Specific early identification of Morganella morganii based on polymerase chain reaction (PCR) method

Background and Aim : Morganella morganii is considered as an opportunistic pathogen and cause of some clinical infections in human with a high degree of mortality. Identification of the organism is mainly performed by tedious conventional biochemical assays or time-consuming culture techniques. Therefore, we developed a molecular method based on conventional PCR assay for the specific detection of M. morganii.Methods : The bacterial strains were purchased in lyophilized form and cultured in the respected bacterial media overnight to reach the log phase in their growth curve. Nucleic acid extraction was performed by a DNA extraction kit. After the analysis of the quality and quantity of the extracted DNA using spectroscopy at ۲۶۰ nm, the target gene was amplified by an optimized PCR protocol using previously designed primers. Furthermore, the specificity of the primers and the developed method was assessed using eight gram-negative bacteria. Finally, the amplification products were investigated by agarose gel electrophoresis and hydroxy naphthol blue indicator.Results : The quantity of the extracted nucleic acid was measured as ۱۰۱ µg mL-۱ which was appropriate for further experiment. A ۲۳۷ bp target product was expected to be amplified by the designed primers. The specificity of the method was determined by observing the supposed band for M. morganii on gel electrophoresis while no amplification product was detectable for any of the other tested bacteria (Shigella boydii, Yersinia enterocolitica, Citrobacter freundii, Klebsiella pneumonia, Enterobacter aerogenes, Pseudomonas aeruginosa, Burkholderia cepacia, Serratia marcescens). The Limit of detection (LOD) was attained to be ۱۰۱ ng ml-۱ of the amplified genomic DNA of M. morganii. Conclusion : The developed method is a molecular amplification-based detection technique to expedite the whole detection process of M. morganii in urgent circumstances.

کلیدواژه ها:

Nucleic acid testing (NAT) ، Morganella morganii ، PCR ، microbial detection ، diagnosis