Analysis of the Pathogenicity and Phylogeny of Colletotrichum Species Associated with Brown Blight of Tea (Camellia sinensis) in Taiwan.

Brown blight, a destructive foliar disease of tea, has become a highly limiting factor for tea cultivation in Taiwan. To understand the population composition of the causal agents (Colletotrichum spp.), the fungal diversity in the main tea-growing regions all over Taiwan was surveyed from 2017 to 2019. A collection of 139 Colletotrichum isolates was obtained from 14 tea cultivars in 86 tea plantations. Phylogenic analysis using the ribosomal internal transcribed spacer, glutamine synthetase gene, Apn2-Mat1-2 intergenic spacer, ß-tubulin, actin, calmodulin, and glyceraldehyde-3-phosphate dehydrogenase genes together with morphological characterization revealed three species associated with brown blight of tea; namely, Colletotrichum camelliae (95.6% of all isolates), C. fructicola (3.7%), and C. aenigma (0.7%). This is the first report of C. aenigma in Taiwan. The optimal growth temperatures were 25°C for C. camelliae and 25 and 30°C for C. fructicola and C. aenigma. Although C. fructicola and C. aenigma were more adapted to high temperature, C. camelliae was the most pathogenic across different temperatures. Regardless of whether spore suspensions or mycelial discs were used, significantly larger lesions and higher disease incidences were observed for wounded than for nonwounded inoculation and for the third and fourth leaves than for the fifth leaves. Wounded inoculation of detached third and fourth tea leaves with mycelial discs was found to be a reliable and efficient method for assessing the pathogenicity of Colletotrichum spp. within 4 days. Preventive application of fungicides or biocontrol agents immediately after tea pruning and at a young leaf stage would help control the disease.

Stemphylium Leaf Blight of Welsh Onion (Allium fistulosum): An Emerging Disease in Sanxing, Taiwan.

Welsh onion (Allium fistulosum L.) is one of the main and oldest vegetable crops grown in Taiwan. A severe epidemic of leaf blight in Welsh onion caused by a Stemphylium-like pathogen was found in Sanxing, Taiwan, from 2018 to 2020. However, correct species identification, biology, and control of Stemphylium leaf blight (SLB) of Welsh onion are not well-established. Therefore, the main objective of this study was to investigate the causal agent of SLB in Sanxing and evaluate the in vitro sensitivity of Stemphylium-like pathogen to commonly used fungicides. A phylogenetic analysis based on combining the internal transcribed spacer (ITS) region and glyceraldedyhe-3-phosphate dehydrogenase (gapdh) and calmodulin (cmdA) gene sequences together with morphological features identified that S. vesicarium is associated with SLB in Sanxing. When inoculated onto Welsh onion leaves, the isolates caused symptoms identical to those observed in the field; therefore, S. vesicarium was reisolated and Koch's postulates were confirmed. We observed a higher incidence of SLB symptoms on the oldest leaves compared with younger leaves. The maximum and minimum temperatures for in vitro mycelial growth and conidial germination (%) of S. vesicarium were 20 to 30°C and 5°C, respectively. Sixteen fungicides were tested for their effectiveness to reduce the mycelial growth and conidial germination of S. vesicarium in vitro. Boscalid plus pyraclostrobin, fluopyram, fluxapyroxad, and fluxapyroxad plus pyraclostrobin were highly effective at reducing mycelial growth and conidial germination in S. vesicarium. However, strobilurin fungicides (azoxystrobin and kresoxim-methyl) commonly used in Welsh onion production in Sanxing were ineffective. This study discusses the emergence of SLB caused by S. vesicarium in the foliar disease complex affecting Welsh onion and the management of the disease using fungicides with different modes of action in Taiwan. The research will support the sustainable management of SLB in Sanxing, Taiwan; however, further field assessments of the fungicides are warranted.

Cryptic Diversity, Molecular Systematics, and Pathogenicity of Genus Pestalotiopsis and Allied Genera Causing Gray Blight Disease of Tea in Taiwan, With a Description of a New Pseudopestalotiopsis Species.

Camellia sinensis (L.) O. Kuntze, commonly known as tea, is widely cultivated around the world in tropical and subtropical areas. Tea is mainly manufactured using young shoots of tea plants. Therefore, it is essential to control foliar diseases. Gray blight disease is caused by pestalotiopsis-like taxa and is known as one of the most destructive tea diseases. Although several studies have provided the groundwork for the fungal diseases associated with C. sinensis in Taiwan, gray blight disease has not been characterized based on diversity, molecular systematics, or pathogenicity. The goal of this study was to identify and characterize the causative agents of tea gray blight disease. A total of 98 pestalotiopsis-like isolates associated with symptomatic leaves of C. sinensis from major tea fields in Taiwan were investigated. Based on phylogenies of single and concatenated DNA sequences (internal transcribed spacer, ß-tubulin, translation elongation factor 1-α) together with morphology, we resolved most of the pestalotiopsis-like species in this study. The study revealed seven well-classified taxa and seven tentative clades in three genera: Pestalotiopsis, Pseudopestalotiopsis, and Neopestalotiopsis. One novel species, Pseudopestalotiopsis annellata, was introduced. Five new records, Pseudopestalotiopsis chinensis, Pseudopestalotiopsis camelliae-sinensis, Pestalotiopsis camelliae, Pestalotiopsis yanglingensis, and Pestalotiopsis trachicarpicola, were introduced for the first time in Taiwan. Pseudopestalotiopsis chinensis was the taxon most frequently isolated from C. sinensis in this study. Furthermore, results of pathogenicity assessments exhibited that, with wound inoculation, all assayed isolates in this study were pathogenic on tea leaves. Pseudopestalotiopsis chinensis and Pseudopestalotiopsis camelliae-sinensis were identified as the major pathogens associated with gray blight disease of tea in Taiwan. To our knowledge, this is the first study of the diversity, pathogenicity, and characterization of pestalotiopsis-like fungi causing tea gray blight disease in Taiwan.

Species diversity of Pleosporalean taxa associated with Camellia sinensis (L.) Kuntze in Taiwan.

Pleosporales species are important plant pathogens, saprobes, and endophytes on a wide range of economically important plant hosts. The classification of Pleosporales has undergone various modifications in recent years due to the addition of many families described from multiple habitats with a high level of morphological deviation. Numerous asexual genera have been described in Pleosporales that can be either hyphomyceteous or coelomycetous. Phoma- or coniothyrium-like species are common and have been revealed as polyphyletic in the order Pleosporales and linked with several sexual genera. A total of 31 pleosporalean strains were isolated in different regions of Taiwan between 2017 and 2018 from the leaves of Camellia sinensis plants with symptoms of leaf spot disease. These strains were evaluated morphologically and genotypically using multi-locus sequence analyses of the ITS, LSU, SSU, rpb2, tef1 and tub2 genes. The results demonstrated the affiliation of these strains with the various families in Pleosporales and revealed the presence of one new genus (Neoshiraia) and eight new species (Alloconiothyrium camelliae, Amorocoelophoma camelliae, Leucaenicola camelliae, L. taiwanensis, Neoshiraia camelliae, N. taiwanensis, Paraconiothyrium camelliae and Paraphaeosphaeria camelliae). Furthermore, to the best of our understanding, Didymella segeticola, Ectophoma pomi and Roussoella mexican were reported for the first time from C. sinensis in Taiwan.