Urinary tract infections (UTIs) are widespread and affect a large proportion of the human population. About 150 million people worldwide develop UTI each year, with high social costs (Flores-Mireles et al., 2015). It is estimated that 40% of women develop at least one UTI during their lifetime (Micali et al., 2014) and that 11% of women over 18 years have an episode of UTI per year (Foxman and Brown, 2003; Foxman, 2014). With roughly eleven-million cases reported in the sole U.S. each year, the costs are estimated $5 billion annually (Figure 1) (Foxman, 2014).

The UTI refers to the presence of a certain number of bacteria in the urine (generally > 10⁵/ml) and symptomatic UTIs are classified in order of severity as urosepsis syndrome, pyelonephritis (or upper UTI, with infection in the kidney) and cystitis (or lower UTI, with bacteria into the bladder; Foxman, 2014; Smelov et al., 2016). Clinically, UTIs classification comprises either uncomplicated or complicated cases, depending on the presence of structural or neurological urinary tract abnormalities (Zacché and Giarenis, 2016). The ORENUC system classifies the risk factors according to the phenotype (Johansen et al., 2011): O, no known risk factors; R, risk of recurrent UTIs without a more severe outcome; E, extraurogenital risk factors; N, relevant nephropathic diseases; U, urologic resolvable (transient) risk factors; C, permanent external urinary catheter and unresolved urologic risk factors (see also Smelov et al., 2016 for a modified classification). The susceptibility to develop an UTI phenotype is related to several factors, as dysfunctions of the urinary tract and/or genetic mechanisms involved in the innate immune response control to infections (Koves and Wullt, 2016). In particular, the innate immune system may respond either to UPEC patterns (pathogen-associated molecular patterns; PAMPs) or to molecules derived from damaged or dying cells (danger/damage-associated molecular patterns; DAMPs). Pattern recognition receptors (PRRs) recognized these patterns in specialized immune cells, epithelia, and other tissues (Purves and Hughes, 2016). Assembling in the cytosol of multimeric protein complexes (inflammasomes) occurs after sensing PAMPs or DAMPs structures that can be formed in both upper and lower urinary tract (Guo et al., 2015). They trigger innate immune responses through mechanisms depending or not from the production of proinflammatory cytokines (Purves and Hughes, 2016).

The bacterial cystitis (also called acute cystitis) can occur in both women and men and some people develop recurrent infections of the urinary tract (Fiore and Fox, 2014). Three or more urinary tract infections within 12 months define the recurring UTI, as well as two or more recurrences within 6 months. The same bacterial species that caused previous infection is typically responsible for relapses. Approximately 20–30% of adult women with an initial UTI will experience a recurrence within 3–4 months; whereas, in children, about one third experiencing a UTI before the age of one, will experience a recurrence within 3 years, and 18% of them will have a recurrence within a few months (Nuutinen and Uhari, 2001). However, these figures are understated; in fact, about 50% of UTI does not come to medical attention. Recurrent UTIs can be introduced from different sources and the same or different UTI-causing strains in the gut are able to (re)inoculate the bladder. Alternatively, bacteria residing in the bladder epithelium are able to re-emerge periodically and cause UTI recurrence (Silverman et al., 2013). In patients suffering from recurrent UTIs, maintenance is ensured by antibiotic prophylaxis; however, in some cases UTI needs to be treated by surgery (Tolg and Bagli, 2012). During pregnancy, recurrent UTIs may be frequent and can cause severe adverse outcomes for the mother and the baby, including preterm birth. The interventions in this setting can be pharmacologic (antibiotics) or non-pharmacological (alternative remedies; Schneeberger et al., 2012). In pre-menopausal women, sexual activities three or more times a week, the use of spermicides, new or multiple sexual partners and having suffered from UTI before age 15 are the main risk factors in UTI development and recurrence. In menopausal women, systemic hormonal therapy is not an effective prevention and usually asymptomatic bacteriuria during this period does not require treatment (Milart et al., 2013). In women after menopause, the risk increases mainly by low estrogen levels after-effects, which are often associated to vaginal atrophy (Arnold et al., 2016). In women over the age of 61–65 years, half have suffered of genital-urinary symptoms while 29% had episodes of urinary incontinence, all symptoms associated with bacteriuria (Raz, 2001).

UPEC is the main cause of community-acquired UTIs (about 80–90%; Foxman, 2014; Flores-Mireles et al., 2015). Four main UPEC phylogroups (A, B1, B2, and D) have been identified on the basis of the occurrence of genomic Pathogenicity Islands (PAI) and the expression of virulence factors, such as adhesins, toxins, surface polysaccharides, flagella, and iron-acquisition systems (Bien et al., 2012). Usually, many of these virulence factors are required for UPEC to cause UTI (Hannan et al., 2012). However, besides UPEC, UTI can be caused by Klebsiella pneumoniae (about 7%), Proteus mirabilis (about 5%), and Pseudomonas aeruginosa, Enterococcus faecalis, Enterobacter cloacae, Streptococcus bovis, and the fungus Candida albicans (for the remaining percentage; Parish and Holliday, 2012; Palou et al., 2013; Hof, 2017). During UTIs, UPEC pathogenesis includes: (a) UPEC colonization of the periurethral and vaginal areas with colonization of the urethra; (b) ascending into the bladder lumen and growth as plantktonic cells in urine; (c) adherence to the surface and interaction with the bladder epithelium defense system (see below); (d) biofilm formation; (e) invasion and replication by forming bladder Intracellular Bacterial Communities (IBCs) where quiescent intracellular reservoirs (QIRs) form and reside in the underlying urothelium; (f) kidney colonization and host tissue damage with increased risk for bacteremia/septicemia.

Replication of bacteria in the IBC can easily reach as many as 10⁵ bacteria per cell; furthermore, bacteria in the IBC undergo morphological changes, flux out of the infected cell, and go onto infect neighboring cells (Dhakal et al., 2008; Flores-Mireles et al., 2015; Spaulding and Hultgren, 2016). The flushing of urine removes most of the invading bacteria, along with UPEC-filled exfoliated bladder epithelium cells (BECs; Kaper et al., 2004).

UPEC colonize the bladder using a variety of virulence factors that therefore play critical roles in UTI pathogenesis. These include surface structural components, such as lipopolysaccharide (LPS), polysaccharide capsule, flagella, outer-membrane vesicles, pili, curli, non-pilus adhesins, outer-membrane proteins (OMPs), as well as secreted toxins, secretion systems, and TonB-dependent iron-uptake receptors, including siderophore receptors (Figure 2). All of these components are attractive candidates for the development of new drugs and vaccines (Klemm et al., 2010; Werneburg et al., 2015; O'Brien et al., 2016).

LPS are molecules with amphipathic properties consisting of fatty acids lined to an oligosaccharide core, which in turn is bound to a long polysaccharide chain commonly called O antigen (Simpson et al., 2015). LPS structural constituents mediate multiple aspects of the UPEC life cycle, including the ability to acutely colonize bladders, form reservoirs, and evoke innate and adaptive immune responses (Aguiniga et al., 2016). LPS provide resistance against hydrophobic antibiotics and hypersensitivity to hydrophobic toxic molecules (such as bile salts and some antibiotics) occurs when the amount of LPS at the cell surface is decreased (Zhang et al., 2013).

In UPEC, the fim operon encodes type 1 pili (expressing an hemagglutination which is mannose-sensitive), whereas the pap operon encodes P- or Pap-pili (which are able to interact with the digalactoside unit in the P-blood group antigen). In UPEC clinical isolates, fim operon is constitutive whereas pap is part of a PAI that is also responsible for other putative virulence determinants. Generally, both types of pili are heteropolymeric consisting of a major pilus protein subunit that provides the pilus stalk and several minor subunit proteins at the distal end, with PapG and FimH representing the actual adhesins. PapG and FimH are composed by two domains, the first allows copolymerization and is made by a pilin domain, whereas the second is a lectin domain able to bind carbohydrates (Kline et al., 2009). The chaperone-usher (CU) pathway assembles pili. More than 1,000 copies of the FimA major pilin form the type 1–pilus rod, while at its distal end the pilus tip contains the FimH adhesin followed by single copies of the FimG and FimF adaptor subunits. Mannosylated proteins that are present on the bladder epithelium bind to FimH in a Rho GTPases (Rac1)-mediated host actin cytoskeleton rearrangement-dependent manner (Eto et al., 2007). This eventually leads to the development of cystitis due to bacterial invasion (Figure 2; Hahn et al., 2002). In addition, the expression of type 1 pili is strictly controlled by phase variation, which reversibly switches between the type 1 pili active expression (Phase-ON, piliated cells) and loss of expression (Phase-OFF, non-piliated cells; Schwan, 2011). Molecular pathways, which are involved in reversible switching between ON-OFF Phases, are strictly regulated by environmental signals within the urinary tract such as acidic pH and salt growth conditions.

Six different subunits which are arranged into two distinct subassemblies (the tip fibrillum and the pilus rod) form the P pilus. At the distal end, the tip fibrillum is composed of one PapG adhesin followed by PapF and PapE subunits. The pilus rod is made by more than 1,000 copies of the PapA subunit. The adaptor subunit PapK connects the above subunits to the PapA rod, which is a superhelical structure at the base of the pilum (Figure 2; Busch and Waksman, 2012).

Curli are bacterial surface appendages that secrete subunits from the cell as soluble monomeric proteins and possess the typical structure and physical characteristics of amyloid fibrils. which are known to be formed in some human degenerative diseases. The bacterial amyloids may facilitate biofilm formation (Goyal et al., 2014). In UPEC, curli formation is coordinated by proteins encoded in the operons csg DEFG. The operon-accessory proteins CsgE, CsgF, and CsgG are required to facilitate the secretion of CsgA whereas CsgB nucleates CsgA subunits into curli fibers (Figure 2; Chapman et al., 2002; Barnhart and Chapman, 2006).

While pili are involved in the initial attachment of UPEC to the urinary tract mucosa, UPEC elaborate numerous other afimbrial ahesins. In fact, the adhesin TosA is present in about 30% of urinary tract isolates and is expressed during UTI (Vigil et al., 2011). Another adhesin, FdeC, is involved in colonization of the bladder and kidneys in a mouse model of infection (Nesta et al., 2012), whereas the iron-regulated adhesin Iha mediates adherence to BECs (Johnson et al., 2005).

Moreover, the large majority of UPEC isolated from women with acute, asymptomatic, or recurrent UTIs shows the presence of flagellum-mediated motility (Wright et al., 2005). Flagella (Figure 2) are organelles that confer adhesive and invasive properties to some EPEC strains (Giron et al., 2002) and play a key role in the dynamic of biofilms (Pratt and Kolter, 1998). It was recently reported that during biofilm formation, flagella play different roles such as adherence, maturation, and dispersal as shown by gene expression and regulation during the growth phase (Nakamura et al., 2016).

On the other hand, UPEC toxins play different pathogenetic roles during infection. The α-hemolysin is in fact associated with renal damage and scarring, induces Ca²⁺ oscillations in renal tubular epithelial cells, thereby potentially enhancing ascension and colonization of ureters and kidney parenchyma by disrupting the normal flow of urine. Recently (Nagamatsu et al., 2015), α-hemolysin was found to induce proinflammatory Caspase-1/Caspase-4-dependent cell death in bladder epithelial cells, resulting in cell exfoliation (see below).

UPEC toxins, adhesins, enzymes, and non-protein antigens like LPS are not released as soluble molecules; rather, they are associated with outer-membrane vesicles, which bud off the surface of Gram-negative bacteria during all stages of growth (Figure 2; Ellis and Kuehn, 2010). The formation of membrane vesicles is considered a “smart” way to protect bacterial toxins and an efficient system to deliver them into host cell (Wiles et al., 2008).

Iron acquisition is a critical requirement for UPEC survival in an environment that is iron-limited as the urinary tract (Skaar, 2010). Thus, is not suprising that IBC UPEC show upregulation of redundant systems for the acquisition of iron (Reigstad et al., 2007). In this regard, siderophores are small-molecule iron chelators that are produced by UPEC strains to scavenge ferric iron (Fe³⁺), thus UPEC express yersiniabactin, salmochelin, and aerobactin. Siderophore receptors require the TonB cytoplasmic membrane-localized complex, a high-affinity iron acquisition system that allows binding and chelation of iron at the cell surface to promote its uptake (O'Brien et al., 2016).

However, uroepithelial cells, to prevent bacterial iron scavenging, upregulate genes for the transferrin receptor and for lipocalin 2.

Lastly, further UPEC factors associated with colonization have been linked to the regulation of metabolic pathways mediated by two-component signaling systems (TCSs). TCSs are main signal transduction pathways by which bacteria sense and respond to a wide array of environmental stimuli, including quorum sensing signals, nutrients, antibiotics. TCSs are composed by a membrane-bound sensor histidine kinase (HK) and a cytoplasmic response regulator (RR) that functions by regulating gene expression (Stock et al., 2000). Among UPEC-associated TCSs involved in UTI pathogenesis, the BarA/UvrY system has been described to regulate switching between glycolytic and gluconeogenic pathways (Tomenius et al., 2006) the EvgS/EvgA and PhoQ/PhoP systems have been involved in acid resistance (Eguchi et al., 2011), while the function of KguS/KguR is in the control of the utilization of α-ketoglutarate. In this way they facilate the adaptation of UPEC in the urinary tract (Cai et al., 2013).

The importance of the above described UPEC virulence factors in UTI pathogenesis has been further supported, in recent years, by the application of multiple “omics” technologies aimed at investigating the UPEC genomic diversity, the global gene expression in different models of infection both in vitro and in vivo, and to define the occurrence of UPEC-specific proteins as new candidate therapeutic and vaccine targets (as recently reviewed by Lo et al., 2017).

Next-generation sequencing (NGS) technologies are providing rapid low-cost determination of UPEC genomes useful to monitor outbreaks, epidemiology of emerging strains, as well as evolution of resistance (Petty et al., 2014; Stoesser et al., 2016). On the other hand, analysis of different UPEC genomes and the comparison with the E. coli genomic database revealed the plasticity of UPEC pan genome, and the presence of UPEC-specific PAIs genes predicted to encode putative virulence factors, such as pilus proteins, adhesins, and iron-uptake systems (Moriel et al., 2016).

Transcriptomics investigations by both microarrays and NGS-based RNA sequencing (RNA-seq), on the other hand, has led to the identification of virulence and fitness UPEC genes, expressed during different in vitro and in vivo infection-relevant conditions. In this regard, RNA-seq-based transcriptome analysis of mouse macrophages infected in vitro with two UPEC strains, allowed to identify strain-specific differentially expressed genes associated to the survival in macrophages, such as those involved in the responses to oxidative stress, as well as those involved in the initial adhesion of UPEC to cells, such as multiple flagella genes (Mavromatis et al., 2015). Moreover, the global gene expression of different UPEC strains has been investigated by RNA-seq of urine samples collected from UTI patients. These transcriptomics studies defined the global transcription profile for UPEC during UTI, highlighted the high genomic diversity of different UPEC strains, and confirmed, on a global scale, the expression during UTI of several genes encoding virulence factors. In fact, it has been observed the transcription of genes associated with the UPEC's adhesion to the uroepithelium (type 1 and P pili), of genes involved in iron uptake (enterobactin, aerobactin, yersiniabactin, and salmochelin), of genes encoding toxins (hemolysins nad cytotoxic factors), as well as those involved in copper efflux (Bielecki et al., 2014; Subashchandrabose et al., 2014).

High-resolution liquid chromatograph-mass spectrometry/mass spectrometry (LC-MS/MS)-based technology has been applied to identify and characterize the surface proteome of UPEC isolates and of strains grown in human urine (Wurpel et al., 2015, 2016). These studies identified several expressed proteins highly conserved among different strains, thus representing the core surface proteome of UPEC. UPEC core surface proteins, such as integral Outer Membrane (OM) proteins (e.g., OmpA, OmpC, OmpF) and several iron-uptake proteins, were in fact detected in more than 80% of strains (Wurpel et al., 2015). Clearly, characterization of those UPEC surface proteins that are conserved among different strains and immunogenic is an essential step for identifying potential vaccine candidates and new therapeutic targets (Cash, 2014).

Moreover, new insights into spatial changes in the UPEC proteome under experimental conditions mimicking bacterial growth in the urinary tract, have been provided by MALDI TOF IMS-based proteome profiling of differentially expressed proteins within UPEC biofilms. The application of this technique, that allows for in situ two-dimensional assessment of protein spatial distribution and abundance, revealed the occurrence of different bacterial subpopulations within biofilms: a type-1 pili-expressing cells localized at the air-exposed region and a curli-equipped population localized to the underlying air-liquid interface (Floyd et al., 2015).

Together, all the above mentioned “omics” approaches have allowed a great deal of new information to be available and that is enabling a more comprehensive understanding of UPEC's pathogenic mechanisms.

The most commonly targeted site of UTIs is the bladder. The bladder epithelium possesses powerful barriers and the BECs show antibacterial activities. Despite their properties, BECs and the bladder epithelium are often circumvented by UPEC (Wu et al., 2017). As discussed, the progressive ascending colonization of bacteria contaminates the urethra and the origin of this infection is usually from the gut (Kaper et al., 2004). Owing to the presence of urine, that represents an ideal growth broth, bacteria proliferate in a relatively short time lapse, while the flushing of urine during urination removes most of the invading bacteria. However, bacterial strains are able of binding tightly to BECs lining the bladder using fimbrial organelles (Duncan et al., 2004; Chahales and Thanassi, 2015).

The multilayered bladder epithelium is also known as “transitional epithelium” and it is composed by three layers: basal cell layer (5–10 μm in diameter), intermediate cell layer (20 μm in diameter), and superficial apical layer with large hexagonal cells (diameters of 25–250 μm), which are also termed “umbrella cells.” A basement membrane lies underneath the basal epithelium (Figures 3A,F). The umbrella cells play a prominent role in maintaining a barrier against most substances found in urine, and show a number of properties, including specialized membrane lipids, asymmetric unit membrane particles, and a plasmalemma with stiff plaques. These plaques may cover up to 90% of the urothelial cell surface, with each plaque being composed of nearly 1,000 subunits. These subunits are made by proteins (uroplakins, UPs), which serve as the major receptors for UPEC adherence to the host cell and are localized within plaques on the apical membranes of the mature umbrella cells (Veranic et al., 2004). There is a correlation between the glycosylation changes in UPs and the different pathological conditions of the urothelium such UTI and interstitial cystitis (Birder, 2005; Katnik-Prastowska et al., 2014; Habuka et al., 2015).

The fusiform vesicles (FVs) are unique cytoplasmic organelles contained in the umbrella cells. FVs deliver preassembled crystalline arrays of UP proteins to the apical cell surface of urothelial umbrella cells. Different Rab GTPases function as regulators of specific steps in membrane traffic pathways and are localized to the cytosolic face of specific intracellular membranes. Rab27b, is a small GTPase regulating intracellular vesicle movement which is expressed at an extraordinary high level (0.1% of total protein) in urothelium. The Rab27b+ FVs are involved in the storage of extra membrane which are necessary when urine accumulates and causes bladder expansion (Wankel et al., 2016). In order to enter epithelial cells, UPEC coopt the superficial epithelial cells by expoiting their bladder volume-regulating properties by stimulating the exocytosis of fusiform vesicles right where the bacterial attach. The adherent bacteria are then internalized when these membranes are subsequently retracted into cells (Figure 3B; Wu et al., 2017). UPEC have been found to reside within Rab27b/CD63/Caveolin-1-positive fusiform vesicles (O'Brien et al., 2016). Internalized UPEC become encased in Rab27b+ fusiform vesicles within the cytosol of the superficial epithelium (Figure 3B; Bishop et al., 2007). Replication of internalized UPEC bacteria rapidly occurs, resulting in the maturation of IBCs, a structure that possesses biofilm-like properties which is protected from innate defenses and antibiotics (Justice et al., 2006; Goller and Seed, 2010). Fusion with lysosomes is thus impaired, because internalized bacteria are mostly encased in Rab27b+ compartments.

Defense mechanisms of bladder epithelial cells against intrusion of bacterial include receptors such as toll-like receptors (e.g., TLR2, TLR4, TLR5, and TLR11) that are able to promptly recognize intruding bacteria (Larue et al., 2013). After UPEC encapsulation within RAB27b+ vesicles in BECs, intracellular UPEC are recognized by TLR4 which increases intracellular cyclic AMP (cAMP) levels (Figure 3B). This triggers the exocytosis of RAB27b+ vesicles harboring UPEC and the intracellular bacterial expulsion back into the bladder lumen (Figure 3C).

However, some UPEC break the RAB27b+ vacuole and cannot be expelled into the urine; thus, these bacteria are targeted by autophagy and delivered into the lysosomes, where they actively neutralize the pH by reducing their acidicity and degradative potential (Abraham and Miao, 2015). These malfunctioning lysosomes are sensed by a lysosomal transient receptor potential mucolipin 3 Ca²⁺ channel (TRPML3), which is localized on the membrane of lysosomes (Miao et al., 2015). The activation of this Ca²⁺channel rapidly fluxes out into the cytosol the Ca²⁺ stored in the lysosome, which induces the spontaneous expulsion into the extracellular space of the defective lysosomes and its contents (Figure 3D).

Pathogen sensing by TLR4 induces the production of various soluble factors which are secreted by BECs, including antimicrobial peptides (AMP, such as cathelicidin and β-defensin 1; Sun et al., 2013; Chromek, 2015), antimicrobial proteins [such as pentraxin 3 (PTX3); (Uzun et al., 2016)] and chemokines [such as CXC-chemokine ligand 1 (CXCL1) and CC-chemokine ligand 5 (CCR5); Schiwon et al., 2014; Figure 3E]. Attachment to the urothelium or bacterial lysis are inhibited by these antimicrobial peptides, which are also induced when bacteria succeed to attach to the urothelium (Spencer et al., 2014). Moreover, excretion in the urine of uromodulin, a major high mannose-containing glycoprotein, exerts a protective effects against UTI by competing with the binding of UPEC FimH to uroplakin Ia (Pak et al., 2001).

When all these export mechanisms fail to clear the urothelium from the invading UPEC, BECs activate the last line of defense. Acute infections are commonly associated with of the exfoliation of the epithelium, with the loss of a large numbers of superficial epithelial cells. Exfoliation is followed by an efficient restoring of superficial BECs through active proliferation of basal progenitor mast cells (MCs). BECs exposed to UPEC release copious amounts of interleukin-1β (IL-1β) that regulates migration of multiple cell types including neutrophils and MCs (Choi et al., 2016). Exfoliation is also triggered by caspase 3- and caspase 8-dependent apoptosis of infected BECs, which shed into the bladder lumen (Figure 3F). However, exfoliation, which is an efficient host defense strategy, may is some cases favor the dissemination of bacteria, by clearing the way to deeper tissues. Indeed, the death of the superficial epithelium is intentionally induced by certain virulent UPEC to better reach deeper tissue where intermediate BECs are located and where they form QIRs and where they can persist for extended period of time. Actually, one of the main reason for high rate recurrence of infections in the bladder and resistance to antibiotics is associated to the presence of QIRs within subepithelium (Leatham-Jensen et al., 2016).

The immune system operates with different and specific strategies to reduce inflammation and to preserve tissue integrity. The direct phagocytosis of bacteria is operated by neutrophils that also clear bacteria through extracellular burst of ROS, which are highly toxic to bacteria (Aubron et al., 2012). Intimate crosstalk between LY6C− and LY6C+ macrophages ensures the precise initiation of neutrophil responses (Figure 3E). Local LY6C− macrophages release CC-chemokine ligand 2 (CCL2), CXC-chemokine ligand 1 (CXCL1) and macrophage migration inhibitory factor (MIF) to recruit LY6C+ macrophages and neutrophils from the bloodstream (Schiwon et al., 2014; Figure 3F). LY6C+ macrophages, as a consequence of infection sensing, secrete tumor necrosis factor (TNF), which acts on local LY6C− macrophages to trigger their production of CXCL2. The last is responsible for spontaneously production of matrix metalloproteinase 9 (MMP9) by neutrophils and their transepithelial movement (Nathan, 2006). The resident LY6C− macrophages play a major role as the main pro-inflammatory cells, whereas the recruited LY6C+ macrophages keep neutrophils in close proximity before targeting the pathogen (Abraham and Miao, 2015).

The efficacy of antibiotic treatment depends on the identification and antimicrobial resistance pattern of uropathogens responsible for UTI (Bartoletti et al., 2016). The practice of prescribing antibiotics to treat UTI without bacterial characterization led to increased resistance among uropathogens and to decreased effectiveness of oral therapies. Despite clinical symptoms of UTIs have been ameliorated by numerous antibiotics, UPEC persistence and resistance to antibiotics represent a serious problem (Blango and Mulvey, 2010). According to the 2015 guidelines of the European Association of Urology, the recommendations for the prevention of recurrent UTI are first aimed at behavioral changes and immediately after toward non-antibiotic measures. If these two recommendations are not sufficiently effective then the antibiotic prophylaxis should be considered, in order to prevent the adverse events and collateral damages that the long-term and not necessary use of antibiotics may cause (Vahlensieck et al., 2016). In Europe, resistance to UPEC isolates shows average values of 11.8% for third-generation cephalosporins and 22.3% for fluoroquinolones. In the U.S., fluoroquinolone-resistant UPEC represented the 31.3% of isolates among hospitalized patients between the years 2007 and 2010 (Edelsberg et al., 2014). These data confirm the general consideration that number of effective antibiotic compounds availability and the prevalence of antibiotic resistance are worsening, as demonstrated by an increased number of clinical studies (Bartoletti et al., 2016).

Antimicrobial prophylaxis for women with recurrent UTI include, for example, 50 mg or 100 mg of nitrofurantoin once a day; 100 mg of Trimethoprim (TMP) once a day; 40/200 mg TMP/sulfamethoxazole (co-trimoxazole) once a day or three times a week; 3 g of fosfomycin trometamol every 10 days and, during pregnancy, for example, 125–250 mg of cephalexin or cefaclor 250 mg once a day (Grabe et al., 2015; Giancola et al., 2017). Among other antibiotics, imipenem represents the best efficient antibiotic against all UPEC strains (100%), followed by ertapenem (99.98%), amikacin (99.94%), and nitrofurantoin (99.91%). Carbapenems like imipenem represent the best option for the treatment of extended-spectrum beta-lactamase (ESBL) strains (Idil et al., 2016). UPEC strains are also susceptible to ciprofloxacin (Tosun et al., 2016), cefotaxime, piperacillin/tazobactam (Dizbay et al., 2016), azithromycin, doxycycline and ceftriaxone (Saha et al., 2015). However, several UPEC isolates are resistant to ampicillin, oral first-generation cephalosporins, TMP-sulfamethoxazole (Moya-Dionisio et al., 2016), cefuroxime (Chang et al., 2016), cotrimoxazole (Saha et al., 2015), amoxicillin-clavulanate, nalidixic acid, cefradine, and aminopenicillins (Narchi and Al-Hamdani, 2010). In some cases, the combined effect of different antibiotics prompted a significant increment in susceptibility, as found for triclosan with amoxicillin and gentamicin (Wignall et al., 2008). A retrospective analysis has identified ciprofloxacin as the most used antibiotic for empirical therapies (76% of cases; Parish and Holliday, 2012).

Due to ecological side effects, the oral cephalosporins and fluoroquinolones are no longer recommended as routine treatments, except for specific clinical situations. Furthermore, the worldwide increment of UPEC strains resistant to TMP questions its use with or without a sulfonamide as an effective prophylactic agent (Idil et al., 2016). High urinary levels of levofloxacin are not enough to cure UTIs and the combination of ceftolozane/tazobactam was more effective as an alternative treatment in settings of increased fluoroquinolone resistance (Huntington et al., 2016). Increased resistance of UPEC strain isolates against ampicillin (96.42 %), tetracycline (85.71 %), amikacin (71.42 %), ciprofloxacin (67.85 %), and gentamycin (58.71 %) has been found in pregnant women with history of recurrent UTIs (Habibi and Khameneie, 2016). Finally, numerous government agencies advise against the long-term use of prophylactic nitrofurantoin because of rare but serious pulmonary and hepatic adverse effects (Vahlensieck et al., 2016).

New antibiotics, such as colistin (Cui et al., 2016), finafloxacin, and cefiderocol (S-649266), which are currently in early clinical development, might be useful in the treatment of UTIs (Zacché and Giarenis, 2016).

Figure 4 shows the structure formulae of the most representative antibiotics for which UPEC resistance has been demonstrated (red background) and those showing susceptibility to UPEC (green background). The yellow background shows antibiotics that already show resistance in some UPEC strains.

Antibiotics will continue to be an unavoidable source for the prevention of UTIs on a case-by-case basis. However, the excessive use of antibiotics and the long-term interference with intestinal microbiota, require to search for alternative remedies. A plethora of molecules has been tested to reduce UPEC infections by exploiting their ability either to stimulate the immune system or to interfere with the UPEC ability to adhere and invade the urothelium. Here we briefly summarize the most effective alternative remedies to fight UPECs.

Uropathogenic Escherichia coli infections pose a serious problem to human health, with societal costs of tens of billion US$ worldwide. The increased resistance to both synthetic and natural antibiotics causes recurrence and chronicity of infection, with the emergence of new and more serious illnesses. There are both direct and indirect strategies against UPEC infections. Direct strategies are targeting bacteria viability, bladder epithelium adhesion and biofilm formation; indirect strategies elicit and enhance immune responses, by stimulating infected tissues and cells to overreact to UPEC invasion.

In this review, we provided a comprehensive picture of the UPEC infection, with the aim of giving the reader elements from both sides of the problem: the pathogen infectivity and the human cell response. The ongoing search for new and more effective antibiotics and the discovery of natural products from plants, fungi and non-pathogenic bacteria shall consider the multiple aspects of UPEC infection. Figure 12 summarizes the four main areas representing the Strengths and the Weaknesses of UPEC, the Opportunities for alternative remedies to antibiotics and, finally, the Treats that UPEC cause to human health. While antibiotics will remain the main drugs to reduce infections, alternative chemicals may offer the opportunity to (a) by-pass antimicrobial resistance and (b) show multi-targeted action, with the potential of targeting both the UPEC and the innate immune system. We cannot exclude that new therapies may include both antibiotics and natural products altogether, as recently demonstrated (Vadekeetil et al., 2016). An increasing body of evidence shows that reduction of adhesion of UPEC to urinary tract tissues reduces recurrence and increases recovery. Future interesting targets might not only be directed to UPEC adhesins, but also to immune-based strategies able to improve cell responses to UPEC infection; in this context, several natural products fit this strategy.

MT, GG and MM wrote the manuscript.