MICs Of Imipenem Research Articles

A Novel Triple Combination To Combat Serious Infections with Carbapenem-Resistant Acinetobacter baumannii in a Mouse Pneumonia Model.

ABSTRACTThe ongoing crisis of antimicrobial resistance demands novel combinations between antimicrobials and nonantimicrobials to manage infections caused by highly resistant pathogens. This study aimed to evaluate the effect of combining sodium ascorbate and/or apo-transferrin with imipenem, forming double and triple combinations, against 20 multiple-carbapenemase-producing Acinetobacter baumannii strains using the checkerboard test, time-kill assay, and disc diffusion test. The results of the checkerboard assay revealed that all double combinations showed indifference, while only triple combination recorded a synergistic effect (fractional inhibitory concentration index [FICI] < 0.8) in 95% the test isolates. Moreover, the MIC of imipenem (MICimp) was strongly reduced (up to 128-fold reduction) after treatment with the triple combination against highly resistant isolates and reached the susceptible range. The time-kill assay revealed that the triple combination led to a 4-log10 reduction in the CFU at 8 h compared with the initial bacterial count, and no viable count was recorded at 10 h. The mouse pneumonia model showed restoration of lung function and structure, with mild to moderate residual inflammation and moderately congested vessels observed 8 h following administration of the triple rescue therapy. Additionally, normal lungs with normal patent alveoli were detected 72 h following treatment. Accordingly, sodium ascorbate and apo-transferrin are promising adjunct biological agents with the potential to restore the effectiveness of critically essential antibiotics like imipenem, commonly used for the treatment of A. baumannii infections.IMPORTANCE Combination therapy provides a perspective to threat multidrug-resistant (MDR) strains. The present study sheds light on a novel and effective triple combination against carbapenem-resistant A. baumannii. Our in vitro results showed that combining imipenem with apo-transferrin and sodium ascorbate yielded synergism in 95% of test isolates, and this was associated with a marked reduction in imipenem MIC, shifting it below the breakpoint. Furthermore, a bactericidal effect was recorded, with no viable count detected at 10 h. An in vivo murine model of pneumonia was induced to mimic human disease. The triple combination therapy restored lung function and structure, with mild to moderate residual inflammation and moderately congested vessels observed 8 h following the initiation of therapy. Therefore, our findings suggest novel insights about a promising new combination therapy against highly resistant carbapenemase-producing A. baumannii to restore the effectiveness of imipenem.

Screening of Carbapenem Resistance Klebsiella pneumoniae and its MIC against Imipenem

Klebsiella pneumoniae is a common opportunistic pathogen causing a wide range of infections; pneumonia, urinary tract infection, bacteremia, and liver abscesses. It infects primarily immunocompromised and immunocompetent individuals. It presents itself as an antibiotic-resistant bacterium, especially for third-generation cephalosporins and carbapenems, creating serious global challenges. Therefore, this cross-sectional study was conducted in B &amp; B Hospital, Lalitpur to screen the distribution of carbapenem resistance K. pneumoniae through ertapenem and to assess the minimum inhibitory concentration of imipenem for screened carbapenem resistance K. pneumoniae. From 3447 different clinical samples collected according to standard guidelines, K. pneumoniae was identified using standard microbiological techniques; staining and a panel of biochemical tests. The antibiotic susceptibility test of the isolates was performed by the Kirby-Bauer disc diffusion method as per CLSI 2018 guidelines. The screening of carbapenem resistance was assessed by using ertapenem disc and the MIC of imipenem for carbapenem resistance and intermediate was performed using an Epsilometer. A total of 85 K. pneumoniae were identified and their antibiotic susceptibility test revealed that ceftriaxone was the least effective antibiotic. The number of MDR, carbapenem-resistant and intermediate isolates was 51, 46, and 3, respectively. The MIC of imipenem through an Epsilometer from ertapenem resistant and intermediate revealed that 31, 5, and 13 isolates were resistant, intermediate, and sensitive respectively. These findings showed the inconsistency in the detection of carbapenem-resistant isolates in routine microbiology laboratories and further support the other tests for the detection of carbapenem resistance as suggested by CLSI.

Imipenem Induced Transcriptional Modulation of Multidrug Efflux Pumps and Porin Genes in Carbapenem Resistant Escherichia coli Isolates of Animal Origin

Background: The emergence of a multidrug resistance phenotype is most likely outcome of co-existing multiple mechanisms. The underlying molecular mechanism of carbapenem resistance is poorly understood in bacterial isolates of animal origin. The present study investigates the resistance mechanism for reduced susceptibility to carbapenem in E. coli. Methods: The study includes 12 multidrug resistant E.coli isolates showing decreased susceptibility to imipenem. Carbapenemase production was evaluated by the mCarba-NP test. The presence of the carbapenem resistance gene was confirmed by multiplex PCR. Efflux pump activity was accessed by broth microdilution method with or without selective efflux pump inhibitor. We analyzed differential expression of the outer membrane porin genes (ompF, ompC) and efflux pump genes (acrA, acrB) by quantitative real-time PCR analysis. Result: We observed increased expression of acrA and acrB genes in all the E. coli isolates under imipenem stress. A positive correlation (p less than 0.001) was evident between imipenem MIC and acrA and acrB over-expression in these isolates. A significant decrease in ompF and ompC expression was observed in these isolates under imipenem stress. The change in MIC was not correlated with the corresponding OmpC and OmpF gene expression. The results suggest the importance of efflux pump and porins proteins in the emergence of multidrug resistance besides carbapenemase production.

In vitro and in vivo activities of a novel β-lactamase inhibitor combination imipenem/XNW4107 against recent clinical Gram-negative bacilli from China

XNW4107 is a novel β-lactamase inhibitor that possesses broad activity against serine-β-lactamases. XNW4107 in combination with imipenem exhibited potent in vitro activity against carbapenem-resistant bacteria and particularly against carbapenem-resistant Acinetobacter baumannii. This study aimed to evaluate the in vitro and in vivo antibacterial activities of imipenem/XNW4107. The minimum inhibitory concentrations, minimum bactericidal concentrations, time-kill curves, post-antibiotic effects, and spontaneous frequency of resistance were used to investigate the imipenem/XNW4107 in vitro activity. A mouse systemic infection model was used to evaluate the imipenem/XNW4107 in vivo efficacy. MIC90 of imipenem/XNW4107 against imipenem-nonsusceptible A. baumannii (n=106) was 8 mg/L, which was 16-fold lower than the MIC90 of imipenem; the resistance rate decreased from 90% to 20% applying the CLSI imipenem breakpoint. MIC90 of imipenem/XNW4107 against imipenem-resistant Klebsiella pneumoniae (n=54) was 2 mg/L, which was 128-fold lower than the MIC90 of imipenem; 80% imipenem-nonsusceptible Pseudomonas aeruginosa (n=101) exhibited MICs of imipenem/XNW4107 from 2 to 8 mg/L, which were 4- to 8-fold lower than the MICs of imipenem. Imipenem/XNW4107 was bactericidal against A. baumannii, K. pneumoniae, and Escherichia coli. The time-kill curves showed that increasing concentrations did not result in progressively increased killing at concentrations >4×MIC. Imipenem/XNW4107 has a low potential for resistance development in tested strains except for K. pneumoniae. Imipenem/XNW4107 provided good protection against imipenem-resistant A. baumannii and K. pneumoniae in vivo. The broad-spectrum profile and potent in vitro and in vivo antibacterial activities support imipenem/XNW4107 as a promising investigational candidate.

Comparison of carbapenem minimum inhibitory concentrations of Oxacillin-48-like Klebsiella pneumoniae by Sensititre, Vitek 2, MicroScan, and Etest

ObjectivesThe aim of this laboratory-based study was to compare carbapenem MICs yielded by Sensititre, Vitek 2, MicroScan WalkAway plus and Etest for Oxacillin (OXA)-48-like Klebsiella pneumoniae isolates. MethodsAnalysis was performed for categorical agreement for ertapenem, meropenem, and imipenem, and the proportion of isolates with MICs ≤8μg/mL and the MIC50/MIC90 for meropenem and imipenem, from a convenience sample of 82 deduplicated blood culture OXA-48-like K. pneumoniae isolates. ResultsThe proportion of isolates testing susceptible to ertapenem by Etest (19/82, 23.1%) differed from Sensititre/Vitek (0/82) and MicroScan (2/82, 2.4%) (p < 0.001 for all). For meropenem, the proportion of isolates susceptible by Etest (31/82, 37.8%) differed from Sensititre/Vitek (16/82, 19.5%) (p = 0.015). There was variation in the proportion of isolates that tested imipenem susceptible when comparing Sensititre (9/82, 11%) and Vitek (8/82, 9.8%) to MicroScan (27/82, 32.9%), p = 0.001 and p < 0.001, respectively, Sensititre and Vitek to Etest (45/82, 54.9%), p < 0.001 for both, and MicroScan to Etest, p = 0.007. The proportion of isolates with meropenem MICs ≤8μg/mL with Sensititre and Vitek differed significantly from Etest, 58.5% and 85.4%, respectively, p < 0.001. A 2-fold difference between the Sensititre and Vitek meropenem and imipenem MIC at which ≥50% of isolates were inhibited compared to the MicroScan, and a 4-fold difference compared to Etest, was present. ConclusionsSubstantial variability in carbapenem MICs for OXA-48-like carbapenemase-producing Enterobacterales isolates by the four methods was demonstrated. Performance characteristics verification of MIC methods in use for the predominant carbapenemase-producing Enterobacterales type is required by laboratories to optimize the accuracy of carbapenem reporting.

Carbapenemase-Encoding Gene Copy Number Estimator (CCNE): a Tool for Carbapenemase Gene Copy Number Estimation.

ABSTRACTCarbapenemase production is one of the leading mechanisms of carbapenem resistance in Gram-negative bacteria. An increase in carbapenemase gene (blaCarb) copies is an important mechanism of carbapenem resistance. No currently available bioinformatics tools allow for reliable detection and reporting of carbapenemase gene copy numbers. Here, we describe the carbapenemase-encoding gene copy number estimator (CCNE), a ready-to-use bioinformatics tool that was developed to estimate blaCarb copy numbers from whole-genome sequencing data. Its performance on Klebsiella pneumoniae carbapenemase gene (blaKPC) copy number estimation was evaluated by simulation and quantitative PCR (qPCR), and the results were compared with available algorithms. CCNE has two components, CCNE-acc and CCNE-fast. CCNE-acc detects blaCarb copy number in a comprehensive and high-accuracy way, while CCNE-fast rapidly screens blaCarb copy numbers. CCNE-acc achieved the best accuracy (100%) and the lowest root mean squared error (RMSE; 0.07) in simulated noise data sets, compared to the assembly-based method (23.4% accuracy, 1.697 RMSE) and the OrthologsBased method (78.9% accuracy, 0.395 RMSE). In the qPCR validation, a high consistency was observed between the blaKPC copy number determined by qPCR and that determined with CCNE. Reverse transcription-qPCR transcriptional analysis of 40 isolates showed that blaKPC expression was positively correlated with the blaKPC copy numbers detected by CCNE (P < 0.001). An association study of 357 KPC-producing K. pneumoniae isolates and their antimicrobial susceptibility identified a significant association between the estimated blaKPC copy number and MICs of imipenem (P < 0.001) and ceftazidime-avibactam (P < 0.001). Overall, CCNE is a useful genomic tool for the analysis of antimicrobial resistance genes copy number; it is available at https://github.com/biojiang/ccne.IMPORTANCE Globally disseminated carbapenem-resistant Enterobacterales is an urgent threat to public health. The most common carbapenem resistance mechanism is the production of carbapenemases. Carbapenemase-producing isolates often exhibit a wide range of carbapenem MICs. Higher carbapenem MICs have been associated with treatment failure. The increase of carbapenemase gene (blaCarb) copy numbers contributes to increased carbapenem MICs. However, blaCarb gene copy number detection is not routinely conducted during a genomic analysis, in part due to the lack of optimal bioinformatics tools. In this study, we describe a ready-to-use tool we developed and designated the carbapenemase-encoding gene copy number estimator (CCNE) that can be used to estimate the blaCarb copy number directly from whole-genome sequencing data, and we extended the data to support the analysis of all known blaCarb genes and some other antimicrobial resistance genes. Furthermore, CCNE can be used to interrogate the correlations between genotypes and susceptibility phenotypes and to improve our understanding of antimicrobial resistance mechanisms.

The Emergence of Carbapenem Resistant Enterobacteriaceae Producing GIM-1 and SIM-1 Clinical Isolates in Khartoum-Sudan.

PurposeThe aim of this study was to detect multidrug resistant GIM-1 and SIM-1 producing Enterobacteriaceae clinical isolates from hospitalized patients across three Khartoum State Teaching Hospitals, Sudan.Patients and MethodsFrom May 2018 to October 2019, Enterobacteriaceae clinical isolates from inpatients admitted to different Khartoum state hospitals. Genes for carbapenemase (GIM-1 and SIM-1) were amplified by polymerase chain reaction (PCR). Agar dilution method was used to determine MICs for imipenem and meropenem after antimicrobial susceptibility testing.ResultsFive (1.29%) isolates of Enterobacteriaceae [2 (0.51%) Escherichia coli isolates produce GIM-1, 2 (0.51%) Klebsiella pneumoniae isolates (one [0.25%] of each produce of GIM-1 and of SIM-1), and 1 (0.25%) Enterobacter cloacae isolate produce GIM-1]. Susceptibility profiling of the isolates showed a low-level resistance to imipenem and meropenem MICs (8, 16 and 32 μg/mL). It also had resistance to ampicillin, extended-spectrum cephalosporin’s, aztreonam, and amoxicillin-clavulanate and with the two K. pneumoniae strains showing resistance to colistin.ConclusionWe report the emergence of four GIM-1 producing Enterobacteriaceae strains and one strain of SIM-1 producing K. pneumoniae genes, isolated from hospitalized patients, with a high resistance pattern to antimicrobial agents. Whole-genome sequencing (WGS) is necessary for precise identification of clonal diversity backgrounds of acquired carbapenemase genes in diagnostic laboratories as the number of cases of carbapenem resistant Enterobacteriaceae infection increases annually.

The predominance of Klebsiella pneumoniae carbapenemase (KPC-type) gene among high-level carbapenem-resistant Klebsiella pneumoniae isolates in Baghdad, Iraq.

The serine carbapenemase enzymes (KPC) which produce from bacteria klebsiella pneumoniae today have been emerged as one of the β-lactamase enzymes that is capable to inactivating the last line of carbapenems. The gene encoding the K. pneumonia (blaKPC) belongs to gene carried on plasmid among Enterobacteriaceae family, which has modulation for the infections control so this study is aimed to spot the presence and evaluate blaKPC gene expression by real-time PCR in local isolates of K. pneumonia. Forty-seven of K. pneumonia isolates were isolated from different clinical samples (blood, sputum, urine, wounds and burns) from patients in separate hospitals in Baghdad., Antimicrobial sensitivity test was carried out by vitik-2 system and Kirby- Bauer method. The PCR was employed to detect carbapenemase gene. The results of this study showed that all explored isolates were resistant to Ertapenem, Meropenem and imipenem 47(100%). Phenotypically, all the isolates had carbapenemase which hydrolyzed the carbapenem antibiotics. Furthermore, the isolates showed (100%) resistance to Cefazolin, Ampicillin and Amoxicillin/ Clavulic acid. However, the most effective antibiotic was Levofloxacin (91.5%). The results of conventional PCR technique for the detection of blaKPC gene showed that 38 (80.9%) isolates of carbapenem-resistant K. pneumoniae harboured blaKPC gene (1010bp), while none carried other carbapenemase genes including blaNDM1, blaVIM and blaIMP genes. High levels of carbapenem resistance was clarified by the imipenem and meropenem MICs determination. All 38 isolates were positive in CNPT. Furthermore, the 38 isolates showed over expression of blaKPC gene compared with housekeeping rpo gene in Real-Time PCR. According to these results, the resistant isolates to carbapenem were belong to the present and high level expression of blaKPC gene in our local isolates.

In vitro activity of nitroxoline against carbapenem-resistant Acinetobacter baumannii isolated from the urinary tract

The old antimicrobial nitroxoline is currently repurposed for oral treatment of uncomplicated urinary tract infections (UTIs). To investigate the in vitro activity of nitroxoline against carbapenem-resistant Acinetobacter baumannii (CRAb). From an international collection of previously well-characterized clinical A. baumannii isolates, 34 isolates from urinary tract sources with different carbapenem-resistance mechanisms were selected. Nitroxoline activity was analysed with broth microdilution (BMD), disc diffusion (DD) and within an in vitro biofilm model. MICs of meropenem and imipenem were assessed with BMD. Susceptibility to ciprofloxacin and trimethoprim/sulfamethoxazole was investigated using DD. Escherichia coli ATCC 25922 and A. baumannii NCTC 13304 were used for quality control. All isolates were carbapenem resistant (MIC90 >32 mg/L for meropenem and imipenem) and most isolates were resistant to ciprofloxacin (33/34) and trimethoprim/sulfamethoxazole (31/34). Nitroxoline yielded MIC50/90 values of 2/2 mg/L (MIC range 1-2 mg/L) and inhibition zone diameters ranging from 20 to 26 mm. In contrast, for definite eradication of biofilm-associated CRAb in vitro, higher nitroxoline concentrations (≥16 to ≥128 mg/L) were necessary for all isolates. Nitroxoline showed excellent in vitro activity against a collection of CRAb despite high resistance rates to other antimicrobials for parental and oral therapy of A. baumannii UTI. Currently, nitroxoline is recommended for the treatment of uncomplicated UTI in Germany with a EUCAST breakpoint limited to uncomplicated UTI and E. coli (S ≤16 mg/L). Nitroxoline could be a promising drug for oral treatment of lower UTI caused by CRAb. More data are warranted to correlate these findings with in vivo success rates.

Diversity of genomic clusters and CfiA/cfiA alleles in Bacteroides fragilis isolates from human and animals

ObjectivesTo compare the phylogeny of cfiA-positive Bacteroides fragilis isolates from diverse human and animal sources. MethodComplete genome sequences were obtained from 42 cfiA-positive B. fragilis isolates (Hong Kong, 2015–2017) and additional 24 genomes deposited in the GenBank (multiple countries, 1985–2019) were included. The genomic clusters were constructed using PopPUNK. The CfiA alleles and polymorphism in the cfiA locus were analyzed in silico. ResultsThe 66 isolates were grouped into 12 genomic clusters (BFSC-1 to 12). Human infection isolates were distributed in diverse clusters, being many of them common to fecal isolates from both human and animals. Thirteen CfiA alleles including 2 novel ones were identified. CfiA-1 (n = 28) is the predominating allele, following by CfiA-13 (n = 8), CfiA-4 (n = 7) and CfiA-14 (n = 6). The other CfiA alleles were identified in 1–3 isolates. Six patterns of gene context were identified in the regions flanking cfiA locus. No consistent association between genomic clusters and CfiA alleles could be detected. Similarly, markedly elevated imipenem MIC was linked to the integration, immediately upstram of cfiA of an IS element but not the CfiA allele or gene context. ConclusionThe phylogeny of cfiA-positive B. fragilis isolates causing human diseases was diverse and overlaped with those from human and animal carriage.

Genomic features of an isolate of Empedobacter falsenii harbouring a novel variant of metallo-β-lactamase, blaEBR-4 gene.

Empedobacter falsenii is an emerging opportunistic pathogen that has been occasionally implicated in various human infections. In this study, we described the genomic features of a multidrug resistant E. falsenii Q1655 obtained from a patient attending a public hospital in Sokoto, northwest Nigeria. The isolate, E. falsenii Q1655, was isolated from the stool sample of a patient in Sokoto, Nigeria. The identity of the isolate was confirmed by MALDITOF-MS. The disc diffusion test and modified Carba-NP test were used for phenotypic antibiotic susceptibility test and carbapenemase enzyme production test, respectively. The whole genome of the strain was sequenced using the Illumina MiSeq technique. Resistome analysis was done by annotation of the WGS against the ARG-ANNOT database. The isolate was resistant to all β-lactam antibiotics with the exception of cefepime. The MICs of imipenem and ertapenem as determined by E-test were 12μg/ml and 2μg/ml, respectively. Modified Carba NP test showed that the strain was carbapenemase producing. Resistome analysis revealed the presence of a novel metallo-β-lactamase, a chromosomal blaEBR-4, which exhibited 94.92% and 97.02% nucleotide and protein sequence identities respectively with blaEBR-3 gene of E. falsenii 174,820. Seven and eight amino-acid substitutions were observed with the blaEBR-1 and blaEBR-2, respectively. We reported the first isolation and genomic description of an extensively drug resistant isolate of Empedobacter falsenii in Nigeria. This report broadens our knowledge of carbapenem resistance in E. falsenii and it will serve as a useful guide in the development of antibiotic use policy.

Invitro susceptibility of Imipenem by E-test in different conditions of Methicillin resistant Staphylococcus aureus

MRSA infections have been confined to health care centres for decades, which is recognised as one of the major pathogen in Hospital as well as community settings. MRSA infections are associated with high morbidity and mortality. However, nowadays MRSA infections are increasingly seen in young healthy individuals with no exposure to health care centres, ie;CA-MRSA infections are seen increasingly. 50% to 70% of Staphylococcus aureus isolates submitted to area labs are methicillin resistant. MRSA is a major threat to patient safety. There are antibiotics that can kill MRSA germs. MRSA are resistant to most of the beta lactam antibiotics, not only to beta lactam but also to several other agents which is used for treatment. Imipenem is the most important antibiotic of Carbapenem, it is bactericidal for most aerobic and anaerobic pathogenic bacteria except MRSA. But in several studies its action against MRSA was shown. Here in this study, invitro susceptibility testing indicated that Imipenem is active against MRSA at different conditions, ie; at a physiological pH 7.4 &amp; pH 5.5 an abscess condition and at two temperature 370c and 300c .Since the methicillin resistance is enhanced at 300c, the MIC of Imipenem detected at two temperature. Imipenem has a low molecular weight so it has a better penetration effect, than vancomycin and Daptomycin which is used for the treatment of MRSA. In, invitro study Imipenem spread in the medium more rapidly, and kill bacteria. So in MIC testing by Epsilometri test there is wide zone will be seen. Imipenem is the most important antibiotic of Carbapenem, it is bactericidal. To detect the in-vitro susceptibility of Imipenem by E test in different conditions of Methicillin Resistant Staphylococcus aureus. Malabar Institute of Medical Sciences, Calicut; Randomized controlled trial. Sample collection, Confirmation of MRSA by latex agglutination and cefoxitin e-test, detection of MIC of Imipenem by E-test at different conditions of MRSA, Detection of combination effect of Imipenem and Daptomycin against MRSA by E-test. paired sample t test

Risedronate and Methotrexate Are High-Affinity Inhibitors of New Delhi Metallo-β-Lactamase-1 (NDM-1): A Drug Repurposing Approach.

Bacteria expressing New Delhi metallo-β-lactamase-1 (NDM-1) can hydrolyze β-lactam antibiotics (penicillins, cephalosporins, and carbapenems) and, thus, mediate multidrug resistance. The worldwide dissemination of NDM-1 poses a serious threat to public health, imposing a huge economic burden in the development of new antibiotics. Thus, there is an urgent need for the identification of novel NDM-1 inhibitors from a pool of already-known drug molecules. Here, we screened a library of FDA-approved drugs to identify novel non-β-lactam ring-containing inhibitors of NDM-1 by applying computational as well as in vitro experimental approaches. Different steps of high-throughput virtual screening, molecular docking, molecular dynamics simulation, and enzyme kinetics were performed to identify risedronate and methotrexate as the inhibitors with the most potential. The molecular mechanics/generalized Born surface area (MM/GBSA) and molecular dynamics (MD) simulations showed that both of the compounds (risedronate and methotrexate) formed a stable complex with NDM-1. Furthermore, analyses of the binding pose revealed that risedronate formed two hydrogen bonds and three electrostatic interactions with the catalytic residues of NDM-1. Similarly, methotrexate formed four hydrogen bonds and one electrostatic interaction with NDM-1’s active site residues. The docking scores of risedronate and methotrexate for NDM-1 were –10.543 kcal mol−1 and −10.189 kcal mol−1, respectively. Steady-state enzyme kinetics in the presence of risedronate and methotrexate showed a decreased catalytic efficiency (i.e., kcat/Km) of NDM-1 on various antibiotics, owing to poor catalytic proficiency and affinity. The results were further validated by determining the MICs of imipenem and meropenem in the presence of risedronate and methotrexate. The IC50 values of the identified inhibitors were in the micromolar range. The findings of this study should be helpful in further characterizing the potential of risedronate and methotrexate to treat bacterial infections.

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.

Non-susceptibilities to antibiotics against important Gram-negative bacteria, and imipenem-relebactam, meropenem-vaborbactam against carbapenem non-susceptible Enterobacterales and Pseudomonas aeruginosa isolates implicated in complicated intra-abdominal and urinary tract infections in Taiwan, 2019

Susceptibility of isolates of top-ranking Enterobacterales species and Pseudomonas aeruginosa implicated in complicated intra-abdominal infections (cIAI) and urinary tract infections (cUTI) to important antibiotics, including imipenem-relebactam (IMR) and meropenem-vaborbactam (MVB), in Taiwan in 2019 were evaluated. MICs to various antibiotics were determined using broth microdilution method. Susceptibility results were interpreted mainly based on the MIC breakpoints of the Clinical and Laboratory Standards Institute (CLSI) 2021, but susceptibilities of IMR and MVB were interpreted based on the CLSI and European Committee on Antimicrobial Susceptibility Testing (EUCAST) 2021. Resistance genes amongst carbapenem-non-susceptible (NS) Gram-negative bacteria (GNB) were investigated using multiplex polymerase chain reaction (PCR). Escherichia coli (n=356), Klebsiella pneumoniae (n=165) and Enterobacter cloacae complex (n=42) isolates accounted for 85.3% of the 660 Enterobacterales isolates. The non-susceptibility rates of imipenem (IPM), IMR against isolates of non-Morganellaceae Enterobacterales, and meropenem (MEM), MVB against all Enterobacterales isolates were 92.2%/94.8%, 98.4-98.7%/98.4-99%, 95%/98.2% and 98.8-100%/99.4-100% for the cIAI/cUTI subgroups, respectively. Amongst the 40 IPM-NS-non-Morganellaceae Enterobacterales isolates, when the CLSI 2021 criteria were applied, 10 were NS to IMR, and four Klebsiella pneumoniae isolates (harbouring blaKPC but neither blaMBL nor blaOXA-48-like genes) were NS to IMR and MVB. Amongst the 93 Pseudomonas aeruginosa isolates under evaluation, the addition of relebactam (4mg/L) resulted in a 4-to-16-fold reduction in the MICs of IPM in all 15 IPM-NS-Pseudomonas aeruginosa isolates (including 10 porin-deficient ones) not harbouring blaMBL/blaOXA-48-like genes. Contrastingly, the addition of vaborbactam (8mg/L) improved the non-susceptibility to MEM in one (20%) of five IPM/MEM-NS Pseudomonas aeruginosa isolates. Continuous monitoring of susceptibility to clinically important GNB is warranted.

Reporting antimicrobial susceptibilities and phenotypes of resistance to vancomycin in vancomycin-resistant Enterococcus spp. clinical isolates: A nationwide proficiency study

IntroductionThe ability of Spanish microbiology laboratories to (a) determine antimicrobial susceptibility (AS), and (b) correctly detect the vancomycin resistance (VR) phenotype in vancomycin-resistant Enterococcus spp. (VRE) was evaluated. MethodsThree VRE isolates representing the VanA (E. faecium), VanB (E. faecium) and VanC (E. gallinarum) VR phenotypes were sent to 52 laboratories, which were asked for: (a) AS method used; (b) MICs of ampicillin, imipenem, vancomycin, teicoplanin, linezolid, daptomycin, ciprofloxacin, levofloxacin and quinupristin–dalfopristin, and high-level resistance to gentamicin and streptomycin; (c) VR phenotype. Results(a) The most frequently used system was MicroScan; (b) according to the system, the highest percentage of discrepant MICs was found with gradient strips (21.3%). By antimicrobial, the highest rates of discrepant MICs ranged 16.7% (imipenem) to 0.7% (linezolid). No discrepant MICs were obtained with daptomycin or levofloxacin. Mayor errors (MEs) occurred with linezolid (1.1%/EUCAST) and ciprofloxacin (5.0%/CLSI), and very major errors (VMEs) with vancomycin (27.1%/EUCAST and 33.3%/CLSI) and teicoplanin (5.7%/EUCAST and 2.3%/CLSI). For linezolid, ciprofloxacin, and vancomycin, discrepant MICs were responsible for these errors, while for teicoplanin, errors were due to a misassignment of the clinical category. An unacceptable high percentage of VMEs was obtained using gradient strips (14.8%), especially with vancomycin, teicoplanin and daptomycin; (c) 86.4% of the centers identified VanA and VanB phenotypes correctly, and 95.0% the VanC phenotype. ConclusionMost Spanish microbiology laboratories can reliably determine AS in VRE, but there is a significant percentage of inadequate interpretations (warning of false susceptibility) for teicoplanin in isolates with the VanB phenotype.

In vitro Eradication of Pseudomonas aeruginosa Persister Cell Producers by Peganum harmala.

Because of increasing antibiotic failure and recurrence of infections in patients with P. aeruginosa, the present study was designed to determine the antibiotic resistance status, presence of persister cells and investigate the antipersister effect of Peganum harmala in P. aeruginosa clinical isolates in vitro in Ilam, Iran. Thirty P. aeruginosa urinary clinical isolates were collected from hospitals in Ilam, Iran and identified by common microbiological and biochemical tests. Afterward, antibiotic susceptibility assay, persister cell assay, P. harmala extraction, cell culture, and cell viability assays were performed. A high rate of antibiotic resistance was observed. All isolates were resistant to co-amoxiclav. Also, 83.3% (n = 25), 90% (n = 27), and 36.6% (n = 11) of isolates showed resistance to ceftazidime, kanamycin, and tobramycin, respectively. The MIC and MBC values for imipenem were ≤ 2 and 2 µg/mL for susceptible isolates, respectively. In addition, 6.66% (n = 2) of isolates were persister cells and were also sensitive to imipenem by MIC but did not show any MBC. IC50 for P. harmala was 35 µg/mL. Eventually, MIC value of P. harmala against two P. aeruginosa persister cell producer isolates was 3 µg/mL and 1 µg/mL, and the value of MBC was 10 µg/mL and 30 µl/mL. Our findings demonstrated that P. harmala may be a suitable antipersister herbal medicine against P. aeruginosa clinical isolates. In this regard, comprehensive research is needed in the future to gain more information in this area.

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.

202. Comparative Activity of Ceftazidime-Avibactam, Imipenem-Relebactam and Meropenem-Vaborbactam Tested Against Carbapenem-Nonsusceptible Enterobacterales that Are Carbapenemase-Negative

202. Comparative Activity of Ceftazidime-Avibactam, Imipenem-Relebactam and Meropenem-Vaborbactam Tested Against Carbapenem-Nonsusceptible <i>Enterobacterales</i> that Are Carbapenemase-Negative

1080. Relebactam Increases Imipenem Activity Against Imipenem-Nonsusceptible and -Susceptible Pseudomonas aeruginosa and Enterobacterales: Assessment of Isolates from RESTORE-IMI 2

BackgroundRelebactam (REL) inhibits class A and C β-lactamases (BLs) and is approved in the US in the combination imipenem/cilastatin/REL (IMI/REL) for hospital acquired bacterial pneumonia (HABP) and ventilator associated bacterial pneumonia (VABP), and also in patients with limited treatment options for complicated urinary tract infections and complicated intraabdominal infections. The objective of this study was to evaluate the potentiation of imipenem (IMI) by REL in baseline respiratory isolates from the recently completed Phase 3 RESTORE-IMI 2 study that demonstrated efficacy and safety of IMI/REL in the treatment of patients with HABP/VABP.MethodsBaseline lower respiratory tract (LRT) isolates were evaluated for IMI MICs in the presence and absence of REL using broth microdilution and CLSI interpretive criteria. All Pseudomonas aeruginosa and Enterobacterales for which IMI/REL is either indicated or the MIC90 is less than or equal to the susceptibility breakpoint were evaluated.ResultsSummary statistics and the MIC distribution for P. aeruginosa are shown in the figure. For P. aeruginosa, REL reduces the IMI mode MIC of IMI-nonsusceptible (IMI-NS) (MIC >2) isolates ≥8-fold (from 16-32 to 2 µg/mL) and that of IMI-susceptible (IMI-S) (MIC ≤2) isolates ≥2-fold (from 1 to ≤0.5 µg/mL). Among Enterobacterales, the IMI mode MIC of IMI-NS (MIC >1) isolates was reduced ≥4-fold (from 2 to ≤0.5 µg/mL). REL enhanced the activity of IMI among IMI-S isolates (MIC ≤1), most notably observed in Enterobacterales species that produce a chromosomal AmpC, increasing the proportion with MIC ≤0.5 µg/mL from 76% to 98%.MIC Distribution and Summary Statistics of RESTORE IMI-2 Isolates ConclusionAmong baseline LRT isolates from RESTORE-IMI 2 the potentiation of IMI by REL results in the restoration of susceptibility among IMI-NS P. aeruginosa and Enterobacterales and enhanced IMI activity among IMI-S isolates. This enhanced activity among IMI-S Enterobacterales is most notable among species with reported chromosomal expression of AmpC. This lowering of IMI MICs upon addition of REL contributes to the high probability of target attainment (≥90%) observed following administration of IMI/REL 1.25g every 6 hours, further supporting the IMI/REL efficacy data observed in RESTORE-IMI 2.Disclosures David W. Hilbert, PhD, Merck (Employee) C. Andrew DeRyke, PharmD, Merck & Co., Inc. (Employee, Shareholder) Maria C. Losada, BA, Merck (Employee) Pamela Moise, PharmD, Merck (Employee) Luke F. Chen, MBBS MPH MBA FRACP FSHEA FIDSA, Merck (Employee) Katherine Young, MS, Merck (Employee)