The whole-genome molecular epidemiology of sequential isolates of Acinetobacter baumannii colonizing the rectum of patients in an adult intensive care unit of a tertiary hospital.

IMPORTANCE: Acinetobacter baumannii is a multidrug-resistant nosocomial pathogen that colonizes and infects debilitated patients in the ICU. There is very little information on the genomic characteristics of colonizing strains. This information is important to understand the evolution of lineages of A. baumannii that develop resistance while patients receive antibiotic treatment in the ICU. Our study demonstrated different patterns of colonization of the rectum of ICU patients with different STs of A. baumannii while one ST colonized all patients. Some STs carried more antibiotic resistance genes compared to others. However, there was a correlation between ST and a particular resistance gene profile. Our results further elucidate the dynamics of enteric colonization of this opportunistic pathogen.

Multiplex gyrB PCR Assay for Identification of Acinetobacter baumannii Is Validated by Whole Genome Sequence-Based Assays.

OBJECTIVE: A multiplex gyrB PCR assay has been used to diagnose Acinetobacter baumannii. However, this assay has not been validated against the gold standard DNA-DNA hybridization assay, which is a laborious method. DNA-DNA hybridization assay is now replaced by whole genome sequence (WGS)-based methods. Two such methods are a k-mer-based search of sequence reads using the Kraken 2 program and average nucleotide identity (ANI). The objective was to validate the gyrB PCR assay with WGS-based methods. SUBJECTS AND METHODS: We cultured 270 sequential A. baumannii isolates from the rectal swabs of 32 adult patients. The identity of the isolates was determined by gyrB PCR. The sequences of 269 isolates were determined by Illumina sequencing and the taxonomy was inferred by the Kraken 2 program and ANI. RESULTS: All the 269 isolates were confirmed as A. baumannii by Kraken 2 and ANI. CONCLUSION: The gyrB PCR assay is now validated for easy identification of A. baumannii in comparison with gold standard WGS-based assays.

Antimicrobial Resistance of Serial Isolates of Acinetobacter baumannii Colonizing the Rectum of Adult Intensive Care Unit Patients in a Teaching Hospital in Kuwait.

Objectives: Outbreak and endemic isolates of Acinetobacter baumannii are known to be polyclonal. In an ongoing study, we hypothesized that the patient gut was the source of the polyclonality where genetic exchanges take place. To test the hypothesis, we collected 270 serial rectal isolates from 32 adult intensive care unit patients over 16 months and investigated their drug resistance profiles. Methods: Antimicrobial susceptibility was determined according to recommended methods. The blaIMP, blaVIM, blaSIM, blaOXA-23, blaOXA-24/40, blaOXA-51, blaOXA-48, blaKPC, blaGES, blaNDM and blaOXA-58 were sought by PCR. A subset of 42 isolates were studied for plasmid-mediated resistance. Results: Most of the 270 isolates were multidrug resistant (MDR; with resistances to meropenem of 85.18% and imipenem of 87.04%), but susceptible to colistin and trimethoprim/sulfamethoxazole. There was no correlation between the pattern of resistance and antibiotics administered to treat infections. There was no consistent pattern of resistance or content of carbapenemase genes in serial rectal isolates suggesting polyclonality of the isolates. Genes mediating production of OXA-23, OXA-24/40, IMP, and GES enzymes were carried on plasmids and they mediated resistance to all carbapenems in conjugation studies. Conclusion: A. baumannii colonizing the rectum were polyclonal, MDR, and carbapenem resistance genes were found on plasmids and some plasmids were transferable.

Genetic relatedness of serial rectal isolates of Acinetobacter baumannii in an adult intensive care unit of a tertiary hospital in Kuwait.

Acinetobacter baumannii is an opportunistic pathogen of intensive care unit (ICU) patients. A. baumannii colonizes many parts of the body including the gastrointestinal tract. Endemic and epidemic strains are polyclonal. There is no clarity on the origin of polyclonality of A. baumannii. The objective of the study was to define the genetic relatedness of serial isolates and the origin of polyclonality. Serial rectal isolates from ICU patients whose rectum was colonized on ≥5 sampling occasions were selected. From a total of 32 eligible colonized patients, isolates from a subgroup of 13 patients (a total of 108 isolates) showing different patterns of colonization as revealed by pulsed-field gel electrophoresis (PFGE) were studied. The isolates were analyzed by PFGE pulsotypes, sequence types (STs) by multi-locus sequence typing (MLST) and clonal complex (CC) by eBURST analysis. Serial isolates constituted a mixture of identical, related and unrelated pulsotypes. Analysis by STs and CCs were less discriminatory. The data suggest a combination of an initial colonizing isolate undergoing mutation as well as colonization by independent isolates. Further clarity on the origin of diversity should be better obtained by whole-genome sequencing.

Antimicrobial resistance among anaerobes isolated from clinical specimens in Kuwait hospitals: comparative analysis of 11-year data.

Our objective was to compare the antimicrobial resistance trends among clinically relevant anaerobes against 9 different antibiotics over two periods, 2008-2012 and 2002-2007. Antimicrobial susceptibility testing was performed by determining the MICs using E test method. The interpretation of results was according to the breakpoints recommended by the Clinical Laboratory and Standard Institute (CLSI) and European Committee on Antimicrobial Susceptibility Testing (EUCAST). A total of 2240 clinically significant isolates were collected between 2008 and 2012 in four teaching hospitals in Kuwait. The commonest isolates were Bacteroides fragilis (40.4%), Prevotella bivia (18.6%), Peptostreptococcus spp. (13.8%) and Bacteroides ovatus (11.1%). According to CLSI and EUCAST breakpoints used for the 2008-2012 and 2002-2007 isolates, high resistance rates to amoxicillin-clavulanic acid, clindamycin, penicillin and piperacillin were noted among the Gram-negative isolates. They ranged between 0 and 0-62.1 and 62.1%, and 0 and 0-59.1 and 62.1%, respectively against clindamycin, 0 and 0-34.5 and 45.3%, and 0 and 0-45 and 57.5%, respectively against piperacillin and 0 and 0-24.2 and 24.2%, and 0 and 0-23.1 and 30.6%, respectively against amoxicillin-clavulanic acid. The mean interpretative results by both CLSI and EUCAST during the 2008-2012 and 2002-2007 periods showed that the B. fragilis isolates were highly resistant to penicillin (100 vs 100%), clindamycin (43.7 vs 44.2%), piperacillin (35.8 vs 42.7%) and amoxicillin-clavulanic acid (13.2 vs 14%), respectively. When compared with 2002-2007, the CLSI, but not EUCAST, demonstrated statistically significant decreased resistance to clindamycin (P < 0.03). However, both interpretative criteria showed demonstrable statistically significant decrease in resistance rates to imipenem (P < 0.00097 vs P < 0.00074), meropenem (P < 0.000006 vs P < 0.0407) and piperacillin (P < 0.000017 vs P < 0.0461). Our data shows that there is a need for periodic monitoring of the susceptibility testing for anaerobic bacteria in the face of increasing resistance rates as well as to guide in the empirical therapy of anaerobic infections.