Source: TEXAS A&M UNIVERSITY submitted to NRP
PARTNERS IN CRIME: PATHOBIOME OF FUSARIUM COTTON WILT PATHOGEN
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
ACTIVE
Funding Source
AFRI COMPETITIVE GRANT
Reporting Frequency
Annual
Accession No.
1030588
Grant No.
2023-67013-40174
Cumulative Award Amt.
$849,535.00
Proposal No.
2022-11111
Multistate No.
(N/A)
Project Start Date
Aug 1, 2023
Project End Date
Jul 31, 2026
Grant Year
2023
Program Code
[A1402]- Agricultural Microbiomes in Plant Systems and Natural Resources
Recipient Organization
TEXAS A&M UNIVERSITY
750 AGRONOMY RD STE 2701
COLLEGE STATION,TX 77843-0001
Performing Department
(N/A)
Non Technical Summary
Microorganisms are never alone, constantly collaborating with other species to survive and thrive in their ecosystems. In this cutting-edge project, we will be studying the cooperative relationship between bacteria and pathogenic fungi, with a particular focus on the devastating Fusarium wilt pathogen of cotton. This pathogen poses a serious threat to the production of Pima and upland cotton, but it cannot survive on its own. It relies on a complex microbial community to support its growth and virulence, and we are going to unravel the mysteries of this community. By characterizing the bacterial microbiome that assists the pathogen and studying the genes involved in these interactions, we will gain a deep understanding of how this deadly disease occurs in nature. We will also be developing metabolic models to shed light on the intricate biochemical processes that enable the pathogen to survive and thrive. This research has the potential to revolutionize our understanding of microbial ecology and pave the way for new strategies to combat devastating plant diseases.
Animal Health Component
0%
Research Effort Categories
Basic
50%
Applied
0%
Developmental
50%
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
10240991160100%
Knowledge Area
102 - Soil, Plant, Water, Nutrient Relationships;

Subject Of Investigation
4099 - Microorganisms, general/other;

Field Of Science
1160 - Pathology;
Goals / Objectives
The primary objective of this project is to decode the pathobionts responsible for Fusarium wilt disease in cotton, caused by the fungal species Fusarium oxysporum. This disease is a significant threat to various crops, and different nominal strains or forma speciales can cause wilt disease in cotton. Among these strains, Fusarium oxysporum f. sp. vasinfectum Race 4 (Fov4) causes devastating damage to Pima and upland cotton plants. The focus of this study is to examine the hyphosphere pathobiome of this pathogen. Initially, we will analyze the core pathobiome of different Fov4 isolates to identify any possible variations within them. Subsequently, we will investigate the functional roles of the hyphosphere pathobionts to attain our objectives.
Project Methods
1. Assess hyphosphere microbiome as a wilt pathobiome using molecular tools - sequencing and qPCR2. Isolate and sequence whole genomes of the bacterial pathobiomes using isolation and dna sequencing methods.3. Characterize the effect of the pathobiome on pathogenicity using multispectrum data.4. Combine and model multiomics data to predictive models.

Progress 08/01/23 to 07/31/24

Outputs
Target Audience:The findings of this research were communicated to: - Scientific audience in attendance at the American Phytopathological Society - Cotton Inc - the cotton commodity board - Undergraduate students attending BESC 203 at Texas A&M Changes/Problems:Firstly, we found the SWIFT whole 16S rRNA sequencing kit to be problematic and did not provide the resolution that we had hoped. Thanks to the new sequencing chemistry in Nanopore Technology, we are now able to continue into obtaining full-length 16S rRNA sequences using this method. Secondly, as mentioned previously, we had issues with using soil-based virulence assays. We were unable to obtain bacteria free soils. Autoclaved soils will start showing bacteria within a couple of weeks. So, we have moved on to use a soil-free gelrite based media for seedling growth and microbial inoculation. What opportunities for training and professional development has the project provided?This project was instrumental in training the following personnel - Dr Gayan Abeysinghe - postdoc - Ms Vanessa Thomas - graduate student - Ms Amrita Gabu - undergraduate student intern - Mr Griffin Featherley - undergraduate student intern - Mr Ettiene Ndem - undergraduate student intern How have the results been disseminated to communities of interest?The results have been presented to: Cotton Inc - Cotton commodity board American Phytopathological Society What do you plan to do during the next reporting period to accomplish the goals?We will assess the hyphosphere core microbiota of Fov4 with different strains and cotton plant genotypes. The genomic and phenotypic data of the bacterial isolates will be characterized. If any novel species are encountered, we will describe them as such. The metagenomics information as well as the metatrascriptomics of the fungal-bacterial interactions during pathogenicity will be obtained. We will ensure that the data are timely published and disseminated.

Impacts
What was accomplished under these goals? Obj1: 1. We have screened different isolates of Fusarium oxysporum for their virulence effects on cotton, and also additionally on watermelon and tomato. While we only proposed to perform tests on cotton fusaria, we included the others since these other fusaria are relevant to objective 2. We had not anticipated the need for virulence measures, but we reckoned while setting up the experiments that we will have to validate non-cotton fusaria as well since we need to ensure that these fungi are virulent in our experimental conditions which will be used in Obj 2. Thus, we screened a range of non-cotton fusaria on their respective hosts. 2. Based on the results, we have selected three Fov4 isolates with high, mid, and low virulence levels. We have also selected Fol and Fov1 strains that are virulent in the selected conditions. 3. The hyphosphere assessment test that covered three levels of virulence of Fov4 and three levels of susceptibility in pima cotton has been collected. The 16S rRNA libraries have been created and are about to be sequenced with Nanopore soon. 4. From the hyphosphere samples, we have also isolated bacteria and are now in the process of identifying them through 16S rRNA, dereplicate using phenotypic and genomic characters, and then will perform whole genome sequencing using long-read technology. 5. We will also attempt metagenomic sequencing from the hyphosphere samples. Obj 2. 1. We encountered issues with using soil based assay like we had mentioned in the proposal. The soils, despite autoclaving, had bacterial growth within a couple of weeks. It was impossible to keep them aseptic. So, we moved on to develop a soil free assay for adding seedlings, fungi and the bacteriome. 2. We have conducted the initial bipartite interaction assays between fungi and individual bacterial isolates, and ensured that they are not antagonistic. 3. We have determined that these bacteria, to the contrary, aid in growth of the fungi. 4. In the soil-free assays, we also found that individual bacteria enhances the virulence of the pathogen. 5. We are currently conducting metratranscriptomic and metabolomic analyses of these combinations.

Publications

  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2024 Citation: Zhou Y, Zhang H, Shim WB. Functional characterization of polyketide synthase (PKS) gene PKS5 in Fusarium oxysporum f.sp. vasinfectum. Fungal Genetics Conference, Asilomar, CA. March 2024.
  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2024 Citation: Thomas VE, Isakeit T, Goetz P, Jo Y, Antony-Babu S. Comparative pathobiome study of Fusarium Wilts. International Conference on Plant Pathogenic Bacteria and 5th International Symposium of Biological Control of Bacterial Plant Diseases (ICPPB & Biocontrol), Virginia Tech, Blacksburg, Virginia, July 2024.
  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2024 Citation: Thomas V, Abeysinghe G, Isakeit T, Antony-Babu S. Tracing back the pathobiont assemblage: core microbiota associated with the hyphal tip of Fusarium wilt of cotton and its relationship with the rhizosphere and endosphere microbiomes of diseased cotton. Plant Health 2024, American Phytopathological Society, Memphis, Tennessee, July 2024.
  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2024 Citation: Abeysinghe G and Antony-Babu S. Unraveling the Molecular Dialogues in Fusarium oxysporum f. sp. vasinfectum Race4 (FOV4) Pathobiome of Pima Cotton (Gossypium barbadense) Wilt Disease. Plant Health 2024, American Phytopathological Society, Memphis, Tennessee, July 2024.
  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2024 Citation: Abeysinghe G, Skelton C, Antony-Babu S. Molecular Dialogues of the Harmonious co-conspirators, Fusarium oxysporum f. sp. vasinfectum Race4 (FOV4) and its bacteriome in Pima Cotton (Gossypium barbadense) Wilt Pathogenicity Network. 8th Annual Postdoctoral Research Symposium, Texas A&M University, College Station, TX, September 2024.
  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2024 Citation: Thomas VE, Goetze PK, Jo Y, Antony-Babu S. The Distinguished Microbiome: Differentiating Microbial Dysbiosis and the Pathobiome of Banana Fusarium Tropical Race 4. Southern Divison Meeting, American Phytopathological Society, South Carolina, February 2024.
  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2024 Citation: Featherly G, Abeysinghe G, Antony-Babu S. Soil free pathogenicity analysis of FOV4: Regulation by the pathobiome. Texas A&M University, College Station, TX, March 2024.
  • Type: Journal Articles Status: Accepted Year Published: 2024 Citation: Thomas V, Antony-Babu S. " Core hyphosphere microbiota of Fusarium oxysporum f. sp. niveum" Environmental Microbiome 19(1), 14