Phenotypic and genotypic antimicrobial resistance profiles of gram-positive cocci and non-fermenting gram-negative rods

Antimicrobial resistance poses a significant global threat, particularly in healthcare settings, due to the spread of gram-positive cocci (GPCs) such as Staphylococcus spp. and Enterococcus spp. and non-fermenting gram-negative rods (NF-GNRs). This study investigates the phenotypic and genotypic resistance profiles of these pathogens. A total of 235 clinical isolates of GPCs and 108 NF-GNRs from a tertiary-care hospital were analyzed for antimicrobial susceptibility and resistance-associated genes. Chi-square tests were applied to assess genotype-phenotype associations, and multivariate analyses (principal component analysis, PCA) were used to explore clustering of isolates and antibiotics according to resistance profiles in each genus. Among GPCs, 69% of Enterococcus spp. and 71% of Staphylococcus spp. isolates were resistant to several first-line antimicrobials. Erythromycin, β-lactams, and tetracyclines showed the highest resistance rates, whereas nitrofurantoin, linezolid and fosfomycin remained largely effective against most isolates. In Enterococcus, PCA separated E. faecalis and E. faecium into two main resistance clusters driven mainly by ampicillin, vancomycin, teicoplanin, and erythromycin loadings, whereas for Staphylococcus no clear clustering by species was observed, and the variance was dominated by fluoroquinolones and selected β‑lactams. Acinetobacter baumannii and Pseudomonas aeruginosa grouped according to resistance, intermediate and susceptible profiles; however no robust grouping pattern could be inferred for Stenotrophomonas maltophilia due to the small sample size. Among NF‑GNRs, 89% of isolates were multidrug-resistant (MDR). Carbapenem resistance in A. baumannii reached 85% and was mainly associated with blaOXA‑23 and blaOXA‑143, while in P. aeruginosa ESBL genes (blaCTX‑M, blaSHV, blaGES, blaBEL‑1) and aac(6')‑Ib‑cr were detected at lower frequencies, and. maltophilia primarily carried sul2 (± sul1). This study highlights the high prevalence of antimicrobial resistance among both gram-positive and gram-negative clinical isolates, with multidrug-resistant phenotypes common in Staphylococcus spp., Enterococcus spp., A. baumannii, and P. aeruginosa. The identified phenotypic and genotypic resistance patterns underscore significant therapeutic challenges and importance for strengthening antimicrobial stewardship and surveillance programs.