Influence of Microbial Interactions in Mixed-community Biofilms on Proteus Mirabilis Encrustation in Catheters Associated With Urinary Tract Infection

Abstract Background Catheter-associated urinary tract infections (CAUTIs) are common healthcare-associated infections worldwide, with Proteus mirabilis a major causative agent. Geographical differences in healthcare, patient populations, catheterization practices, and antimicrobial resistance may influence biofilm composition, P. mirabilis interactions with coexisting uropathogens, and catheter encrustation. This study aimed to explore microbial interactions within P. mirabilis -dominated urinary catheter biofilms and evaluate associations between patient-related factors and P. mirabilis biofilm positivity. Methods 76 CAUTI patients at Colombo South Teaching Hospital, Sri Lanka, were recruited for the study, following the Centers for Disease Control and Prevention (CDC) 2024 criteria. CAUTI-associated uropathogens in urine and catheter tip samples were identified using HiCrome™ UTI agar. Biofilm formation was assessed using 3-(4,5-dimethylthiazol-2-yl)-2,5 diphenyltetrazolium bromide (MTT), crystal violet (CV) assays, and the qualitative tube method. Selected catheter samples were subjected to scanning electron microscopy with energy dispersive X-ray spectroscopy (SEM-EDX) for ultrastructure and elemental analysis, and atomic absorption spectroscopy (AAS) was used to measure Ca²⁺ and Mg²⁺ levels in biofilms with and without P. mirabilis . Results A total of 197 microbial isolates were identified from 76 catheter samples, comprising 95.43% bacteria and 4.57% Candida spp. E. faecalis (65.79%), E. coli (55.23%), Klebsiella spp. (55.23%), and P. mirabilis (47.37%) were the most frequently isolated organisms. Among the organisms co-isolated with P. mirabilis , E. faecalis (72.22%), E. coli (58.33%), and Klebsiella spp. (44.44%) were the predominant species. Biofilm positivity involving P. mirabilis showed statistically significant associations with males (p = 0.030; Chi-square test), long-term catheterization (≥ 30 days) (p = 0.030; Chi-square test), and alkaline urine (p < 0.001; Chi-square test). Mineral analysis showed increased calcium and magnesium accumulation in P. mirabilis biofilms, with calcium identified as the predominant element. In short-term catheterization, co-existing species such as E. faecalis and Klebsiella spp. reduced P. mirabilis biofilm formation, whereas long-term mixed-species biofilms enhanced biofilm maturation and mineral accumulation, indicating potential synergistic interactions. Conclusions P. mirabilis biofilms in CAUI are significantly associated with male sex, prolonged catheterization, and alkaline urine. Calcium-dominated mineral deposition was also associated with prolonged catheterization, indicating its potential contribution to catheter encrustation. Co-existing bacterial species exhibited variable effects on P. mirabilis biofilm development, indicating complex microbial interactions that influence biofilm maturation and catheter blockage.