Location: Floral and Nursery Plants ResearchTitle: Real-time and conventional PCR tools for detection and discrimination of Calonectria pseudonaviculata and C. henricotiae causing boxwood blight
|GUO, HENRY - Rutgers University|
Submitted to: Plant Disease
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 7/8/2020
Publication Date: 11/16/2020
Citation: Guo, H., Pooler, M.R. 2020. Real-time and conventional PCR tools for detection and discrimination of Calonectria pseudonaviculata and C. henricotiae causing boxwood blight. Plant Disease. https://doi.org/10.1094/PDIS-09-19-2053-RE.
Interpretive Summary: Boxwoods are an important landscape plant in the U.S., with an annual market value of over $100 million. A new fungal disease, boxwood blight, caused by two species of a fungal pathogen, was identified in Europe in the mid 1990s and first reported in the U.S. in 2011. This disease causes leaf drop, stem lesions, and plant death, and threatens the boxwood industry and native boxwood populations worldwide. Fast, accurate, and inexpensive methods to detect and differentiate these fungal species is critical in stopping the spread of the disease. ARS scientists from the U.S. National Arboretum developed PCR primers that can be used with either conventional PCR or SYBR-based real-time PCR to specifically detect and differentiate the two fungal species. This assay will be useful for growers, extension agents, regulatory agencies, and managers of plant collections to enable rapid screening of field-grown plant samples test for the presence of the disease.
Technical Abstract: Calonectria pseudonaviculata and C. henricotiae are the causal agents of boxwood blight, a devastating disease of boxwood that has caused significant economic impact on the nursery and landscape industries in the U.S. and in Europe. The two species are genetically distinct and are found in different geographic areas, but are difficult to distinguish based on morphology and pathogenicity. Fast, accurate, and inexpensive methods to detect and differentiate these species is critical in stopping the spread of the disease. We designed primer pairs based on available sequences of three genes - calmodulin, histone H3, and ß-tubulin – and tested their ability to differentiate the two Calonectria species. Here we report three primer pairs derived from sequence differences in the Histone H3 region that can be used to specifically detect C. pseudonaviculata, C. henricotiae, or both species. Specificity of these primers was tested against nine isolates of C. pseudonaviculata, three isolates of C. henricotiae, 13 other Calonectria species, and five isolates from related genera using conventional and real-time PCR. These are the first primers available that can be used with either conventional PCR or SYBR-based real-time PCR to specifically detect and differentiate the two fungal species.