Figure 1: Xf symptoms on almond.
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Agdia, Inc (Elkhart, IN) has commercialized a rapid and user-friendly DNA test kit, based on their AmplifyRP® XRT+ platform, for detection of Xylella fastidiosa (Xf).
Xylella fastidiosa (Xf) is the causal agent of multiple crop diseases including Pierce's disease in grape, citrus variegated chlorosis (CVC), almond leaf scorch disease (Figure 1) and olive quick decline syndrome. Severe Xf infections can decimate entire crops and cause enormous financial losses to growers.
The test is a real-time isothermal DNA amplification assay that utilizes recombinase polymerase amplification (RPA) technology. RPA facilitates rapid nucleic acid (DNA or RNA) amplification at a single operating temperature of 39°C using crude sample extracts. RPA can provide the same level of sensitivity and specificity as laboratory based PCR methods, but offers some clear advantages. The total assay time is less than 30 minutes, including sample preparation. This is a stark contrast to PCR methods which can take hours or days to complete. PCR analysis must also be completed in a laboratory whereas AmplifyRP XRT+ can be performed on-site.
Figure 2: Real-time results on AmpliFire®Agdia’s AmplifyRP XRT+ platform works similarly to the recently launched XRT platform in that the results can be obtained using a portable fluorometer. However, XRT+ provides greater flexibility because the test can alternatively be performed using a small portable heat block with results being visualized post-amplification on an Amplicon Detection Chamber. This allows end-users who don’t want to invest in a fluorometer an alternative method of running the test.
To run the test, an end-user first extracts their sample by adding it, along with extraction buffer, to a micro centrifuge tube that is provided in the kit. After the sample has been extracted a small volume is added to a solution that is used to rehydrate a lyophilized reaction pellet. Once the reaction pellet has been rehydrated it can be added to a portable fluorometer or a small heat block for 20 minutes. During this time, if present, Xf DNA will be amplified. If using the fluorometer results will automatically be displayed in real-time. Alternatively, if using a heat block, the reaction is added to an Amplicon Detection Chamber following the amplification step where results will be visible on a lateral flow strip.
Agdia’s AmplifyRP XRT+ kit for Xf includes 48 reactions and the necessary buffers to perform the test. If performing the test as a real-time assay, the end-user will need to purchase a portable fluorometer separately. Agdia claims the test is compatible with several commercially available fluorometers but recommends the AmpliFire® (Figure 2), manufactured by Douglas Scientific. The AmpliFire is battery operated and has a small footprint, making it possible to test in remote locations. Setup is simple because all the assay parameters are loaded into the AmpliFire using a barcode that is affixed to the test’s packaging. It also has a large, intuitive, touch screen interface that is used to initiate reactions and view results. More information on the AmpliFire can be found on Agdia’s website.
Figure 3: Portable heat-block with post-amplification detection using Amplicon Detection Chambers.Alternatively, if the end-user runs the test as an end-point detection assay they will need to purchase Amplicon Detection Chambers (Figure 3), as well as a starter pack, separately. The starter pack includes a heat block (necessary for the amplification step) as well as pipettes, pipette tips, and a tube rack.
The test is also offered as a service by Agdia Testing Services for those who wish to send samples for diagnosis. For more information on the test contact Agdia at 574-264-2014 or via email at info@agdia.com
About Agdia
A leading provider of diagnostic solutions for agriculture, Agdia, Inc. has been serving plant breeders, propagators, growers, universities, and private testing laboratories since 1981. The company offers a comprehensive portfolio of validated, easy-to-use diagnostics for identifying plant pathogens, hormones, and transgenic traits, as well as in-house testing services. Agdia’s quality management system is ISO 9001:2008 certified and their Testing Services Laboratory is ISO 17025:2005 accredited.
About AmplifyRP
AmplifyRP XRT Test Kits employ recombinase polymerase amplification (RPA) technology licensed by Agdia, Inc (licensee) from TwistDx Limited, U.K (licensor). Use of the RPA process and probe technologies are protected by US patents 7,270,981 B2, 7,399,590 B2, 7,435,561 B2, 7,485,428 B2 and foreign equivalents in addition to pending patents.
AmplifyRP® is a registered Trademark of Agdia, Inc.
AmpliFire® is a registered Trademark of Douglas Scientific.
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