Prevalence of Transferable OXA-1 β-Lactamase Associated with Carbapenem-Resistant Klebsiella pneumoniae Isolates in Iraq

This study was designed to explore the incidence of blaOXA-1 amongst Klebsiella pneumoniae isolates with resistant to carbapenem. Between December 2014 and April 2015, one hundred samples were taken from two hospitals: Babylon Teaching Hospital for Maternity and Pediatric / Babylon Province (clinical, umbilical infections, n= 40; environmental, n=20) and Karbala Hospital for Pediatric / Karbala Province (40 stool samples). All patients were hospitalized or attended these hospitals, all under 1 year of age. Seventeenth (17%) isolates were identified as Klebsiella pneumoniae. The antibiotic resistance profile of isolates was tested using disk diffusion method. High-level of resistance was recorded with ampicillin (94.1%) and piperacillin (88.2%) antibiotics. Resistance to carbapenem was reported in two K.pneumoniae isolates, these were investigated for the existence of OXA-1b-lactamase using Polymerase Chain Reaction (PCR) technique. Two (100%) isolates gave positive result. Transference of this gene was studied by conjugation experiment. The blaOXA-1 gene conjugated successfully in 1 (50%) isolate only.

Prevalence of 16S rRNA Methylation Enzyme Gene armA in Salmonella From Outpatients and Food

Salmonella is the primary cause of community-acquired foodborne infections, so its resistance to antimicrobials, such as aminoglycosides, is a public health issue. Of concern, aminoglycoside resistance in Salmonella is increasing rapidly. Here, we performed a retrospective study evaluating the prevalence of Salmonella harboring armA-mediated aminoglycoside resistance in community-acquired infections and in food or environmental sources. The prevalence rates of armA-harboring Salmonella strains were 1.1/1,000 (13/12,095) and 8.7/1,000 (32/3,687) in outpatient and food/environmental isolates, respectively. All the armA-harboring Salmonella strains were resistant to multiple drugs, including fluoroquinolone and/or extended-spectrum cephalosporins, and most (34/45) belonged to serovar Indiana. The armA gene of these strains were all carried on plasmids, which spanned five replicon types with IncHI2 being the dominant plasmid type. All the armA-carrying plasmids were transferable into Escherichia coli and Acinetobacter baumannii recipients. The conjugation experiment results revealed that the armA-harboring S. Indiana strains had a relatively higher ability to acquire armA-carrying plasmids. The low similarity of their pulsed field gel electrophoresis patterns indicates that the armA-harboring Salmonella strains were unlikely to have originated from a single epidemic clone, suggesting broad armA spread. Furthermore, the genetic backgrounds of armA-harboring Salmonella strains isolated from outpatients exhibited higher similarity to those isolated from poultry than to those isolated from swine, suggesting that poultry consumption maybe an infection source. These findings highlight an urgent need to monitor the prevalence and transmission of armA-harboring Salmonella, especially S. Indiana, to better understand the potential public health threat and prevent the further spread of these strains.

Genomic Characterization of VIM and MCR Co-Producers: The First Two Clinical Cases, in Italy

Background: the co-production of carbapenemases and mcr-genes represents a worrisome event in the treatment of Enterobacteriaceae infections. The aim of the study was to characterize the genomic features of two clinical Enterobacter cloacae complex (ECC) isolates, co-producing VIM and MCR enzymes, in Italy. Methods: species identification and antibiotic susceptibility profiling were performed using MALDI-TOF and broth microdilution methods, respectively. Transferability of the blaVIM- and mcr- type genes was verified through conjugation experiment. Extracted DNA was sequenced using long reads sequencing technology on the Sequel I platform (PacBio). Results: the first isolate showed clinical resistance against ertapenem yet was colistin susceptible (EUCAST 2020 breakpoints). The mcr-9.2 gene was harbored on a conjugative IncHI2 plasmid, while the blaVIM-1 determinant was harbored on a conjugative IncN plasmid. The second isolate, resistant to both carbapenems and colistin, harbored: mcr-9 gene and its two component regulatory genes for increased expression on the chromosome, mcr-4.3 on non-conjugative (yet co-transferable) ColE plasmid, and blaVIM-1 on a non-conjugative IncA plasmid. Conclusions: to our knowledge, this is the first report of co-production of VIM and MCR in ECC isolates in Italy.

Discovery of mcr-1 Harboring Incl2 Plasmids from Clinical Isolates of Multiclonal E. Coli prevalent in Pakistan.

Background: Colistin is the last resort antibiotic against multiple drug-resistant (MDR) bacteria found in clinical infections; however, the emergence of plasmid-mediated mcr-1 gene annulled the efficacy of Colistin. This study was planned to determine the prevalence of mcr genes in clinical isolates collected in Pakistan. The molecular types and plasmids of mcr-1 bearing isolates were analysed. Methods: A total of 545 E. coli isolates collected from two major cities of Pakistan were screened for colistin-resistance and mcr genes from June 2018 to September 2019. All positive strains were subjected to antimicrobial susceptibility testing, ESBL, and MBL detection via DDST and CDT. ESBL genes detection, molecular typing, conjugation experiment, plasmid replicon typing, S1 PFGE, and southern hybridization were performed. Results: Four (0.73%) strains of mcr-1 positive isolates were susceptible to meropenem, fosfomycin, and chloramphenicol, including one that showed moderate level resistance to chloramphenicol and fosfomycin. All four strains were ESBL positive and harbored the blaCTX-M-15 gene, while three of the isolates also harbored the blaTEM-1 gene. Molecular typing revealed that four isolates belonged to diverse clonal types, mostly (75%) from avian pathogenic E. coli lineage. The mcr-1 gene was present on ~ 60 kb Incl2 plasmid, which was successfully transconjugated. Conclusion: The mcr-1 gene's detection in diverse clonal types from MDR clinical isolates enforces priority basis large scale surveillance studies followed by corrective actions to prevent the spread of the mcr-1 gene in clinical, poultry, and environmental setting.

Spectrum of Aminoglycoside Modifying Enzymes in Gram-Negative Bacteria Causing Human Infections

Introduction Aminoglycosides are formidable broad-spectrum antibiotics used in clinical settings; woefully their usage has been reduced by the emergence and distribution of resistance mainly due to aminoglycoside modifying enzymes (AME). Purpose This study was performed to determine the diverse prevalence of AME and their pattern of occurrence in the clinical isolates of gram-negative bacteria. This study also aimed to detect the presence of AMEs that are prevalent in gram-positive bacteria, among gram negatives. Materials and Methods A total number of 386 clinical isolates were included in this study. Polymerase chain reaction revealed the prevalence rate of AMEs screened [aac(6′)-lb, aac(3′)-I, aac(3′)-II, aac(3′)-VI, ant(2′)-I, ant(4′)-IIb, aac(3′)-III, aac(3′)-IV, aph(2′)-Ib, aph(2′)-Ic, aph(2′)-Id, aac (6′)-Ie- aph(2′)-Ia, and aph(3′)-IIIa]. Conjugation experiment was performed for the clinical isolates which harbored any one of the AME which was prevalent in gram-positive bacteria [aph(3′)-IIIa, aac(6′)-Ie-aph(2′)-Ia]. Results aac(6′)-lb is the most prevalent AME, followed by aac(3′)-I, aph(3′)-VI, aac(3′)-VI, and aac(3′)-II. The AMEs such as ant (2′)-I, ant(4′)-IIb, aac(3′)-III, aac(3′)-IV, aph(2′)-Ib, aph(2′)-Ic, and aph(2′)-Id were not established in our study isolates. The rate of prevalence of aph(3′)-IIIa, aac(6′)-Ie-aph(2′)-Ia—the AMEs encountered in gram-positive and their co-existence was 19.68% and the conjugation experiment revealed their transfer via plasmids. Conclusion This is the first report from India revealing the presence and prevalence of AMEs which are often encountered among gram-positive bacteria in gram negatives and their presence on conjugative plasmids.

Co occurrence of two 16S rRNA methyltrasferases along with NDM and OXA 48 like carbapenamases on a single plasmid in Klebsiella pneumoniae

BACKGROUND: The carbapenemase-encoding genes, bla NDM- and bla OXA-48 - like , confer resistance to all the known beta-lactams and are encountered along with other beta-lactamase-encoding genes and/or 16S ribosomal RNA (rRNA)-methylating genes. The co-occurrence of bla NDM and bla OXA-48 - like on a single plasmid is a rare occurrence. AIM AND OBJECTIVE: The purpose of the study was to characterize the plasmids in Klebsiella pneumoniae isolates producing 16S rRNA methyltransferase along with bla NDM , bla OXA-48-like , and other resistance encoding genes. MATERIALS AND METHODS: One-hundred and seventeen K. pneumoniae clinical isolates which were resistant to aminoglycosides were collected. Polymerase chain reaction-based screening for 16S rRNA methyltransferase genes armA, rmtB, and rmtC; carbapenamase genes bla NDM , bla OXA-48-like , bla IMP, bla VIM, and bla KPC ; and other resistance genes such as bla TEM, bla SHV, bla CTX-M , and qnr (A, B, and S) determinants acc (6') Ib-cr was performed. Conjugation experiment was carried out for seven isolates that anchored bla NDM and bla OXA-48-like along with any one of the 16S rRNA methyltransferases. The plasmid-based replicon typing for different plasmid-incompatible (Inc) group was performed on the conjugatively transferable plasmids. RESULTS: Among the 16S rRNA methyltransferases, armA was more predominant. bla NDM and bla OXA-48 -like were present in 56 (47.86%) and 22 (18.80%) isolates, respectively. Out of seven isolates which were conjugatively transferable, only four had bla NDM and bla OXA-48 - like on the same plasmid and they belonged to Inc N and A/C replicon. Three isolates co-harbored 16S rRNA methyltransferases armA, rmtB, and rmtC, and out of the them, one isolate harbored two 16S rRNA methyltransferases armA and rmtB, on the single-plasmid replicon A/C. CONCLUSION: This is the first report revealing the coexistence of bla NDM and bla OXA-48 - like co-harboring two 16S rRNA methylases on a single conjugative plasmid replicon belonging to incompatibility group A/C.

Transfer of CMY-2 Cephalosporinase from Escherichia coli to Virulent Klebsiella pneumoniae Causing a Recurrent Liver Abscess

ABSTRACTA CMY-2-producing capsular type K2Klebsiella pneumoniaestrain (TVGHKP93) with multidrug resistance was isolated from a recurrent liver abscess in a patient who also carried a CMY-2-producingEscherichia colistrain (TVGHEC01) in the stool. TVGHKP93 retained its high virulence compared with that of the isogenic strain (TVGHKP60) with wild-type resistance from the first liver abscess. Our conjugation experiment showed the successful transfer of theblaCMY-2-carrying plasmid from TVGHEC01 into TVGHKP60. The transconjugant showed both high virulence and the multidrug-resistant phenotype, as did TVGHKP93.

Detection of clonal KPC-2-producing Klebsiella pneumoniae ST258 in Korea during nationwide surveillance in 2011

This study analysed the characteristics and genetic similarity of recent Klebsiella pneumoniae carbapenemase (KPC-2)-producing Klebsiella pneumoniae isolates from Korea. Recent laboratory surveillance detected an increase in carbapenemase-producing Enterobacteriaceae in Korea. A total of 6 KPC-2-producing K. pneumoniae were identified from 277 Enterobacteriaceae clinical isolates. All were sequence type (ST) 258 and they had the same pulsotype. They had high MICs for carbapenems and multi-drug resistance. TEM-1, SHV-11 and OXA type β-lactamases were detected in all isolates, whereas CTX-M type β-lactamases and plasmid-mediated AmpC β-lactamase (PABL) were not present. A conjugation experiment failed, but bla KPC-2-harbouring plasmids from the six isolates were used to transform Escherichia coli DH5-α by electroporation. Each of the transformants harboured a bla KPC-2-positive approximately 95 kb plasmid, which was typed in the IncFII incompatibility group and co-harboured TEM-1 and OXA-9 β-lactamases. They shared the same restriction profile. This study confirms the emergence of clonal ST258 KPC-2-producing K. pneumoniae in some regions of Korea.

Emergence of Proteus mirabilis HarboringblaKPC-2andqnrDin a Chinese Hospital

ABSTRACTNineteen carbapenem-nonsusceptibleProteus mirabilisisolates were recovered from intensive care units in the Second Affiliated Hospital of Zhejiang University during a 3-month period. The isolates showed a high level of resistance against ciprofloxacin, in addition to their resistance against the carbapenems. Pulsed-field gel electrophoresis (PFGE) analysis showed that these isolates belonged to three clonal strains. PCRs and DNA sequence analysis of the carbapenemase and other β-lactamase genes indicated that all the isolates harbored theblaKPC-2gene. Twelve of 19 isolates harbored the plasmid-mediated quinolone resistance (PMQR) genes, both theqnrDandaac(6′)-Ib-crgenes. Eight representative isolates with high levels of quinolone resistance carried the similar mutation profiles of S83I ingyrA, E466D ingyrB, and S80I inparC. Reduced carbapenem susceptibility was transferred toEscherichia coli(EC600) in a conjugation experiment, while the quinolone resistance was not. DNA hybridization showed thatqnrDwas located on a plasmid of approximately 4.5 kb. In summary, large clonally related isolates of KPC-2-producingP. mirabilisemerged in a Chinese hospital, andqnrDwas detected in KPC-producingP. mirabilisfor the first time.