil. is a south American species of agronomic interest with studies supporting its medicinal properties. As the investigation of active ingredients with antimicrobial effect from medicinal plants is a suitable approach to the current antibacterial resistance problem, the aim of the present study was to determine the antibacterial activity of yerba mate ethanolic extracts against carbapenemase-producing gram-negative bacteria (reference strains and clinical isolates). Extracts showed antibacterial activity against Klebsiella pneumoniae ATCC® BAA-2342™ (KPC producing), Providencia rettgeri (NDM producing), Pseudomonas aeruginosa (MBL producing) and P. aeruginosa (VIM producing) at the concentrations tested. The Minimal-Inhibitory-Concentration and Minimal-Bactericidal-Concentration values ranged between 1 and 32 mg.ml-1 for the reference strains, and between 0.125 and 1 mg.ml-1 for the clinical isolates. The MBC/MIC index characterized the extracts as bactericidal. The combinations of commercial antibiotics and extracts showed a synergistic action on the reference strains studied. The lethal concentration 50 obtained using the Artemia salina toxicity assay were higher than 1 mg.ml-1 for all the extracts, indicating a low toxicity. The in vitro activity and low toxicity suggest that ethanolic I. paraguariensis leaf extracts constitute an outstanding source for new antibacterial compounds, and further studies should be carried out to understand their mechanism of action.

Prevalence of AmpC beta-lactamase and extended spectrum beta-lactamase co-producer in Escherichia coli and Klebsiella species in a teaching hospital.

INTRODUCTION: Beta-lactamase producing bacterial infection has been on surge due to selection pressure and injudicious antibiotics usage. Organisms that co-produced more than one beta lactamase enzyme posed diagnostic challenges which may result in inadequate treatment. To date, there is no standardised guideline offering phenotypic detection of AmpC ß-lactamase. The purpose of this study was to determine the prevalence of ESBLs, AmpC ß-lactamase and co-producer organisms in a teaching hospital. MATERIALS AND METHODS: Three hundred and four isolates of E. coli and Klebsiella sp. had been selected via convenient sampling. These isolates were identified using conventional laboratory methods and their antimicrobial susceptibilities were determined using disc diffusion method. Those isolates were then proceeded with ESBL confirmatory test, cloxacillin-containing Muller Hinton confirmatory test, modified double disk synergy test and AmpC disk test. RESULTS: Out of 304 isolates, 159 isolates were E. coli and 145 were Klebsiella sp. The prevalence of organisms which co-produced AmpC ß-lactamase and ESBL enzymes were 3.0%. Besides that, 39 cefoxitin resistant and three cefoxitin susceptible isolates (13.8%) were proven to produce AmpC ß-lactamase through AmpC disk test. Through the CLSI confirmatory test, 252 (82.9%) isolates were identified as ESBLs producers and the prevalence increased slightly when cloxacillin-containing Muller Hinton were used. Only three ESBLs positive organisms were positive for modified double disk synergy test. CONCLUSION: Distinguishing between AmpC ß-lactamase and ESBL-producing organisms has epidemiological significance as well as therapeutic importance. Moreover, AmpC ß-lactamase and ESBLs co-producing organisms can lead to false negative ESBL confirmatory test. Therefore, knowing the local prevalence can guide the clinician in navigating the treatment.

Uso de ceftazidima-avibactam en una recién nacida prematura con infección urinaria bacteriémica por Klebsiella pneumoniae productora de carbapenemasa / Use of ceftazidime-avibactam in a preterm infant with urinary tract infection with bacteremia caused by carbapenemase-producing Klebsiella pneumoniae

Los recién nacidos tienen un alto riesgo de morbimortalidad asociada a infecciones durante su estancia en unidades de cuidado intensivo neonatal, a lo que se asocia un aumento progresivo de infecciones por microorganismos multi-resistentes que requiere el uso de nuevos antimicrobianos. Presentamos el caso de una recién nacida de pretérmino de 36 semanas que cursó con una infección del tracto urinario bacteriémica por Klebsiella pneumoniae productora de carbapenemasa tratada de forma efectiva con 14 días de cefazi- dima-avibactam, sin efectos adversos observados. Según nuestro conocimiento, este es el primer caso reportado en nuestro país del uso de este antimicrobiano en población neonatal. Se necesita más información sobre la eficacia y seguridad de ceftazidima-avibactam en este grupo de pacientes.

Neonates are high risk patients regarding morbimortality secondary to infections during their neonatal intensive care unit stay, which is associated to a progressive increase in the report of multidrug resistant organism infections, that require the use of new antimicrobial. We report the case of a 36-week preterm with an urinary tract infection with bacteriemia caused by carbapenemase- producing Klebsiella pneumoniae treated effectively with 14 day of ceftazidime-avibactam, without observed adverse effects. To our knowledge, this is the first case report in our country of the use of this antibiotic in neonatal population. More information is needed regarding efficacy and safety of ceftazidime-avibactam in this group of patients.

Plasmid-mediated colistin resistance and ESBL production in Escherichia coli from clinically healthy and sick pigs.

This study aimed to determine the percentage of colistin resistant and ESBL-producing Escherichia coli from clinically sick and healthy pigs and understand the molecular mechanisms underlying colistin resistance and ESBL production. A total of 454 E. coli isolates from healthy pigs (n = 354; piglets, n = 83; fattening pigs, n = 142 and sows, n = 100) and sick pigs (n = 100) were examined for antimicrobial susceptibility, chromosomal and plasmid-mediated colistin resistance mechanisms and ESBL genes. The healthy (41%) and sick pig (73%) isolates were commonly resistant to colistin. Three mcr genes including mcr-1 (10.4%), mcr-2 (1.1%) and mcr-3 (45%) were detected, of which mcr-3 was most frequently detected in the healthy (33%) and sick pig (57%) isolates. Coexistence of mcr-1/mcr-3 and mcr-2/mcr-3 was observed in piglets (23%), fattening pig (3.5%) and sick pig (13%) isolates. Three amino acid substitutions including E106A and G144S in PmrA and V161G in PmrB were observed only in colistin-resistant isolates carrying mcr-3. The percentage of ESBL-producing E. coli was significantly higher in the sick pigs (44%) than the healthy pigs (19.2%) (P = 0.00). The blaCTX-M group was most prevalent (98.5%), of which blaCTX-M-14 (54.5%) and blaCTX-M-55 (42.9%) were predominant. The blaTEM-1 (68.8%) and blaCMY-2 (6.3%) genes were identified in ESBL-producers. All ESBL producers were multidrug resistant and the majority from piglets (97%), fattening pigs (77.3%) and sick pigs (82%) carried mcr gene (s). ESBL producers from piglets (n = 5) and sick pig (n = 1) simultaneously transferred blaTEM-1 (or blaCTX-M-55) and mcr-3 to Salmonella. In conclusion, pigs are important reservoirs of colistin-resistant E. coli that also produced ESBLs, highlighting the need for prudent and effective use of antimicrobials in pigs and other food-producing animals.

Detection of beta-lactamase-producing Enterobacteriaceae in a veterinary hospital environment / Detecção de Enterobactérias produtoras de beta-lactamases em ambiente hospitalar veterinário

Due to the strong selective pressure resulting from the misuse of antibiotics, the natural process of bacterial resistance has been accelerated, leading to the increasingly constant appearance of multiresistant isolates. The high number of multi-resistant bacteria is a one health problem. Enterobacteriaceae are usually commensal bacteria of the gastrointestinal tract. However, they can cause infections, and the most important resistance characteristic among them is the production of ß-lactamases. This study aimed to identify ESBL-producing Enterobacteriaceae of types of TEM, SHV, and the CTX-Mgroups. To isolate the enterobacteria, swabs were collected by swiping objects that had contact with the patients and professionals, and the water of the hospital environment. Ten collections were carried out, yielding 306 samples, from which 118 enterobacteria were identified: Escherichia coli, Enterobacter spp., Klebsiella spp., Proteus mirabilis, Serratiaspp., and Citrobacter spp. Isolates. The genes TEM and CTX-M, for the production of ß-lactamases, were detected in 12.7% of the 118 enterobacterial isolates. It is very important to know the bacterial population circulating in the veterinary hospital environment and its resistance to antimicrobials so that professionals can take appropriate measures to minimize the risks of transmission, especially from cages and consultation tables. In addition, the correct control of the microbiological quality of the supply water, as well as environmental cleaning procedures, are essential to prevent the transmission of these microorganisms.(AU)

Devido à grande pressão seletiva decorrente do uso indevido de antibióticos, tem se acelerado o processo natural de resistência das bactérias, levando ao aparecimento cada vez mais constante de isolados multirresistentes. O elevado número de bactérias multirresistentes identificadas é um problema da saúde única. As enterobactérias são bactérias geralmente comensais do trato gastrointestinal, entretanto podem causar infecções, e a característica de resistência mais importante entre elas é a produção de ß-lactamases. Buscando caracterizar melhor os microrganismos circulantes e potencialmente causadores de infecções em ambiente hospitalar veterinário, este estudo objetivou identificar as enterobactérias produtoras de ESBL do tipo TEM, SHV e os cinco grupos de CTX-M presentes em isolados circulantes em hospital veterinário. Foi realizada coleta de suabes de arrasto de objetos que entram em contato com os pacientes e com os profissionais que ali trabalham, bem como de água, para a identificação das enterobactérias. Foram realizadas 10 coletas, obtendo-se 306 amostras, dessas, 118 enterobactérias foram identificadas: Escherichia coli, Enterobacter, Klebsiella, Proteus mirabilis, Serratia e Citrobacter. Dentre as enterobactérias identificadas, alguns isolados possuíam genes para a produção de ß-lactamases, do tipo TEM e CTX-M. É de grande importância conhecer a população bacteriana circulante no ambiente hospitalar veterinário, e a sua resistência aos antimicrobianos, para que os profissionais possam tomar medidas apropriadas para minimizar os riscos de transmissão, principalmente a partir de gaiolas e mesas de atendimento. Além disso, o correto controle da qualidade microbiológica da água de abastecimento, bem como dos procedimentos de higienização do ambiente, são fundamentais para evitar a transmissão destes microrganismos.(AU)

Replicative transposition contributes to the evolution and dissemination of KPC-2-producing plasmid in Enterobacterales.

ABSTRACTKlebsiella pneumoniae carbapenemase (KPC)-producing Enterobacterales are prevalent worldwide and pose an alarming threat to public health. The incidence and transmission of blaKPC-2 gene via horizontal gene transfer (e.g. transposition) have been well documented. However, the dynamics of transposon structure bearing blaKPC-2 and their exact effects on the evolution and dissemination of blaKPC-2 gene are not well characterized. Here, we collected all 161 carbapenem-resistant Enterobacterales (CRE) isolates during the early stage of CRE pandemic. We observed that the prevalence of KPC-2-producing Enterobacterales was mediated by multiple species and sequence types (STs), and that blaKPC-2 gene was located on three diverse variants of Tn1721 in multi-drug resistance (MDR) region of plasmid. Notably, the outbreak of KPC-2-producing plasmid is correlated with the dynamics of transposon structure. Furthermore, we experimentally demonstrated that replicative transposition of Tn1721 and IS26 promotes horizontal transfer of blaKPC-2 and the evolution of KPC-2-producing plasmid. The Tn1721 variants appearing concurrently with the peak of an epidemic (A2- and B-type) showed higher transposition frequencies and a certain superior ability to propagation. Overall, our work suggests replicative transposition contributes to the evolution and transmission of KPC-2-producing plasmid and highlights its important role in the inter- and intra-species dissemination of blaKPC-2 gene in Enterobacterales.

Rapid Detection of Carbapenemase-producing Enterobacteriaceae From Blood Culture Bottles of Known CPE Carriers: Real-world Experience.

We used a rapid antigen test for the detection of carbapenemases directly from positive blood culture bottles of pediatric hemato-oncologic patients, known carriers of carbapenemase-producing enterobacteriaceae. Resistance mechanism was detected within 15 minutes of observing Gram-negative bacilli from a positive bottle, leading to treatment modification. This simple-to-use, inexpensive assay shortens the interval between empiric to tailored antimicrobial therapy.

A hospital-wide outbreak of IMI-17-producing Enterobacter ludwigii in an Israeli hospital.

ABSRACT: BACKGROUND: To describe the course and intervention of an hospital-wide IMI-Producing Enterobacter ludwigii outbreak. METHODS: This was an outbreak interventional study, done at a tertiary care center in Tel-Aviv, Israel. Data was collected on the course of the outbreak and the demographic and clinical characteristics of all patients involved in the outbreak. The intervention measures included patients' cohorting, contact isolation precautions, environmental cleaning and screening of contacts. The molecular features and phylogeny of outbreak-related isolates were studied by whole-genome based analysis. RESULTS: The outbreak included 34 patients that were colonized by IMI-Producing E. ludwigii and were identified in 24 wards throughout the hospital. Colonization was identified in the first 72 h of admission in 13/34 patients (38.2%). Most patients (91.2%) were admitted from home and had relatively low level of comorbidities. The majority of them (88%) had no recent use of invasive catheters and none had previous carriage of other multi-drug resistant bacteria. All available isolates harbored the blaIMI-17 allele and belonged to Sequence-Type 385. With the exception of two isolates, all isolates were closely related with less than a 20-SNP difference between them. CONCLUSIONS: This outbreak had most likely originated in the community and subsequently disseminated inside our institution. More studies are required in order to elucidate the epidemiology of IMI-Producing E. ludwigii and the possible role of environmental sources in its dissemination.

National surveillance pilot study unveils a multicenter, clonal outbreak of VIM-2-producing Pseudomonas aeruginosa ST111 in the Netherlands between 2015 and 2017.

Verona Integron-encoded Metallo-beta-lactamase (VIM) is the most frequently-encountered carbapenemase in the healthcare-related pathogen Pseudomonas aeruginosa. In the Netherlands, a low-endemic country for antibiotic-resistant bacteria, no national surveillance data on the prevalence of carbapenemase-producing P. aeruginosa (CPPA) was available. Therefore, in 2016, a national surveillance pilot study was initiated to investigate the occurrence, molecular epidemiology, genetic characterization, and resistomes of CPPA among P. aeruginosa isolates submitted by medical microbiology laboratories (MMLs) throughout the country. From 1221 isolates included in the study, 124 (10%) produced carbapenemase (CIM-positive); of these, the majority (95, 77%) were positive for the blaVIM gene using PCR. Sequencing was performed on 112 CIM-positive and 56 CIM-negative isolates (n = 168), and genetic clustering revealed that 75/168 (45%) isolates were highly similar. This genetic cluster, designated Group 1, comprised isolates that belonged to high-risk sequence type ST111/serotype O12, had similar resistomes, and all but two carried the blaVIM-2 allele on an identical class 1 integron. Additionally, Group 1 isolates originated from around the country (i.e. seven provinces) and from multiple MMLs. In conclusion, the Netherlands had experienced a nationwide, inter-institutional, clonal outbreak of VIM-2-producing P. aeruginosa for at least three years, which this pilot study was crucial in identifying. A structured, national surveillance program is strongly advised to monitor the spread of Group 1 CPPA, to identify emerging clones/carbapenemase genes, and to detect transmission in and especially between hospitals in order to control current and future outbreaks.

Risk factors for colonization with multiple species of extended-spectrum beta-lactamase producing Enterobacterales: a case-case-control study.

BACKGROUND: Approximately 11% of patients colonized with extended-spectrum beta-lactamase producing Enterobacterales (ESBL-PE) are colonized with more than one ESBL-producing species. We investigated risk factors associated with colonization with multiple ESBL-PE species. METHODS: We performed a case-case-control study at the University Hospital Basel, Switzerland, including hospitalized patients colonized with ESBL-PE between 01/2008 and 12/2018. Patients colonized with multiple species of ESBL-PE during the same hospitalization were assigned to group 1. Group 2 consisted of patients with ESBL-PE and a newly acquired ESBL-PE-species identified during subsequent hospitalization. Controls (i.e., group 3) were patients with only one species of ESBL-PE identified over multiple hospitalizations. Controls were frequency-matched 3:1 to group 2 cases according to time-at-risk (i.e., days between ESBL-PE detection during first and subsequent hospitalizations) to standardize the duration of colonization. ESBL was identified with phenotypic assay and the presence of ESBL genes was confirmed by whole genome sequencing. RESULTS: Among 1559 inpatients, 154 cases met eligibility criteria (67 in group 1, 22 in group 2, 65 in group 3). International travel within the previous 12 months (OR 12.57, 95% CI 3.48-45.45, p < 0.001) and antibiotic exposure within the previous 3 months (OR 2.96, 95% CI 1.37-6.41, p = 0.006) were independently associated with co-colonization with multiple ESBL-PE species. Admission from another acute-care facility was the only predictor of replacement of one ESBL-PE species with another during subsequent hospitalizations (OR 6.02, 95% CI 1.15-31.49, p = 0.003). CONCLUSION: These findings point to strain-related factors being the main drivers of co-colonization with different ESBL-PE and may support stratification of infection prevention and control measures according to ESBL-PE species/strains.

Long-term Follow-up of Preterm Infants Having Been Colonized With Extended Spectrum Beta-lactamase-producing Enterobacterales Over the First 6 Years of Life.

We performed a retrospective case-control cohort study following 146 preterm infants (≤32 weeks of gestation) who had been colonized with extended spectrum beta-lactamase producing Enterobacterales and compared them with 1:1 matched controls regarding rates of hospitalizations and outpatient visits because of infectious and gastrointestinal diseases and developmental impairment up to school age. Preterm infants with extended spectrum beta-lactamase producing Enterobacterales colonization did have neither higher rates of gastrointestinal or infectious diseases nor higher rates of developmental impairments up to the age of 6 years.

Comparison of outcomes in urinary tract infections caused by AmpC-harboring organisms treated with AmpC stable versus AmpC susceptible agents.

There is minimal data on the optimal treatment of lower inoculum infections such as urinary tract infections (UTIs) caused by SPICE organisms which encode the betalactamase enzyme, AmpC. This single-center, retrospective review of adult hospitalized patients with UTIs caused by a SPICE organism compared outcomes amongst patients treated with drugs susceptible to AmpC hydrolysis versus drugs stable against AmpC. Of 156 patients, similar rates of clinical response, 30-day infection related readmission, 30-day infection recurrence, 30-day mortality rates, and median length of hospital stay were found between the two groups. Notably, 44% of patients with ceftriaxone resistance reported had recent ß-lactam exposure versus only 11% of patients without ceftriaxone resistance (P = 0.002). Based on our data, there does not appear to be a difference in clinical response or any of the secondary outcomes in patients with UTIs treated with AmpC stable and AmpC susceptible agents.

Presence of ß-Lactamase-producing Enterobacterales and Salmonella Isolates in Marine Mammals.

Marine mammals have been described as sentinels of the health of marine ecosystems. Therefore, the aim of this study was to investigate (i) the presence of extended-spectrum ß-lactamase (ESBL)- and AmpC-producing Enterobacterales, which comprise several bacterial families important to the healthcare sector, as well as (ii) the presence of Salmonella in these coastal animals. The antimicrobial resistance pheno- and genotypes, as well as biocide susceptibility of Enterobacterales isolated from stranded marine mammals, were determined prior to their rehabilitation. All E. coli isolates (n = 27) were screened for virulence genes via DNA-based microarray, and twelve selected E. coli isolates were analyzed by whole-genome sequencing. Seventy-one percent of the Enterobacterales isolates exhibited a multidrug-resistant (MDR) pheno- and genotype. The gene blaCMY (n = 51) was the predominant ß-lactamase gene. In addition, blaTEM-1 (n = 38), blaSHV-33 (n = 8), blaCTX-M-15 (n = 7), blaOXA-1 (n = 7), blaSHV-11 (n = 3), and blaDHA-1 (n = 2) were detected. The most prevalent non-ß-lactamase genes were sul2 (n = 38), strA (n = 34), strB (n = 34), and tet(A) (n = 34). Escherichia coli isolates belonging to the pandemic sequence types (STs) ST38, ST167, and ST648 were identified. Among Salmonella isolates (n = 18), S. Havana was the most prevalent serotype. The present study revealed a high prevalence of MDR bacteria and the presence of pandemic high-risk clones, both of which are indicators of anthropogenic antimicrobial pollution, in marine mammals.

A rabbit monoclonal antibody-mediated lateral flow immunoassay for rapid detection of CTX-M extended-spectrum ß-lactamase-producing Enterobacterales.

Infections of CTX-M extended-spectrum ß-lactamase-producing Enterobacterales are a severe threat in clinical settings. CTX-M genes on plasmids have been transferred to many Enterobacterales species, and these species have spread, leading to the global problem of antimicrobial resistance. Here, we developed a lateral flow immunoassay (LFIA) based on an anti-CTX-M rabbit monoclonal antibody. This antibody detected CTX-M variants from the CTX-M-9, CTX-M-2, and CTX-M-1 groups expressed in clinical isolates. The LFIA showed 100% sensitivity and specificity with clinical isolates on agar plates, and its limit of detection was 0.8 ng/mL recombinant CTX-M-14. The rabbit monoclonal antibody did not cross-react with bacteria producing other class A ß-lactamases, including SHV. In conclusion, we developed a highly sensitive and specific LFIA capable of detecting CTX-M enzyme production in Enterobacterales. We anticipate that our LFIA will become a point-of-care test enabling rapid detection of CTX-M in hospital and community settings as well as a rapid environmental test.

Establishing a quantitative index of meropenem hydrolysis for the detection of KPC- and NDM-producing bacteria by MALDI-TOF MS.

BACKGROUND: Matrix Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry (MALDI-TOF MS), commonly used for microorganism identification, can also be applied for the detection of carbapenemase-producing bacteria by the evaluation of carbapenem hydrolysis. Since KPC- and NDM-producing bacteria are related to high mortality rates, diagnostic assays for its detection are essential. The aim of this study was to develop and evaluate a method to establish a quantitative measure (hydrolysis index - HI) to detect meropenem hydrolysis by MLADI-TOF MS. METHODS: blaKPC and blaNDM positive and negative Klebsiella pneumoniae isolates and Escherichia coli ATCC 25922 (control) were incubated in a meropenem solution for 2 h. Protein extraction from these suspensions were submitted to MALDI-TOF MS analysis. The intensity of peaks at 384 m/z and 379 m/z of each isolate were used to establish the HI as follows: HI = (Peak intensity384 Test / Peak intensity379 Test) / (Peak intensity384 Control / Peak intensity379 Control). Receiver Operating Characteristic curve was used to determine a cutoff value to differentiate carbapenemase-producing from carbapenemase non-producing bacteria. RESULTS: As all carbapenemase-producing K. pneumoniae presented HI ≤0.55 and all carbapenemase non-producing isolates presented a HI ≥0.57, the index of 0.56 was established as a cutoff value to differentiate carbapenemase (KPC and NDM) producing and non-producing bacteria.

Impact of antibiotic usage on extended-spectrum ß-lactamase producing Escherichia coli prevalence.

An increase in antibiotic usage is considered to contribute to the emergence of antimicrobial resistance. Although experts are counting on the antimicrobial stewardship programs to reduce antibiotic usage, their effect remains uncertain. In this study, we aimed to evaluate the impact of antibiotic usage and forecast the prevalence of hospital-acquired extended spectrum ß-lactamase (ESBL)-producing Escherichia coli (E. coli) using time-series analysis. Antimicrobial culture information of E. coli was obtained using a text processing technique that helped extract free-text electronic health records from standardized data. The antimicrobial use density (AUD) of antibiotics of interest was used to estimate the quarterly antibiotic usage. Transfer function model was applied to forecast relationship between antibiotic usage and ESBL-producing E. coli. Of the 1938 hospital-acquired isolates, 831 isolates (42.9%) were ESBL-producing E. coli. Both the proportion of ESBL-producing E. coli and AUD increased over time. The transfer model predicted that ciprofloxacin AUD is related to the proportion of ESBL-producing E. coli two quarters later. In conclusion, excessive use of antibiotics was shown to affect the prevalence of resistant organisms in the future. Therefore, the control of antibiotics with antimicrobial stewardship programs should be considered to restrict antimicrobial resistance.

Antisense peptide nucleic acid inhibits the growth of KPC-producing Klebsiella pneumoniae strain.

Klebsiella pneumoniae causes common and severe hospital- and community-acquired infections with a high incidence of multidrug resistance (MDR) and mortality. In this study, we investigated the ability of the antisense peptide nucleic acids (PNA) conjugated to the (KFF)3K cell-penetrating peptide (CPP) to target the gyrA KPC-producing K. pneumoniae and inhibit bacterial growth in vitro. The inhibitory effect on gyrA gene was evaluated by measuring 16s gene amplification in KPC-producing K. pneumoniae treated with the antisense PNA conjugate. The hemolytic property of the antisense PNA conjugate was accessed toward mice red blood cells. Finally, molecular modeling and dynamics simulations analyses in aqueous solutions were performed to predict the PNA conformation alone in contact with DNA (gyrA gene sequence). PNA was capable of inhibiting bacterial growth at 50 µM, also reducing 16S gene amplification in 96.7%. Besides, PNA presented low hemolytic activity (21.1% hemolysis) at this same concentration. Bioinformatics analysis demonstrated that the structure of the PNA is stable in water without major changes in its secondary structure. The ability of PNA and its conjugated CPP ((KFF)3K) to inhibit bacterial growth demonstrates the potential of this new class of antibacterial agents, encouraging further in vivo studies to confirm its therapeutic efficacy.

Selection of ESBL-producing Escherichia coli in the gut of calves experimentally fed with milk containing antibiotic residues.

In the bovine sector, the spread of Enterobacterales producing extended-spectrum and AmpC ß-lactamases (ESBL/AmpC) mostly concerns veal calves, and the use of waste milk containing antibiotic residues has been recurrently incriminated. In this study, calves were experimentally fed with milk containing either 2,000 µg/L or 20,000 µg/L of the critically important antibiotic cefquinome. The total counts of enterobacterales and ESBL-producing E. coli were monitored using non-selective and selective media. Our data highlighted the important combination of two main factors (cefquinome exposure and initial ESBL colonization level) in the ESBL selection and amplification process in the gut of calves. Results also proved the dose-independent effect of cefquinome administration on the selection and amplification of ESBL-producing E. coli. Finally, the blaCTX-M-1/IncI1 ST3 plasmid was systematically recovered after cefquinome exposure, highlighting its epidemic success. Altogether, this work is one of the rare experimental studies providing quantitative information on the impact of waste milk containing antimicrobials on the ESBL load in calves' microbiota, and the first one using cefquinome. These data emphasise the need for global guidelines on the use of waste milk on dairy farms in order to decrease the antimicrobial resistance burden in this sector.

Molecular epidemiology of NDM-5-producing Escherichia coli high-risk clones identified in two Italian hospitals in 2017-2019.

Between November 2018 and October 2019, carbapenem-resistant Enterobacterales carrying New Delhi Metallo-ß-lactamase (NDM) caused one of the largest and persistent outbreaks occurred in Italy and intensified surveillance measures have been taken in all Italian hospitals. In this study we analyzed NDM-5- producing Escherichia coli identified in 2 hospitals of the Lazio region in Italy. Epidemiological and microbiological data demonstrated that in 2018-2019 the NDM-5-producing high-risk E. coli ST167 clone circulated in patients from both hospitals. In 2019, another NDM-5-producing E. coli clone, identified by MLST as ST617 was introduced in one of the 2 hospitals and caused an outbreak. This study describes an application of genomics as a useful method to discern endemic and outbreak clones when applied to strains of the same species (E. coli) with the same resistance determinant (NDM-5) and the relevance of screening patients admitted in critical units for carbapenemase producers to prevent outbreaks.