Molecular study of tetracycline resistance genes in Shigella sonnei isolated from clinical samples and the effect of spring grass extract on the expression of resistance gene by Real time-PCR method

Document Type : Research Paper

Authors

1 Master of Microbiology, Department of Biology, Faculty of Basic Sciences, Islamic Azad University, Central Tehran Branch, Tehran, Iran

2 Professor,Department of Pathobiology,Faculty of Veterinary Specialized Sciences,Islamic Azad University,Science and Research Branch,Tehran,Iran

3 Associate Professor, Department of Microbiology, Faculty of Basic Sciences, Saveh Branch, Islamic Azad University, Saveh, Iran

Abstract

Introduction: Shigella is a gram-negative intestinal bacillus whose infections have caused serious problems in developed and developing countries. The prevalence of Shigella infections is very low due to the low dose of pathogenicity of this bacterium as well as its easy transmission from one person to another and also the indirect contamination of people through the consumption of contaminated food and water. Objective of isolation of Shigella sonnei from children's feces, Effect and isolation of spring grass extract, Molecular isolation of tetracycline genes by multiplex PCR, Effect of extract on tetracycline gene expression by multiplex PCR.
Methods: Out of 60 Shigella sonnei strains isolated from children with diarrhea after isolation of strains on different media and MacConkey Agar (MAC) medium in terms of shape and color of colonies were examined and recorded. First, the DNA of each sample was extracted by a DNA extraction kit, and after extraction, the DNA of the template was amplified by PCR. To investigate the effect of antibiotic resistance of spring grass to the extracted gene resistant to tetracycline, MIC of plant extract on different concentrations of Shigella sonnei was investigated. Strains with tetracycline-resistant genes were treated with MIC, using both treatment and non-treatment groups, and after incubation for 18 hours, stationary lag log was performed using RNA extraction kit x plus RNA extraction. Then, using RT-PCR ABI plus device, the expression of tet A and tet B genes in the treated and untreated groups under the influence of spring grass extract was read quantitatively (numerically) CT and the expression of the gene under the influence of treatment was interpreted.
Results: Shigella sonnei isolated results showed a high percentage of the highest resistance to streptomycin, Nalidixic acid, tetracycline and ampicillin and chloramphenicol with 100%, 100%, 86.7%, 91.8% and 50%. Also, the highest sensitivity to gentamicin and ciprofloxacin was observed with 85% and 86.6%, respectively. Multiple PCR results showed that out of all 60 Shigella sonnei isolates, 3 isolates had only tetB gene (5%) and 55 isolates had tetA gene (91.6%).
Conclusion: The results of Fisher test indicate that there was a significant relationship between tetA genes and antibiotic resistance at 5% error level (p-value <0.05). The results showed that at the 5% error level there was no significant relationship between tetB genes and antibiotic resistance (p-value = 0.05). The results showed that at the 5% error level, there was no significant relationship between positive and negative genes of all genes with antibiotic resistance (p-value <0.05).

Keywords

References

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medical journal of mashhad university of medical sciences
Volume 64, Issue 1
March and April 2021
Pages 2340-2352

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