Enterococci are among Gram-positive cocci and are common inhabitants of the human gastrointestinal tract and enough potent to cause serious infections such as bacteremia, urinary tract infections (UTIs) and endocarditis. Enterococci are not generally considered as highly infectious bacterium. However, the main reason for treatment failure in enterococcal infections is acquired resistance to glycopeptide antibiotics, specifically vancomycin. Most enterococcal infections in human such as gastroenteritis, intestinal infections, and endocarditis have been caused by E. faecalis and E. facieum. They are holding the second place of most frequent cause of hospital infections since 1990. In present study g yrA S83R polymorphism rate in vancomycin resistant Enterococcus faecium isolated from environment, food industries, and hospitals of Tehran were examined by ARMS-PCR technique. DNA was extracted from the bacterial colonies using standard column method, after separating the samples into two groups of antibiotic resistant and antibiotic susceptible by antibiogram test. A fragment of gyrA gene was amplified using PCR method to investigate point mutation of S83R position. ARMS-PCR technique was applied to detect the presence or absence of mutation using a set of specified primers which can be annealed when the mutation is present. The results were statistically analyzed by chi-square test (p<0.05) using SPSS 19th version. The results showed that there was a significant correlation between the presences of S83R polymorphism with vancomycin resistance trait in Entrococcus faecium. Therefore, this technique could be used as a diagnostic tool to detect vancomycin resistance cases of E. faecium in patients and environment.

Background: Multi-drug resistant (MDR) Klebsiella pneumoniae strains cause the majority of community ac-quired and life-threatening infections. We aimed to detect the gyrA mutations in the clinical strains of nalidixic acid and ciprofloxacin resistant K. pneumoniae isolated from the patients with urinary tract infections. Methods: Bacterial strains were isolated from the patients with urinary tract infections admitted to a major hospital in Tehran, Iran (2017-2018). Bacterial identification was done according to standard microbiological tests. Antimicrobial susceptibility testing for quinolones and fluoroquinolone antibiotics was done using both disc diffusion and minimal inhibitory concentrations (MICs) methods. PCR-RFLP was used to detect the probable mutation in the gyrA gene in nalidixic acid and ciprofloxacin resistant strains. Finally sequencing was performed to detect point mutations in isolated K. pneumoniae strains. Results: One hundred K. pneumonia isolates were recovered from the urine samples of the clinical cases. Anti-biotic resistance testing showed that among all K. pneumoniae isolates, 26% and 19% of the strains were re-sistant to nalidixic acid and ciprofloxacin respectively. MIC value was ≥, 4 μ, g/ml for ciprofloxacin resistant isolates. The results of RFLP on gyrA PCR amplicons using HinfI restriction enzyme showed point mutation in this gene in 46% of nalidixic acid and ciprofloxacin resistant K. pneumonia. The data obtained from the se-quencing confirmed the RFLP results and indicated the presence of point mutations in codons 83 and 87 in the gyrA gene which leads to the substitution of different amino acids in gyrA protein. Conclusion: Our findings indicated a relative increased rate of resistance against quinolones and fluoroquino-lone antibiotics that raised a concern about extensive dissemination of clinical strains of nalidixic acid and ciprofloxacin resistant K. pneumonia. Point mutation of gyrA gene was responsible for the resistance in our strains however to gain more insight into the molecular characterization of quinolone-resistant isolates, other possible mechanisms of the resistance should also be investigated.

Quinolones are a large and widely consumed class of synthetic drugs. Expanded-spectrum quinolones, like ciprofloxacin are highly effective against Gram-negative bacteria, especially Escherichia coli. In E. coli the major target for quinolones is DNA gyrAse. This enzyme is composed of two subunits, gyrA and GyrB encoding by gyrA and gyrB, respectively. Mutations in either of these genes cause quinolone resistance. Mutations in QRDR section of gyrA are more common in quinolone resistant clinical isolates. However, a mutation outside of this region was also reported. Thus, this study was aimed to provide more information on mutations sites in gyrA. For this purpose, spontaneous ciprofloxacin resistant mutants arisen in cultures of E. coli ATCC 25922 and MG1655 were isolated on LB agar containing ciprofloxacin. Next, the MICs of these clones were measured and the presence of mutation in gyrA was investigated. Results showed that the frequency of ciprofloxacin resistant mutants in cultures of E. coli strains was low. However, these mutants had different MICs, depending on the day of isolation. Most of ciprofloxacin-resistant mutants possess mutations in QRDR region and precisely at Ser-83. However, mutations outside of this region were also found at Tyr-50 and Ala-119. In conclusion, the presence of mutations at amino acids 50 and 119 suggests that in addition to QRDR section and Tyr-122, these sites are also essential for DNA gyrAse activity.