Chronic bacterial prostatitis (CBP) is one of the important factors affecting male reproductive health that is characterized by a high prevalence, low cure rate, frequent recurrence, and severely affect patient’s quality of life. An increased risk of benign prostatic hyperplasia and prostate cancer have found to be correlated with chronic prostatitis [12] [13]. Expressed prostatic secretions (EPSs) were used as the study samples because they are produced from the prostate and could acquire bacteria while they pass through the urethra, so they reflect the composition of bacterial communities inhabiting both prostate and the urethra. The 16S rRNA gene positive rate in CBP/CPPS is similar to that among patients with localized prostate cancer [14], so future studies should be performed to investigate the role of bacterial community agents causing CBP/ CPPS in prostate cancer development.
Identifying the causative bacterium is important for antibiotic selection and the determination of a treatment strategy in the management of any infective disease such as chronic prostatitis, in general, clinical features and treatment methods vary according to the causative microorganism.
In this study, we found that the most prevalent bacterial pathogens from EPS of 10 patients diagnosed with CBP were belong to genus Enterobacter. Species level identification of isolated Enterobacter strains indicated their relation to species of Enterobacter cloacae complex (ECC), three of them (MAG-06, MAG-07 and MAG-010) are related to E. cloacae while Pb1 and Pb2 are related to E. kobei. The remaining two isolates were related to Providencia stuartii (MAG-8) and pseudomonas stutzeri (MAG-09).
The Enterobacter cloacae complex (ECC) includes common nosocomial pathogens (E. cloacae is the most important within this taxon) capable of producing a wide variety of infections such as pneumonia, urinary tract infections, and septicemia [15] [16]. Seven species are included in the Enterobacter cloacae complex includes Enterobacter asburiae, Enterobacter carcinogenus, Enterobacter cloacae, Enterobacter hormaechei, Enterobacter kobei, Enterobacter nimipressuralis, and Enterobacter mori. All these species are genotypically very close, with more than 60% DNA-DNA homology [17]. Our results clearly indicated that the genotypic relatedness between the three isolated bacterial strains (MAG-06, MAG-07 and MAG-010), that their genomic identification using 16S rDNA sequence homology indicated their grouping with Enterobacter cloacae, and the closest species in the assembled Neighbor-Joining phylogenetic tree, Enterobacter kobei, is 59% (Fig. 1). This reflects the close genomic relatedness between the three isolated E. cloacae strains to the other isolated two Enterobacter strains Pb1 and Pb2 (biochemically identified as E. kobei).
Most clinical laboratories in developed countries routinely identify Enterobacter spp.by phenotypic methods employing commercially available kits or semiautomated systems that are limited to E. cloacae and E. asburiae, while for further identification and discrimination of the other species in this genus, biochemical tests or molecular methods such as 16S rRNA, rpoB and hsp60 gene sequencing should be used [18]. A case study of a nosocomial urosepsis caused by E. kobei that was previously identified as E. cloacae [19]. The accurate identification of species and subspecies remains a challenge. The development of genome sequencing has rapidly modified the phylogeny of the genus particularly that of the E. cloacae complex[20] [21].
Enterobacter species are members of the ESKAPE group (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter species), which are described as the leading cause of resistant nosocomial infections [22] [23] [24]. Escherichia coli and Enterobacter cloacae complex are very common human enterobacteriaceae pathogens. Due to their intestine colonization, they can be a reservoir for infection in patients upon long-term hospitalization and antimicrobial treatment. Enterobacter cloacae is very often isolated in samples of clinical origin: urine, sputum, and blood culture [25] [26]. It considered difficult to apply a generalized strategies of infection control for chronically colonized patients because these intestine resident bacteria can acquire resistance determinants and so becoming Multiple drug resistant [27].
Due to the intensive use of β-lactams antibiotic, rate of Enterobacteriaceae resistance to ß-lactams highly increased in French hospitals [28]. The emergence of multidrug resistance (MDR) has led to an increased interest in these bacteria. In our study, results indicated that, the resistance to the monobactam, Aztreonam, was prevalent in the isolated bacteria from EPS (5 of the 6 enterobacteriaceae strains) and also for the non-enteric P. stutzeri MAG − 09.
Clinical strains of Enterobacter kobei have been isolated from various clinical samples: blood, sputum, urine. Enterobacter kobei was previously in a group included in E. cloacae [29]. Strains of E. kobei differ from those of E. cloacae by a negative Voges–Proskauer (VP) test. However, a new VP-positive biotype causing urinary tract infection (UTI) has been characterized [19]. However, the species is now also of concern because they can produce enzymes called extended-spectrum beta-lactamases (ESBLs). ESBL enzymes break down and destroy some commonly used antibiotics, including penicillins and cephalosporins making these drugs ineffective for infections treatment [30].
Providencia stuartii is an opportunistic, biofilm-forming pathogen from the Enterobacteriacae family [31]. Providencia species are common uropathogens in patients with long-term indwelling urinary catheters [32]. Providencia species isolated from catheter-associated urinary tract infections usually exhibit multiple resistance to antibiotics, which contributes to the high mortality of patients with Providencia bacteremia [33], [34]. P. stuartii is generally the second most common species isolated from blocked indwelling catheters after proteus mirabilis[31] [35]. As shown from Table 1, the isolated Providencia stuartii MAG-08 was resistant to cephaclor, aztreonam, gentamycin and exhibited low sensitivity to azithromycin. The Enterobacteriaceae, are the predominant pathogens in acute and chronic bacterial prostatitis. [36].
Pseudmonas. stutzeri is widely distributed in the environment and rarely causes infections, but it has been isolated as an opportunistic pathogen in clinical conditions. P. stutzeri infection have been reported in association with bacteremia/septicemia [37]; bone infection [38]; eye infection [39]; meningitis [40]; pneumonia [41]; urinary tract infection [42]. P. stutzeri is sensitive to many more antibiotics than P. aeruginosa, its most closely related species and a well-known human pathogen [43]. The higher sensitivity of P. stutzeri can be explained by its reduced exposure to antibiotics because of its low incidence rate in clinical environments. Interestingly, with the exception of fluoroquinolones, resistant P. stutzer strains have been isolated for almost all antibiotic families [44]. In current study, P. stutzeri strain MAG-09 recorded resistance against some tested antibiotics such as cephaclor, azithromycin, aztreonam and showed low sensitivity to rifampicin.
Treating CBP is challenging, because only few oral antibiotics could penetrate the prostate and achieve sufficient effective bactericidal concentration there [3] [45]. Effective therapy for bacterial prostatitis should afford or deliver a sufficient high concentration of antibiotics at prostatic secretion and tissues that would inhibit bacterial growth there or eradicate them [46]. In addition, with the emergence of multidrug resistance (MDR) and/or ESBLs-producing bacteria, biofilm-producing bacteria and the shift in bacterial etiology, the therapeutic schemes of CBP become more complex and trickier.
Clinicians should consider local drug resistance patterns in selecting antibiotics, especially with the emergence of extended spectrum beta-lactamase (ESBL)-producing strains in complicated UTIs and therapy adjustment should be based on culture results. The evaluation of microbiological characteristic for each CBP patients to perform a correct, appropriate, and personalized treatment schedule is essential [47].
In contrast to ß-lactam antibiotics, quinolone (discovered in the early 1960s) concentrations in prostatic fluid and prostatic tissue are high. Because of their unique and favorable pharmacokinetic properties, broad antibacterial spectrum and high activity especially against the Enterobacteriaceae. [48]. The coverage of the quinolone class was expanded significantly by the breakthrough development of fluoroquinolones, which show a much broader spectrum of activity and improved pharmacokinetics compared to the first-generation quinolone [49].
Fluoroquinolones have become a common first-line agent. Ciprofloxacin has been one of the most widely used fluoroquinolones for bacterial prostatitis treatment because of its broad-spectrum bactericidal activity, that commonly covers bacterial prostatitis causing pathogens and its advantageous pharmacokinetics in prostatic tissue [3].
Researchers have developed different strategies to improve the therapeutic effectiveness of ciprofloxacin and overcome the resistance issue, of them, targeting ciprofloxacin more precisely to the site of infection by using carriers. Ciprofloxacin–nanoparticles conjugation, ciprofloxacin nanoencapsulation, loading ciprofloxacin in liposomes, and polymeric nanoformations are among the different promising nanotechnology-based approaches to combat ciprofloxacin resistance [50]. In conclusion, isolates of Genus Enterobacter sp. represent the mor common bacterial pathogen in tested EPS of CBP diagnosed patients, of them Enterobacter cloacae was the mor common. Strain E. cloacae MAG-06 was chosen for a second part of this study as an infective CBP causing bacterium for the evaluation of targeted nano carriers of ciprofloxacin in the treatment of bacterial prostatitis".