Location: Sustainable Perennial Crops Laboratory
2023 Annual Report
Objectives
Objective 1: Characterize the genetic diversity of pathogens of cacao and other tropical perennial crops to improve understanding of how pathogen diversity impacts plant/pathogen interactions. (NP303, C1, PS1A, C2, PS2B).
Sub-objective 1.A: Sequence the genome and transcriptome of fungal species causing thread blight on cacao in Ghana (years 1-3). (Non-hypothesis-driven).
Sub-objective 1.B: Sequence the genomes and transcriptomes of Rosellinia species causing black root rot of cacao in Colombia, Ecuador and Peru (years 1-3). (Non-hypothesis-driven).
Objective 2: Integrate genetic resources of pathogens and hosts for use in breeding disease tolerance in cacao and other tropical perennial crops. (NP303, C2, PS2B, C3, PS3A, NP301, C2, PS2A).
Sub-objective 2.A: Detailed genome assembly and analysis of cacao clones showing tolerance and susceptibility to P. palmivora/P. megakarya. (years 1-3). (Non-hypothesis-driven).
Sub-objective 2.B: Transient gene expression of candidate cacao defense genes and tissue-specific gene promoters for use in the GAANTRY system in cacao. (years 2-3). (Non-hypothesis-driven).
Sub-objective 2.C: Stable transformation of cacao using the GAANTRY gene stacking system. (years 3-5). (Non-hypothesis-driven).
Objective 3: Develop novel strategies for managing diseases of cacao and other tropical perennial crops. (NP303, C2, PS2B, C3, PS3A). (Hypothesis-driven).
Approach
Black pod rot of cacao, caused by Phytophthora spp., has the greatest negative impact on cacao production globally while coffee leaf rust, caused by Hemileia vastatrix, causes severe yield reduction wherever it occurs on coffee. Although these two diseases are being studied extensively because of their global concerns, other diseases like thread blight (basidiomycete disease complex) and black root rot (Rosellinia species) of cacao are much less studied because of their local/regional distributions. Disease mitigation methodologies continue to have limited success in managing these and other diseases in cacao and coffee. While progress is being made in breeding tolerance to some diseases in these crops, these efforts continue to be hindered by our limited knowledge of pathogen genetics and host genes conferring disease tolerance. Therefore, this project will continue developing tools with applications to managing plant diseases. The genome and transcriptomes of established and emerging cacao pathogens, including those causing thread blight and black root rot, will be sequenced. New knowledge of cacao genes that contribute to disease resistance against Phytophthora spp. will be acquired through functional analysis. A transient gene expression system will be exploited to identify condition and tissue-specific cacao gene promoters and candidate cacao defense genes for inclusion in a stable multigene transformation system (GAANTRY) validating its use in cacao. New methodologies with potential for mitigating cacao and coffee disease losses will be developed. LED-based UV light technologies will be tested for efficacy in mitigating of both black pod rot in cacao and coffee leaf rust. The objectives of this project target specific plant/pathogen interactions using multiple approaches. Still, there is a high probability the findings of these studies will have applicability to additional plant/pathogen interactions.
Progress Report
Currently project staffing is going through significant changes. The lead scientist on the project is retiring as of July 1, 2023, and active recruitment of a replacement is underway. As the result of additional funds being added to the project, a third scientist is also being recruited along with the search for one new laboratory technician.
Cacao Gene Promoters: Research on project objectives continue. Using RNA-Seq data generated from RNA extracted from cacao pods inoculated with the black pod rot pathogen Phytophthora palmivora, disease-resistance promoter motifs for cacao genes were identified. Differential gene expression analysis and de novo promoter motif discovery was used to identify 76 novel sequence motifs associated with cacao response to P. palmivora. Further analysis identified 17 candidate cis-regulatory modules likely to be important to cacao resistance against P. palmivora. In addition, 7 chromosomal lengthen genomes from genetically diverse population groups of Theobroma cacao have been sequenced providing haploid genomes to improve gene function and promoter analysis.
Thread blight genomes. To better understand the genomics of thread blight disease-causing species in the family Marasmiaceae, DNA was extracted, and long-read sequencing data obtained for three species: Marasmius tenussimus, Marasmiellus palmivorus, and Marasmiellus scandens. Using the newly generated data and previously generated short-read sequencing data a hybrid assembly approach was used to produce a high-quality reference genome for M. tenuissimus GH-37, and the sequenced published and released to the NCBI GenBank repository. The genome assemblies and analysis for M. palmivorus and M. scandens are being completed using the pipeline generated in producing the Marasmius tenussimus assembly.
Moniliophthora roreri and M. perniciosa genome assembly and analysis. A manuscript describing five high quality genome assemblies and 22 short sequence assemblies for M. roreri was recently accepted for publication. Data in this manuscript adds substantially to our understanding of M. roreri biology and evolution. Genome structural rearrangements were identified in association with the pathogens spread and several new mating types were identified as originating from within Colombia, further supporting Colombia as the pathogens center of origin. Similarly, a manuscript describing 5 high quality genome assemblies for M. perniciosa is under analysis and preparation, since the project has acquired all the needed sequencing data and completed the sequence assemblies.
Accomplishments
1. Comparative genomes of Moniliophthora roreri, reveal new biological features and exposes evolutionary changes. The fungus, Moniliophthora roreri, causes Frosty Pod Rot, a devastating disease of Theobroma cacao, the source of chocolate. Scientists at USDA ARS in Beltsville, Maryland, completed 5 M. roreri genome assemblies from a diverse collection of pathogen isolates. Comparative analysis identified new mating type alleles that expands what was previously considered a 2-allele mating system. These new mating types came from Colombia, South America. Additional effector candidates were identified and a number of these are unique to Moniliophthora species. This new genomic data supports the hypothesis that M. roreri is a clonally propagated pathogen with its center of diversity and possible center of origin in Colombia, and that the fungus is evolving through significant changes in genome structure that is propagated by clonal linages. This work is being used collaboratively with scientists in the USA and South America to improve disease mitigation activities to improve production.
Review Publications
Boncristiani, D.L., Tauber, J.P., Palmer-Young, E., Cao, L., Chen, Y., Grubbs, K.F., Lopez, J.A., Meinhardt, L.W., Nguyen, V., Oh, S., Peterson, R.J., Zamora, H., Evans, J.D. 2021. Impacts of diverse natural products on honey bee viral loads and health. Journal of Insect Science. 11:10732. https://doi.org/10.3390/app112210732.
Pirro, S., White, D., Bailey, B.A., Meinhardt, L.W. 2022. Genome assemblies of Erythroxylum. coca var. ipadu and Erythroxylum novogranatense var. truxillense. F1000Research. 11:457. https://doi.org/10.12688/f1000research.108549.1.
Ali, S., Amoako-Attah, I., Shao, J.Y., Meinhardt, L.W., Bailey, B.A. 2021. Mitochondrial genomics of six cacao pathogens from the basidiomycete family Marasmiaceae. Frontiers in Microbiology. 12:752094. https://doi.org/10.3389/fmicb.2021.752094.
Akpertey, A., Padi, F., Meinhardt, L.W., Zhang, D. 2022. Relationship between genetic distance based on single nucleotide polymorphism markers and hybrid performance in C. canephora. Plant Breeding. https://doi.org/10.1111/pbr.13005.
Kronmiller, B.A., Feau, N., Shen, D., Tabima, J.F., Ali, S.S., Armitage, A.D., Arredondo, F., Bailey, B.A., Bollmann, S.R., Dale, A., Harrison, R., Hrywkiw, K., Kasuga, T., Mcdougal, R., Nellist, C.F., Panda, P., Tripathy, S., Williams, N.M., Ye, W., Wang, Y., Hamelin, R.C., Grunwald, N.J. 2023. Comparative genomic analysis of 31 Phytophthora genomes reveals genome plasticity and horizontal gene transfer. Molecular Plant-Microbe Interactions. 36:26-46. https://doi.org/10.1094/MPMI-06-22-0133-R.
Leung, J., Cohen, S.P., Baruah, I., Ali, S., Shao, J.Y., Bukari, Y., Amoako-Attah, I., Meinhardt, L.W., Bailey, B.A. 2023. A draft genome resource for Marasmius tenuissimus, an emerging causal agent of thread blight disease in cacao. PhytoFrontiers. https://doi.org/10.1094/PHYTOFR-03-23-0027-A.
Puig, A.S., Cohen, S.P., Koch Bach, R.A., Arciniegas Leal, A., Gutierrez, O.A., Bailey, B.A. 2023. Comparative RNA-seq analysis of resistant and susceptible Theobroma cacao fruits in response to infection with frosty pod pathogen (Moniliophthora roreri). PhytoFrontiers. https://doi.org/10.1094/PHYTOFR-09-22-0101-R.
