Types Of Carbapenemases Research Articles

The Predominance of Klebsiella aerogenes among Carbapenem-Resistant Enterobacteriaceae Infections in Japan.

The emergence of carbapenem-resistant Enterobacteriaceae (CRE) is an important public health issue worldwide, not only due to the potential of these pathogens for widespread dissemination, but also due to the limited antimicrobial therapy options, and the elevated mortality rates associated with these infections. As with other multidrug-resistant organisms (MDROs), active surveillance via timely testing, early diagnosis, and contact isolation is an important strategy to control the occurrence and spread of CRE bacteria. Here we summarize the epidemiology of CRE infections in Japan from 2015 to 2019. Data were extracted from a public dataset collected by the nationwide surveillance system via the National Institute of Infectious Diseases (NIID) of Japan. The annual number of reported CRE infections has remained relatively stable, with a tendency to increase in the last two years (1671 cases reported in 2015 and 2333 cases reported in 2019). The majority of patients who presented CRE infections over this five year period were older than 65 years (~80%, mean age 75), 60% of them were men, and mortality rates were around 3.5%. Importantly, about 60% of infections are caused by both Enterobacter cloacae and Klebsiella aerogenes (previously known as Enterobacter aerogenes), the former being the most common pathogen in 2015 and 2016 (~30%), and the latter the leading pathogen since 2017 (~40%). The most common carbapenemase isolated was the IMP carbapenemase type. Further studies are needed to determine the prevalence of CRE colonization, especially in the healthcare setting, and to elucidate the mechanisms behind the local predominance of Klebsiella aerogenes and Enterobacter cloacae.

Carbapenemase investigation with rapid phenotypic test (RESIST-4 O.K.N.V) and comparison with PCR in carbapenem-resistant Enterobacterales strains.

RESİST-4 O.K.N.V. assay is a lateral immunochromatocraphic test for the identification of oxacillinase (OXA)-48-like, Klebsiella pneumoniae carbapenemase (KPC), New Delhi metallo-β-lactamase (NDM), and Verona integron-encoded metallo-β-lactamase (VIM) producing strains. It was aimed to evaluate the performance of the RESIST-4 O.K.N.V. test and to compare it with the reference method polymerase chain reaction (PCR). Also, the objective was to determine the distribution of carbapenemase types of CRE strains isolated in our hospital. Between January 2016-October 2019, 187 strains isolated from clinical samples were included in this study. Bacterial identification was done using MALDI-TOF MS. Antibiotic susceptibility tests were studied with the VITEK-2 automated system. Meropenem minimum inhibitory concentrations (MICs) were determined by the gradient test. All strains were studied with the RESIST-4 O.K.N.V. test and then the strains were selected for the PCR test. bla OXA-48, bla NDM, bla KPC, and bla VIM were investigated with PCR. K. pneumoniae NCTC® 13438 (KWIKSTIKTM, Microbiologics®, USA) was used as the positive control, E. coli ATTC® 25922 TM (Microbiologics®, USA) and three carbapenem-sensitive clinical isolates were also used as the negative control. Meropenem MIC50 and MIC90 values were determined to be >32 mg/L. With PCR bla OXA-48, bla NDM, bla KPC and bla VIM were detected in 79, 63, 20, and 4 strains, respectively. bla OXA-48 and bla NDM were found together in 51 of the isolates. bla OXA-48, bla NDM, bla KPC, and bla VIM were not detected in two strains with carbapenem resistance in susceptibility tests. The sensitivity of the immunochromatographic test was 100% for OXA-48, KPC, and VIM but 84.1% for NDM. Specificity was determined as 100% for OXA-48, NDM, KPC, and VIM. RESIST-4 O.K.N.V. test showed high sensitivity and specificity in detecting OXA-48, KPC, NDM, and VIM type carbapenemases. However, it should be kept in mind that there may be false-negative results related to NDM.

Association Between Types of Carbapenemase and Clinical Outcomes of Infection Due to Carbapenem Resistance Enterobacterales.

PurposeCompared with non-carbapenemase producing carbapenem-resistant Enterobacterales (non-CP-CRE), carbapenemase-producing carbapenem-resistant Enterobacterales (CP-CRE) are associated with considerable mortality. However, given that the patients are treated with various therapeutic options, it remains unclear whether differences in types of carbapenemase genes yield different mortality rates. Therefore, this study aims to identify carbapenemase genes and identify whether clinical outcomes differ according to the prevalence of genotype and phenotype of carbapenemase among Enterobacterales clinical isolated.Patients and MethodsA retrospective cohort study was performed to determine whether types of carbapenemase genes have an impact on clinical outcomes. Carbapenem-resistant clinical isolates were collected at a tertiary care university hospital in Songkhla, Thailand, between June 2018 and February 2020. Demographic and microbiological data such as antimicrobial susceptibility, carbapenemase genes, and overall mortality were evaluated.ResultsA total of 121 Enterobacterales clinical isolated were evaluated. The blaNDM-1 gene was detected in 44% of the isolates, followed by blaOXA-48 (28%) and blaNDM-1/OXA-48 (28%). NDM-1- or NDM-1/OXA-48- producing isolates were more likely to require meropenem MICs of ≥16 mg/L, while OXA-48-producing isolates were more likely to require meropenem MICs of <16 mg/L. The patients with NDM-1 or NDM-1/OXA-48 had a higher 14 days mortality rate than those with OXA-48 after treating with carbapenem-containing regimens (P-value 0.001) or colistin-containing regimens (P-value < 0.001).ConclusionOur findings suggest that the mortality for CP-CRE infection in patients with NDM-1 or NDM-1/OXA-48 was higher than the mortality in those with OXA-48, which It seems that the type of carbapenemase gene may affect meropenem MIC levels. Hence, in treatment decisions involving the use of either carbapenem-containing regiment or colistin-containing regiment in patients with CP-CRE infection, especially those in the NDM-1 and NDM-1/OXA-48 groups, the patient symptoms should be closely monitored.

In vitro activity of sulbactam-durlobactam against carbapenem-resistant Acinetobacter baumannii and mechanisms of resistance

Multidrug-resistant Acinetobacter baumannii (MDR-Ab), particularly strains producing oxacillinase (OXA)-type carbapenemases, have rapidly emerged in health care settings as a frequent cause of serious infections with limited treatment options. This study evaluated the in vitro activity of sulbactam (SUL) combined with durlobactam (DUR) against a collection of carbapenemase-producing A. baumannii, and investigated the mechanisms of resistance. Susceptibility testing was performed on 100 isolates by either broth microdilution or by the Epsilometer test. Isolates were screened for the insertion sequence ISAba1 upstream of the intrinsic chromosomal blaADC by polymerase chain reaction (PCR). Whole genome sequencing was performed on 25 SUL-DUR resistant isolates, and analyses were performed using the Center for Genomic Epidemiology platform. Target gene sequences were compared to A. baumannii American Type Culture Collection (ATCC) 17978. SUL-DUR exhibited excellent activity against A. baumannii isolates with susceptibility levels as follows: amikacin, 18%; colistin, 91%; cefepime, 5%; imipenem, 0%; minocycline, 46%; SUL, 3%; sulbactam-cefoperazone, 8%; SUL-DUR, 71% (based on a breakpoint at 4 mg/L). Twenty-five non-New Delhi metallo-ß-lactamase (NDM)-producing isolates had SUL-DUR MIC values >4 mg/L, amongst which 14 isolates showed substitutions in penicillin-binding protein (PBP)3, previously shown to be associated with SUL-DUR resistance. Substitutions that have not previously been described were detected in SUL-DUR targets, namely PBP1a, PBP1b, PBP2, and PBP3. By contrast, there was no evidence of the involvement of permeability or efflux. SUL-DUR exhibited excellent in vitro antibacterial activity against carbapenemase-producing A. baumannii isolates. Amongst the 25 resistant isolates, we identified a number of mechanisms which may be contributing factors, in particular PBP substitutions and the production of specific beta-lactamases.

Carbapenem-resistant Enterobacterales and Pseudomonas aeruginosa causing infection in Africa and the Middle East: a surveillance study from the ATLAS programme (2018–20)

ObjectivesTo determine the in vitro susceptibility of Enterobacterales (n = 5457) and Pseudomonas aeruginosa (n = 1949) isolated from hospitalized patients in Africa (three countries) and the Middle East (five countries) in 2018–20 to a panel of 11 antimicrobials and to identify β-lactamase/carbapenemase genes in isolates with meropenem-non-susceptible and/or ceftazidime/avibactam-resistant phenotypes.MethodsCLSI broth microdilution testing generated MICs that were interpreted using CLSI (2021) breakpoints. β-Lactamase/carbapenemase genes were identified using multiplex PCR assays.ResultsEnterobacterales isolates were highly susceptible to amikacin (96.7%), ceftazidime/avibactam (96.6%) and tigecycline (96.0%), and slightly less susceptible to meropenem (94.3%). In total, 337 Enterobacterales isolates (6.2% of all Enterobacterales isolates) carried one or more carbapenemase genes: 188 isolates carried a serine carbapenemase (178 OXA, 10 KPC) and 167 isolates carried an MBL (18 isolates carried both an MBL and an OXA). NDM-1 was the most common MBL identified (64.1% of NDM enzymes; 59.9% of all MBLs). OXA-48 (47.8%) and OXA-181 (38.8%) were the most common OXAs detected. P. aeruginosa isolates were most susceptible to ceftazidime/avibactam (89.1%) and amikacin (88.9%). Only 73.1% of P. aeruginosa isolates were meropenem susceptible. The majority (68.1%) of P. aeruginosa isolates tested for carbapenemase/β-lactamase genes were negative. In total, 88 isolates (4.5% of all P. aeruginosa isolates) carried one or more carbapenemase genes: 81 isolates carried an MBL and 8 carried a GES carbapenemase (1 isolate carried genes for both).ConclusionsCarbapenemase detection was closely associated with meropenem-non-susceptible phenotypes for Enterobacterales (89.1%) but not for P. aeruginosa (24.2%). Wide geographic variation in carbapenemase type and frequency of detection was observed.

Evaluation of In Vitro Efficacy of Ceftazidime-Avibactam, Meropenem, and Colistin Single and Binary Combinations Against Carbapenem Resistant Klebsiella pneumoniae Strains Isolated from Various Clinical Specimens

It has been reported that many antibiotics used today, including the carbapenem group, fail to treat Klebsiella pneumoniae infections effectively. Despite many studies in recent years, the definitive treatment for carbapenem-resistant Klebsiella pneumoniae (CRKP) infections is still uncertain. In this study, it was aimed to investigate in vitro activities of colistin (COL) and meropenem (MEM), which are frequently used in the treatment of CRKP infections, and ceftazidime-avibactam (CZA), which is recently used in our country, alone or in combination against different CRKP isolates having different carbapenem resistance mechanisms andto analyze whether the presence of colistin resistance, which is an important problem in CRKP strains, influences the drug interaction results. This study was carried out in 42 K.pneumoniae isolates, which were isolated from various clinical samples as an infectious agent in Süleyman Demirel University Faculty of Medicine, Department of Medical Microbiology, Bacteriology Laboratory and whose carbapenem resistance was confirmed by carbapenemase inactivation test. The carbapenemase genes of the isolates were determined by the polymerase chain reaction (PCR) method. Antimicrobial susceptibilities of CRKP strains to CZA, MEM, and COL were determined by the broth microdilution method and in vitro synergy activities of dual combinations of these drugs were evaluated by checkerboard and time-kill methods. Statistical evaluation of categorical data was performed using Fisher's exact test, and p-value of less than 0.05 was considered statistically significant in terms of difference between the groups. Of the 42 CRKP isolates 34 (81%) were only OXA-48 positive, 5 (11.9%) were OXA-48+NDM and 3 (7.1%) were OXA-48+KPC positive. In the checkerboard test, synergy was detected against 97.6% of the isolates both with CZA+MEM and CZA+COL combinations, whereas this rate was 50% with MEM+COL. In the time-kill test, synergy was detected with CZA+MEM and CZA+COL combinations in the OXA-48 positive isolate and OXA-48+KPC positive isolate, while synergy was detected with CZA+COL and MEM+COL combinations in the OXA-48+NDM positive isolate. There was no significant relationship between whether the isolates were resistant to colistin or not and the checkerboard test results of antibiotic combinations (pCZA+MEM= 0.33, pCZA+COL= 0.11, pMEM+COL= 0.61). Results of our study revealed that the most common carbapenemase type in CRKP isolates was OXA-48 in our hospital, and the combinations of CZA with MEM and COL had high potential for synergism against these isolates.

The first nationwide surveillance of carbapenem-resistant Enterobacterales in the United Arab Emirates – increased association of Klebsiella pneumoniae CC14 clone with Emirati patients

ObjectivesTo assess the current prevalence, distribution, and main clonal types of carbapenem-resistant Enterobacterales (CRE) in the United Arab Emirates. MethodsA total of 504 CRE collected over a 9-month period in 15 hospitals were studied. Antibiotic susceptibility and the presence of common carbapenemase, 16S methylase, and mobile colistin resistance genes were assessed. Selected strains forming larger clusters by pulsed field gel electrophoresis were subjected to whole genome sequencing to identify their sequence types and core genome MLST. ResultsStrains expressing OXA and NDM type carbapenemases and 16S methylases were present in all major hospitals. Considerable interhospital differences were noticed, suggesting the role of specific clones. A total of three major Klebsiella pneumoniae clones (CC14, ST231, and CC147) were identified, accounting for 48.6% of all CRE. All clones were significantly more resistant than sporadic isolates. CC14 strains exhibited a significant association with Emirati patients. ConclusionsNearly half of CRE infections in the country are due to a limited number of clones. The data indicate the possibility of interhospital transmission, combined in some hospitals with inadequate stewardship practices. The study also revealed an association of the largest, most resistant clone (CC14) with Emirati patients. The specific reasons for it should be clarified by further investigations.

Rapid LC–MS/MS detection of different carbapenemase types in carbapenemase-producing Enterobacterales

Klebsiella pneumoniae carbapenemase (KPC)-2, metallo-beta-lactamases (MBL), and oxacillinase (OXA)-48-like carbapenemases are considered the most important carbapenemases. In vitro studies have demonstrated that the carbapenemase activity of KPC-2 and MBL can be inhibited by 3-aminophenylboronic acid and ethylenediaminetetraacetic acid (EDTA), respectively. Understanding the carbapenemase types expressed in carbapenem-resistant Enterobacterales (CRE) is of great significance to clinical therapies. Liquid chromatography-coupled tandem mass spectrometry (LC-MS/MS) is fast, stable, and specific; and is considered the gold standard method for measuring small molecules. In this study, we developed carbapenemase inhibition tests combined with LC-MS/MS to rapidly identify carbapenemase types. A total of 295 clinical isolates were examined, including 212 KPC-2 producers, 29 MBL producers, 15 OXA-48-like producers, 3 KPC-2 + OXA-232 producers, and 36 carbapenem-sensitive strains. We used LC-MS/MS to determine the carbapenemase types by measuring the ratio of the hydrolyzed meropenem peak areas in the presence and absence of different inhibitors. The sensitivity and specificity of LC-MS/MS in detecting single KPC-2 producers were 97.64% and 100.00%, respectively, and 96.55% and 100.00% for MBL producers, respectively. In addition, the sensitivity and specificity of LC-MS/MS in detecting single OXA-48-like producers were both 100.00% when extending incubation time up to 2.5h. LC-MS/MS showed excellent agreement in detecting carbapenemase types using the modified carbapenem inactivation (mCIM)/EDTA-carbapenem inactivation assay (eCIM) (kappa = 0.93 for serine carbapenemases and kappa = 0.98 for MBL carbapenemases). In this study, LC-MS/MS demonstrated excellent detection of different carbapenemase types, showing potential reliability in future clinical applications.

Investigation on the transmission rate of carbapenemase-producing carbapenem-resistant Enterobacterales among exposed persons in a tertiary hospital using whole-genome sequencing

To reduce transmission of carbapenemase-producing carbapenem-resistant Enterobacterales (CP-CRE), screening is recommended for patients sharing rooms with CP-CRE-detected patients and healthcare workers caring for them. The aim of this study was to investigate the transmission rate of CP-CRE among exposed people in a tertiary hospital using whole-genome sequencing. This study was conducted in a 1751-bed tertiary teaching hospital from January 2017 to December 2019. Index patients were defined as those with positive results in CP-CRE tests during hospitalization. When an index patient was detected in a shared room, we performed CRE screening tests for patients whose stay overlapped with an index patient's stay for at least one day. Where a second case was found, healthcare worker contacts were also screened. CP-CRE were confirmed, and the carbapenemase type identified, by PCR. Whole-genome sequencing was used to compare isolates from index and exposed patients. During the study period, 47 index patients were identified, and they had been in contact with 152 patients in shared rooms and 54 healthcare workers. None of the healthcare workers had CRE. Among the 152 exposed patients, four patients had the same type of carbapenemases as their CP-CRE index patients and all of them were KPC. Whole-genome sequencing revealed that three of these four pairs showed genotypic accordance between the index and the exposed. The CP-CRE transmission rate among the exposed patients was calculated as 2.0% (= 3/152).

Effectiveness of a double-carbapenem combinations against carbapenem-resistant Gram-negative bacteria

The emergence of carbapenem-resistant organisms posed considerable threat to global health while only limited treatment options are available and led to efforts to discover a novel way to treat them. To evaluate in vitro synergistic activity of meropenem plus ertapenem, a total of 203 carbapenem-resistant strains, collected from 12 provinces and municipalities in China, were examined with a dual carbapenem combination therapy. The statistical software R was used for analysis. Two hundred and one (201) of carbapenem-resistant strains mainly produced four types of carbapenemase: KPC-2 (n = 142, 69.95%), OXA-232 (n = 7, 3.45%), NDM (n = 38, 18.72%; 36 NDM-1, 1 NDM-4, 1 NDM-5), and IMP (n = 15, 7.39%; 1 IMP-26, 10 IMP-30, 4 IMP-4). Fifty-one out of two hundred and three (51/203 or 25.12%) of the examined strains showed a synergistic effect for the meropenem plus ertapenem combination throughout the checkerboard method, while only three isolates showed potential clinically relevant synergy (3/203, 1.48%). An additive effect was observed in 55/203 (27.09%) of the examined strains. Ninety-seven of the examined isolates (47.78%) showed fractional inhibitory concentration (FIC) greater or equal to 2 (indicating antagonism). The synergistic activity of meropenem plus ertapenem combination suggests this combination can be a possible way to treat the infection caused by the carbapenem-resistant organisms, especially for IMP or NDM producer with a lesser minimum inhibitory concentration (MIC) and the infected individual who was not recommended to use colistin or tigecycline.

Antimicrobial Activity Profiles and Potential Antimicrobial Regimens against Carbapenem-Resistant Enterobacterales Isolated from Multi-Centers in Western Thailand.

The spread of carbapenem-resistant Enterobacterales (CRE) constitutes a global health burden. Antimicrobial susceptibility and types of carbapenemase differ by geographic region. This study aimed to (1) examine the minimum inhibitory concentrations (MICs) and antibiotic resistance genes and (2) investigate antibiotic dosing regimens against CRE using Monte Carlo simulation. Clinical carbapenem-resistant Klebsiella pneumoniae (CRKP), Escherichia coli (CREC), and Enterobacter cloacae (CREclo) isolates were collected from various hospitals in western Thailand. Broth microdilution was performed, and the types of carbapenemase and mcr-1 genes were detected using polymerase chain reaction (PCR). Monte Carlo simulation was used to establish optimal antimicrobial dosing regimens meeting the criterion of a cumulative fraction of response (CFR) >90%. A total of 150 CRE isolates from 12 hospitals were included. The proportion of CRKP (76%) was greater than that of CREC (22%) and CREclo (2%). Regional hospitals reported higher rates of resistance than general hospitals. Most isolates were resistant to aztreonam and ceftazidime/avibactam, whereas they were highly susceptible to aminoglycosides. Most carbapenemases were NDM (47.33%), OXA-48 (43.33%) and NDM plus OXA-48 (6.67%); five OXA-48 positive isolates carried mcr-1 genes. Currently, high-dose tigecycline is the only optimal regimen against CRE isolates. Further extensive research on antibiotic synergism or new antibiotics should be conducted.

A retrospective observational cohort study of the clinical epidemiology of bloodstream infections due to carbapenem-resistant Klebsiella pneumoniae in an OXA-48 endemic setting

This study aimed to characterize the epidemiology and clinical outcomes of patients with bloodstream infections (BSIs) due to carbapenem-resistant Klebsiella pneumoniae (CRKP) in an OXA-48-predominant environment. This was a retrospective single-centre cohort study including all consecutive patients with CRKP BSIs treated between 1 January 2014 and 31 December 2018. Multivariate analysis, subgroup analysis and propensity-score-matched analysis were employed to analyse 30-day mortality as the primary outcome. Clinical cure at day 14 was also analysed for the whole cohort. In total, 124 patients with unique isolates met all the inclusion criteria. OXA-48 was the most common type of carbapenemase (85.5%). Inappropriate therapy was significantly associated with 30-day mortality [70.6% vs 39.7%, adjusted odds ratio (aOR) 4.65, 95% confidence interval (CI) 1.50-14.40, P=0.008] and 14-day clinical failure (78.5% vs 56.2%, aOR 3.14, 95% CI 1.09-9.02, P=0.033) in multivariate analyses. Among those treated appropriately, the 30-day mortality rates were similar in monotherapy and combination therapy arms (OR 2.85, 95% CI 0.68-11.95, P=0.15). INCREMENT CPE mortality score (aOR 1.16, 95% CI 1.01-1.33, P=0.029), sepsis at BSI onset (aOR 2.90, 95% CI 1.02-8.27, P=0.046), and inappropriate therapy (aOR 4.65, 95% CI 1.50-14.40, P=0.008) were identified as independent risk factors for 30-day mortality. Colistin resistance in CRKP had no significant impact on 30-day mortality. These results were also confirmed in all propensity-score-matched analyses and sensitivity analyses. Appropriate regimens were associated with better clinical outcomes than inappropriate therapies for BSIs with CRKP predominantly possessing OXA-48.

Analysis of carbapenemases genes of carbapenem-resistant Klebsiella pneumoniae isolated from Tehran heart center.

Background and Objectives:Emerging of carbapenem-resistant Klebsiella pneumoniae (CRKP) is one of the major concerns among healthcare systems. This study aimed to investigate the antibiotic susceptibility pattern and carbapenemase genes of carbapenemase-producing K. pneumoniae isolates obtained from Iranian hospitalized patients.Materials and Methods:This study was performed on 71 CRKP strains isolated from different clinical specimens collected in Tehran Heart Center (Tehran, Iran). A Modified Hodge test (MHT) was done for the detection of carbapenemase-producing K. pneumoniae. The presence of blaKPC, blaVIM, blaIMP, blaNDM, and blaOXA-48 -type carbapenemases was evaluated by the PCR method.Results:We identified 8.82% (71/805) of K. pneumoniae isolates as CRKP by MHT test. The antibiotic susceptibility indicated that all isolates were resistant to imipenem, meropenem, cefotaxime, ceftazidime, cefepime, ceftriaxone, cephalothin, ciprofloxacin, and augmentin, and then mostly resistant to aztreonam, cefoxitin, gentamicin, and trimethoprim/sulfamethoxazole with 98.6%, 98.6%, 97.2%, and 94.4%, respectively. The lowest resistance was related to amikacin with 46.5% (33/71 isolates). The level of imipenem MIC for all carbapenem-resistant isolates was estimated ≥32 μg/mL. Among positive isolates for carbapenemase genes, the most frequent gene was blaOXA-48. It was found in 48 (67.6%) isolates followed by blaVIM in 28 (39.4%) isolates. bla IMP, blaNDM, and bla KPC genes were identified in 19 (26.8%), 13 (18.3%) and 5 (7.0%) isolates, respectively. These genes were not detected in nine isolates.Conclusion:The relatively high frequency of some carbapenemase genes suggests major concern about the emergence of isolates containing carbapenem resistance genes as a potential health threat.

Multicenter surveillance of in vitro activities of cefepime-zidebactam, cefepime-enmetazobactam, omadacycline, eravacycline, and comparator antibiotics against Enterobacterales, Pseudomonas aeruginosa, and Acinetobacter baumannii complex causing bloodstream infection in Taiwan, 2020

ABSTRACT Objectives To determine the in vitro activities of novel and comparator antibiotics against Gram-negative bacteria (GNB) in Taiwan. Methods Isolates of Escherichia coli (n = 335), Klebsiella pneumoniae (n = 316; 144 isolates with hyperviscosity characteristics), Pseudomonas aeruginosa (n = 271), Acinetobacter baumannii complex (n = 187), and non-typhoidal Salmonella species (n = 226), Shigella species (n = 13) from miscellaneous culture sources were collected in 2020 in Taiwan. The MICs of the isolates to test antibiotics were determined using the broth microdilution method. GeneXpert was used to detect genes encoding carbapenemases among the carbapenem-non-susceptible (NS) Enterobacterales isolates. Results The MIC values of the cefepime-enmetazobactam combination against extended-spectrum β-lactamase-producing E. coli and K. pneumoniae isolates (MIC90 ≤ 0.5 mg/L), bla KPC-harboring E. coli isolates (0.25 mg/L; n = 2), and 80% of bla OXA-48-like gene-harboring K. pneumoniae isolates (≤2 mg/L) were low. The MIC ranges of the cefepime-zidebactam against carbapenemase-producing Enterobacterales isolates (irrespective of the carbapenemase type [MIC90 ≤ 4 mg/L]) and carbapenem-NS or ceftolozane-tazobactam-NS P. aeruginosa isolates (MIC90 value, 8 mg/L) were significantly lower than those of the cefepime-enmetazobactam. Conclusions The efficacy of novel antibiotics against important drug-resistant GNB must be monitored and validated during the clinical treatment of patients.

Investigation of ceftazidime-avibactam susceptibility in clinical isolates of gram-negative bacteria.

Our study investigated the susceptibility rate of ceftazidime-avibactam and the risk factors associated with its resistance by analyzing gram-negative bacteria isolated from various patient samples. Between March and November 2020, 1119 gram-negative bacteria strains were isolated from patient samples in Acıbadem Healthcare Group hospitals; ceftazidime-avibactam susceptibility results were evaluated using a 10/4µg (Oxoid, UK) disc and evaluated according to Eucast 2020 recommendations. Patient and isolate characteristics that could be risk factors were retrospectively investigated and statistically analyzed using SPSS 25.0. Male patients made up 52% (n = 581) of the study's total patient population, and they averaged 55.5 ± 24.9 years old. Of 1119 gram-negative strains culture and antibiogram, 1023 (91.4%) were sensitive to ceftazidime-avibactam. An increased risk of resistance was observed with female gender (OR = 2.29; CI 95% [1.45-3.61]; p < 0.05), Pseudomonas aeruginosa (OR = 1.67, CI 95% [1.03-2.7]; p < 0.05), the presence of multidrug-resistance (MDR) (OR = 4.07, CI 95% [2.47-6.7]; p < 0.05) pandrug-resistance (PDR) (OR = 12, (CI) 95% [9.9-14.7] ]; p < 0.05) and admission to intensive care unit (ICU) (OR = 1.89, CI 95% [1.22-2.93]; p < 0.05). The resistance rate of ceftazidime-avibactam was found to be 8.6%, and it was thought that resistant strains produced metallo-ß-lactamase (MBL) type carbapenemase. Risk factors were female gender, Pseudomonas aeruginosa, MDR, PDR, and admission to ICU. Therefore, studying the ceftazidime-avibactam susceptibility test together with gram-negative bacteria identification, especially in groups at risk for resistance, is one of the important factors that can positively affect the success of treatment.

Phenotypical and genotypical assessment of resistance of carbapenemase producing Klebsiella pneumoniae strains

Introduction. The emergence and global spread of antimicrobial resistance genes, such as genes for extended-spectrum beta-lactamases and carbapenemases in Klebsiella pneumoniae strains, poses a significant threat to public health. The reason for this is that carbapenems are the latest therapeutic agent used to treat diseases caused by multi-resistant gram-negative bacteria. The detection of carbapenemases is epidemiologically important because ESBL-genes are plasmid-mediated and can be transmitted horizontally among different bacterial species. The aim of this research was to assess the spectrum of antibiotic resistance of carbapenemase-producing strains of K. pneumoniae circulating in hospitals in St. Petersburg using phenotypic and genotypic methods. Materials and methods. The susceptibility of K. pneumoniae strains (n=182), isolated from samples from patients admitted to hospitals in St. Petersburg in the period from 2016 to 2018, was assessed with 16 antimicrobials. Resistance to antibiotics was assessed by the gradient diffusion method. Carbapenemase genes were detected by molecular (PCR) and phenotypic (CIM) methods. Results. Most of the strains demonstrated a high percentage of resistance to ticarcillin/clavulanate (96.7%), quinolones (nalidixic acid (91.8%), norfloxacin (91.2%), ciprofloxacin (90.1%), ofloxacin (89%)), III and IV generation cephalosporins (cefotaxime (92.9%), ceftazidime (88.5%), cefoperazone (87.9%), ceftriaxone (86.8%), cefepime (91.2%)), netilmicin (83.5%) and gentamicin (80.2%). The investigated strains showed moderate resistance to amikacin (74.2%), meropenem (72.5%) and imepenem (65.9%). The main type of carbapenemases produced by the studied isolates of K. pneumoniae were NDM metallo-beta-lactamases (60.4%) and OXA-48 serine carbapenemases (49.5 %). KPC carbapenemases were found in only 1.1% of the strains. Discussion. The results of the study demonstrate a high resistance level of K. pneumoniae nosocomial isolates to various antibiotics. Therefore, the range of antimicrobials for treatment of Klebsiella infections is significantly reduced, which can lead to a decrease in the effectiveness of clinical therapy. Acknowledgments. The study had no sponsorship. Conflict of interest. The authors declare no apparent or potential conflicts of interest related to the publication of this article.

The Antimicrobial Resistance Characteristics of Imipenem-Non-Susceptible, Imipenemase-6-Producing Escherichia coli.

Imipenemase-6 (IMP-6) type carbapenemase-producing Enterobacteriaceae is regarded as dangerous due to its unique lack of antimicrobial susceptibility. It is resistant to meropenem (MEPM) but susceptible to imipenem (IPM). In addition to carbapenemase, outer membrane porins and efflux pumps also play roles in carbapenem resistance by reducing the antimicrobial concentration inside cells. Extended-spectrum β-lactamase (ESBL) is transmitted with IMP-6 by the plasmid and broadens the spectrum of antimicrobial resistance. We collected 42 strains of IMP-6-producing Escherichia coli and conducted a molecular analysis of carbapenemase, ESBL, porin, efflux, and epidemiological characteristics using plasmid replicon typing. Among the 42 isolates, 21 strains were susceptible to IPM (50.0%) and 1 (2.4%) to MEPM. Seventeen strains (40.5%) co-produced CTX-M-2 type ESBL. We found that the relative expression of ompC and ompF significantly correlated with the MIC of IPM (p = 0.01 and p = 0.03, respectively). Sixty-eight% of CTX-M-2-non-producing strains had IncI1, which was significantly different from CTX-M-2-producing strains (p < 0.001). In conclusion, 50.0% of our IMP-6-producing strains were non-susceptible to IPM, which is different from the typical pattern and can be attributed to decreased porin expression. Further studies investigating other types of carbapenemase are warranted.

In vitro Bactericidal Activities of Combination Antibiotic Therapies Against Carbapenem-Resistant Klebsiella pneumoniae With Different Carbapenemases and Sequence Types.

Carbapenem-resistant Klebsiella pneumoniae (CRKP) is becoming increasingly problematic due to the limited effectiveness of new antimicrobials or other factors such as treatment cost. Thus, combination therapy remains a suitable treatment option. We aimed to evaluate the in vitro bactericidal activity of various antibiotic combinations against CRKP with different carbapenemase genotypes and sequence types (STs). Thirty-seven CRKP with various STs and carbapenemases were exposed to 11 antibiotic combinations (polymyxin B or tigecycline in combination with β-lactams including aztreonam, cefepime, piperacillin/tazobactam, doripenem, meropenem, and polymyxin B with tigecycline) in static time-kill studies (TKS) using clinically achievable concentrations. Out of the 407 isolate-combination pairs, only 146 (35.8%) were bactericidal (≥3 log10CFU/mL decrease from initial inoculum). Polymyxin B in combination with doripenem, meropenem, or cefepime was the most active, each demonstrating bactericidal activity in 27, 24, and 24 out of 37 isolates, respectively. Tigecycline in combination with β-lactams was rarely bactericidal. Aside from the lower frequency of bactericidal activity in the dual-carbapenemase producers, there was no apparent difference in combination activity among the strains with other carbapenemase types. In addition, bactericidal combinations were varied even in strains with similar STs, carbapenemases, and other genomic characteristics. Our findings demonstrate that the bactericidal activity of antibiotic combinations is highly strain-specific likely owing to the complex interplay of carbapenem-resistance mechanisms, i.e., carbapenemase genotype alone cannot predict in vitro bactericidal activity. The availability of WGS information can help rationalize the activity of certain combinations. Further studies should explore the use of genomic markers with phenotypic information to predict combination activity.

Occurrence of blaOXA-48 type carbapenemase in Escherichia coli with coexisting resistance determinants: A report from India

The Ambler class D β-lactamase, OXA-48 has emerged as one of the most frequently reported and clinically significant carbapenemase. The study aims to investigate the carriage of blaOXA-48 within Escherichia coli of clinical origin along with their resistance profile, carriers and associated genes from a tertiary referral hospital of Northeast India. From January 2019 through December 2019, 179 Escherichia coli isolates with reduced susceptibility to carbapenem were collected and screened for the presence of blaOXA-48 gene via PCR assay. OXA-48 positive isolates were further checked for coexistence of different carbapenemases and ESBLs genes followed by their horizontal transferability, plasmid incompatibility by PBRT, phylogroup analysis and strain typing via ERIC PCR. PCR assay result revealed that 20.67% (n = 37) multidrug resistant Escherichia coli isolates were harbouring the class D carbapenemase gene blaOXA-48. 43.24% (n = 16) of the blaOXA-48 positive isolates were co-harbouring other resistance genes namely blaNDM (n = 11), blaOXA-23 (n = 2), blaOXA-58 (n = 1), blaTEM (n = 1) and blaCTX-M-15 (n = 1). The study identified diverse conjugatively transferable Inc type plasmids were carrying this gene within 11 different haplotypes of Escherichia coli. In addition to Inc L/M plasmid, the study identified four Inc types (FREPB, A/C, K/B and FIB) with the carriage of blaOXA-48 among the phylogenetic groups A, B1 and B2 of Escherichia coli. Genetic analysis of blaOXA-48 showed the association with two different insertion sequences in the upstream region. The need of the hour is monitoring of this plasmid mediated gene along with its potential carriers to check their further dissemination so that they don't act as future source to spread carbapenem resistance in this region.

Molecular Detection of blaOXA -type Carbapenemase Genes and Antimicrobial Resistance Patterns among Clinical Isolates of Acinetobacter baumannii.

Acinetobacter baumannii is a bacterium found in most places, especially in clinics and hospitals, and an important agent of nosocomial infections. The presence of class D enzymes such as OXA-type carbapenemases in A. baumannii is proven to have a key function in resistance to carbapenem. The aim of the current study is to determine the blaOXA -type carbapenemase genes and antimicrobial resistance among clinically isolated samples of A. baumannii. We assessed 100 clinically isolated specimens of A. baumannii from patients in intensive care units of educational hospitals of Hamadan, West of Iran. The A. baumannii isolates' susceptibility to antibiotics was performed employing disk diffusion method. Multiplex polymerase chain reaction was used to identify the bla OXA-24-like , bla OXA-23-like , bla OXA-58-like , and bla OXA-51-like genes. The bla OXA-23-like , bla OXA-24-like , and bla OXA-58-like genes' prevalence were found to be 84, 58, and 3%, respectively. The highest coexistence of the genes was for bla OXA-51/23 (84%) followed by bla OXA-51/24-like (58%). The bla OXA-51/23- like pattern of genes is a sort of dominant gene in resistance in A. baumannii from Hamadan hospitals. The highest resistance to piperacillin (83%) and ciprofloxacin (81%) has been observed in positive isolates of bla OXA-23-like . The A. baumannii isolates with bla OXA-58-like genes did not show much resistance to antibiotics. Based on the results of the phylogenetic tree analysis, all isolates have shown a high degree of similarity. This study showed the high frequency of OXA -type carbapenemase genes among A. baumannii isolates from Hamadan hospitals, Iran. Thus, applying an appropriate strategy to limit the spreading of these strains and also performing new treatment regimens are necessary.