Use of culture and molecular methods for identification and characterization of dry bean fungal root rot pathogens in Zambia

Abstract

Root and crown rot (RCR) of dry bean is a disease complex caused by soil-borne pathogens that can limit bean production in Zambia. Identifying the primary pathogen in the complex would facilitate the control as well as breeding for resistance to the disease. Plant tissue samples and DNA embedded FTA® Cards were collected to isolate, detect and molecularly identify RCR pathogens using genus/species specific primers as well as high throughput deep sequencing with illumina Miseq using universal eukaryote bar coded primers Euk7. A total of 204 isolates were characterized using morphological features and Sanger sequencing. Analysis of reads and operational taxonomic units from illumina sequencing, detection frequencies from polymerized chain reaction as well as morphological characterization revealed Fusarium species complex as the main component of RCR complex of dry bean in Zambia, primarily F. oxysporum, followed by F. solani. A sensitivity chi-square test comparing detection methods of the four major RCR pathogens showed statistical differences. A highly significant positive correlation (p < 0.001) confirmed by a good agreement (kappa (k) 0.5 to 0.6, p < 0.05) was found between FTA® Card and direct plant tissue. Use of FTA® Cards can facilitate sampling, storing and analysis of DNA for RCR pathogen identification.