Phenotypic, PCR, and whole-genome sequencing characterization of carbapenem-resistant Acinetobacter baumannii in Makkah and Riyadh, Saudi Arabia: AST discordance and local genomic micro-clusters

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Background Carbapenem-resistant Acinetobacter baumannii (CRAB) is a World Health Organization (WHO) critical-priority pathogen, yet hospitals need practical workflows that link rapid rule-in testing, definitive minimum inhibitory concentration (MIC) confirmation, and actionable genomic thresholds for outbreak response. Integrated phenotype–polymerase chain reaction (PCR)–whole-genome sequencing (WGS) characterization studies with decision-ready performance metrics remain limited for settings in Saudi Arabia. Methods We analyzed 71 non-duplicate multidrug-resistant (MDR) A. baumannii clinical isolates from two cities in Saudi Arabia (Riyadh, n = 36; Makkah, n = 35), collected from 2024 to 2025. We compared VITEK II antimicrobial susceptibility testing (AST) to Clinical and Laboratory Standards Institute (CLSI) broth microdilution (BMD) MICs for imipenem (IMI), meropenem (MEM), ceftazidime (CAZ), and ciprofloxacin (CIP); evaluated targeted PCR for blaOXA-23 and blaNDM (with blaOXA-51-like confirmation); and performed whole-genome sequencing (WGS; n = 59 passing quality control [QC]) for recombination-filtered core-single nucleotide polymorphism (SNP) phylogeny, multilocus sequence typing (MLST), capsular locus (KL)/lipooligosaccharide outer-core locus (OCL) typing, resistome profiling, and SNP-threshold micro-cluster definition. Results By BMD, carbapenem resistance was essentially universal (imipenem [IMI] 100% in both cities; meropenem [MEM] 100% in Makkah and 97.2% in Riyadh). VITEK II software showed high categorical agreement with BMD (94–100% across drug city strata) but non-zero very major error (VME) rates for carbapenems (up to 5.9%), supporting reflex BMD confirmation for decision-changing susceptible calls. The composite PCR marker ( blaOXA-23 or blaNDM ) achieved high sensitivity for imipenem resistance (100% in Makkah; 91.7% in Riyadh). WGS revealed global clone 2 (GC2)/sequence type 2 (ST2) predominance with interleaved Riyadh–Makkah clades (regional circulation) but predominantly single-city ≤10-SNP micro-clusters (0 mixed-city clusters; largest: 22 isolates in Makkah; largest Riyadh cluster: 5 isolates), indicating putative recent local transmission or shared-source signals. Conclusion A combined same-day PCR rule-in, reference MIC confirmation, and WGS micro-cluster surveillance strategy operationalizes rapid CRAB triage and outbreak triggering in hospitals in Saudi Arabia, enabling proactive infection-prevention containment.

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