High prevalence of oxacillinases in clinical multidrug-resistant Acinetobacter baumannii isolates from the Tshwane region, South Africa - an update

Abstract

Background: Acinetobacter baumannii is an important hospital-acquired pathogen in healthcare facilities that frequently causes bacteraemia and ventilator-associated pneumonia in intensive care units. Acinetobacter baumannii can be isolated from various sites in the hospital environment like medical equipment, bed linen, medical personnel and indwelling catheters. It is difficult to treat A. baumannii infections because of their highly resistant antimicrobial profiles. The purpose of this study was to determine the prevalence of β-lactamase genes in multidrug-resistant (MDR) clinical A. baumannii isolates using Multiplex-PCR (M-PCR) assays.

Methods: One hundred MDR A. baumannii isolates were collected from the diagnostic division of the Department of Medical Microbiology after routine analysis of the submitted specimens. All collected isolates were identified and tested for susceptibility using the VITEK 2® system (bioMérieux, France). Six isolates were excluded from this study because the isolates were incorrectly identified as A. baumannii with the VITEK 2® system (bioMérieux, France). Molecular tests, namely M-PCR assays, pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST) were performed. MLST analyses were performed on representative isolates from the four major pulsotypes (≥5 isolates with 80 % similarity) and selective isolates from each minor pulsotype.

Results: All the A. baumannii isolates showed 100 % resistance to ampicillin, amoxicillin, cefuroxime, cefuroximine axetil, cefoxitin, cefotaxime and nitrofurantoin. Seven percent of the isolates were resistant to amikacin. Two percent of the isolates were classified as having intermediate susceptibility to tigecycline. A. baumannii isolates showed an antibiotic resistance profile of 67 % and higher to antibiotics, such as ceftazidime, cefepime, imipenem, meropenem, gentamicin, ciprofloxacin and trimethoprim/sulfamethoxazole. None of the isolates were resistant to colistin. The M-PCR assays showed that 99 % of the isolates contained the OXA-51 gene and 77 % contained the OXA-23 gene. None of the isolates contained the GES, GIM, IMP, KPC, NDM, OXA-24, OXA-58, PER, SIM, SPM, VEB and VIM genes. Representative A. baumannii isolates were grouped into five existing sequence types (ST): ST106, ST258, ST339, ST502, ST758 and ST848. Isolates belonging to the pan-European clonal lineages I and II (EUI and EUII) were identified.

Conclusion: The high prevalence of MDR A. baumannii isolates has a severe impact on available treatment choices and this in return impacts on treatment outcomes in the studied healthcare facilities. The most dominant ST among the collected isolates was ST758, member of the EUI group. The presence of the OXA-23 gene was not restricted to a specific ST. Continuous research and surveillance is necessary to monitor the circulating β-lactamase genes in clinical settings to guide infection control policies in order to try and curb the spread of this bacterium.

Fig. 1

The percentage of antibiotic resistance in clinical A. baumannii isolates in 2008 and 2013 in the Pretoria region

Fig. 2

Results obtained after the M-PCR I assay, using a 1.8% (m/v) Metaphor agarose gel electrophoresis for the detection of oxacillinase (OXA) genes in A. baumannii isolates. The M-PCR assay amplified the OXA-23 (501 bp) and OXA-51 (353 bp) genes. Lanes 3, 5 and 7 were negative for all of the OXA genes. Lanes 2, 4, 8, 9, 10, 11 and 12 were positive for OXA-23 and OXA-51. Lane 6 was positive for OXA-51 only. Lane 1 was the negative control. Lane “L” represented the O’Range Ruler™ 100 bp DNA ladder

Fig. 3

PFGE patterns of A. baumannii isolates. The bandings patterns were analysed using the GelCompar II software programme (GelCompar II, Applied Maths, Belgium). A distance matrix was constructed using the Dice coefficient and a dendogramme was constructed from the distance matrix using UPGMA. Major pulsotype designation was based on five or more isolates showing ≥80 % relatedness. Isolate name, ST and the carriage of bla _OXA-23 (Y, positive; N, negative) are indicated