Real-Time PCR for Diagnosing and Quantifying Co-infection by Two Globally Distributed Fungal Pathogens of Wheat

Frontiers in Plant Science
Araz S AbdullahAyalsew Zerihun

Abstract

Co-infections - invasions of a host-plant by multiple pathogen species or strains - are common, and are thought to have consequences for pathogen ecology and evolution. Despite their apparent significance, co-infections have received limited attention; in part due to lack of suitable quantitative tools for monitoring of co-infecting pathogens. Here, we report on a duplex real-time PCR assay that simultaneously distinguishes and quantifies co-infections by two globally important fungal pathogens of wheat: Pyrenophora tritici-repentis and Parastagonospora nodorum. These fungi share common characteristics and host species, creating a challenge for conventional disease diagnosis and subsequent management strategies. The assay uses uniquely assigned fluorogenic probes to quantify fungal biomass as nucleic acid equivalents. The probes provide highly specific target quantification with accurate discrimination against non-target closely related fungal species and host genes. Quantification of the fungal targets is linear over a wide range (5000-0.5 pg DNA μl-1) with high reproducibility (RSD ≤ 10%). In the presence of host DNA in the assay matrix, fungal biomass can be quantified up to a fungal to wheat DNA ratio of 1 to 200. The utili...Continue Reading

References

Aug 15, 1991·Proceedings of the National Academy of Sciences of the United States of America·P M HollandD H Gelfand
Nov 10, 2001·Annual Review of Phytopathology·J Zadoks
Jul 13, 2006·Nature Genetics·Timothy L FriesenRichard P Oliver
Mar 29, 2012·Proceedings of the National Academy of Sciences of the United States of America·Conrad L SchochUNKNOWN Fungal Barcoding Consortium Author List
Jan 26, 2013·Ecology Letters·Samuel AlizonYannis Michalakis
Dec 15, 2015·Trends in Plant Science·Charlotte TollenaereAnna-Liisa Laine
Nov 10, 2017·Frontiers in Plant Science·Araz S AbdullahAyalsew Zerihun

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Citations

Jun 14, 2019·Frontiers in Plant Science·Maria Doroteia CamposMaria do Rosário Félix
Jan 29, 2021·Frontiers in Cellular and Infection Microbiology·Ganeshamoorthy Hariharan, Kandeeparoopan Prasannath

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Methods Mentioned

BETA
PCR
Fluorescence
electrophoresis

Software Mentioned

CFX Manager
Geneious

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