Development of a risk prediction model for subsequent infection after colonization with carbapenem-resistant Enterobacterales: a retrospective cohort study.

BACKGROUND: Colonization of carbapenem-resistant Enterobacterale (CRE) is considered as one of vital preconditions for infection, with corresponding high morbidity and mortality. It is important to construct a reliable prediction model for those CRE carriers with high risk of infection. METHODS: A retrospective cohort study was conducted in two Chinese tertiary hospitals for patients with CRE colonization from 2011 to 2021. Univariable analysis and the Fine-Gray sub-distribution hazard model were utilized to identify potential predictors for CRE-colonized infection, while death was the competing event. A nomogram was established to predict 30-day and 60-day risk of CRE-colonized infection. RESULTS: 879 eligible patients were enrolled in our study and divided into training (n = 761) and validation (n = 118) group, respectively. There were 196 (25.8%) patients suffered from subsequent CRE infection. The median duration of subsequent infection after identification of CRE colonization was 20 (interquartile range [IQR], 14-32) days. Multisite colonization, polymicrobial colonization, catheterization and receiving albumin after colonization, concomitant respiratory diseases, receiving carbapenems and antimicrobial combination therapy before CRE colonization within 90 days were included in final model. Model discrimination and calibration were acceptable for predicting the probability of 60-day CRE-colonized infection in both training (area under the curve [AUC], 74.7) and validation dataset (AUC, 81.1). Decision-curve analysis revealed a significantly better net benefit in current model. Our prediction model is freely available online at https://ken-zheng.shinyapps.io/PredictingModelofCREcolonizedInfection/ . CONCLUSIONS: Our nomogram has a good predictive performance and could contribute to early identification of CRE carriers with a high-risk of subsequent infection, although external validation would be required.

A challenging case of carbapenem resistant Klebsiella pneumoniae-related pyogenic liver abscess with capsular polysaccharide hyperproduction: a case report.

BACKGROUND: Carbapenem-resistant Klebsiella pneumoniae (CRKP) infections are a major public health problem, necessitating the administration of polymyxin E (colistin) as a last-line antibiotic. Meanwhile, the mortality rate associated with colistin-resistant K. pneumoniae infections is seriously increasing. On the other hand, importance of administration of carbapenems in promoting colistin resistance in K. pneumoniae is unknown. CASE PRESENTATION: We report a case of K. pneumoniae-related pyogenic liver abscess in which susceptible K. pneumoniae transformed into carbapenem- and colistin-resistant K. pneumoniae during treatment with imipenem. The case of pyogenic liver abscess was a 50-year-old man with diabetes and liver transplant who was admitted to Abu Ali Sina Hospital in Shiraz. The K. pneumoniae isolate responsible for community-acquired pyogenic liver abscess was isolated and identified. The K. pneumoniae isolate was sensitive to all tested antibiotics except ampicillin in the antimicrobial susceptibility test and was identified as a non-K1/K2 classical K. pneumoniae (cKp) strain. Multilocus sequence typing (MLST) identified the isolate as sequence type 54 (ST54). Based on the patient's request, he was discharged to continue treatment at another center. After two months, he was readmitted due to fever and progressive constitutional symptoms. During treatment with imipenem, the strain acquired blaOXA-48 and showed resistance to carbapenems and was identified as a multidrug resistant (MDR) strain. The minimum inhibitory concentration (MIC) test for colistin was performed by broth microdilution method and the strain was sensitive to colistin (MIC < 2 µg/mL). Meanwhile, on blood agar, the colonies had a sticky consistency and adhered to the culture medium (sticky mucoviscous colonies). Quantitative real-time PCR and biofilm formation assay revealed that the CRKP strain increased capsule wzi gene expression and produced slime in response to imipenem. Finally, K. pneumoniae-related pyogenic liver abscess with resistance to a wide range of antibiotics, including the last-line antibiotics colistin and tigecycline, led to sepsis and death. CONCLUSIONS: Based on this information, can we have a theoretical hypothesis that imipenem is a promoter of resistance to carbapenems and colistin in K. pneumoniae? This needs more attention.

[Carbapenemase Genes, Virulence Genes, and Molecular Epidemiology of Carbapenem-Resistant Klebsiella pneumoniae Derived From Bloodstream Infections]. / è¡€æµæ„ŸæŸ“ç¢³é’éœ‰çƒ¯è€è¯è‚ºç‚Žå ‹é›·ä¼¯èŒçš„è€è¯åŸºå› å’Œæ¯’åŠ›åŸºå› åŠåˆ†å­æµè¡Œç— å­¦ç ”ç©¶.

Objective: To investigate the clinical characteristics and molecular epidemiology of carbapenem-resistant Klebsiella pneumoniae (CRKP) isolated from patients with bloodstream infections in a large tertiary-care general hospital in Southwest China. Methods: A total of 131 strains of non-repeating CRKP were collected from the blood cultures of patients who had bloodstream infections in 2015-2019. The strains were identified by VITEK-2, a fully automated microbial analyzer, and matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry. The minimum inhibitory concentration (MIC) was determined by microbroth dilution method. The common carbapenemase resistant genes and virulence factors were identified by PCR. Homology analysis was performed by multilocus sequencing typing. Whole genome sequencing was performed to analyze the genomic characteristics of CRKP without carbapenemase. Results: The 131 strains of CRKP showed resistance to common antibiotics, except for polymyxin B (1.6% resistance rate) and tigacycline (8.0% resistance rate). A total of 105 (80.2%) CRKP strains carried the Klebsiella pneumoniae carbapenemase (KPC) resistance gene, 15 (11.4%) strains carried the New Delhi Metallo-ß-lactamase (NDM) gene, and 4 (3.1%) isolates carried both KPC and NDM genes. Sequence typing (ST) 11 (74.0%) was the dominant sequence type. High detection rates for mrkD (96.2%), fimH (98.5%), entB (100%), and other virulence genes were reported. One hypervirulent CRKP strain was detected. The seven strains of CRKP that did not produce carbapenemase were shown to carry ESBL or AmpC genes and had anomalies in membrane porins OMPK35 and OMPK36, according to whole genome sequencing. Conclusion: In a large-scale tertiary-care general hospital, CRKP mainly carries the KPC gene, has a high drug resistance rate to a variety of antibiotics, and possesses multiple virulence genes. Attention should be paid to CRKP strains with high virulence.

Enterobacterales resistentes a carbapenemes: estudio epidemiológico de aislamientos en un hospital público de Santa Fe / Carbapenem-resistant Enterobacterales: Epidemiological study of isolates in a public hospital in Santa Fe

Introducción: La incidencia de Enterobacterales resistentes a carbapenemes (ERC) se elevó en la última década, y en especial durante la pandemia de COVID-19. Objetivo: Conocer el perfil de resistencia antimicrobiana, así como la frecuencia y tipo de carbapenemasas presen-tes en los aislamientos de ERC en un hospital regional.Materiales y métodos: Estudio epidemiológico, observa-cional y retrospectivo. Incluyó ERC aislados en muestras clínicas durante 2021 en un hospital regional de Santa Fe, Argentina. El cálculo de la incidencia (aislamientos/pacien-tes-día) e intervalo de confianza 95% (IC 95%), y las pruebas estadísticas se realizaron con OpenEpi.Resultados: 348 ERC aislados (11,9 aislamientos/1000 pacientes-día; IC95% 10,7-13,2). La incidencia se correla-cionó con los casos de COVID-19 (rho=0,874, p<0,001) y fue a expensas de la Unidad de Cuidados Intensivos (76,4%). El principal ERC aislado fue Klebsiella pneumoniae (71,4%, n=260). KPC fue el principal mecanismo de resistencia (61,2%). Se aislaron dos doble productores de carbapene-masas. La tasa global de resistencia a los antibióticos no betalactámicos evaluados fue superior en Klebsiella pneu-moniae que en el resto de los aislamientos resistentes a carbapenemes (60,6% vs. 38,5%, p<0,001). En KPC hubo mayor resistencia a colistin (44,6% vs. 23,9%, p=0,001) y menor a amikacina (23,9% vs. 72,6%, p<0,001).Conclusión: Frente a las escasas opciones terapéuticas en infecciones por ERC se destaca la importancia de conocer los mecanismos de resistencia implicados y la epidemiología local

Introduction: The incidence of carbapenem-resistant Enterobacterales (CRE) rose in the last decade, and especially during the COVID-19 pandemic.Objective: To identify the antimicrobial resistance profile, as well as the frequency and type of carbapenems that were present in CRE isolations in a tertiary care hospital.Materials and methods: Epidemiological, observational and retrospective study. It included CRE isolated in clinical samples during 2021 in a tertiary care hospital in Argentina. Incidences (isolations/patients-day), confidence intervals of 95% (CI 95%) and statistical comparisons were made with OpenEpi.Results: 348 CRE were isolated (11.9 isolations/1,000 patients-day, IC95% 10.7-13.2). Incidence correlated to COVID-19 cases (rho=0.874, p<0.001). Most isolations were from the Intensive Care Unit (76.4%) and the from respiratory samples (27.6%, n=96) and blood cultures (24.4%, n=92). The main isolated CRE was Klebsiella pneumoniae (71.4%, n=260), with a general carbapenem resistance of 53.4%. KPC was the main resistance mechanism (61.2%). Two double carbapenemase-producing Enterobacterales were isolated. Klebsiella pneumoniae presented a higher overall resistance rate to non-betalactam antibiotics (60.6% vs 38.5%, p<0.001). Among CRE, a higher colistin resistance rate was found in KPC isolations (44.6% vs 23.9%, p=0.001) and lower resistance to amikacin (23.9% vs 72.6%, p<0.001).Conclusion: The difficulty in the selection of antibiotic regimens for CRE forces the treating physicians to put emphasis on the knowledge of resistance mechanisms to optimize them

Diagnostic Performance of BD Phoenix CPO Detect Panels for Detection and Classification of Carbapenemase-Producing Gram-Negative Bacteria.

BD Phoenix CPO Detect panels can identify and classify carbapenemase-producing organisms (CPOs) simultaneously with antimicrobial susceptibility testing (AST) for Gram-negative bacteria. Detection and classification of carbapenemase producers were performed using the BD Phoenix CPO Detect panels NMIC/ID-441 for Enterobacterales, NMIC/ID-442 for nonfermenting bacteria, and NMIC-440 for both. The results were compared with those obtained using comparator methods. A total of 133 strains (32 Klebsiella pneumoniae, 37 Enterobacter cloacae complex, 33 Pseudomonas aeruginosa, and 31 Acinetobacter baumannii complex strains), including 60 carbapenemase producers (54 imipenemases [IMPs] and 6 OXA type), were analyzed. Using panels NMIC-440 and NMIC/ID-441 or NMIC/ID-442, all 54 IMP producers were accurately identified as CPOs (positive percent agreement [PPA], 100.0%; 54/54). Among the 54 IMP producers identified as CPOs using panels NMIC-440 and NMIC/ID-441, 12 and 14 Enterobacterales were not resistant to carbapenem, respectively. Among all 54 IMP producers, 48 (88.9%; 48/54) were correctly classified as Ambler class B using panel NMIC-440. Using panels NMIC-440 and NMIC/ID-442, all four OXA-23-like carbapenemase-producing A. baumannii complex strains (100.0%, 4/4) were correctly identified as CPOs, and three (75.0%, 3/4) were precisely classified as class D using panel NMIC-440. Both carbapenemase producers harboring the blaISAba1-OXA-51-like gene were incorrectly identified as non-CPOs using panels NMIC-440 and NMIC/ID-442. For detecting carbapenemase producers, the overall PPA and negative percent agreement (NPA) between panel NMIC-440 and the comparator methods were 96.7% (58/60) and 71.2% (52/73), respectively, and the PPA and NPA between panels NMIC/ID-441 or NMIC/ID-442 and the comparator methods were 96.7% (58/60) and 74.0% (54/73), respectively. BD Phoenix CPO Detect panels can successfully screen carbapenemase producers, particularly IMP producers, regardless of the presence of carbapenem resistance and can be beneficial in routine AST workflows. IMPORTANCE Simple and efficient screening methods of detecting carbapenemase producers are required. BD Phoenix CPO Detect panels effectively screened carbapenemase producers, particularly IMP producers, with a high overall PPA. As the panels enable automatic screening for carbapenemase producers simultaneously with AST, the workflow from AST to confirmatory testing for carbapenemase production can be shortened. In addition, because carbapenem resistance varies among carbapenemase producers, the BD Phoenix CPO Detect panels, which can screen carbapenemase producers regardless of carbapenem susceptibility, can contribute to the accurate detection of carbapenemase producers. Our results report that these panels can help streamline the AST workflow before confirmatory testing for carbapenemase production in routine microbiological tests.

Epidemiological characteristics and molecular features of carbapenem-resistant Enterobacter strains in China: a multicenter genomic study.

Epidemiological characteristics and molecular features of carbapenem-resistant Enterobacter (CR-Ent) species remain unclear in China. In this study, we performed a genomic study on 92 isolates from Enterobacter-caused infections from a multicenter study in China. Whole genome sequencing (WGS) was used to determine the genome sequence of 92 non-duplicated CR-Ent strains collected from multiple tertiary health centres. The precise species of Enterobacter strains were identified by average nucleotide identity (ANI) and in silico DNA-DNA hybridization (isDDH). Molecular features of high-risk CR-Ent sequence type (ST) lineages and carbapenemase-encoding plasmids were determined. The result revealed that the most common human-source CR-Ent species in China was E. xiangfangensis (66/92, 71.93%), and the proportion of carbapenemase-producing Enterobacter (CP-Ent) in CR-Ent was high (72/92, 78.26%) in comparison to other global regions. Furthermore, ST171 and ST116 E. xiangfangensis were the major lineages of CP-Ent strains, and ST171 E. xiangfangensis was more likely to cause infections in older patients. Genomic analysis also highlighted the likelihood of intra-hospital/inter-hospital clonal transmission of ST171 and ST116 E. xiangfangensis. In addition, the blaNDM-harbouring IncX3-type plasmid was identified as the prevalent carbapenemase-encoding plasmid carried by CR-Ent strains, and was experimentally confirmed to be able to self-transfer with high frequency. This study detailed the genomic and clinical characteristics of CR-Ent in China in the form of multicenter for the first time. The high risk of carbapenemase-producing ST171 and ST116 E. xiangfangensis, and the blaNDM-harbouring IncX3-type plasmid were detected and emphasized.

Phenotypic and genotypic distribution of ESBL, AmpC ß-lactamase and carbapenemase-producing Enterobacteriaceae in community-acquired and hospital-acquired urinary tract infections in Sri Lanka.

OBJECTIVES: Although Sri Lanka belongs to a region with a high prevalence of extended-spectrum ß-lactamase (ESBL), AmpC ß-lactamase and carbapenemase-producing Enterobacteriaceae, data regarding antimicrobial resistance (AMR) is limited. We studied the prevalence and diversity of ß-lactamases produced by Enterobacteriaceae urinary pathogens from two hospitals in the Western Province of Sri Lanka. METHODS: ESBL, AmpC ß-lactamase and carbapenemase production was detected by phenotypic testing followed by genotyping. RESULTS: The species responsible for urinary tract infections (UTI) were Escherichia coli (69%), Klebsiella pneumoniae (16%) and Enterobacter sp (6%). The prevalence of ESBL (50%), AmpC ß-lactamase (19%) and carbapenemase (11%) phenotypes was high, and greater in hospital-acquired (HA-UTI) (75%) than in community-acquired UTI (CA-UTI) (42%). Identification of CA-UTI caused by carbapenemase-producing Enterobacteriaceae (5%) is alarming. Only one ESBL gene, blaCTX- M-15, was detected. AmpC ß-lactamase genes found in E. coli and K. pneumoniae were blaCMY-2, blaCMY-42 and blaDHA-1, while Enterobacter sp. carried blaACT-1. Carbapenemase genes were blaNDM-1, blaNDM-4, blaOXA-181 and blaOXA-232, while blaKPC, blaIMP and blaVIM were absent. Co-occurrence of multiple bla genes, with some isolates harbouring six different bla genes, was common. Carbapenem-resistant isolates without carbapenemase genes displayed mutations in the outer membrane porin genes, ompF of E. coli and ompK36 of K. pneumoniae. Factors associated with UTI with ß-lactamase-producing Enterobacteriaceae were age ≥50 years, previous hospitalization, presence of an indwelling urinary catheter, history of diabetes mellitus or other chronic illness and recurrent urinary tract infections. CONCLUSION: This study adds to the currently scarce data on AMR in Sri Lanka.

Prevalence of Carbapenem-Resistant Hypervirulent Klebsiella pneumoniae and Hypervirulent Carbapenem-Resistant Klebsiella pneumoniae in China Determined via Mouse Lethality Tests.

Objective: To investigate the epidemiology of carbapenem-resistant hypervirulent Klebsiella pneumoniae (CR-HvKP) and hypervirulent carbapenem-resistant Klebsiella pneumoniae (Hv-CRKP). Methods: Totally 436 K. pneumoniae strains were collected from 7 hospitals in mainland China between 2017.01 and 2018.02. Sequence types, serotypes, antimicrobial-resistance and virulence genes were analyzed. Additionally, string test, capsule stain, Periodic Acid Schiff stain, fitness analysis, quantitative real-time PCR and mouse lethality test were also performed. Molecular combinations were used to screen putative blaKPC(+)-HvKP and Hv-blaKPC(+)-KP, followed by the confirmation of mouse lethality test. Results: Diverse detection rates were found for the virulence genes, ranging from c-rmpA (0.0%) to entB (100.0%). According to the molecular criteria, 127, 186, 9 and 26 strains were putatively denoted as HvKP, blaKPC(+)-KP, blaKPC(+)-HvKP and Hv-blaKPC(+)-KP. Mouse lethality test confirmed 2 blaKPC(+)-HvKP strains (JS184 and TZ20) and no Hv-blaKPC(+)-KP. JS184 showed K2 serotype, thin capsule, positive exopolysaccharid and string test. TZ20 presented K20 serotype, thin capsule, negative exopolysaccharide and string test. Compared with the positive control NTUH-K2044, equal galF expression and growth curves were confirmed for JS184 and TZ20. Conclusions: Molecular determination of CR-HvKP and Hv-CRKP brings remarkable bias compared with mouse lethality test. The exact prevalence of CR-HvKP is less than 1.0%, which of Hv-CRKP is much lower.

Rising incidence of carbapenem resistant isolates: an Argentinian hospital's experience. More trouble in the aftermath of the COVID-19 pandemic / Incidencia creciente de aislamientos resistentes a carbapenémicos: la experiencia de un hospital argentino. Más problemas tras la pandemia de COVID-1

Introduction: During COVID-19 outbreaks, disproportionate use of antibiotics, high Intensive Care Units burden and longer in-hospital stays may have aggravated the emergency posed by carbapenem-resistant isolates. Therefore, we set out to determine whether the incidence of carbapenem-resistant isolates rose in a tertiary care center in Santa Fe, Argentina during the period with active cases of COVID-19. Material and methods: In this retrospectively designed analytic epidemiologic study, two periods were defined: Period 1 (without active cases of COVID-19) from September 2019 to August 2020 and Period 2 (starting at the onset of the first wave of COVID-19 in this Institution) from September 2020 to June 2021. All clinically relevant microbiological samples taken during these periods in the Internal Medicine, Surgical and Intensive Care Unit wards were included. The primary analysis of interest was the differential incidence between the two periods, overall and in the Intensive Care Units wards in particular. Results: 9,135 hospitalizations, 50,145 patient-days of analysis. 7,285 clinical samples were taken, with an overall positivity for carbapenem-resistant isolates of 12.1% (n=883). Overall carbapenem-resistant isolates incidence during Period 2 was 2.5 times higher than in Period 1 (2.52 vs 0.955/100 patient-days, p<0.001). Intensive Care Units' carbapenem-resistant isolates incidence raised from 6.78 to 8.69/100 patient-days in Period 2 (p=0.006). Conclusions: We found alarming rates of carbapenem-resistant isolates in our center, 2.5 times higher in the period following the first wave of COVID-19. This rise was due to a higher amount of clinically relevant microbiological samples taken and to a higher carbapenem resistance among Enterobacteria and non-fermentative Gram-negative bacilli. To our knowledge, this is one of the few Latin-American reports on the effect of the COVID-19 pandemic on carbapenem-resistant isolates incidence (AU)

Introducción: Durante los brotes de COVID-19, el uso desproporcionado de antibióticos, la alta carga de las Unidades de Cuidados Intensivos y las estancias hospitalarias más prolongadas pueden haber agravado la emergencia planteada por los aislados resistentes a carbapenémicos. Por lo tanto, nos propusimos determinar si la incidencia de aislamientos resistentes a carbapenémicos aumentó en un centro de tercer nivel de atención en Santa Fe, Argentina, durante el período con casos activos de COVID-19. Material y métodos: En este estudio epidemiológico analítico de diseño retrospectivo se definieron dos periodos: Periodo 1 (sin casos activos de COVID-19) de septiembre de 2019 a agosto de 2020, y Periodo 2 (a partir del inicio de la primera ola de COVID-19. en esta Institución) desde septiembre de 2020 hasta junio de 2021. Se incluyeron todas las muestras microbiológicas clínicamente relevantes tomadas durante estos períodos en las salas de Medicina Interna, Quirúrgica y Unidad de Cuidados Intensivos. El principal análisis de interés fue la incidencia diferencial entre los dos períodos, en general y en las salas de las Unidades de Cuidados Intensivos en particular. Resultados: 9.135 hospitalizaciones, 50.145 pacientes-día de análisis. Se tomaron 7.285 muestras clínicas, con una positividad global para aislados resistentes a carbapenémicos del 12,1% (n=883). La incidencia general de aislamientos resistentes a carbapenémicos durante el Período 2 fue 2,5 veces mayor que en el Período 1 (2,52 frente a 0,955/100 pacientes-día, p<0,001). La incidencia de aislamientos resistentes a carbapenémicos en Unidades de Cuidados Intensivos aumentó de 6,78 a 8,69/100 pacientes-día en el Período 2 (p=0,006). Conclusiones: Encontramos tasas alarmantes de aislamientos resistentes a carbapenémicos en nuestro centro, 2,5 veces mayores en el período posterior a la primera ola de COVID-19 (AU)

Carbapenem-resistant Enterobacteriaceae in sink drains of 40 healthcare facilities in Sindh, Pakistan: A cross-sectional study.

In Pakistan, antimicrobial resistance (AMR) is expected to greatly increase the already high mortality and morbidity rates attributed to infections, making AMR surveillance and prevention a priority in the country. The aims of the project were to characterize the prevalence of carbapenem-resistant Enterobacteriaceae (CRE) in healthcare facility sink drains in Pakistan and to characterize how physical characteristics of sinks and healthcare facility rooms were associated with CRE in those sinks. The study took place in 40 healthcare facilities in Jamshoro Pakistan. Swabs were collected from sink drains in each facility that had a sink, and structured observations of sinks and facilities were performed at each facility. Swabs were plated on CHROMagar KPC to screen for carbapenem-resistant Enterobacteriaceae, which were then isolated on Mueller-Hinton agar plates. Antibiotic susceptibility was determined using the disk diffusion method to assess resistance to carbapenems, cephalosporins, and fluoroquinolones. Thirty-seven of the healthcare facilities had at least one sink, and thirty-nine total sinks were present and sampled from those healthcare facilities. Sinks in these facilities varied in quality; at the time of sampling 68% had water available, 51% had soap/alcohol cleanser at the sink, 28% appeared clean, and 64% drained completely. Twenty-five (64%) of the sink samples grew Enterobacteriaceae on CHROMagar KPC, sixteen (41%) of which were clinically non-susceptible to ertapenem. Seven of the 39 sampled sinks (18%) produced Enterobacteriaceae that were resistant to all three antibiotic classes tested. Several facilities and sink characteristics were associated with CRE. Sinks and drains can serve as undetected reservoirs for carbapenem-resistant Enterobacteriaceae. Control and remediation of such environments will require both systemic strategies and physical improvements to clinical environments.

Emergence of High Prevalence of Extended-Spectrum Beta-Lactamase and Carbapenemase-Producing Enterobacteriaceae Species among Patients in Northwestern Ethiopia Region.

BACKGROUND: World Health Organization identified some Enterobacteriaceae as superbugs because of their high production and spread of extended-spectrum beta-lactamases (ESBL) and carbapenemases. Moreover, their resistance against commonly prescribed antibiotics left few choices of drugs to treat infection. This study is aimed at determining the magnitude of ESBL and carbapenemase-producing Enterobacteriaceae pathogens and their antimicrobial resistance pattern. MATERIALS AND METHODS: A hospital-based cross-sectional study was carried out from February to April 2019 in the Northwestern Ethiopia region. A total of 384 patients presumptive for bacterial infections were conveniently enrolled in the study. Specimens were collected and processed following standard bacteriological procedures. Drug susceptibility tests were performed using disk diffusion technique. ESBL and carbapenemase enzymes were tested by double disk diffusion and modified carbapenem inhibition methods, respectively. The data obtained were analyzed using SPSS version 22 software, and descriptive statistics were summarized in tables and graphs. RESULTS: Out of 384 clinical specimens processed 100 (26%) were culture positive for Enterobacteriaceae. The proportion of Enterobacteriaceae infection was relatively higher among in-patients 86 (32.6%) than out-patients 14 (11.7%). Overall, Escherichia coli 35 (9.1%) was the leading isolate followed by Klebsiella pneumoniae 31 (8.1%). Klebsiella pneumoniae 15 (15.6%) was the most frequent isolate from bloodstream infection and is the leading isolate from intensive care unit patients 15 (38.3%). Overall, 44 (44%) of Enterobacteriaceae were extended-spectrum beta-lactamase producers. Among them, Citrobacter spp. was the leading one 4 (80%) followed by Enterobacter cloacae 6 (60%) and K. pneumoniae 18 (58.1%). Furthermore, 6 (6%) of Enterobacteriaceae were carbapenemase-producers, in which 5 (50%) of E. cloacae and 3 (9.7%) of K. pneumoniae had highest percentage. Conclusions. ESBL and carbapenemase-producing isolates of Enterobacteriaceae are alarmingly spreading in the study area. Thus, improving the infection prevention strategy and further screening at the national level is recommended to develop the optimal use of antibiotics.

Methodology Establishment and Application of VITEK Mass Spectrometry to Detect Carbapenemase-Producing Klebsiella pneumoniae.

The ability of VITEK mass spectrometry (MS) in detection of bacterial resistance is currently under exploration and evaluation. In this study, we developed and validated a VITEK MS method to rapidly test carbapenemase-producing Klebsiella pneumoniae (CPKP). Solvents, antibiotic concentrations, crystal conditions and times, centrifugation speeds, and other factors were optimized to design a rapid sample pretreatment process for CPKP detection by VITEK MS. The related parameters of the mass spectrum were adjusted on the instrument to establish an CPKP detection mode. 133 clinically isolated strains of CPKP in the microbiology laboratory at the Shenzhen People's Hospital from 2004 to 2017 were selected for accuracy evaluation. The fresh suspected strains from the microbiology laboratory in 2020 were used to complete the clinical verification. Two antibiotics, meropenem (MEM) and imipenem (IPM), were used as substrates. These two substrates were incubated with suspected CPKP, and the results were obtained by VITEK MS detection. Using this method, different types of CPKP showed different detection results and all the CPKP strains producing KPC-2 and IMP-4 carbapenemase were detected by VITEK MS. Thus, VITEK MS can be used for rapid detection of CPKP, especially for some common types of CPKP. This method provides high accuracy and speed of detection. Combined with its cost advantages, it can be intensely valuable in clinical microbiology laboratories after the standard operating procedures are determined.

The importance of active surveillance of carbapenem-resistant Enterobacterales (CRE) in colonization rates in critically ill patients.

OBJECTIVE: This study aimed to demonstrate the importance of active carbapenem-resistant Enterobacterales (CRE) surveillance and evaluate the prevalence of invasive infections, risk factors, and mortality risk in CRE-colonized patients. METHODS: Retrospective cohort study analyzing 1,920 patients identified using an active CRE surveillance protocol, admitted to an adult intensive care unit in southeastern Brazil from January 2014 to December 2018. RESULTS: There were 297 (15.47%) CRE colonized patients, with one colonized for every six control patients. CRE-colonized patients demonstrated an increased chance of infection (odds ratio [OR] 7.967, p < 0.001). Overall, 20.54% of the colonized patients presented invasive infection (81.96% due to Klebsiella pneumoniae). The colonization and infection ratio demonstrated the important role of the active CRE surveillance protocol. There were identified multiple risk factors for CRE colonization, including long-term mechanical ventilation (OR 1.624, p = 0.019) and previous exposure to aminopenicillins (OR 5.204, p < 0.001), carbapenems (OR 3.703, p = 0.017), cephalosporins (OR 12.036, p < 0.001), and fluoroquinolones (OR 5.238, p = 0.012). The mortality risk was significantly higher among colonized (OR 2.356, p < 0.001) and colonized-infected (OR 2.000, p = 0.009) patients and in those with Enterobacter cloacae colonization (OR 5.173, p < 0,001) and previous aminopenicillins exposure (OR 3.452, p = 0.007). CONCLUSIONS: Early detection of CRE colonization through screening testing proved to be an important tool to control CRE spread. However, observation over the years has shown no effective control of colonization and infection. The prevalence rates of CRE colonization and colonization-infection were high, as were the mortality rates. In conclusion, an active CRE surveillance protocol is essential, but its impact depends on the effective implementation of preventive measures and feedback between team members.

The Magnitude of Carbapenemase and ESBL Producing Enterobacteriaceae Isolates from Patients with Urinary Tract Infections at Tikur Anbessa Specialized Teaching Hospital, Addis Ababa, Ethiopia.

BACKGROUND: The emergence of multidrug-resistant organisms, such as extended-spectrum beta-lactamase-producing Enterobacteriaceae (ESBL-PE), and carbapenemase-producing Enterobacteriaceae (CPE) is a public health concern. Therefore, this study aimed to determine the magnitude of carbapenemase and ESBL producing bacteria isolated from patients affected by Urinary Tract Infection (UTI). METHODS: A cross-sectional study was conducted from December 2018 to March 2019 at Tikur Anbessa Specialized Hospital. A total of 120 Enterobacteriaceae isolates from UTI patients were collected and identified on species level using standard microbiological methods. Antimicrobial susceptibility test was determined according to the guidelines of the Clinical and Laboratory Standards Institute. Detection of ESBL production was carried out by using ESBL ChromoSelect Agar medium and the combined disk diffusion. Production of carbapenemase was determined by using Hodge-test and modified carbapenem inactivation method as described in CLSI guidelines. RESULTS: Out of the total 120 Enterobacteriaceae isolates, 74 (61.7%) were ESBL-producers, and 8 (6.7%) were carbapenemase producers. The most common ESBL producing isolate was E.coli 38 (51.4%) and the most common carbapenemase-producing isolate was K.pneumoniae five (62.5%). Most of the ESBL and carbapenemase-producing isolates were recovered from hospitalized patients 46 (62.2%) and 7 (87.5%) respectively. The rate of ESBL and CPE production was observed high among patients taking antibiotics 64.8% (59/91) and 7.7% (7/91) respectively, but no significant association was observed p > 0.05. Furthermore, about 1.7% (2/120) isolates were found both ESBL and carbapenemase producers. Significant resistances rates were observed in ESBL and CPE isolates. CONCLUSION: Enterobacteriaceae isolates showed a significantly higher rate of ESBL production. A significant figure of carbapenemase production was observed among Enterobacteriaceae isolates causing UTI. The production of ESBL and CPE enhanced for an increased rate of MDR patterns. Efforts need to be made to introduce a system for tracking and detecting ESBL-PE and CPE-producing bacteria in hospitals, and monitoring dissemination of ESBL and CPE-producing Enterobacteriaceae is strongly recommended.

Clinical and molecular characteristics and risk factors for patients acquiring carbapenemase-producing and non-carbapenemase-producing carbapenem-nonsusceptible-Enterobacterales bacteremia.

BACKGROUND/PURPOSE: Carbapenem-nonsusceptible Enterobacterales (CNSE) are a growing global threat. Carbapenemases are often produced by plasmids, which allow rapid transmission. This study aimed to investigate (1) the bacterial type (2) resistant genes (3) antimicrobial susceptibility and (4) risk factors for acquisition of carbapenemase-producing carbapenem-nonsusceptible Enterobacterales (CP-CNSE) and non-carbapenemase-producing carbapenem-nonsusceptible Enterobacterales (non-CP-CNSE) bacteremia. METHODS: There were a total of 113 isolates of Enterobacterales from 2013 to 2018. After excluding nonblood isolates and including only one sample from each patient, 99 isolates were analyzed and the medical charts of these patients were reviewed. Carbapenemase genes, ß-lactamase genes and antimicrobial susceptibility of the isolates were determined. Multilocus sequence typing (MLST) was performed on CP-CNSE isolates. RESULTS: CP-CNSE carried more blaSHV (P = 0.004) and were more resistant to imipenem than non-CP-CNSE (P < 0.001). In the univariate analyses, we found that CP-CNSE bloodstream infection was associated with patient <65 years of age (odds ratio, 3.90; 95% confidence interval [CI], 1.16 to 13.10; P = 0.027), mechanical ventilation at the time of bloodstream infection (BSI) (odds ratio, 3.85; 95% CI, 1.16-12.78; P = 0.028) and exposure to piperacillin/tazobactam (odds ratio, 3.96; 95% CI, 1.09-14.38; P = 0.037). However, on multivariate analyses, no independent predictor for CP-CNSE was identified in this study. CONCLUSION: CP-CNSE carried more blaSHV and were more resistant to imipenem when compared to non-CP-CNSE. No independent predictor for CP-CNSE was identified after multivariate analysis. This is the first study conducted in Taiwan comparing risk factors between CP-CNSE and non-CP-CNSE from both clinical and molecular aspects.

Rapid detection of KPC-producing Klebsiella pneumoniae in China based on MALDI-TOF MS.

Klebsiella pneumoniae carbapenemase-producing K. pneumoniae (KPC-Kp) represent a serious threat to public health and their timely detection is essential for patient management and the prevention of nosocomial infections. Here, matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) was used to rapidly identify dominant KPC-Kp in China, by using an automated detection of a KPC-specific peak (at 4521 m/z) by a genetic algorithm using ClinProTools software. Whole-genome sequencing (WGS) was used to understand the genetic environment of the blaKPC-2 gene. In this study, we analyzed 235 K. pneumoniae Chinese clinical isolates, of which 175 (93 KPC-positive isolates and 82 KPC-negative isolates) isolates were used to build a model to select a KPC-specific peak, and another 60 isolates for external validation. In addition, all the spectra were visually inspected by the FlexAnalysis software to evaluate the accuracy of the automated detection. The results showed a 4521 m/z peak found in all blaKPC-2-positive isolates but absent in blaKPC-2-negative isolates. Interestingly, all KPC-Kp belonged to ST11, the dominant clone in China. WGS analysis of a representative isolate showed that the genetic environment of KPC-2 was IS26-ISKpn27-blaKPC-2-ΔISKpn6-Tn1721, similar to the KPC-2 genetic environment of ST11 KPC-Kp previously reported in China. Therefore, the 4521 m/z peak is closely related to ST11 KPC-Kp. In summary, we used MALDI-TOF MS to quickly detect KPC-Kp in the process of routine bacterial identification without increasing costs or requiring further knowledge, which has broad application prospects in drug resistance analysis and infection control.

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.

Distribution of ß-lactamases and emergence of carbapenemases co-occurring Enterobacterales isolates with high-level antibiotic resistance identified from patients with intra-abdominal infection in the Asia-Pacific region, 2015-2018.

PURPOSE: In this study, we aimed to assess the geographic distribution and molecular characteristics of ß-lactamases among Enterobacterales isolates causing intra-abdominal infections (IAIs) from 2015 to 2018 in the Asia-Pacific region. METHOD: Isolates were investigated for extended-spectrum ß-lactamases (ESBLs), AmpC ß-lactamases, and carbapenemases using multiplex PCR assays and full-gene DNA sequencing. RESULT: A total of 832 Enterobacterales isolates from 8 different countries with ß-lactamase genes were analysed. Plasmid-mediated ESBLs and AmpC ß-lactamases were encoded in 598 (71.9 %) and 314 (37.7 %) isolates, respectively. In 710 (85.3 %) carbapenemase-negative isolates, positivity for both AmpC ß-lactamases and ESBLs was identified in 51 (8.5 %) Escherichia coli and 24 (3.4 %) Klebsiella pneumoniae isolates. The most prevalent countries were Taiwan and Vietnam, and the co-occurrence of CMY/CTX-M in E. coli and DHA-1/ESBLs in K. pneumoniae was predominant. All isolates showed high susceptibility to colistin, but susceptibility to carbapenems varied among different resistance mechanism combinations. Among 122 (14.7 %) isolates encoding carbapenemase, NDM (n = 67, including 64.2 % NDM-1) was the most common, followed by the OXA-48-type (n = 49), KPC (n = 24) and IMP (n = 4). The most prevalent country was Thailand (n = 44), followed by Vietnam (n = 35) and the Philippines (n = 21). Twenty-two isolates were found to encode multiple carbapenemases, 16 of which were collected from Thailand and harbored NDM-1, OXA-232 and CTX-M-15. Despite high susceptibility to amikacin, susceptibility to colistin was only 56 %. CONCLUSION: The emergence of carbapenem-non-susceptible AmpC/ESBL co-occurring Enterobacterales and colistin non-susceptible carbapenemases co-occurring K. pneumoniae highlights potential therapeutic challenges in the Asia-Pacific region.

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.

Transmission of NDM-5-Producing and OXA-48-Producing Escherichia coli Sequence Type 648 by International Visitors without Previous Medical Exposure.

Carbapenemase-producing Escherichia coli sequence type (ST) 648 strains were isolated from two international visitors without previous medical exposure from Southeast Asian countries in a hospital in Japan. One isolate, FUJ80154, carried blaNDM-5 in a complex class 1 integron on an IncFIB/FII plasmid; the other isolate, FUJ80155, carried two copies of blaOXA-48 on the chromosome flanked by IS1R on both sides. The core-genome based-phylogenetic analysis with publicly available genome data of E. coli ST648 carrying blaNDM-5 or blaOXA-48-like demonstrated high genetic similarity between FUJ80154 and NDM-5-prooducing E. coli ST648 strains isolated in South and Southeast Asian countries. On the other hand, no closely related isolates of FUJ80155 were identified. In the absence of prior hospitalization overseas, neither patient had qualified for routine screening of multidrug-resistant organisms, and the isolates were incidentally identified in cultures ordered at the discretion of the treating physician. IMPORTANCE Although patients with history of international hospitalization are often subject to screening for multidrug-resistant organisms, it is unclear whether patients who reside in countries where carbapenemase-producing Enterobacterales (CPE) is endemic but have no history of local hospitalization contribute to the transmission of CPE. In this study, NDM-5-producing and OXA-48-producing Escherichia coli sequence type (ST) 648, a recently recognized high-risk, multidrug-resistant clone, were detected from two overseas visitors without previous medical exposure. The findings of this study suggest that active surveillance culture on admission to hospital may be considered for travelers from countries with endemicity of carbapenem-resistant organisms even without history of local hospitalization and underscore the need to monitor cross-border transmission of high-risk clones, such as carbapenemase-producing E. coli ST648.