Detection of intracellular bacterial communities and biofilms in urinary tract infections with Escherichia coli using staining protocols

BACKGROUND Urinary tract infections (UTIs) are prevalent worldwide, and Escherichia coli (E. coli ) is the most common causative agent. The ability of the bacteria to form intracellular bacterial communities (IBCs) and biofilm is a major reason for UTIs. Studies have indicated that the persistence of uropathogenic E. coli as IBCs and biofilms has been implicated in UTIs. However, IBCs are not routinely identified by standard diagnostic methods. AIM To compare the various staining techniques for the detection of IBCs in urine samples from E. coli culture-positive UTI patients with the biofilm-forming capability of the isolates. METHODS The study included 73 patients with E. coli culture-confirmed UTI. Before antibiotic treatment midstream urine sample was collected, and the sediment was obtained by centrifugation. The samples were visualized using Sternheimer-Malbin, Wright-Giemsa, Safranin, and immunofluorescence staining to detect IBCs. Formation of biofilms was analyzed by the tube method. Descriptive statistics were used. RESULTS E. coli clusters were seen by light microscopy using various stains. However, immunofluorescence staining showed a better picture in the form of bright intracellular signals, which indicate bacterial aggregates. Biofilm assay showed an association with intracellular colonization. CONCLUSION The various staining techniques help in the identification of uropathogenic E. coli as IBCs inside superficial epithelial cells. These bacteria are also capable of forming biofilms, which resists action of antibiotics. Thus, IBCs and biofilms are rich reservoirs of organisms in the urinary bladder, paving the way for chronic treatment-resistant UTIs. This study requires further larger studies to substantiate these findings.