Carbapenemase Enzymes Research Articles

Molecular detection of blaOXA-23 gene and blaOXA-51 gene in carbapenem resistant strains of Acinetobacter baumannii in patients with ventilator associated pneumonia at tertiary care hospitals.

To evaluate carbapenem resistance and to detect blaOXA-23 and blaOXA-51 genes in carbapenem-resistant Acinetobacter baumanii isolates recovered from patients having pneumonia secondary to ventilation. The cross-sectional study was conducted from July 2017 to June 2018 at the Department of Microbiology, University of Health Sciences, Lahore, Pakistan, and comprised endotracheal aspirates / tracheobroncheal lavage samples from patients irrespective of age and gender who developed pneumonia after being on the ventilator for 48 hrs at the Combined Military Hospital, and Jinnah Hospital, Lahore. The samples were inoculated on MacConkey and blood agar and aerobically incubated at a temperature of 370C for 18-24 hours. The isolated organisms were further assessed by standard morphological, cultural and biochemical profile. Antibiotic susceptibility was done by Kirby-Bauer disc diffusion method. Carbapenem-resistant Acinetobacter baumannii were checked for carbapenemase production using Modified Hodge Test. Conventional polymerase chain reaction and agarose gel electrophoresis were performed to detect blaOXA-23 and blaOXA-51 genes. Data was analysed using SPSS 17. Out of 157 samples, 92(58.6%) yielded growth of bacteria, and, among them, 39(42.4%) were identified as Acinetobacter baumannii. All (100%) Acinetobacter baumannii cases showed resistance to carbapenem, were producing carbapenemase enzyme, and were positive for blaOXA-51 gene. The blaOXA-23 gene was amplified in 38(97.4%) isolates. BlaOXA-23 gene appeared to be the major cause of carbapenem resistance.

Phenotypic and molecular characterization of antimicrobial resistance in clinical species of Enterobacter, Serratia, and Hafnia in Northeast Iran

PurposeExtended-spectrum beta-lactamases (ESBLs) and carbapenemase enzymes are the most common and significant mechanisms for bacterial resistance to extended-spectrum cephalosporins and carbapenems, respectively. The present study was conducted to investigate the prevalence and molecular epidemiology of ESBL coding genes and carbapenem-resistant genes among clinical isolates of Enterobacter, Serratia, and Hafnia species in Northeast Iran. MethodsA total of 143 clinical samples suspected as Enterobacter species were collected from three hospitals in Mashhad (northeast of Iran) during November–January 2018–2020. Bacterial identification was done using VITEK 2 system. Antimicrobial susceptibility was determined by the Kirby–Bauer disk diffusion test. Molecular epidemiology of ESBLs and carbapenemase coding genes were assessed by PCR. ResultsResults of VITEK 2 system showed that Serratia marcescens was the predominant isolate (n = 37, 25.87%) followed by Enterobacter cloacae (n = 30, 20.97%), Cronobacter sakazakii (n = 26, 18.18%), and Hafnia alvei (n = 16, 11.19%). The evaluation of antimicrobial susceptibility showed high resistance rates to trimethoprim/sulfamethoxazole, ampicillin-sulbactam, and ceftriaxone (59.44%, 55.24%, and 48.25%, respectively). Phenotypic ESBLs were positively detected in 52 isolates [36.4%]. Moreover, eight isolates (5.59%) were resistant to carbapenems. The frequency of the genes blaTEM, blaSHV, blaCTX-M15, was 71.15%, 23.08%, and 80.77%, respectively, in the 52 ESBL-producing isolates, while the frequency of the blaNDM gene was 25% (2/8) in carbapenem resistant-isolates. ConclusionsA Highly frequency distribution of ESBL producer clinical bacterial species such as Serratia, and Enterobacter may be has been acquired by horizontal gene transfer phenomenon from other bacteria and this issue become an important public health issue.

Occurrence of Multiple, Extensive and Pan Drug-Resistant Pseudomonas aeruginosa and Carbapenemase Production from Presumptive Isolates Stored in a Biobank at Ethiopian Public Health Institute.

PurposePseudomonas aeruginosa is a common cause of nosocomial infections with associated morbidity and mortality because the organism is unresponsive to commonly available antimicrobials. This study was undertaken to determine the multiple drug-resistant (MDR), extensive drug-resistant (XDR) and pan drug-resistant (PDR) phenotype of P. aeruginosa and its carbapenemase production rate from presumptive isolates stored in the biobank at the Ethiopian Public Health Institute (EPHI).MethodsA cross-sectional study was conducted at the EPHI laboratory, Addis Ababa, Ethiopia from March to June 2021. Stored isolates of Pseudomonas spp. which had been characterized by manual identification methods were further processed for species-level identification (ID) and antimicrobial susceptibility testing (AST) using a Becton Dickinson (BD) Phoenix automated system. The isolates were analyzed for carbapenemase enzyme production using the modified Carbapenem Inactivation Method (mCIM). The data analysis was done using SPSS version 20 software.ResultsIn this study, 110 presumptive Pseudomonas isolates from a biobank were re-analyzed, 100 of them were found to be Pseudomonas and among these P. aeruginosa accounted for 98% and P. putida accounted for 2%. The majority of isolates were recovered from wound (46%) specimens followed by ear swabs (18%). The highest level of resistance was observed against ceftazidime (35%) and the lowest level of resistance was observed against amikacin (2%). Twenty-seven isolates were identified as candidates for carbapenemase enzyme production testing, of which only 3/27 (11%) isolates were detected as carbapenemase enzyme producers.ConclusionThis study shows an increasing rate of MDR and XDR isolates and the appearance of PDR in P. aeruginosa strains; this is a serious problem in Ethiopia. The lack of newer anti-pseudomonal antibiotics adds to the problem. In order to alleviate this, infection prevention activities should be promoted, and treatment of bacterial infections should be guided by antibiotic susceptibility test results.

Emergence of rmtD1 gene in clinical isolates of Pseudomonas aeruginosa carrying blaKPC and/or blaVIM-2 genes in Brazil.

The aim of the present study is to describe clinical aminoglycoside- or carbapenem-resistant Pseudomonas aeruginosa isolates collected between 2018 and 2019 in a hospital in Recife City, Northeastern Brazil. It was done based on phenotypic and molecular markers of antimicrobial resistance, as well as on the clonal diversity of the investigated isolates. Thirty-four carbapenem- and/or aminoglycoside-resistant P. aeruginosa isolates were collected in a hospital in Recife City-PE, Brazil. Their antimicrobial susceptibility profile was identified based on the automated BD Phoenix ™ system. In addition, broth microdilution was performed to determine the MICs of tobramycin and polymyxin B. Eventually, isolates were subjected to PCR and sequencing in order to detect the carbapenemase enzyme (blaKPC, blaNDM, blaVIM, blaSPM-1, and blaIMP) and 16S rRNA methylase (armA, rmtB, rmtD, rmtF, and rmtG) genes; ERIC-PCR was conducted for clonal profile determination purposes. Thirty-four of the 64 isolates evaluated in the present study were selected for complementary molecular phenotypic tests, based on sample inclusion criteria. The blaKPC and blaVIM-2 genes were identified in 32.4% (11/34) and 38.2% (13/34) of tested isolates, respectively. The rmtD1 gene was detected in 32.4% (11/34) of analyzed isolates. Eight isolates carried both the blaKPC and rmtD1 genes, whereas blaVIM-2 and rmtD1 genes co-occurrence was detected in three strains; one isolate had all blaKPC, blaVIM-2, and rmtD1 genes. ERIC-PCR molecular typing has evidenced cross-transmission of three pathogenic clones among patients in the hospital. The present study is a pioneer in describing isolates harboring both blaVIM-2 and rmtD1 genes. Moreover, it emphasizes the need of conducting local molecular epidemiology studies at different time intervals in order to monitor measures adopted to prevent nosocomial infections in different hospital units.

Phenotypic and Molecular Detection of BlaNDM Gene Among Drug-Resistant Klebsiella Isolates.

In the past few centuries, a widespread increase in antimicrobial resistance has been observed among Klebsiella species. The antibiotic- resistant strains of the genus Klebsiella are becoming a serious threat in clinical settings due to their involvement in severe invasive and non-invasive infections. The emergence of resistance among these strains is associated with their strong enzymatic activity against several broad-spectrum antibiotics. These enzymes include beta-lactamases, extended-spectrum beta-lactamases (ESBL), AmpC beta-lactamases, and carbapenemases. These resistance enzymes are capable of hydrolyzing various broad-spectrum drugs like extended-spectrum cephalosporin and carbapenems. The present study was conducted to determine the emerging resistance among Klebsiella strains by identifying the production of carbapenemase enzyme phenotypically and the frequency of the NDM resistance gene by a polymerase chain reaction. In this study, 236 Gram-negative isolates from different clinical laboratories were identified. Out of which, 125 isolates were found as Klebsiella species by using standard microbiological techniques. Minimum inhibitory concentrations (MIC) were determined using eight representative antibiotics by the Macro broth dilution method. Phenotypic detection of carbapenemase producing Klebsiella species was performed by Modified Hodge Test. Phenotypic findings were then checked and compared with genotypic results obtained by using the Polymerase chain reaction (PCR) for the detection of resistance genes responsible for the production of carbapenemase. In this study, carbapenemase production was found only in 6 (5%) Klebsiella isolates by using the phenotypic method; however, 3 isolates out of 125 were screened positive for the gene NDM-1. Since we are considering carbapenems as the last therapeutic option for treating infections, mainly caused by Gram-negative isolates, the prevailing resistance against this drug is widely disseminating. It is better to evaluate the antibiotic susceptibility, phenotypic screening as well genotypic screening (where possible) for implementing strict antibiotic control policies in health care settings, hospitals, laboratories, etc.

Activity of meropenem/vaborbactam and comparators against Gram-negative isolates from Eastern and Western European patients hospitalized with pneumonia including ventilator-associated pneumonia (2014-19).

ObjectivesMeropenem/vaborbactam has been approved in Europe for the treatment of hospital-acquired bacterial pneumonia, ventilator-associated pneumonia (VAP) and bacteraemia among other indications. Vaborbactam is an inhibitor of class A and C β-lactamases, including Klebsiella pneumoniae carbapenemase (KPC) enzymes, but not class B or D carbapenemases. We analysed the activity of meropenem/vaborbactam and comparators against 6846 Enterobacterales and 3567 Pseudomonas aeruginosa isolates from patients hospitalized with pneumonia (PHP), including VAP.MethodsIsolates from PHP were consecutively collected during 2014–19 from 42 European hospitals located in 21 countries and susceptibility tested using the broth microdilution method. Carbapenem-resistant Enterobacterales (CRE) isolates were molecularly characterized to identify their carbapenem-resistance mechanisms. EUCAST (2020) interpretive criteria were used.ResultsThe most common Gram-negative pathogens isolated from PHP were P. aeruginosa (n = 3567), K. pneumoniae (n = 1877) and Escherichia coli (n = 1646). Overall, 98.0% of Enterobacterales and 82.1% of P. aeruginosa were susceptible to meropenem/vaborbactam, with 99.8% of Enterobacterales and 89.7% of P. aeruginosa in Western Europe (WE) and 92.7% of Enterobacterales and 69.1% of P. aeruginosa in Eastern Europe (EE). CRE were more common in EE (15.1%) than WE (2.1%). KPC was the most common carbapenemase in WE, while OXA-48-like was the most common carbapenemase in EE. Meropenem/vaborbactam susceptibility was 63.0% for all CRE (92.2% in WE and 51.5% in EE). Meropenem/vaborbactam inhibited 99.1% of KPC-producing isolates and 40.5% of OXA-48-like-producing isolates.ConclusionsThese in vitro data demonstrate that meropenem/vaborbactam has potent activity against isolates from PHP, including isolates producing KPC, and may be a useful treatment option for PHP, including VAP.

Neonatal Sepsis: The Impact of Carbapenem-Resistant and Hypervirulent Klebsiella pneumoniae.

The convergence of a vulnerable population and a notorious pathogen is devastating, as seen in the case of sepsis occurring during the first 28 days of life (neonatal period). Sepsis leads to mortality, particularly in low-income countries (LICs) and lower-middle-income countries (LMICs). Klebsiella pneumoniae, an opportunistic pathogen is a leading cause of neonatal sepsis. The success of K. pneumoniae as a pathogen can be attributed to its multidrug-resistance and hypervirulent-pathotype. Though the WHO still recommends ampicillin and gentamicin for the treatment of neonatal sepsis, K. pneumoniae is rapidly becoming untreatable in this susceptible population. With escalating rates of cephalosporin use in health-care settings, the increasing dependency on carbapenems, a “last resort antibiotic,” has led to the emergence of carbapenem-resistant K. pneumoniae (CRKP). CRKP is reported from around the world causing outbreaks of neonatal infections. Carbapenem resistance in CRKP is largely mediated by highly transmissible plasmid-encoded carbapenemase enzymes, including KPC, NDM, and OXA-48-like enzymes. Further, the emergence of a more invasive and highly pathogenic hypervirulent K. pneumoniae (hvKP) pathotype in the clinical context poses an additional challenge to the clinicians. The deadly package of resistance and virulence has already limited therapeutic options in neonates with a compromised defense system. Although there are reports of CRKP infections, a review on neonatal sepsis due to CRKP/ hvKP is scarce. Here, we discuss the current understanding of neonatal sepsis with a focus on the global impact of the CRKP, provide a perspective regarding the possible acquisition and transmission of the CRKP and/or hvKP in neonates, and present strategies to effectively identify and combat these organisms.

Carbapenem resistance in Acinetobacter baumannii clinical isolates from northwest Iran: high prevalence of OXA genes in sync.

Background and Objectives:Carbapenem treatment for Acinetobacter baumannii infections presently faces threats owing to the production of several types of carbapenemase enzymes, prevalence of which varies among different countries. We explored the current trend of antibiotic resistance in A. baumannii clinical isolates from North West Iran, sought the mechanism of carbapenem resistance and addressed the sequence type groups in carbapenem resistant A. baumannii (CRAB).Materials and Methods:A. baumannii (n=112) isolates were recovered from various clinical specimens of patients admitted in internal, surgery, burn, infectious diseases and various ICUs wards. Genetically confirmed A. baumannii isolates were screened for carbapenem resistance by the Kirby-Bauer and E-test and the presence of bla MBL, bla OXA-like, ISAba1 genes by PCR. Sequence groups were identified by multiplex PCR.Results:Multidrug-resistance (MDR) was a characteristic feature of all A. baumannii isolates. Frequency of oxacillinase genes in combination including bla OXA-51-like/bla OXA-23-like, bla OXA-51-like/blaOXA-24/40-likeand bla OXA-51-like/bla OXA-23-like/bla OXA-24/40-like was 82.1%, 36.6% and 25.8% respectively. Blending of oxacillinase and MBL genes was evident in eight bla OXA-23-like positive and 7 bla OXA-24-like positive isolates thereby depicting synchronous etiology of carbapenem resistance. Amongst CRAB isolates, 97.3% contained ISAba1 element and 50.9% belonged to the European clone II.Conclusion:Synchronicity among bla OXA-like with bla MBL and ISAba1 gene was a hallmark of this investigation. Though origin or route of transmission was not elucidated in this study but co-existence among OXA and MBL producing genes is a therapeutic concern demanding strict surveillance strategies and control programs to halt the dissemination of these isolates in the hospital setting.

β-lactam resistance associated with β-lactamase production and porin alteration in clinical isolates of E. coli and K. pneumoniae.

β-lactam resistance represents a worldwide problem and a serious challenge for antimicrobial treatment. Hence this research was conducted to recognize several mechanisms mediating β-lactam resistance in E. coli and K. pneumoniae clinical isolates collected from Mansoura University hospitals, Egypt. A total of 80 isolates, 45 E. coli and 35 K. pneumoniae isolates, were collected and their antibiotic susceptibility was determined by the Disc diffusion method followed by phenotypic and genotypic detection of extended-spectrum β-lactamases (ESBLs), AmpC β-lactamase, carbapenemase enzymes. The outer membrane protein porins of all isolates were analyzed and their genes were examined using gene amplification and sequencing. Also, the resistance to complement-mediated serum killing was estimated. A significant percentage of isolates (93.8%) were multidrug resistance and showed an elevated resistance to β-lactam antibiotics. The presence of either ESBL or AmpC enzymes was high among isolates (83.75%). Also, 60% of the isolated strains were carbapenemase producers. The most frequently detected gene of ESBL among all tested isolates was blaCTX-M-15 (86.3%) followed by blaTEM-1 (81.3%) and blaSHV-1 (35%) while the Amp-C gene was present in 83.75%. For carbapenemase-producing isolates, blaNDM1 was the most common (60%) followed by blaVIM-1 (35%) and blaOXA-48 (13.8%). Besides, 73.3% and 40% of E. coli and K. pneumoniae isolates respectively were serum resistant. Outer membrane protein analysis showed that 93.3% of E. coli and 95.7% of K. pneumoniae isolates lost their porins or showed modified porins. Furthermore, sequence analysis of tested porin genes in some isolates revealed the presence of frameshift mutations that produced truncated proteins of smaller size. β-lactam resistance in K. pneumoniae and E. coli isolates in our hospitals is due to a combination of β-lactamase activity and porin loss/alteration. Hence more restrictions should be applied on β-lactams usage to decrease the emergence of resistant strains.

PROPORTION OF ESBL AND CARBAPENEMASE PRODUCING ENTEROBACTERIACEAE CAUSING BLOODSTREAM INFECTIONS AND SHOWING MULTI DRUG RESISTANCE IN INFECTED PATIENTS IN TERTIARY CARE HOSPITAL AT AGARTALA, TRIPURA

BACKGROUND: Enrerobacteriaceae species are the major leading cause of bloodstream infections in many developing countries. Moreover, ESBL and Carbapenemase-producing Enterobacteriaceae species are often associated with high resistance to a wide class of antibiotics. There are few studies regarding the bloodstream infections causing by Enterobacteriaceae with the production of Extended-spectrum β-lactamases and Carbapenemase enzymes in Tripura, North-East India. This study aimed to determine the “ proportion of ESBL and carbapenemase-producing Enterobacteriaceae causing bloodstream infection and showing Multi-drug resistance (MDR) in infected patients'' in Tertiary care Hospital at Agartala, Tripura. METHOD: Blood samples were collected from all indoor and outdoor patients suspected of Bloodstream infection. Then specimens were inoculated in various culture media after that with this culture isolates we performed gram staining and many biochemical tests (as per CLSI guidelines) to identies the Enterobacteriaceae species. And the production of β-lactamases and Carbapenemase was conrmed by the combined disk test and Modied Hodge method. RESULTS: In this study out of 760 samples processed in the laboratory 77 (10.13%) was tested positive for bacteremia from which 42 (54.54%) blood specimens were infected by Enterobacteriaceae. The members of the Enterobacteriaceae family isolated in patient samples are E.coli (22/42, 52.38%), K. pneumoniae (11/42, 52.38%) others are Enterobacter spp. (8/42, 52.38%) and S.typhi(1/42, 2.38%). In all 42 Enterobacteriaceae species, 17 (40.47%) isolates were found ESBL positive on antibiotic screening which is conrmed by Combined disc diffusion test, and out of 17 Beta-lactamase producers 8 (47.05%) were E. coli, 5 (29.41%) were K. pneumoniae and 4 (23.52%) were Enterobacter spp. And among 42 isolates of Enterobacteriaceae 16 (38.09%) isolates showed Carbapenemase producers, in that 8 (50%) were E.coli, 5 (31.25%) were K. pneumoniae, and 3 (18.75%) were Enterobacter spp. CONCLUSION: This study aims to provide an early, rapid, and effective phenotypic method for identifying Multi-drug resistant (MDR) Bloodstream infections (BSIs) causing by the species of the Enterobacteriaceae family

Multi-Drug Resistant (MDR) and carbapenemase co-producing Gram-negative bacilli in northwest of Iran

Carbapenemase enzymes confer clinically significant resistance to Gram-negative bacteria, only a few antibiotics with adverse side effects and/or unfavorable in vivo efficacy remains. This study sought to determine the prevalence of carbapenemase producing gram-negative organisms and also investigate the antibacterial susceptibility profile of carbapenem-resistant isolates. In this study, over a two-year period (2017–2019) a total of 140 imipenem and/or meropenem resistant gram negative bacilli were collected from various specimens sent to microbiology laboratory of university hospital. Disk diffusion antimicrobial susceptibility test was used to evaluate MDR isolates; also multiplex polymerase chain reaction and sequencing were performed for confirmation of carbapenemase production. In disk diffusion method, all isolates were resistant to cefazolin. High rate of resistance was observed against imipenem (98.6%), followed by cefotaxime (97.1%), trimethoprim-sulfamethoxazole (91.4%). Colistin was found as the most effective agent with only 5.7% of resistance, followed by amikacin. Based on PCR results and subsequent sequencing, 46.4% of the isolates were genetically recognized to be carrying at least one carbapenemase gene. Encoded carbapenemases were OXA-48-like (18.6%), VIM-2 (15%), IMP-1 (9.3%), KPC (5.7%), NDM-1 (3.6%), and AIM-1 (2.1%). All isolates were negative for SIM, BIC, SPM, GIM and DIM enzymes. Five isolates were co-harbored blaVIM-2 and blaOXA-48. Moreover, co-harbored carbapenemase genes were observed in six isolates as 6 two-genes-harbored profiles. Our results indicate a relatively high incidence of carbapenemase genes. In addition, the emergence of isolates harboring multiple carbapenemases is of great concern, since these isolates will result in a broad-spectrum antibiotic resistance. Molecular characterization of carbapenemase producers can be helpful tool to offer an effective treatment policy. The worrisome high rate of antibiotic resistance highlights the necessity of active surveillance to antibiotic prescription and\\or self-medication by patients.

Carbapenem Resistance in Non-Fermentative Gram-Negative Bacilli Isolated from Intensive Care Unit Patients of a Referral Hospital.

Non-fermentative Gram-negative bacilli or non-fermenters are opportunistic pathogens associated with serious infections in intensive care unit patients. Although carbapenems were considered as a backbone of treatment for life-threatening infections, these bacteria are increasingly acquiring resistance to carbapenems. Carbapenem-resistant Acinetobacter baumannii and Pseudomonas aeruginosa are prioritized as critical pathogens by the World Health Organization. The objective of the study was to document the status of carbapenem-resistant and carbapenemase-producing non-fermenters isolated from intensive care unit patients. This study was conducted at Tribhuvan University Teaching Hospital, Kathmandu, Nepal. The clinical specimens collected from intensive care unit patients were processed for isolation and identification of non-fermenters and antibiotic susceptibility profile of bacterial isolates was determined. The multidrug-resistant isolates were identified and carbapenemase enzyme was detected in the carbapenem-resistant isolates. A total of 157 non-fermenters were isolated from 1063 samples which included Acinetobacter species (n=85), Pseudomonas aeruginosa (n=55), Burkholderia cepacia complex (n=15), and Stenotrophomonas maltophilia (n=2). Carbapenem resistance was reported in 85.9%, 72.7%, and 33.3% of Acinetobacter species, Pseudomonas aeruginosa, and Burkholderia cepacia complex, respectively. Among total non-fermenters, 91.1% isolates were multidrug-resistant and 60.8% carbapenem-resistant isolates were carbapenemase producers. The carbapenem-resistant isolates demonstrated an extremely high degree of resistance than carbapenem-susceptible isolates towards other antimicrobial classes. This study reported high rates of carbapenem-resistant, carbapenemase-producing, and multidrug-resistant non-fermenters isolates. Therefore, preventing the spread of these superbugs among the critically ill patients in intensive care units should be a major initiative in hospitals.

Bacteriological profile and antibiotic susceptibility patterns of wound infections in a tertiary care hospital in South India

Infections caused due to injuries are due to microbial proliferation at the wound site following skin damage. Initial testing of micro-organisms in terms of culturing and sensitivity leads to appropriate antibiotic selection and prevents escalation of antimicrobial resistance.To evaluate the bacteriological profile and antibiotic susceptibility patterns of wound infections in this single-centered study.The study included 160 patients suspected to have wound infection. Pus or tissue samples collected from patients were subjected to microbiological processing including Gram staining, culture and antibiotic susceptibility testing. Their demographic data and wound related factors (duration, nature, type) were recorded. The isolated organisms were evaluated for β-lactamase production using Extended spectrum β-lactamase (ESBL) test, Modified Hodge test for Carbapenemase and AmpC β-lactamase enzyme detection tests. Most participants were 41-60 years old (45.63%). The majority had surgical site infections (SSI, 91.25%), early infected (91.1%) and clean (67.12%) wounds. The bacterial isolation rate was 80% and 45.27% (n=67) were Gram positive isolates, out of which 47 (70.14%) were multidrug resistant. was identified as the predominant organism (n=40), where 33 among 40 were methicillin sensitive, followed by (n=30, 23.43%). Amongst the Gram negative isolates (n=81, 54.73%), 60 (74.07%) were multidrug resistant with majority being susceptible to imipenem, meropenem and amikacin. The most common pathogen associated with wound infection was Methicillin sensitive with SSI being the most common type of wound infection.

Selective digestive decontamination with oral colistin plus gentamicin for persistent bacteraemia caused by non-carbapenemase-producing carbapenem-resistant Klebsiella pneumoniae in a neutropenic patient.

BackgroundCarbapenem-resistant Klebsiella pneumoniae (CRKp) have become an increasing public health problem worldwide. While most CRKp around the world harbour a carbapenemase enzyme, the clinical relevance of non-carbapenemase-producing CRKp (non-CP-CRKp) is increasingly recognized. Selective digestive decontamination (SDD) has been proven successful as a decolonization strategy for patients colonized with Gram-negatives in the ICU. However, it is not regularly used to treat invasive infections.ObjectivesTo report the use of SDD as a useful strategy for managing recalcitrant CRKp bloodstream infections.Patients and methodsWe present a neutropenic patient with a recalcitrant bloodstream infection with non-CP-CRKp treated with SDD. Besides, genomic analyses of five isolates of non-CP-CRKp was performed.ResultsAfter 11 days of SDD treatment with oral colistin and gentamicin, bacteraemia was successfully eradicated. Genomic analysis indicates a fully carbapenem-resistant phenotype evolved in vivo and suggests that the mechanism of carbapenem resistance in our strains relates to gene amplification of narrow-spectrum β-lactamases.ConclusionsOur report highlights that SDD might be a useful strategy to manage CRKp bloodstream infections, when intestinal translocation is the likely source of the bacteraemia. In addition, the development of a resistant phenotype during therapy is worrisome as therapies directed against these organisms are likely to favour the amplification process.

Molecular characterization of carbapenemases in Peru during 2019.

Resistance to carbapenems is a public health problem. This study presents the identification of carbapenemase enzymes in Enterobacteriaceae, Pseudomonas spp. and Acinetobacter spp. present in strains from 30 institutions that provide health services in Peru as part of the quality control process in diagnoses. Phenotypic confirmation and enzymatic identification were performed using the Blue CARBA test and the synergy test with phenylboronic acid and ethylenediaminetetraacetic acid/sodium mercaptoacetic acid discs. 185 strains with carbapenemases were identified: 78 in Enterobacteriaceae, 61 in P. aeruginosa and 46 in Acinetobacter spp. The types of carbapenemases identified were: blaKPC, blaNDM, blaIMP, blaVIM, blaOXA-23, blaOXA-24, blaOXA-51 and the blaVIM/IMP co-production. It is important to strengthen the promotion of the rational use of antimicrobials and epidemiological surveillance in the country's hospitals.

Phenotypic Characterisation of Klebseilla pneumoniae Carbapenemase and Metallo Beta-lactamase in Carbapenem Resistant Gram-negative Bacteria

Background: Antibiotic resistance is an emerging concern in the therapy of clinical infections worldwide. Previous studies conducted in our laboratory have confirmed an increase in the prevalence of extended spectrum beta-lactamase (ESBL) among the Gram-negative bacterial pathogens associated with dogs, which could act as a potential source for the transfer of these resistant pathogens or their genetic determinants to human. Since carbapenems are the last resort drugs against these resistant pathogens, the study was aimed to isolate and characterise carbapenem resistance among Escherichia coli (E. coli), Klebsiella pneumoniae (K. pneumoniae) and Pseudomonas aeruginosa (P. aeruginosa) associated with common clinical infections in dogs.Methods: A total of 100 samples were collected from lesional skin, urine and anterior vagina of dogs presented to the Veterinary Hospitals of Kerala Veterinary and Animal Sciences University at Mannuthy and Thrissur. The samples were cultured onto Brain Heart Infusion Agar (BHIA), Eosin Methylene Blue (EMB) and Mac Conkey (MAC) for isolation of bacteria. Identification of the isolates was performed based on cultural, morphological and biochemical characteristics. The isolates were subjected to antimicrobial susceptibility test (ABST) against the 12 commonly used beta-lactam and non–beta-lactam group of antibiotics by disc diffusion method and further subjected to screening for ESBL double disc diffusion method. Carbapenem-resistant isolates were subjected to phenotypic confirmatory test for carbapenemase production employing Imipenem-EDTA and Ertapenem-boronic acid minimum inhibitory concentration (MIC) strip method.Result: Forty four Gram-negative bacterial isolates obtained were viz., E. coli (30), K. pneumonia (11) and P. aeruginosa (3) from the 100 samples. Apart from these, other isolates obtained were Staphylococcus spp. (53) and Bacillus spp. (2). All the Gram-negative isolates were subjected to ABST employing 12 common antibiotics belonging to beta-lactam and non-beta-lactam groups. Multidrug resistance (MDR) could be observed in 28 E. coli, 11 K. pneumoniae and three P. aeruginosa isolates. All the 42 MDR isolates showed positive results for ESBL production. A total of 14 isolates out of the 44 Gram-negative bacilli were found to be resistant to carbapenem either to imipenem, meropenem or ertapenem. Among the 14 Gram-negative isolates, nine turned out to be positive for metallo-beta-lactamase (MBL) and none for K. penumoniae carbapenemase (KPC) on phenotypic confirmatory test for detecting major carbapenemase enzymes. The present study documented that Gram- negative bacteria like E. coli, K. pneumoniae and P. aeruginosa isolated from dogs are showing an increase rate of resistance against carbapenems which are the last resort drugs against ESBL producers. Hence, there is an urgent need to curb the irrational and excessive use of antibiotics in veterinary sector.

Antibacterial and COX-2 Inhibitory Tetrahydrobisbenzylisoquinoline Alkaloids from the Philippine Medicinal Plant Phaeanthus ophthalmicus.

Phaeanthus ophthalmicus (Roxb. ex G.Don) J.Sinclair (previously known as P. ebracteolatus (Presl) Merr) is a Philippine medicinal plant occurring as evergreen shrub in the lowland forests of Luzon islands. It is used traditionally by Filipinos to treat bacterial conjunctivitis, ulcer and wound infections. Based on previous investigations where cyclooxygenase-2 (COX-2) functions as immune-linked factor in infectious sensitivities to bacterial pathogens by triggering pro-inflammatory immune-associated reactions, we investigated the antimicrobial and COX inhibitory activities of the extracts and tetrahydrobisbenzylisoquinoline alkaloids of P. ophthalmicus in vitro and in silico to validate its ethnomedicinal uses. Thus, the dichloromethane–methanol (DCM–MeOH) crude extract and alkaloid extracts exhibiting antibacterial activities against drug-resistant bacterial strains such as methicillin-resistance Staphylococcus aureus (MRSA), vancomycin-resistant Enterococcus (VRE), Klebsiella pneumoniae + CRE and Pseudomonas aeruginosa + MBL afforded (+)-tetrandrine (1) and (+)-limacusine (2) as the major biologically active tetrahydrobisbenzylisoquinoline alkaloidal constituents after purification. Both tetrahydrobisbenzylisoquinoline alkaloids 1 and 2 showed broad spectrum antibacterial activity with strongest inhibition against the Gram-negative bacteria MβL-Pseudomonas aeruginosa Klebsiella pneumoniae + CRE. Interestingly, the alkaloid limacusine (2) showed selective inhibition against ovine COX-2 in vitro. These results were ascertained by molecular docking and molecular dynamics simulation experiments where alkaloid 2 showed strong affinity in the catalytic sites of Gram-negative bacterial enzymes P. aeruginosa elastase and K. pneumoniae KPC-2 carbapenemase (enzymes involved in infectivity mechanisms), and of ovine COX-2. Overall, our study provides credence on the ethnomedicinal use of the Philippine medicinal plant P. ophthalmicus as traditional plant-based adjuvant to treat bacterial conjunctivitis and other related infections. The antibacterial activities and selective COX-2 inhibition observed for limacusine (2) point to its role as the biologically active constituent of P. ophthalmicus. A limited number of drugs with COX-2 inhibitory properties like celecoxib also confer antibacterial activity. Thus, tetrahydrobisbenzyl alkaloids, especially 2, are promising pharmaceutical inspirations for developing treatments of bacterial/inflammation-related infections.

Continuous Evolution: Perspective on the Epidemiology of Carbapenemase Resistance Among Enterobacterales and Other Gram-Negative Bacteria.

The global emergence of carbapenemase-producing bacteria capable of hydrolyzing the once effective carbapenem antibiotics is considered a contemporary public health concern. Carbapenemase enzymes, once constrained to isolates of Klebsiella pneumoniae, are now routinely reported in different bacteria within the Enterobacterales order of bacteria, creating the acronym CRE which now defines Carbapenem-Resistant Enterobacterales. CRE harboring different types of enzymes, including the most prevalent types KPC, VIM, IMP, NDM, and OXA-48, are now routinely reported and more importantly, are now frequently present in many infections world-wide. Defining and updating the contemporary epidemiology of both the US and global burden of carbapenem-resistant infections is now more important than ever. This review describes the global distribution and continued evolution of carbapenemases which continue to spread at alarming rates. Informed understanding of the current epidemiology of CRE, coupled with advances in antibiotic options, and the use rapid diagnostics offers the potential for rapid identification and management of carbapenem-resistant infections.

Karbapenem Dirençli Enterobacterales İzolatlarında Karbapenemaz Genlerinin Araştırılması: Dokuz Eylül Üniversitesi Hastanesi’nden İlk KPC Bildirimi

Objective: In recent years, increasing carbapenem resistance of Enterobacterales bacteria limits treatment options, considerably. The main mechanism of this resistance is the production of carbapenemase enzymes. The aim of this study is to determine carbapenemase gene types in Enterobacterales isolates from our hospitalized patients and assess the clonal associations of the isolates with KPC gene. Method: A total of 48 clinical Enterobacterales isolates resistant to at least one carbapeneme and received between January 2019 and March 2019 were included in the study. Sample types were consisted of urine, blood, tracheal aspirate, wound and sputum. Of these isolates, three were Escherichia coli while 45 were Klebsiella pneumoniae. Types of carbapenemases were investigated by polymerase chain reaction, using specific primers for VIM, IMP, NDM, KPC and OXA-48 genes. PFGE was performed to determine the clonal associations between blaKPC positive K. pnemoniae isolates. Results: According to the results, blaOXA-48 (n=2) and blaKPC (n=1) were found to be present among E. coli isolates. Regarding 45 K. pneumoniae isolates; only blaOXA-48 and only blaNDM were present in 30 and two isolates, respectively. Seven K. pneumoniae isolates were found positive for both blaOXA-48 and blaNDM. Remaining K. pneumoniae isolates (n=6) harboured only blaKPC. None of the isolates were positive for blaIMP and blaVIM. PFGE analysis showed four isolates had the same pulsotype (A), while two had different pulsotypes (B-C). Conclusion: To our knowledge, this is the first report of KPC gene isolated in Dokuz Eylul University Hospital.

1432. Population-based Surveillance of Carbapenem-Resistant Enterobacteriaceae (CRE) in Alameda County, 2017-2020

BackgroundInfections caused by carbapenem-resistant Enterobacteriaceae (CRE), especially those that are carbapenemase-producing (CP), are difficult to treat and result in high mortality— the Centers for Disease Control and Prevention (CDC) designates CRE an urgent level threat to public health. Alameda, a northern California county with 1.67 million residents, mandates submission of all carbapenem-resistant isolates Escherichia coli, Klebsiella spp., and Enterobacter spp. We assessed the genetic profiles of CRE isolates and compared to aggregate US data from the same time period.MethodsIsolates are submitted to the Alameda County Public Health Laboratory (ACPHL), where antimicrobial resistance genetic markers are identified by whole genome sequencing (WGS) using single-end, 150-cycle reactions in a MiSeq (Illumina). Resistance genes were identified using pipelines built in Geneious and confirmed with Resfinder. All epidemiological analyses were conducted using R (Version 4.0).ResultsACPHL performed WGS on 226 CRE isolates submitted between June 2017 and February 2020. A total of 34/95 (36%) Klebsiella spp., 17/60 (28%) E. coli, and 10/71 (14%) Enterobacter spp. a carbapenemase enzyme. Among all Enterobacteriaceae, 21/226 (9%) produced the New Delhi-metallo-β-lactamase (NDM) carbapenemase (Table 1). Among all CRE, 17/226 (8%) were Klebsiella pneumoniae with the multilocus sequence type (MLST) of ST-258. All six Klebsiella pneumoniae ST-35 and ST-11 isolates produced a carbapenemase (Table 2).Table 1: Carbapenemases, Extended Spectrum β-Lactamases (ESBL) and Other β-Lactamases by Organism Table 2: Organism, Multilocus Sequence Type (MLST) and Carbapenemases Present ConclusionCompared to US data, Alameda County Klebsiella spp. were nearly half as likely to produce any carbapenemase, and E. coli were three times more likely to produce an NDM carbapenemase. CP-CRE represented a wide array of MLST, with the globally distributed KPC-producing outbreak strain ST-258 being the most frequently identified. Two strains that have been associated with hypervirulent and multidrug resistant infections in Asia, K. pneumoniae ST-35 and ST-11, were also reported.DisclosuresSam Horwich-Scholefield, MPH CIC, Merck (Research Grant or Support)