J Clin Microbiol
Free-Living and Intestinal Amebas (FLIA) Laboratory, Waterborne Disease Prevention Branch, Division of Foodborne, Waterborne, and Environmental Diseases, Centers for Disease Control and Prevention, Atlanta, Georgia, USA.
Published: December 2020
There are over 40 species within the genus , eight of which infect humans. Of these, four species (, , , and ) are morphologically indistinguishable from each other, and yet differentiation is important for appropriate treatment decisions. Here, we developed a hydrolysis probe-based tetraplex real-time PCR assay that can simultaneously detect and differentiate these four species in clinical samples. In this assay, multicopy small-subunit (SSU) ribosomal DNA (rDNA) sequences were used as targets. We determined that the tetraplex real-time PCR can detect amebic DNA corresponding to as little as a 0.1 trophozoite equivalent of any of these species. We also determined that this assay can detect DNA in the presence of 10-fold more DNA from another species in mixed-infection scenarios. With a panel of more than 100 well-characterized clinical samples diagnosed and confirmed using a previously published duplex real-time PCR (capable of detecting and ), our tetraplex real-time PCR assay demonstrated levels of sensitivity and specificity comparable with those demonstrated by the duplex real-time PCR assay. The advantage of our assay over the duplex assay is that it can specifically detect two additional species and can be used in conventional PCR format. This newly developed assay will allow further characterization of the epidemiology and pathogenicity of the four morphologically identical species, especially in low-resource settings.
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http://dx.doi.org/10.1128/JCM.01986-20 | DOI Listing |
Sci Rep
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Faculty of Biotechnology, October University for Modern Sciences and Arts, 6th of October, Egypt.
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