Agronomy and Horticulture, Department of


Date of this Version

Summer 8-2016

Document Type



A THESIS Presented to the Faculty of The Graduate College at the University of Nebraska In Partial Fulfillment of Requirements For the Degree of Master of Science, Major: Agronomy, Under the Supervision of Professor James R. Steadman. Lincoln, Nebraska: August 2016

Copyright (c) 2016 Chikoti Mukuma


Dry bean is among the most important food legume crops for direct human consumption in Latin America and Africa. Recently, root and crown rot (RCR) has emerged as an important production constraint. Root and crown rot often involve fungal complexes. Thus, the straw, detached leaf, cup and stem tests were compared on their ability to detect the most common pathogens reported to be associated with RCR: Fusarium solani, Fusarium oxysporum, Pythium ultimum, Rhizoctonia solani and Macrophomina phaseolina.Significant differences (P

Next Generation Sequencing (NGS), conventional Polymerase Chain Reaction (PCR), and classical fungal culturing methods were compared in identification of the primary pathogens causing RCR of dry bean in Zambia. Analysis of DNA from plant tissue and ground tissue extracts spotted on FTA cards by NGS identified F. oxysporum, F. solani and other species of the Fusarium complex as the most abundant reads and Operation Taxonomic Units (OTU’s). Fusarium spp. were also detected in over 70% of the samples analyzed by conventional PCR using specific primers and also had the highest frequency of recovery (>0.8) of which over 90% were pathogenic. Thus it appears that, in Zambia, the Fusarium complex causes RCR in dry bean. Significant correlations at 0.05 level and high inter-method agreements >0.7 observed between plant tissue and FTA cards support the hypothesis that FTA cards can substitute for sampling plant tissue and preserving DNA for RCR pathogen identification. Significant correlations also were observed among the different methods of identification of primary pathogens responsible for RCR where F. oxysporum is the most likely target for initial screening for resistance in Zambia.

Identification of a primary pathogen associated with RCR of dry bean in Zambia provides evidence to direct initial breeding for RCR resistance to breeding for resistance to Fusarium oxysporum.

Advisor: James R. Steadman