Click for new scientific resources and news about Corona[COVID-19]

Paper Information

Journal:   IRANIAN JOURNAL OF PUBLIC HEALTH   AUGUST 2014 , Volume 43 , Number SUPPLEMENT 2; Page(s) 69 To 69.
 
Paper: 

ESCHERICHIA COLI AND SHIGELLA GENUS DETECTION IN WATER SAMPLE BY PENTAPLEX PCR

 
 
Author(s):  AKHTARI LEILI*, AKHAVAN SEPAHY ABBAS, DAVARI KAMBIZ
 
* DEPARTMENT OF MICROBIOLOGY, FACULTY OF SCIENCE, ISLAMIC AZAD UNIVERSITY SCIENCE AND RESEARCH BRANCH OF KORDESTAN, KORDESTAN, IRAN
 
Abstract: 

Background: The aim of this study was to determine the most ubiquitous Escherichia coli specific genes to apply as a genetic marker to speed up the detection time and the sensitivity and introduce a multiplex PCR to detect and screen this marker organism in water sample as soon as possible. Since Shigella species share great sequence similarity, so we included a specific gene corresponding to putative integrase enzyme to differentiate Shigella species from Escherichia coli.
Methods: Out of 1956 water samples, one hundred and six Escherichia coli strains, isolated from different water sources of Tehran province (Jalalieh water treatment center, kan water treatment center, number 3, 4 and 5 water treatment centers) within a 1 year were characterized by standard methods based on fast lactose fermentation and IMViC tests. All the isolates were kept in glycerol at -70oC until the final procedure.We extracted DNA from all isolates applying Accuprep Bioneer column kit. New specific sets of primers including structural genes like uspA, phoE and enzyme coding ones such as gadA/B, uidA for Escherichia coli and put int for Shigella were designed in a novel pentaplex PCR by Allele ID 7.6 (Table 1). Following checking the specificity of any set of primers in NCBI, dimer formation and primers interaction, applying Oligo analyzer 1.03, we optimized the multiplex at different temperatures and DNA concentration gradients.
Results: Among 978 water samples all tested for E. coli, we characterized 106 strains by standard methods such as lactose fermentation and IMViC test. Following pentaplex PCR development and optimization, we looked for the uspA, phoE, gadA/B, uidA for Escherichia coli and put int for Shigella genus. While detecting all 4 genes of E. coli, our results showed this multiplex PCR is a great tool to detect such organism in water samples within a short period of time. Our results showed that all 106 strains carried the above-mentioned genes which confirms and verifies the accuracy of our approach.
Conclusion: Our results showed the ubiquitous specific genes of Escherichia coli, as well as the other cells, are the structural ones encoding the structural proteins. These results are highly justifiable because the structural proteins serve as cell components and if absent or deficient, the cells have no chance to survive because of the selection pressure. Besides the structural ones, the above mentioned genes encoding the enzymes (gadA/B, uidA) are also prevalent because they belong to essential or housekeeping genes.

 
Keyword(s): ESCHERICHIA COLI, SHIGELLA, PENTAPLEX PCR, WATER SAMPLE
 
References: 
  • ندارد
 
  Yearly Visit 51
 
Latest on Blog
Enter SID Blog