Agronomy and Crop Science

Background: Alfalfa, Medicago sativa L. is the most important and widely grown leguminous fodder crop in temperate and tropical regions of the world. The production of alfalfa crop is limited by several biotic stresses, among which witches’ broom disease (AWB) was reported to cause significant economic losses. Methods: The phytoplasma infected alfalfa plants were collected from a central research farm, ICAR-IGFRI, Jhansi, U.P. Qualitative parameters such as crude protein content, acid detergent fibre and neutral detergent fibre were estimated in diseased and healthy plants. Phytoplasma universal primer (P1/P7) and nested primer (R16mF2/R16mR1) were used for the molecular characterization of AWB infected plants and phytoplasma infected, Parthenium hysterophorus. Result: The incidence of AWB disease ranged from 8-10%. The quantitative analysis of disease plants showed reduced plant height (-35%), fresh weight (-46.89%) and dry weight (-50.08%) compared to healthy plants. The diseased plant recorded low crude protein content (-21.38%) and higher dry matter content (+0.68%), acid detergent fibre (+33.72%) and neutral detergent fibre (+13.06%). The association of phytoplasma in diseased alfalfa and parthenium samples was confirmed by using P1/P7 and R16mF2/R16mR1 primer pair and Blastn analysis shared 99.6-100% similarity with ‘Candidatus Phytoplasma australasia’ belongs to the 16Sr group II-D.

Las variedades de papa (Solanum tuberosum L.) producidas en México son susceptibles a Candidatus Liberibacter solanacearum (CaLso), causante de la enfermedad conocida como ‘manchado interno de la pulpa’, por lo que se requiere conocer la respuesta de genotipos experimentales a la bacteria. El presente estudio tuvo como objetivo evaluar el efecto de la infección de los haplotipos LsoA + LsoB de CaLso en el follaje, la biomasa seca y la calidad de tubérculo de papa, variedad Fianna, una colecta de Solanum demissum y los clones experimentales T90-1-63 y T05-13-21 de Solanum spp. El manchado interno de la pulpa del tubérculo se determinó mediante análisis de imágenes de tubérculos. Las plantas de Fianna mostraron la mayor severidad de daño foliar; en cambio, los clones experimentales presentaron 17 % menos daño foliar que Fianna y 8 % más daño foliar que S. demissum. Este último fue el genotipo con la mayor biomasa seca de hoja y produjo tubérculos de un tamaño pequeño; las plantas infectadas de S. demissum presentaron mayor número de tubérculos y mayor rendimiento de tubérculo fresco que las plantas sin inoculación, aunque tuvieron la menor proporción de superficie sana del tubérculo. El clon T90-1-63 presentó los porcentajes más altos de superficie sana de tubérculo (> 79 %) y las menores intensidades del manchado interno de la pulpa del tubérculo.

‘Candidatus Phytoplasma trifolii’ is a cell wall-less phytopathogenic bacterium that infects many agriculturally important plant species such as alfalfa, clover, eggplant, pepper, potato, and tomato. The phytoplasma is responsible for repeated outbreaks of potato purple top (PPT) and potato witches’ broom (PWB) that occurred along the Pacific Coast of the United States since 2002, inflicting significant economic losses. To effectively manage these phytoplasmal diseases, it is important to develop diagnostic tools for specific, sensitive, and rapid detection of the pathogens. Here we report the development of a DNA endonuclease targeted CRISPR trans reporter (DETECTR) assay that couples isothermal amplification and Cas12a transcleavage of fluorescent oligonucleotide reporter for highly sensitive and specific detection of ‘Candidatus Phytoplasma trifolii’-related strains responsible for PPT and PWB. The DETECTR assay was capable of specifically detecting the 16S-23S ribosomal DNA intergenic transcribed spacer sequences from PPT- and PWB-diseased samples at the attomolar sensitivity level. Furthermore, the DETECTR strategy allows flexibility to capture assay outputs with fluorescent microplate readers or lateral flow assays for potentially high-throughput and/or field-deployable disease diagnostics.