ABSTRACT
Objective:To analyse molecular detection of coliforms and shorten the time of PCR. Methods:Rapid detection of coliforms by amplification of lacZ and uidA genes in a multiplex PCR reaction was designed and performed in comparison with most probably number (MPN) method for 16 artificial and 101 field samples. The molecular method was also conducted on isolated coliforms from positive MPN samples; standard sample for verification of microbial method certificated reference material; isolated strains from certificated reference material and standard bacteria. The PCR and electrophoresis parameters were changed for reducing the operation time. Results:Results of PCR for lacZ and uidA genes were similar in all of standard, operational and artificial samples and showed the 876 bp and 147 bp bands of lacZ and uidA genes by multiplex PCR. PCR results were confirmed by MPN culture method by sensitivity 86%(95%CI:0.71-0.93). Also the total execution time, with a successful change of factors, was reduced to less than two and a half hour. Conclusions:Multiplex PCR method with shortened operation time was used for the simultaneous detection of total coliforms and Escherichia coli in distribution system of Arak city. It’s recommended to be used at least as an initial screening test, and then the positive samples could be randomly tested by MPN.
ABSTRACT
Extended-spectrum beta-lactamase [ESBLs] producing bacteria are issued as a serious problem considering their ability to hydrolyze most of beta-lactam antibiotics. The outbreak of infections derived by ESBL-producing enterobacteriaceae is increasing throughout the world. Therefore, this study aims to determine a pattern of antibiotic resistance and investigate the extended-spectrum beta-lactamases production of enterobacteriaceae isolates separated from clinical specimens. In this study, 170 various strains of enterobacteriaceae isolated from clinical specimens in teaching hospitals of Arak cultured and identified applying standard methods during one year [2010-2011]. The antibiotic resistance of isolates was investigated through disk Agar diffusion according to CLSI criteria. The resistant isolates against ceftazidime and cefotaxime antibiotics were studied through the combined disk test for the final confirmation of ESBL-production. The minimum inhibitory concentration was determined through micro broth dilution. In this study, the resistance rate of various strains of enterobacteriaceae against amoxiclav, cefotaxime, ceftriaxone, ceftazidime, cefoxitin, cefotetan, meropenem and imipenem were respectively, 91.1%, 70%, 68.8%, 62.9%, 28.2%, 11.1%, 11.1% and 1.7%. Among 125 resistant enterobacteriaceae isolates against ceftazidime or cefotaxime, 108 isolates [86.4%] had ESBL-positive phenotype and 17 isolates [13.6%] had ESBLnegative phenotype. The MICs of the resistant isolates were indicated within a range of 16 to 512 micro g/ml for ceftazidime and 64 to 512 micro g/ml for cefotaxime. According to the results of this study, imipenem is the most effective antimicrobial antibiotic. On the other hand, the present study indicates that the bacteria within the family of ESBL producing enterobacteriaceae are highly prevalent among the patients. The increase in rate of such cases is often resulted by irrational antibiotic prescription. Application of new antimicrobials, limitation of the use of antimicrobial factors and increasing the utilization of infection control tools are all required in order to solve this problem.