Biofilm Formation and Antibiotic Susceptibility Patterns in Contact Lens-Associated Bacteria

Purpose To characterize biofilm-forming capacity, extracellular polymeric substance (EPS), and antibiotic susceptibility (AST) of bacteria isolated from contact lenses (CL) and their accessories.

Methods Bacterial isolates from 20 CLs and their cases from asymptomatic participants were assessed for biofilm formation using crystal violet (CV) staining in tubes and tissue culture plates (TCP). AST was performed by the Kirby-Bauer method. EPS production was detected using Congo red agar (CRA) and broth (CRB), with the contents of carbohydrate, protein, and eDNA assessed. Biofilm structures were visualized, and the effect of different physiological conditions on biofilm was studied.

Results The bacterial isolates included A. xylosoxidans, K. pneumoniae, S. aureus, S. cohnii, S. epidermidis, and S. saprophyticus, identified and confirmed by MALDI TOF MS. Out of 110 isolates, biofilm production by CV staining showed 28.5% strongly positive, 38.09% moderately positive, and 23.33% weakly positive by the tube method, and 32.07%, 18.57%, and 29.04% respectively by TCP method; 14.28% were non-biofilm producers. The selected isolates were susceptible to the tested antibiotics, but resistance was noted in A. xylosoxidans. The EPS production was confirmed by black coloration in CRA plates, and CRB tubes with carbohydrate content was found to increase, whereas protein content was slightly reduced, and the presence of eDNA was detected after 48 hrs. By Maneval's stain, a pink bacterial cell embedded in blue-colored extracellular matrix (ECM) is observed. The SEM images depicted the presence of branched and sheet-like ECM structures. Physiological conditions like acidic pH and the presence of ribose sugar showed reduced biofilm formation; conversely, incubation at 28°C enhanced bacterial attachment. In Staphylococcus species, increased salt concentration supported biofilm formation.

Conclusion This study shows that bacteria present on contact lenses of asymptomatic users possess significant biofilm-forming capacity. We conclude that A. xylosoxidans and coagulase-negative Staphylococcus were predominant and capable biofilm formers. CRB was found to be a convenient qualitative method for detecting EPS production in this study. The bacterial attachment on CL may vary with physiological conditions. These findings reflect presence of bacteria rather than active infection and highlight the importance of hygiene and preventive strategies during contact lens use.