MOLECULAR CHARACTERIZATION OF BIOFILM FORMATION IN MULTIDRUGRESISTANT PSEUDOMONAS AERUGINOSA ISOLATED FROM CLINICAL AND ENVIRONMENTAL SOURCES

Abstract

Pseudomonas aeruginosa is a major opportunistic pathogen implicated in persistent
healthcare-associated infections with poor treatment outcomes. Its capacity to develop multidrug
resistance (MDR) and to establish biofilms significantly complicates clinical management. This
study investigated the biofilm-forming ability, antimicrobial resistance profile, and distribution
of selected biofilm-associated genes among MDR P. aeruginosa isolates recovered from clinical
and environmental sources. A total of 414 samples comprising clinical specimens (urine, wound
swabs, ear swabs, high vaginal swabs, and catheter swabs) and environmental samples (soil, air,
water, toilet seats, window sills, hospital beds, and door handles) were collected from three
tertiary hospitals. Isolation and identification of P. aeruginosa were performed using standard
microbiological methods in accordance with CLSI guidelines and confirmed with the VITEK 2
system. Antimicrobial susceptibility testing was conducted using the modified Kirby–Bauer disc
diffusion method. Multidrug resistance was defined as resistance to at least one agent in three or
more antimicrobial classes. Biofilm formation was quantified using the microtiter plate crystal
violet assay, while polymerase chain reaction (PCR) was employed to detect biofilm-associated
genes (algD, pslD, lasR, pelF, and ppgL). Out of 101 confirmed P. aeruginosa isolates, 43
(42.6%) were of clinical origin and 58 (57.4%) were environmental. Wound swabs yielded the
highest number of clinical isolates, while soil samples accounted for the highest environmental
recovery. Multidrug resistance was observed in 20 (19.8%) isolates, with a higher proportion
among clinical sources. Biofilm production was detected in 31 (30.6%) isolates, comprising 5%
strong, 11% moderate, and 14.9% weak biofilm producers. Among biofilm-producing isolates,
10 (32.3%) were MDR. Molecular analysis revealed the presence of algD (80%), pslD (60%),
lasR (40%), and ppgL (30%), while pelF was not detected. A statistically significant association
was observed between strong biofilm formation and multidrug resistance (p < 0.05). The
findings highlight the clinical and environmental significance of biofilm-forming MDR P.
aeruginosa. Targeted infection-control strategies and biofilm-directed therapeutic approaches are
essential to mitigate the public health burden posed by this pathogen.