Abstract
Eutypa dieback (ED) and botryosphaeria dieback (BD) are important grapevine trunk diseases causing significant yield reduction and threatening the sustainability of Australian vineyards. Laboratory testing using conventional and PCR-based techniques are often required for accurate diagnosis of these pathogens. While PCR-based techniques allow the detection and quantification of very low amounts of DNA in a wide range of samples, they are labour intensive, require expensive equipment and highly skilled staff to perform the analysis. Development of rapid, simple but sensitive and accurate molecular assays for field use can allow rapid diagnostics for infected vines in the field and nurseries without the need for diagnostic lab services. The objective of this study was to develop and evaluate LAMP assays for their suitability and sensitivity to detect the ED and BD pathogens for on-site diagnostics of infected grapevine plant materials. Two genus-specific and five species-specific LAMP primers designed for BD and ED pathogens, respectively, were developed. Preliminary results showed these assays were suitable for real time LAMP assays using the Genie II instrument and colorimetric LAMP. A commercial DNA extraction kit was also found suitable for rapid DNA extraction from infected wood. The real time LAMP assays were suitable for detecting and discriminating different ED and BD pathogens, but were less sensitive compared to the existing qPCR methods, and at least two times more sensitive compared to conventional fungal isolation methods. The development of LAMP assays may offer simple, cost effective and robust field-based diagnostics that will allow on-site testing for ED and BD pathogens in low resource environments.
1. Billones-Baaijens R, Úrbez-Torres JR, Liu M, Ayres M, Sosnowski M, Savocchia S (2018). Molecular methods to detect and quantify Botryosphaeriaceae inocula associated with grapevine dieback in Australia. Plant Disease 102-1489-1499
2. Notomi T, Okayama H, Masubuchi H, Yonekawa T, Watanabe K, Amino N, Hase T. 2000. Loop-mediated isothermal amplification of DNA. Nucleic Acids Research 28(12): E63
1. Billones-Baaijens R, Úrbez-Torres JR, Liu M, Ayres M, Sosnowski M, Savocchia S (2018). Molecular methods to detect and quantify Botryosphaeriaceae inocula associated with grapevine dieback in Australia. Plant Disease 102-1489-1499
2. Notomi T, Okayama H, Masubuchi H, Yonekawa T, Watanabe K, Amino N, Hase T. 2000. Loop-mediated isothermal amplification of DNA. Nucleic Acids Research 28(12): E63
| Original language | English |
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| Pages | 106 |
| Number of pages | 1 |
| Publication status | Published - 2021 |
| Event | 23rd Australasian Plant Pathology Society Online Conference - Online Duration: 23 Nov 2021 → 26 Nov 2021 https://appsconference.com.au/2021-home/program#1631147137230-aab6493a-1634 (Program) https://appsconference.com.au/ (Conference website) |
Conference
| Conference | 23rd Australasian Plant Pathology Society Online Conference |
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| Abbreviated title | Stay connected for plant health |
| Period | 23/11/21 → 26/11/21 |
| Other | We welcome the Australasian plant pathology community to what will be both the 23rd Biennial Australasian Plant Pathology Society Conference and our 1st fully online conference! This event was originally planned to be held in Hobart, Tasmania, however, due to the potential for ongoing impacts from COVID-19, we have decided to hold a fully online conference. The theme for the conference is “Stay Connected for Plant Health”. A fully online format will allow us to plan with certainty and we are confident that all the key aspects of previous conferences (keynote speakers in plenary sessions, oral presentations across concurrent sessions and, poster presentations), can be well supported online. The system will provide user-friendly virtual networking and options to engage with exhibitors. |
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