Introduction Enterococci commonly inhabit the gastrointestinal tract of both human and animals; however, they have emerged as a leading cause of several infections with substantial morbidity and mortality. Their ability to acquire resistance combined with intrinsic resistance to various antimicrobials makes treatment of enterococcal infections challenging. Materials and methods The aim of the study was to evaluate the antimicrobial resistance pattern, and assess the prevalence of multidrug resistance (MDR) and extensive drug resistance (XDR) among enterococcal isolates, collected from different clinical sources, in Mansoura University Hospitals, Egypt. Results Antibiotic sensitivity testing revealed elevated levels of resistance among enterococcal clinical isolates (N=103). All E. faecium (N=32) and 74.6% of E. faecalis isolates (N=71) were MDR, while two E. faecalis and four E. faecium isolates were XDR. High level gentamicin resistance was detected in 79.6%, most of them carried the aac(6’)-Ie-aph(2’’)-Ia gene. High level streptomycin resistance was seen in 36.9%, of which 52.6% carried the ant(6’)-Ia gene. Resistance to macrolides and lincosamides were mediated by ermB (92.2%) and msrA/B (42.7%). tetK , tetL , and tetM genes were detected among tetracyclines resistant isolates. Resistance to vancomycin was detected in 15.5%, where vanB and vanC 1 gene clusters were detected in VRE isolates. Ten isolates (9.7%) were resistant to linezolid, eight of which harbored the optrA gene. Vancomycin and linezolid resistant enterococci were more likely to exhibit strong/moderate biofilm formation than vancomycin and linezolid sensitive ones. Conclusion Elevated levels of resistance to different classes of antimicrobial agents and emergence of MDR and XDR strains pose a major threat with limited therapeutic options for infections caused by this emerging pathogen.

The incidence of fluconazole-resistant Candida albicans has been increasing worldwide. Both biofilm and fungal morphogenesis are main virulence factors of C. albicans cells. Extracellular fungal prostaglandins are synthesized during biofilm adhesion and development and through yeast-hypha conversion. Hence, we targeted prostaglandin synthesis with various cyclooxygenase (COX) inhibitors (aspirin, diclofenac, ketoprofen, tenoxicam, and ketorolac) and assessed their effect on fungal adhesion, biofilm formation, and yeast-hypha conversion in clinical isolates of Fluconazole resistant C. albicans. Significant reduction in fungal adhesion and detachment of mature biofilm was attained down to 1 mM concentrations of anti-inflammatory agents. Microscopical examination of fungal cells in the presence of the tested drugs showed significant reduction of germ tube formation. Therefore, COX inhibitors have a significant effect on reduction of Candida adhesion and biofilm development in correlation with fungal morphogenesis. Moreover, inhibition of C. albicans by COX inhibitors gave synergistic activity with fluconazole suggesting that combination therapeutic strategies may be fruitful for management of infection of Fluconazole resistant C. albicans.

β-lactam resistance represents a worldwide problem and a serious challenge for antimicrobial treatment. Hence this research was conducted to recognize several mechanisms mediating β-lactam resistance in E. coli and K. pneumoniae clinical isolates collected from Mansoura University hospitals, Egypt. A total of 80 isolates, 45 E. coli and 35 K. pneumoniae isolates, were collected and their antibiotic susceptibility was determined by the Disc diffusion method followed by phenotypic and genotypic detection of extended-spectrum β-lactamases (ESBLs), AmpC β-lactamase, carbapenemase enzymes. The outer membrane protein porins of all isolates were analyzed and their genes were examined using gene amplification and sequencing. Also, the resistance to complement-mediated serum killing was estimated. A significant percentage of isolates (93.8%) were multidrug resistance and showed an elevated resistance to β-lactam antibiotics. The presence of either ESBL or AmpC enzymes was high among isolates (83.75%). Also, 60% of the isolated strains were carbapenemase producers. The most frequently detected gene of ESBL among all tested isolates was blaCTX-M-15 (86.3%) followed by blaTEM-1 (81.3%) and blaSHV-1 (35%) while the Amp-C gene was present in 83.75%. For carbapenemase-producing isolates, blaNDM1 was the most common (60%) followed by blaVIM-1 (35%) and blaOXA-48 (13.8%). Besides, 73.3% and 40% of E. coli and K. pneumoniae isolates respectively were serum resistant. Outer membrane protein analysis showed that 93.3% of E. coli and 95.7% of K. pneumoniae isolates lost their porins or showed modified porins. Furthermore, sequence analysis of tested porin genes in some isolates revealed the presence of frameshift mutations that produced truncated proteins of smaller size. β-lactam resistance in K. pneumoniae and E. coli isolates in our hospitals is due to a combination of β-lactamase activity and porin loss/alteration. Hence more restrictions should be applied on β-lactams usage to decrease the emergence of resistant strains.

Uricase is an important microbial enzyme that can be used in the clinical treatment of gout, hyperuricemia, and tumor lysis syndrome. A total of 127 clinical isolates of Pseudomonas aeruginosa were tested for uricase production. A Pseudomonas strain named Ps43 showed the highest level of native uricase enzyme expression. The open reading frame of the uricase enzyme was amplified from Ps43 and cloned into the expression vector pRSET-B. Uricase was expressed using E. coli BL21 (DE3). The ORF was sequenced and assigned GenBank Accession No. KJ718888. The nucleotide sequence analysis was identical to the coding sequence of uricase gene puuD of P. aeruginosa PAO1. We report the successful expression of P. aeruginosa uricase in Escherichia coli. E. coli showed an induced protein with a molecular mass of about 58 kDa that was confirmed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blotting. We also established efficient protein purification using the Ni-Sepharose column with activity of the purified enzyme of 2.16 IU and a 2-fold increase in the specific activity of the pure enzyme compared with the crude enzyme.

The increasing incidence of β-lactam resistance due to AmpC β-lactamases in Egypt necessitated this study which aimed to evaluate four different phenotypic methods for detection of AmpC β-lactamases among some clinical isolates of Enterobacteriaceae and compare these results with those obtained using polymerase chain reaction. The distribution of five AmpC β-lactamases genes (AmpC, CIT-M, Fox-1, ACC-1, ACT-1) were determined among the clinical isolates. Among 180 clinical isolates of Enterobacteriaceae, only 57 isolates were AmpC producers by phenotypic methods and 108 were AmpC producers by polymerase chain reaction. Phenotypic methods adapted in this study gave variable results with the most discriminatory results given by direct inoculation of both the enzyme extract and the bacterial culture in the wells. Of these, the best results were given by enzyme inoculation methods where 43 isolates exhibited positive result by this method. The distribution of AmpC β-lactamases gene among the clinical isolates showed that AmpC gene predominated in Escherichia coli. Fox gene was predominantly present in Enterobacter cloacae and E. coli isolates. ACT-1 predominated in E. cloacae. In contrast, enzymes from CIT-M and ACC-1 group were rarely present in Enterobacteriaceae.