An investigation of Fusarium verticillioides infection in maize using physiological and molecular approaches

Simple item page
dc.contributor.advisorRafudeen, Suhailen_ZA
dc.contributor.advisorMurray, Shaneen_ZA
dc.contributor.authorLambarey, Humairaen_ZA
dc.date.accessioned2017-09-23T06:35:12Z
dc.date.available2017-09-23T06:35:12Z
dc.date.issued2017en_ZA
dc.description.abstractAbstract Maize (Zea mays L.) is an important staple food crop in sub-Saharan Africa providing food security to millions of people. Fusarium verticillioides is an important fungal pathogen that infects maize and causes 'Fusarium Ear Rot' which decreases maize kernel yield and quality. In addition, the fungus produces mycotoxins which contaminate the kernel and upon ingestion have negative health consequences for both people and livestock. To this date, there is still no African maize line completely resistant to infection by F. verticillioides. In this study, an African maize line, Zea mays CML144, was infected with F. verticillioides using a soak-seed inoculation method and grown for two weeks under controlled conditions. Analysis of the morphological characteristics showed that compared to the control (mock-infected) maize, infected maize seedlings displayed signs of stunting with leaves shorter & thinner while roots were shorter and displayed visible signs of rotting. Control and infected maize plants were also characterised physiologically and biochemically. Electrolyte leakage experiments were conducted on the meristem regions of the plants after week one and two of infection and showed that leakage increased over time in both control and infected samples with no significant difference observed between the two groups. Biochemical characterisation by analysing superoxide dismutase (SOD), catalase (CAT) and glutathione reductase (GR) antioxidant enzymes showed an increase after the two weeks of infection, indicating a defense response by the plants in response to infection by the fungal pathogen. RNA-sequencing, the main aim of this study was conducted on control and infected plants after two weeks of infection to identify differentially expressed genes (DEGs) involved in F. verticillioides infection. The Illumina NextSeq 500 platform was used to sequence the transcriptome and quantify changes in gene expression. Analysis of the RNA-seq data using the Tuxedo suite of protocols revealed significant DEGs that were both up- and down-regulated in the infected samples compared to the control. Data analysis was conducted using the DNA subway online bioinformatics tool and these results were compared to those obtained using a separate analysis which also incorporated the Tuxedo suite of protocols. Bioinformatic analysis on the RNA-seq DEGs were performed using the agriGO analysis tool which revealed three significant Gene Ontology (GO) terms for both the up- and down-regulated genes, respectively, with the 'response to stimulus' GO-term (within the down-regulated genes) being of specific interest. Other GO-terms included response to chemical stimulus, carbohydrate metabolic process and ion bonding, which also played a role in the defense response when plants were infected by the fungal pathogen. Quantitative Real-Time PCR was performed on five DEGs that were either up- or down-regulated in response to F. verticillioides infection to validate RNA-seq data as well as the GO-analysis results. Quantitative Real-Time PCR was also used as a pre-validation (before RNA-seq) on shrunken-1, a down-regulated gene found in a previously conducted study. We observed that in response to infection by F. verticillioides, expression of shrunken-1 was down-regulated, however, this was not shown to be significant (p>0.05). The results in the current study and the identification of the genes in Zea mays CML144 responding to fungal infection will aid in the goal to develop a maize line completely resistant to F. verticillioides in Africa and in particular South Africa. This would provide improved food security and minimise health risks to the population in the long term. To our knowledge, this is the first study investigating F. verticillioides infection in the African maize line Zea mays CML144 using the soak-seed inoculation method.en_ZA
dc.identifier.apacitationLambarey, H. (2017). <i>An investigation of Fusarium verticillioides infection in maize using physiological and molecular approaches</i>. (Thesis). University of Cape Town ,Faculty of Science ,Department of Molecular and Cell Biology. Retrieved from http://hdl.handle.net/11427/25348en_ZA
dc.identifier.chicagocitationLambarey, Humaira. <i>"An investigation of Fusarium verticillioides infection in maize using physiological and molecular approaches."</i> Thesis., University of Cape Town ,Faculty of Science ,Department of Molecular and Cell Biology, 2017. http://hdl.handle.net/11427/25348en_ZA
dc.identifier.citationLambarey, H. 2017. An investigation of Fusarium verticillioides infection in maize using physiological and molecular approaches. University of Cape Town.en_ZA
dc.identifier.ris TY - Thesis / Dissertation AU - Lambarey, Humaira AB - Abstract Maize (Zea mays L.) is an important staple food crop in sub-Saharan Africa providing food security to millions of people. Fusarium verticillioides is an important fungal pathogen that infects maize and causes 'Fusarium Ear Rot' which decreases maize kernel yield and quality. In addition, the fungus produces mycotoxins which contaminate the kernel and upon ingestion have negative health consequences for both people and livestock. To this date, there is still no African maize line completely resistant to infection by F. verticillioides. In this study, an African maize line, Zea mays CML144, was infected with F. verticillioides using a soak-seed inoculation method and grown for two weeks under controlled conditions. Analysis of the morphological characteristics showed that compared to the control (mock-infected) maize, infected maize seedlings displayed signs of stunting with leaves shorter & thinner while roots were shorter and displayed visible signs of rotting. Control and infected maize plants were also characterised physiologically and biochemically. Electrolyte leakage experiments were conducted on the meristem regions of the plants after week one and two of infection and showed that leakage increased over time in both control and infected samples with no significant difference observed between the two groups. Biochemical characterisation by analysing superoxide dismutase (SOD), catalase (CAT) and glutathione reductase (GR) antioxidant enzymes showed an increase after the two weeks of infection, indicating a defense response by the plants in response to infection by the fungal pathogen. RNA-sequencing, the main aim of this study was conducted on control and infected plants after two weeks of infection to identify differentially expressed genes (DEGs) involved in F. verticillioides infection. The Illumina NextSeq 500 platform was used to sequence the transcriptome and quantify changes in gene expression. Analysis of the RNA-seq data using the Tuxedo suite of protocols revealed significant DEGs that were both up- and down-regulated in the infected samples compared to the control. Data analysis was conducted using the DNA subway online bioinformatics tool and these results were compared to those obtained using a separate analysis which also incorporated the Tuxedo suite of protocols. Bioinformatic analysis on the RNA-seq DEGs were performed using the agriGO analysis tool which revealed three significant Gene Ontology (GO) terms for both the up- and down-regulated genes, respectively, with the 'response to stimulus' GO-term (within the down-regulated genes) being of specific interest. Other GO-terms included response to chemical stimulus, carbohydrate metabolic process and ion bonding, which also played a role in the defense response when plants were infected by the fungal pathogen. Quantitative Real-Time PCR was performed on five DEGs that were either up- or down-regulated in response to F. verticillioides infection to validate RNA-seq data as well as the GO-analysis results. Quantitative Real-Time PCR was also used as a pre-validation (before RNA-seq) on shrunken-1, a down-regulated gene found in a previously conducted study. We observed that in response to infection by F. verticillioides, expression of shrunken-1 was down-regulated, however, this was not shown to be significant (p>0.05). The results in the current study and the identification of the genes in Zea mays CML144 responding to fungal infection will aid in the goal to develop a maize line completely resistant to F. verticillioides in Africa and in particular South Africa. This would provide improved food security and minimise health risks to the population in the long term. To our knowledge, this is the first study investigating F. verticillioides infection in the African maize line Zea mays CML144 using the soak-seed inoculation method. DA - 2017 DB - OpenUCT DP - University of Cape Town LK - https://open.uct.ac.za PB - University of Cape Town PY - 2017 T1 - An investigation of Fusarium verticillioides infection in maize using physiological and molecular approaches TI - An investigation of Fusarium verticillioides infection in maize using physiological and molecular approaches UR - http://hdl.handle.net/11427/25348 ER - en_ZA
dc.identifier.urihttp://hdl.handle.net/11427/25348
dc.identifier.vancouvercitationLambarey H. An investigation of Fusarium verticillioides infection in maize using physiological and molecular approaches. [Thesis]. University of Cape Town ,Faculty of Science ,Department of Molecular and Cell Biology, 2017 [cited yyyy month dd]. Available from: http://hdl.handle.net/11427/25348en_ZA
dc.language.isoengen_ZA
dc.publisher.departmentDepartment of Molecular and Cell Biologyen_ZA
dc.publisher.facultyFaculty of Scienceen_ZA
dc.publisher.institutionUniversity of Cape Town
dc.subject.otherMolecular and Cell Biologyen_ZA
dc.titleAn investigation of Fusarium verticillioides infection in maize using physiological and molecular approachesen_ZA
dc.typeMaster Thesis
dc.type.qualificationlevelMasters
dc.type.qualificationnameMScen_ZA
uct.type.filetypeText
uct.type.filetypeImage
uct.type.publicationResearchen_ZA
uct.type.resourceThesisen_ZA
Files
Original bundle
Now showing 1 - 1 of 1
Thumbnail Image
Name:
thesis_sci_2017_lambarey_humaira.pdf
Size:
2.76 MB
Format:
Adobe Portable Document Format
Description:
Download
Collections
Masters