Taxonomy and phylogenetic studies of botryosphaeriaceae, diaporthe and endophytic fungi with emphasis on grapevine

Name: Dissanayake, Asha Janadaree

Organization : Mae Fah Luang University

keyword: Fungi

LCSH: Fungi -- Genetics

Classification :.LCCS: QK603

LCSH: Phylogeny

Abstract: Grapevine (Vitis vinifera L.) is one of the most widely cultivated fruit crops in the world. The top five grape growing countries are Spain, China, France, Italy and Turkey. These five leading countries represent 50% of the world vineyards. Grapes have been grown in China for more than 2000 years and after the founding of People’s Republic of China, grape production increased rapidly since 1980. During last two decades China became a leading country for producing grape. Studies related to pathogenic grapevine fungi in Chinese vineyards incorporating DNA sequence data have been carried out since 2010. This study focused on two main fungal pathogen groups that belong to the family Botryosphaeriaceae and the genus Diaporthe, which are considered as the most important pathogens causing grapevine dieback. This study revealed six Botryosphaeriaceae species: Botryosphaeria dothidea, Diplodia seriata, Lasiodiplodia theobromae, L. pseudotheobromae, Neofusicoccum parvum, N. mangiferae and nine Diaporthe species: Diaporthe eres, D. guangxiensis, D. gulyae, D. hongkongensis, D. hubeiensis, D. pescicola, D. phaseolorum, D. sojae and D. unshiuensis causing grapevine dieback throughout China. Their characterization was based on both morphological and multi-gene molecular phylogenies. Pathogenicity tests were carried out to check their virulence and Koch’s postulates were fulfilled successfully, re-isolating the pathogens from all inoculated substrates. This study also focused on Botryosphaeriaceae and Diaporthe fungi from various other hosts from China, Thailand and Italy. The pathogenic Diaporthe species: D. eres, D. momicola, D. pescicola and D. taoicola causing peach tree dieback in Hubei province was studied. Botryosphaeria auasmontanum, B. dothidea, Diplodia alatafructa, Di. iberica, Di. mutila, Di. sapinea, Di. seriata, Di. symphoricarposicola, Di. vidmadera, Dothiorella omnivora, Do. rhamni, Do. sarmentorum, Do. sempervirentis, Eutiarosporella dactylidis and Neofusicoccum parvum were studied including several novel species: Di. crataegicola, Di. galiicola, Dothiorella italica, Mucoharknessia anthoxanthi and Sardiniella celtidis from Italy. Both sexual and asexual morphs of the novel species Barriopsis thailandica, were collected from Thailand. During last five years, seven new species of Diaporthe, D. acericola, D. cichorii, D. dorycnii, D. lonicerae, D. pseudotsugae, D. schoeni and D. torilicola and few other known Diaporthe species: D. eres, D. foeniculina, D. gulyae, D. novem, D. oncostoma, D. ravennica, D. rhusicola, D. rudis and D. sterilis were identified from a collection of 54 isolates in Italy. Two novel species D. aseana, D. garethjonesii were isolated from Thailand. Recently, some taxa in Botryosphaeriaceae have undergone nomenclatural changes and this study provided the first comprehensive overview of Botryosphaeriaceae since 2013. Twenty three genera and 187 species of Botryosphaeriaceae were listed alphabetically, brief notes for species in each genus were provided. Information was provided for the location of ex-type isolates, new host records and geographical distribution of species. Available DNA sequences from ex-type isolates and multilocus phylogenetic trees for the whole order Botryosphaeriales and major genera in Botryosphaeriaceae are provided. An account of the species in the genus Diaporthe was provided focusing the 169 species for which ex-type/ex-epitype/ex-isotype/ex-neotype isolates and corresponding molecular data are available and these species were listed alphabetically. Existence of sexual or asexual morph were noted under each species, detailed descriptions of type materials, host records and geographic distribution were provided. An endophytic study was carried out to determine the fungi inhabiting grapevine stems. To acquire this goal, two different approaches: culture-dependent method and culture-independent method were applied. There is no any study conducted to link next generation sequencing results with taxa detected from a culturing approach, assimilating morphological and multi-gene phylogenetic analysis of endophytes. A total of 94 endophyte isolates were obtained from culture-dependent approach and they were identified into species level using subsets of combined ITS, LSU, SSU, TEF, GPDH, RPB2, HSP60, TUB and ACT sequence data. Each multi-gene phylogenetic analysis allowed the identification of 28 species belong to 19 genera (Acremonium, Alternaria, Arthrinium, Ascorhizoctonia, Aspergillus, Aureobasidium, Bipolaris, Botryosphaeria, Botrytis, Chaetomium, Cladosporium, Curvularia, Hypoxylon, Lasiodiplodia, Mycosphaerella, Nigrospora, Penicillium, Phoma, Scopulariopsis). Species diversity was higher in 13 year-old vines, followed by eight and three years. Fifty nine fungal OTUs were obtained from a sequencing coverage of 55, 822 sequences per sample, being highest in 13 year-old grapevines, followed by eight and three years. Though the results of both methods are consistent related to stem age on fungal community composition, a difference in fungal richness was detected (culture-dependent < culture-independent approaches). An overlap of 53% of fungal genera were detected by a comparison of fungal taxa from both methods. In many cases the OTUs (even with the highly abundant ones) were only assigned to order, family or genus level due to the short fragment of sequences from NGS (in this study the ITS2). To support the taxonomic identification of NGS, the current study sets the standard of applying multigene phylogenies. This study successfully identified many of the NGS derived OTUs with poor taxonomic information in reference databases to the genus or species levels. A taxonomic revision of family Myriangiaceae was also carried out as part of the PhD, but the published manuscript is not included in this thesis since it is unrelated with the thesis title. It is provided as an attachment in the appendix.

Mae Fah Luang University

Address: CHIANG RAI

Email: library@mfu.ac.th

Name: Kevin David Hyde

Role: Advisor

Name: Ekachai Chukeatirote

Role: Co-Advisor

Created: 2017

Issued: 2561

Modified: 2020-05-01

Issued: 2018-02-15

วิทยานิพนธ์/Thesis

application/pdf

CallNumber: Thesis QK603 D613t 2017

eng

DegreeName: Dotor of Philosophy

Level: Doctoral Degree

Descipline: Biosciences

Grantor: Mae Fah Luang University

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