Bacterial characteristics of importance for recurrent urinary tract infections caused by Escherichia coli.

Urinary tract infections (UTIs) are among the most common bacterial infectious diseases encountered in clinical practice and account for significant morbidity and high medical costs. Escherichia coli is the most predominant pathogen causing 80-90% of community-acquired UTIs and 30-50% of nosocomially-acquired UTIs. Recurrent UTIs (RUTIs) are reported in 25% of women within 6 months of an acute UTI episode and pose a major problem. The aim of the present thesis was to look for bacterial characteristics of importance for recurrence of UTI caused by E. coli. The thesis is based on three papers. The study is based on E. coli from 236 Swedish women with community-acquired symptomatic lower UTI from a large study of 1162 patients treated with one of three different dosing regimens of pivmecillinam or placebo. The women were evaluated clinically and bacteriologically at the initial visit and at two scheduled follow-up visits. According to pulsed-field gel electrophoresis (PFGE) and culture results all primary infecting E. coli (initial isolates, pretherapy) were assigned into whether the initial infection was followed by cure, persistence, reinfection or relapse during follow-up. The prevalence of virulence factor genes (VFGs), phylogenetic groups, biofilm formation, plasmids and resistance to antimicrobials among primary infecting E. coli causing persistence or relapse at the follow-up visits were compared with the prevalence of these among E. coli followed by cure or reinfection. Previous studies of RUTI using phenotypically based typing methods or less specific DNA based typing methods have concluded, that RUTIs are mainly attributable to reinfection with new strains. However, applying PFGE showed that 77% of RUTIs were caused by a relapse with the primary infecting E. coli (Paper I). This may support the recent observation that E. coli can invade and replicate within the murine bladder forming biofilm-like intracellular bacterial communities (IBCs) and establish quiescent intracellular reservoirs that may represent stable reservoirs for RUTIs. The IBC pathogenic cycle has not been studied in humans; however, recently exfoliated IBCs were detected in urine from women with acute uncomplicated cystitis supporting the presence of the IBC pathway and occurrence of an intracellular bacterial niche in some women with UTI. Based on a triplex PCR E. coli can be divided into four main phylogenetic groups (A, B1, B2 and D). Phylogenetic group B2 was the most predominant group among the primary infecting E. coli followed by group D, A and B1. The majority of the tested 29 VFGs were associated with phylogenetic group B2, whereas only a few VFGs were more broadly distributed among the phylogenetic groups (Paper III). Primary infecting E. coli causing persistence or relapse of the infection were associated with phylogenetic group B2, whereas primary infecting E. coli followed by cure or reinfection were associated with group D (Paper II). Phylogenetic group B2 was associated with susceptibility to many of the tested antimicrobials, whereas group A was associated with resistance to many of these antimicrobials and multidrug resistant (MDR) strains, and group D with MDR strains. Phylogenetic group A and D were associated with carriage of IncH and IncI plasmids, respectively. Resistance patterns or plasmid profiles of the primary infecting E. coli were not associated with outcome during follow-up (cure, persistence, reinfection or relapse) (Paper II). Resistance to ampicillin, sulfamethizole, streptomycin and tetracycline was associated with a lower prevalence of some VFGs (sfa/focDE, agn43bCFT073, chuA, iroN, cnf1, hlyD, ibeA, malX, usp) and higher prevalence of other VFGs (afa/draBC, agn43aCFT073, iha, iutA, sat) but the aggregate VFG score did not differ among the resistant and susceptible strains of these antimicrobials (Paper III). Primary infecting E. coli causing persistence or relapse showed to have a higher biofilm formation capacity in vitro than those being followed by cure or reinfection (Paper II). This indicates that biofilm may be an important determinant for developing RUTI and may support the observation of IBCs. Primary infecting E. coli causing relapse or persistence had a higher aggregate VFG score and higher prevalence of hemolysis and of many of the VFGs than those followed by cure or reinfection. The VFGs associated with persistence or relapse included: adhesins (sfa/focDE, papAH), a biofilm related factor (agn43), iron-uptake systems (chuA, fyuA, iroN), protectins (kpsM II, kpsMII K2), toxins (cnf1, hlyD), a marker of a pathogenicity-associated island from CFT073 (malX), and a bacteriocin-like factor (usp). No specific combination of VFGs could predict persistence or relapse (Paper III). A regimen of three days pivmecillinam therapy for primary infecting E. coli positive for at least one of a number of traits (phylogenetic group B2, sfa/focDE, papAH, agn43, chuA, fyuA, iroN, kpsM II, kpsM II K2, traT, cnf1, hlyD, ibeA, malX, usp and being hemolytic) gave a significantly higher prevalence of persistence or relapse as opposed to primary infecting E. coli subjected to three days therapy with absence of these traits or primary infecting E. coli subjected to seven days therapy irrespective of these traits (Paper III). In conclusion, our results may support the hypothesis of an intracellular reservoir of E. coli in the bladder. The recognition of uropathogenic E. coli as a potential intracellular pathogen challenges our current treatment regimens of UTI and argues for the development of new antimicrobials or treatment regimens/strategies. No distinct virulence profile could predict RUTI. However, we found VFGs associated with persistence or relapse that may be potential targets for prevention and treatment of UTI. Furthermore we identified potential markers that may be used to select a more differentiated and optimal treatment. Future studies must explore the function of these VFGs and other putative and novel VFGs in relation to persistence or relapse of UTI and their possible role in IBC formation. Defining the repertoire and mechanism of VFGs could facilitate the development of new diagnostic tools, regimens and drugs for prevention and treatment of RUTI.