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The white-rot fungus Phellinus noxius is known to cause brown root rot disease (BRRD) in woody trees and shrubs. To understand the pathogenicity of P. noxius in herbaceous plants, we investigated 23 herbaceous weed and turfgrass species in 32 BRRD-infested sites in Taiwan and/or tested them by artificial inoculation. In the field survey, P. noxius was isolated from seven symptomless herbaceous species (i.e., Typhonium blumei, Paspalum conjugatum, Paspalum distichum, Oplismenus compositus, Bidens pilosa, Digitaria ciliaris, and Zoysia matrella). Potted plant inoculation assays suggested that P. noxius is able to infect Artemisia princeps, O. compositus, and Z. matrella but not Axonopus compressus, Eremochloa ophiuroides, Ophiopogon japonicus, or Cynodon dactylon. A. princeps plants wilted within 2 weeks postinoculation, but inoculated O. compositus and Z. matrella were asymptomatic, and P. noxius could be isolated from only inoculated sites. The colonization of P. noxius in the cortex and vascular cylinder of roots was visualized by paraffin sectioning and trypan blue staining of juvenile seedlings grown on water agar. To evaluate the effect of replantation for the remediation of BRRD-infested sites, P. noxius-inoculated wood strips were buried in soil with or without vegetation. After 4 weeks, P. noxius could be detected only in the bare soil group. For the control of BRRD, the herbaceous hosts should be removed around the diseased trees/stumps and non-host turfgrasses (e.g., A. compressus, E. ophiuroides, O. japonicus, or C. dactylon) planted to accelerate the degradation of P. noxius.

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, β-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.

Rice blast caused by Magnaporthe oryzae is a dangerous threat to rice production and food security worldwide. Breeding and proper deployment of resistant varieties are effective and environmentally friendly strategies to manage this notorious disease. However, a highly dynamic and quickly evolved rice blast pathogen population in the field has made disease control with resistance germplasms more challenging. Therefore, continued monitoring of pathogen dynamics and application of effective resistance varieties are critical tasks to prolong or sustain field resistance. Here, we report a team project that involved evaluation of rice blast resistance genes and surveillance of M. oryzae field populations in Taiwan. A set of International Rice Research Institute-bred blast-resistant lines (IRBLs) carrying single blast resistance genes was utilized to monitor the field effectiveness of rice blast resistance. Resistance genes such as Ptr (formerly Pita2) and Pi9 exhibited the best and most durable resistance against the rice blast fungus population in Taiwan. Interestingly, line IRBLb-B harboring the Pib gene with good field protection has recently shown susceptible lesions in some locations. To dissect the genotypic features of virulent isolates against the Pib resistance gene, M. oryzae isolates were collected and analyzed. Screening of the AvrPib locus revealed that the majority of field isolates still maintained the wild-type AvrPib status but eight virulent genotypes were found. Pot3 insertion appeared to be a major way to disrupt the AvrPib avirulence function. Interestingly, a novel AvrPib double-allele genotype among virulent isolates was first identified. Pot2 repetitive element-based polymerase chain reaction (rep-PCR) fingerprinting analysis indicated that mutation events may occur independently among different lineages in different geographic locations of Taiwan. This study provides our surveillance experience of rice blast disease and serves as the foundation to sustain rice production.

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.

Brown root rot disease (BRRD), caused by

The white-rot fungus

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

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, β-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.

The Fusarium solani species complex (FSSC) comprises fungal pathogens responsible for mortality in a diverse range of animals and plants, but their genome diversity and transcriptome responses in animal pathogenicity remain to be elucidated. We sequenced and compared six chromosome-level FSSC clade 3 genomes of aquatic animal and plant host origins and revealed a spectrum of conservation patterns in chromosomes categorised into three compartments: core, fast-core (FC), and lineage-specific (LS). Each chromosome type varied in structural architectures, with FC and LS chromosomes containing significantly higher proportions of repetitive elements and methylation levels than core chromosomes, with genes exhibiting higher dN/dS and enriched in functions related to pathogenicity and niche expansion. Mesosynteny were detected between FC chromosomes of Fusarium genomes, indicating that these chromosomes were present in a common ancestor that predated FSSC species. These findings provide evidence that genome compartmentalisation was the outcome of multi-speed evolution amongst FSSC chromosomes. We further demonstrated that F. falciforme and F. keratoplasticum are opportunistic pathogens by inoculating Pelodiscus sinensis eggs and identified differentially expressed genes also associated with plant pathogenicity. These included the most upregulated genes encoding the CFEM (Common in Fungal Extracellular Membrane) domain. The study establishes genomic resources and an animal model for fungal pathogens of trans-kingdom hosts.

Strawberry is a small fruit crop with high economic value. Anthracnose caused by Colletotrichum spp. poses a serious threat to strawberry production, particularly in warm and humid climates, but knowledge of pathogen populations in tropical and subtropical regions is limited. To investigate the diversity of infectious agents causing strawberry anthracnose in Taiwan, a disease survey was conducted from 2010 to 2018, and Colletotrichum spp. were identified through morphological characterization and multilocus phylogenetic analysis with internal transcribed spacer, glyceraldehyde 3-phosphate dehydrogenase, chitin synthase, actin, beta-tubulin, calmodulin, and the intergenic region between Apn2 and MAT1-2-1 (ApMAT). Among 52 isolates collected from 24 farms/nurseries in Taiwan, a new species, Colletotrichum miaoliense sp. nov. (6% of all isolates), a species not previously known to be associated with strawberry, Colletotrichum karstii (6%), and three known species, Colletotrichum siamense (75%), Colletotrichum fructicola (11%), and Colletotrichum boninense (2%), were identified. The predominant species C. siamense and C. fructicola exhibited higher mycelial growth rates on potato dextrose agar and caused larger lesions on wounded and non-wounded detached strawberry leaves. Colletotrichum boninense, C. karstii, and C. miaoliense only caused lesions on wounded leaves. Understanding the composition and biology of the pathogen population will help in disease management and resistance breeding.

Background Bakanae is a seedborne disease caused by Fusarium fujikuroi. Rice seedlings emerging from infected seeds can show diverse symptoms such as elongated and slender stem and leaves, pale coloring, a large leaf angle, stunted growth and even death. Little is known about rice defense mechanisms at early stages of disease development. Results This study focused on investigating early defenses against F. fujikuroi in a susceptible cultivar, Zerawchanica karatals (ZK), and a resistant cultivar, Tainung 67 (TNG67). Quantitative PCR revealed that F. fujikuroi colonizes the root and stem but not leaf tissues. Illumina sequencing was conducted to analyze the stem transcriptomes of F. fujikuroi-inoculated and mock-inoculated ZK and TNG67 plants collected at 7 days post inoculation (dpi). More differentially expressed genes (DEGs) were identified in ZK (n = 169) than TNG67 (n = 118), and gene ontology terms related to transcription factor activity and phosphorylation were specifically enriched in ZK DEGs. Among the complex phytohormone biosynthesis and signaling pathways, only DEGs involved in the jasmonic acid (JA) signaling pathway were identified. Fourteen DEGs encoding pattern-recognition receptors, transcription factors, and JA signaling pathway components were validated by performing quantitative reverse transcription PCR analysis of individual plants. Significant repression of jasmonate ZIM-domain (JAZ) genes (OsJAZ9, OsJAZ10, and OsJAZ13) at 3 dpi and 7 dpi in both cultivars, indicated the activation of JA signaling during early interactions between rice and F. fujikuroi. Differential expression was not detected for salicylic acid marker genes encoding phenylalanine ammonia-lyase 1 and non-expressor of pathogenesis-related genes 1. Moreover, while MeJA did not affect the viability of F. fujikuroi, MeJA treatment of rice seeds (prior to or after inoculation) alleviated and delayed bakanae disease development in susceptible ZK. Conclusions Different from previous transcriptome studies, which analyzed the leaves of infected plants, this study provides insights into defense-related gene expression patterns in F. fujikuroi–colonized rice stem tissues. Twelve out of the 14 selected DEGs were for the first time shown to be associated with disease resistance, and JA-mediated resistance was identified as a crucial component of rice defense against F. fujikuroi. Detailed mechanisms underlying the JA-mediated bakanae resistance and the novel defense-related DEGs are worthy of further investigation.

Secondary branch number per panicle plays a crucial role in regulating grain number and yield in rice. Here, we report the positional cloning and functional characterization for SECONDARY BRANCH NUMBER7 (qSBN7), a quantitative trait locus affecting secondary branch per panicle and grain number. Our research revealed that the causative variants of qSBN7 are located in the distal promoter region of FRIZZLE PANICLE (FZP), a gene previously associated with the repression of axillary meristem development in rice spikelets. qSBN7 is a novel allele of FZP that causes an ∼56% decrease in its transcriptional level, leading to increased secondary branch and grain number, and reduced grain length. Field evaluations showed that qSBN7 increased grain yield by 10.9% in a temperate japonica variety, TN13, likely due to its positive effect on sink capacity. Our findings suggest that incorporation of qSBN7 can increase yield potential and improve the breeding of elite rice varieties.

Phellinus noxius causes brown root rot (BRR) of diverse trees. Basidiospores and diseased host tissues have been recognized as important sources of P. noxius inoculum. This study aimed to understand whether P. noxius could occur or survive in soil without host tissues in the natural environment. Soil was sampled before and after the removal of diseased trees at eight BRR infection sites (total of 44 samples). No P. noxius colonies were recovered in soil plating assays, suggesting that no or little viable P. noxius resided in the soil. To know whether P. noxius could disseminate from decayed roots to the surrounding soil, rhizosphere and non-rhizosphere soils were sampled from another two infection sites. Although P. noxius DNA was detectable with specific primers, no P. noxius could be isolated, even from the rhizosphere soils around decayed roots covered with P. noxius mycelial mats. The association between viable P. noxius and the presence of its DNA was also investigated using field soil mixed with P. noxius arthrospores. After P. noxius was exterminated by flooding or fumigation treatment, its DNA remained detectable for a few weeks. The potential of onsite soil as an inoculum was tested using the highly susceptible loquat (Eriobotrya japonica). Loquats replanted in an infection site that had been cleaned up by simply removing the diseased stump and visible residual roots remained healthy for a year. Taken together, P. noxius is not a soilborne pathogen, and diseased host tissues should be the focus of field sanitation and detection for BRR.

Phellinus noxius, the cause of brown root-rot disease, is an invasive fungal pathogen that causes a white rot among woody plants in Asia, Oceania, and Africa. Because the origin and diversity of this pathogen are unknown, it is difficult to predict its behavior and invasive capacity, especially under future climate-change scenarios. We characterized genetic relationships and ecological differences among P. noxius lineages across eastern Asia and Oceania to better understand evolutionary responses of the pathogen to environmental changes. Sequences of four loci (nuclear large subunit, internal transcribed spacers, partial RNA polymerase II, and partial translation elongation factor – 1 alpha) from 95 P. noxius isolates were used for genetic analyses. Our analyses revealed three genetically distinct lineages of P. noxius: 1) eastern Asia (Japan, Taiwan, Hong Kong, and Malaysia); 2) western Oceania/Japan/Taiwan (including Australia, Palau, Guam, Saipan, Yap, Pohnpei, and Kosrae) with some isolates from Japan and Taiwan; and 3) a distinct group from American Samoa. Population genetic analyses highlighted admixture and migration among the three lineages. Climate-based, species distribution models were used to predict ecological patterns of P. noxius for two genetic lineages: eastern Asia and western Oceania/Japan/Taiwan. Contemporary bioclimatic models depicted potential areas at high risk for P. noxius invasion, and predicted that suitable climate space (potential distribution) is lineage specific. Trade of important economic crops worldwide coupled with changing climates could exacerbate the spread of P. noxius into new geographic areas with suitable habitats for brown root-rot disease.

Brown root rot (BRR), caused by the white rot fungus Phellinus noxius, is an epidemic disease of diverse broadleaved and coniferous tree species in many tropical and subtropical regions. Flooding and trenching control measures are difficult to implement, and chemical controls can have an adverse impact on ecosystems. Previous studies have provided in vitro evidence for the potential use of Trichoderma spp. for biocontrol of BRR. Here, we analyzed the in vitro antagonistic and mycoparasitic abilities of four Trichoderma spp. isolates against four P. noxius isolates in dual culture and Ficus microcarpa wood blocks. A convenient inoculation system based on root inoculation of a highly susceptible loquat (Eriobotrya japonica) with P. noxius-colonized wheat-oat grains was developed to examine the effect of Trichoderma treatment in planta. Preventive application of Trichoderma asperellum TA, the isolate showing high antagonistic activity in vitro, was effective in preventing and delaying the wilting of P. noxius-inoculated loquat cuttings in greenhouse trials. To understand the specific niche in which T. asperellum TA interacts with P. noxius, KOH-aniline blue fluorescence microscopy was used to investigate the colonization of loquat roots by P. noxius and/or T. asperellum TA. Dilution plating assays were also conducted to quantify Trichoderma populations in the rhizosphere and potting mix. T. asperellum TA was able to robustly establish in the rhizosphere and potting mix but with scarce root penetration limited to the superficial layer. We discuss the timing and strategy for applying antagonistic Trichodema sp. on living trees or in BRR-infested areas for BRR management.

Brown root rot (BRR) caused by Phellinus noxius is a destructive tree disease in tropical and subtropical areas. To understand how BRR affects the composition of the plant rhizoplane-enriched microbiota, the microbiomes within five root-associated compartments (i.e., bulk soil, old/young root rhizosphere soil, old/young root tissue) of Ficus trees naturally infected by P. noxius were investigated. The level of P. noxius infection was determined by quantitative PCR. Illumina sequencing of the internal transcribed spacer and 16S rRNA revealed that P. noxius infection caused a significant reduction in fungal diversity in the bulk soil, old root tissue, and rhizosphere soil. Interestingly, Cosmospora was the only fungal genus positively correlated with P. noxius. The abundance and composition of dominant bacterial taxa such as Actinomadura, Bacillus, Rhodoplanes, and Streptomyces differed between BRR-diseased and healthy samples. Furthermore, 838 isolates belonging to 26 fungal and 35 bacterial genera were isolated and tested for interactions with P. noxius. Antagonistic activities were observed for isolates of Bacillus, Pseudomonas, Aspergillus, Penicillium, and Trichoderma. Cellophane overlay and cellulose/lignin utilization assays suggested that Cosmospora could tolerate the secretions of P. noxius and that the degradation of lignin by P. noxius may create suitable conditions for Cosmorpora growth. This article is protected by copyright. All rights reserved.

Welsh onion (

Rice blast is a serious threat to global rice production. Large-scale and long-term cultivation of rice varieties with a single blast resistance gene usually leads to breakdown of resistance. To effectively control rice blast in Taiwan, marker-assisted backcrossing was conducted to develop monogenic lines carrying different blast resistance genes in the genetic background of an elite japonica rice cultivar, Kaohsiung 145 (KH145). Eleven International Rice Research Institute (IRRI)-bred blast-resistant lines (IRBLs) showing broad-spectrum resistance to local Pyricularia oryzae isolates were used as resistance donors. Sequencing analysis revealed that the recurrent parent, KH145, does not carry known resistance alleles at the target Pi2/9, Pik, Pita, and Ptr loci. For each IRBL × KH145 cross, we screened 21 to 370 (average of 108) plants per generation from the BC1F1 to BC3F1/BC4F1 generation. A total of 1,499 BC3F2/BC4F2 lines carrying homozygous resistance alleles were selected and self-crossed for four to six successive generations. The derived lines were also evaluated for background genotype using genotyping by sequencing, for blast resistance under artificial inoculation and natural infection conditions, and for agronomic performance in multiple field trials. In Chiayi and Taitung blast nurseries in 2018 to 2020, Pi2, Pi9, and Ptr conferred high resistance, Pi20 and Pik-h moderate resistance, and Pi1, Pi7, Pik-p, and Pik susceptibility to leaf blast; only Pi2, Pi9, and Ptr conferred effective resistance against panicle blast. The monogenic lines showed agronomic traits, yield, and grain quality similar to those of KH145, suggesting the potential of growing a mixture of lines to achieve durable resistance in the field.

Comparative genomics of fungal mitochondrial genomes (mitogenomes) have revealed a remarkable pattern of rearrangement between and within major phyla owing to horizontal gene transfer (HGT) and recombination. The role of recombination was exemplified at a finer evolutionary time scale in basidiomycetes group of fungi as they display a diversity of mitochondrial DNA (mtDNA) inheritance patterns. Here, we assembled mitogenomes of six species from the Hymenochaetales order of basidiomycetes and examined 59 mitogenomes from two genetic lineages ofPyrrhoderma noxium. Gene order is largely colinear while intergene regions are major determinants of mitogenome size variation. Substantial sequence divergence was found in shared introns consistent with high HGT frequency observed in yeasts, but we also identified a rare case where an intron was retained in five species since speciation. In contrast to the hyperdiversity observed in nuclear genomes ofP. noxium, mitogenomes’ intraspecific polymorphisms at protein coding sequences are extremely low. Phylogeny based on introns revealed turnover as well as exchange of introns between two lineages. Strikingly, some strains harbor a mosaic origin of introns from both lineages. Analysis of intergenic sequence indicated substantial differences between and within lineages, and an expansion may be ongoing as a result of exchange between distal intergenes. These findings suggest that the evolution in mtDNAs is usually lineage specific but chimeric mitotypes are frequently observed, thus capturing the possible evolutionary processes shaping mitogenomes in a basidiomycete. The large mitogenome sizes reported in various basidiomycetes appear to be a result of interspecific reshuffling of intergenes.

The high-power ultrasonic cleaning system is used in the manufacturing of clean product contaminants. The pulse density modulation (PDM) switching strategy is more efficient for cleaning the rough surface of equipment. This paper proposes a novel pulse density modulation duty cycle stepping (PDMDCS) method that incorporates a proportional–integral–derivative controller and fuzzy logic for the mechanical resonance frequency tracking and PDM duty cycle stepping for ultrasonic cleaning system power control. The experimental results show that based on the same hardware architecture, the proposed PDMDCS method increases the power control range and resolution of power control without adding more hardware that compares to mechanical resonance frequency control method.

For the past 30 years, the most predominant strawberry cultivar in Taiwan has been 'Taoyuan No. 1', which produces fruit with rich flavor and aroma but is highly susceptible to anthracnose (Chung et al. 2019). Because epidemics of anthracnose became more destructive, farmers switched to an anthracnose-tolerant cultivar 'Xiang-Shui' (~50% and ~80% of the cultivation area in 2018 and 2019, respectively). Since 2018, severe leaf blight and crown rot symptoms have been observed all year in 'Xiang-Shui' in Miaoli, Nantou, Hsinchu, Taipei, Taoyuan, and Chiayi Counties. The disease became more prevalent and severe during 2019 to 2020 and caused up to 30% plant loss after transplanting. Symptoms appeared as brown necrotic lesions with black acervuli on leaves, slightly sunken dark-brown necrosis on stolons, and sunken reddish-brown necrosis on fruit. The diseased crown tissue showed marbled reddish-brown necrosis with a dark-brown margin, and plants with severe crown rot usually showed reddish-brown discoloration on leaves (the leaves initially turned reddish-brown between the veins and could become entirely scorched at later stages). To isolate the causal agent, small fragments of diseased leaves, crowns, stolons, and fruits were surface-disinfested with 0.5% sodium hypochlorite for 30 seconds, rinsed with sterile water then placed on 1.5% water agar. Single hyphal tips extended from tissues were transferred to potato dextrose agar and cultured for 7 days at 25°C under a 12-h/12-h photoperiod. Total 20 isolates were obtained from diseased leaves, crowns, stolons, and fruits. Colonies were white with cottony aerial mycelium, irregular margins, and black acervuli distributed in concentric rings. Conidia were fusiform to ellipsoid (five cells) with one basal appendage and three or four (usually three) apical appendages. From colony and conidial morphology, the causal agent was identified as

Bakanae disease in rice can cause abnormal elongation of the stem and leaves, development of adventitious roots, a larger leaf angle, and even death. Little is known about the infection, colonization, and distribution of

Secondary branch number per panicle plays a crucial role in regulating grain number and yield in rice. Here, we report the positional cloning and functional characterization for SECONDARY BRANCH NUMBER7 (qSBN7), a quantitative trait locus affecting secondary branch per panicle and grain number. Our research revealed that the causative variants of qSBN7 are located in the distal promoter region of FRIZZLE PANICLE (FZP), a gene previously associated with the repression of axillary meristem development in rice spikelets. qSBN7 is a novel allele of FZP that causes an ∼56% decrease in its transcriptional level, leading to increased secondary branch and grain number, and reduced grain length. Field evaluations showed that qSBN7 increased grain yield by 10.9% in a temperate japonica variety, TN13, likely due to its positive effect on sink capacity. Our findings suggest that incorporation of qSBN7 can increase yield potential and improve the breeding of elite rice varieties.

The order Hymenochaetales of white rot fungi contain some of the most aggressive wood decayers causing tree deaths around the world. Despite their ecological importance and the impact of diseases they cause, little is known about the evolution and transmission patterns of these pathogens. Here, we sequenced and undertook comparative genomics analyses of Hymenochaetales genomes using brown root rot fungus Phellinus noxius, wood-decomposing fungus Phellinus lamaensis, laminated root rot fungus Phellinus sulphurascens, and trunk pathogen Porodaedalea pini. Many gene families of lignin-degrading enzymes were identified from these fungi, reflecting their ability as white rot fungi. Comparing against distant fungi highlighted the expansion of 1,3-beta-glucan synthases in P. noxius, which may account for its fast-growing attribute. We identified 13 linkage groups conserved within Agaricomycetes, suggesting the evolution of stable karyotypes. We determined that P. noxius has a bipolar heterothallic mating system, with unusual highly expanded ~60 kb A locus as a result of accumulating gene transposition. We investigated the population genomics of 60 P. noxius isolates across multiple islands of the Asia Pacific region. Whole-genome sequencing showed this multinucleate species contains abundant poly-allelic single-nucleotide-polymorphisms (SNPs) with atypical allele frequencies. Different patterns of intra-isolate polymorphism reflect mono-/heterokaryotic states which are both prevalent in nature. We have shown two genetically separated lineages with one spanning across many islands despite the geographical barriers. Both populations possess extraordinary genetic diversity and show contrasting evolutionary scenarios. These results provide a framework to further investigate the genetic basis underlying the fitness and virulence of white rot fungi.

Angular leaf spot of strawberry, considered an A2 quarantine pest by the European and Mediterranean Plant Protection Organization (EPPO 2019), is an important bacterial disease in many regions. Since 2017, symptoms similar to angular leaf spot were observed in several strawberry cultivars including 'Taoyuan No. 1' and 'Xiang-Shui'. Early symptoms were angular, water-soaked lesions on the abaxial leaf surface, and later, reddish-brown irregular spots and coalesced lesions developed on the adaxial surface. In the humid conditions, sticky bacterial ooze exuding from lesions was observed. To isolate the causal agent, leaves showing water-soaked lesions were surface sterilized, cut into small pieces and soaked in 5 ml sterile water for at least 15 min. The supernatant from the cut-up pieces was serially diluted followed by spreading on sucrose peptone agar (SPA) (Hayward 1960). After incubating at 20°C for 4-5 days, single colonies grown on SPA were transferred to a new SPA plate and cultured at 20°C until colonies appeared. The yellow, glossy and mucoid colonies, which resembled the colony morphology of Xanthomonas fragariae, were selected as candidates for further confirmation. First, bacterial DNA of four candidate isolates, B001, B003 and B005 from Miaoli County and B004 from Taoyuan City, was PCR amplified with X. fragariae-specific primers: XF9/XF12 (Roberts et al. 1996) and 245A/B and 295A/B (Pooler et al. 1996). All four isolates could be detected by XF9/XF12 primer. Furthermore, isolates B003 and B004 could be detected by both 245A/B and 295A/B primers, while B001 and B005 could be detected by 295A/B only. Next, DNA gyrase subunit B (gyrB) was PCR amplified with the primers XgyrB1F/XgyrB1R (Young et al. 2008). The gyrB sequences of these four isolates were deposited in GenBank with accession numbers MT754942 to MT754945. The gyrB phylogenetic tree was constructed based on Bayesian inference analysis and maximum likelihood analysis. The gyrB sequences of the four isolates from Taiwan clustered in the clade containing the type strain of X. fragariae ICMP5715, indicating that they belong to X. fragariae. B001 and B005 formed a sub-group separated from B003 and B004, suggesting genetic differences between these isolates. To fulfill Koch's postulates, the abaxial surface of strawberry leaves were syringe infiltrated (KJP Silva et al., 2017) or wounded inoculated (Wang et al., 2017) with bacterial suspensions (final OD600 = 1.0-2.0) prepared from colonies of B001 and B003 washed from SPA plates. Inoculated plants were enclosed in a plastic bag (> 90% RH) at 25/20°C (day/night) under a 12-h/12-h photoperiod. After 7-14 days, water-soaked lesions similar to those observed in the field were developed on all inoculated leaves. The bacteria were successfully re-isolated from lesions of inoculated leaves and confirmed by specific primers XF9/XF12, 245A/B and 295A/B. We also found that the disease commonly occurs in the strawberry fields/nurseries with sprinkler irrigation during winter or early spring, and was particularly serious in the windward side or near riverside. To our knowledge, this is the first report of X. fragariae causing angular leaf spot on strawberry in Taiwan. Currently, the disease only occurs severely in certain regions, but establishment of effective management strategies will be needed to prevent spreading of this disease and potential economic loss in the future.

Secondary branch number (SBN) is an important component affecting spikelet number per panicle (SPP) and yield in rice. During recurrent backcross breeding, four BC2F4 populations derived from the high-yield donor parent IR65598-112-2 and the recurrent parent Tainan 13 (a local japonica cultivar) showed discontinuous variations of SPP and SBN within populations. Genetic analysis of 92 BC2F4 individuals suggested that both SPP and SBN are controlled by a single recessive allele. Two parents and 37 BC2F4 individuals showing high- and low-SBN type phenotypes were analyzed by restriction-site associated DNA sequencing (RAD-seq). Based on 2,522 reliable SNPs, the qSBN7 was mapped to a distal region of the long arm of chromosome 7. Trait-marker association analysis with an additional 166 high-SBN type BC2F4 individuals and 8 newly developed cleaved amplified polymorphic sequence markers further delimited the qSBN7 locus to a 601.4-kb region between the markers SNP2788 and SNP2849. Phenotype evaluation of two BC2F5 backcross inbred lines revealed that qSBN7 increased SPP by 83.2% and SBN by 61.0%. The qSBN7 of IR65598-112-2 could be used for improving reproductive sink capacity in rice.

Brown root rot (BRR), caused by the white rot fungus

For the past 30 years, the most predominant strawberry cultivar in Taiwan has been 'Taoyuan No. 1', which produces fruit with rich flavor and aroma but is highly susceptible to anthracnose (Chung et al. 2019). Because epidemics of anthracnose became more destructive, farmers switched to an anthracnose-tolerant cultivar 'Xiang-Shui' (~50% and ~80% of the cultivation area in 2018 and 2019, respectively). Since 2018, severe leaf blight and crown rot symptoms have been observed all year in 'Xiang-Shui' in Miaoli, Nantou, Hsinchu, Taipei, Taoyuan, and Chiayi Counties. The disease became more prevalent and severe during 2019 to 2020 and caused up to 30% plant loss after transplanting. Symptoms appeared as brown necrotic lesions with black acervuli on leaves, slightly sunken dark-brown necrosis on stolons, and sunken reddish-brown necrosis on fruit. The diseased crown tissue showed marbled reddish-brown necrosis with a dark-brown margin, and plants with severe crown rot usually showed reddish-brown discoloration on leaves (the leaves initially turned reddish-brown between the veins and could become entirely scorched at later stages). To isolate the causal agent, small fragments of diseased leaves, crowns, stolons, and fruits were surface-disinfested with 0.5% sodium hypochlorite for 30 seconds, rinsed with sterile water then placed on 1.5% water agar. Single hyphal tips extended from tissues were transferred to potato dextrose agar and cultured for 7 days at 25°C under a 12-h/12-h photoperiod. Total 20 isolates were obtained from diseased leaves, crowns, stolons, and fruits. Colonies were white with cottony aerial mycelium, irregular margins, and black acervuli distributed in concentric rings. Conidia were fusiform to ellipsoid (five cells) with one basal appendage and three or four (usually three) apical appendages. From colony and conidial morphology, the causal agent was identified as Neopestalotiopsis sp. (Maharachchikumbura et al. 2014). The internal transcribed spacer (ITS) region, β-tubulin (TUB), and translation elongation factor 1-alpha (TEF-1α) of three isolates (ML1664 from diseased crown tissue collected in Hsinchu County; ML2147 and ML2411 from diseased leaves collected in Miaoli County) were sequenced (GenBank nos. MT469940 to MT469948). All three isolates clustered with the ex-type strain of Neopestalotiopsis rosae in the multilocus (ITS+TUB+TEF-1α) phylogenetic tree. To fulfill Koch's postulates, spore suspensions of ML1664 and ML2147 at 1×106 conidia/mL were used to spray-inoculate 'Xiang-Shui' seedlings at the 3 to 4 leaf stage until run-off (two trials, five seedlings per trial). Inoculated plants were put in a plastic bag (> 90% RH) at 25°C under a 12-h/12-h photoperiod. After 10-14 days, 80% of inoculated plants showed leaf or crown symptoms similar to those in the field. Control plants sprayed with sterile water showed no symptoms (4-5 seedlings per trial). The fungi were re-isolated from necrotic lesions with 100% frequency (n ≥ 3 isolates per trial), and morphological characters and ITS sequences were identical to the original ones. This is the first report of N. rosae causing leaf blight and crown rot in strawberry in Taiwan. N. rosae and N. clavispora have been reported as new threats to strawberry in several other countries (Rebollar-Alviter 2020; Gilardi 2019). Clarification of the pathogen provides a basis for developing strategies to control the emerging disease. Further studies are needed to evaluate the resistance/susceptibility of major strawberry cultivars and the fungicide sensitivity of the pathogen.

A microscope system was developed for acquiring high resolution grain images of 30 rice varieties.The morphological, textural, and color traits of the rice grains were quantified using image processing.Trait discrepancies among varieties were observed and explained.Sparse-representation-based classifier was developed to identify the varieties of the grains.The classification achieved an accuracy of 89.1% and a standard deviation of 7.0%. Rice (Oryza sativa L.) is a major staple food worldwide, and is traded extensively. The objective of this study is to distinguish the rice grains of 30 varieties nondestructively using image processing and sparse-representation-based classification (SRC). SRC uses over-complete bases to capture the representative traits of rice grains. In the experiments, rice grain images were acquired by microscopy. The morphological, color, and textural traits of the grain body, sterile lemmas, and brush were quantified. An SRC classifier was subsequently developed to identify the varieties of the grains using the traits as the inputs. The proposed approach could discriminate rice grain varieties with an accuracy of 89.1%.

Rice bacterial leaf blight, caused by Xanthomonas oryzae pv. oryzae [(Ishiyama) Swings et al. 1990] (Xoo), is a major rice disease of the second crop season in Taiwan. A total of 88 Xoo strains collected from 10 major rice cultivating areas in Taiwan from 1986, 1997, 2000, 2004, and 2011 were characterized by repetitive-element PCR (REP-PCR) fingerprinting and virulence analyses. Among the five genetic clusters identified by the pJEL1/pJEL2 (IS1112-based) and REP1R-Dt/REP2-D [repetitive extragenic palindromic (REP)-based] primer sets, clusters A, C and D contained Xoo strains from geographically distant regions, which suggests a high frequency of Xoo dispersal in Taiwan. The 88 Xoo strains were evaluated by inoculations on IRBB near-isogenic lines and five Taiwan rice cultivars. A subset of 45 moderately or highly virulent strains were classified into 15 pathotypes by their compatible or incompatible reactions on IR24 and 12 IRBB near-isogenic lines, each containing a single resistance gene. Analysis of molecular haplotypes and pathotypes revealed a partial relationship. IRBB5, IRBB21 and IRBB4 were incompatible with 96%, 96% and 73% of the strains, so xa5, Xa21 and Xa4 can recognize most of the Xoo strains in Taiwan and elicit resistance. In contrast, IRBB3 (Xa3), IRBB8 (xa8), IRBB10 (Xa10), IRBB11 (Xa11), IRBB13 (xa13) and IRBB14 (Xa14) were susceptible to almost all of the 45 Xoo strains. Inoculation trials revealed significant differences in the susceptibility of five Taiwan cultivars to Xoo (from high to low susceptibility: Taichung Sen 10 > IR24, Taichung Native 1 > Taichung 192, Taikeng 9, Tainan 11). This study provides useful information for resistance breeding and the development of disease management strategies against bacterial blight disease of rice.

The rice disease bakanae, caused by Fusarium fujikuroi Nirenberg, has been present in Taiwan for over a century. To better understand the genetic diversity and structure of F. fujikuroi, a set of 16 polymorphic simple sequence repeat (SSR) markers were newly developed and used to analyze 637 F. fujikuroi isolates collected in 14 cities or counties around Taiwan from 1996 to 2013. On the basis of Bayesian clustering, the isolates were classified into four highly differentiated clusters: cluster B likely derived from the more widespread and genetically diversified clusters A or C, and cluster D was restricted to four cities or counties and may have been introduced from unknown sources genetically distinct from clusters A, B, and C. The coexistence of both mating types (MAT1-1:MAT1-2 = 1:1.88) and the highly diversified vegetative compatibility groups (VCG) (16 VCG among the 21 assessed isolates) suggest the likelihood of sexual reproduction in the field. However, the biased mating type ratios and linkage disequilibrium in the population suggest nonrandom mating between individuals. A significant pattern of isolation by distance was also detected, which implies a geographical restricted gene flow and low dissemination ability of F. fujikuroi. Evaluation of 24 representative isolates on eight rice varieties revealed differential levels of virulence, however no clear pattern of specific variety x isolate interaction was observed. Investigations of the differences in virulence and fungicide sensitivity between 8 early isolates (1998 and 2002) and 52 recent isolates (2012) indicate the evolution of increased resistance to the fungicide prochloraz in F. fujikuroi in Taiwan.

Rice blast, caused by Magnaporthe oryzae, is one of the most devastating diseases in rice worldwide. We aimed to develop an integrated approach for convenient collection, quantification and characterisation of M. oryzae spores (airborne inoculum) in the field. We developed an easy-to-use cyclone-based spore trap (the AirSampler) and a standard procedure for handling a small amount of airborne spores. Using a specific primer pair or a probe designed for the single-copy gene mif23, SYBR Green and TaqMan assays could quantify 10 and 4 copy numbers, respectively, of M. oryzae DNA. During 2012 and 2013, the AirSampler and SYBR Green quantitative real-time polymerase chain reaction were used to monitor temporal dynamics of M. oryzae spores in nursery fields of rice showing symptoms of blast disease. During four cropping seasons, the new techniques could detect M. oryzae spores before the appearance of rice blast symptoms. The amount of spores was low in the early season, then increased, with high fluctuations during the mid-season and decreased to low levels at the heading stage in the late season. To improve the handling and storage of spore samples, we tested the effect of different treatments on the preservation of spore DNA. DNA loss was reduced with samples protected from ultraviolet B radiation, suspended in CTAB buffer, kept at room temperature or 4°C and used for DNA extraction in 2 weeks. Finally, we demonstrated that the high resolution melting analysis could be used for rapid determination of A, D, A + D and C alleles of the avirulence gene pex31 (Avr-Pik/kp/km) in M. oryzae.

Hyperspectral imaging has proven to be an effective non-destructive method for assessing strawberry foliar anthracnose.The incubation stage, in which symptoms are not yet visible, can be distinguished.Several hyperspectral imaging analysis methods were investigated using 3 duplicate sets of experiments.Significant wavelengths for strawberry foliar anthracnose are 551, 706, 750 and 914nm. Hyperspectral imaging provides comprehensive spectral and spatial information about observed objects. This technology has been applied to many fields, such as geology, mining, surveillance and agriculture. Strawberry qualities have been examined using hyperspectral imaging in several studies. However, none of the previous literature presented a non-destructive method for diagnosing the infection stages of anthracnose, a devastating disease for strawberries. This study examined strawberry foliar anthracnose using three different hyperspectral imaging analyzing methods: spectral angle mapper (SAM), stepwise discriminant analysis (SDA) and self-developed correlation measure (CM). Three different infection stages, including healthy, incubation and symptomatic stages, were investigated using these methods. The incubation stage is a stage at which the symptoms are still not yet visible. The three infection stage classification results were promising, with a classification accuracy of approximately 80%. For two infection stage classification (healthy and symptomatic stages), an average accuracy of high 80% was attained. In fact, an average accuracy of 93% was achieved by SDA for two-stage classification. This study not only proves the feasibility of hyperspectral imaging for diagnosing strawberry foliar anthracnose infection, but also identifies a smaller set of significant wavelengths at which similar classification performance was accomplished. For significant wavelength selection, partial least squares (PLS) regression is an standard wavelength selection method and it was applied to be compared with SDA and CM. Wavelengths of 551, 706, 750 and 914nm formed the multispectral imaging observing bands that showed an accuracy of 80% when classifying the three infection stages. Therefore, using either hyperspectral or multispectral imaging to detect anthracnose infected foliar areas is more practical and efficient than classic destructive methods. In particular, early detection (the incubation stage), something that cannot be seen via naked eyes, reaches 80% classification accuracy with both SDA and CM. Strawberry farmers could profit greatly from this technology.

Rice seedlings infected by Fusarium fujikuroi show complex diseased phenotypes.The morphological and color traits of 3-week-old seedlings are examined.SVM classifiers are developed to screen diseased seedlings.Genetic algorithm is implemented for selecting significant attributes.The classification accuracy is 87.9% and the positive predictive value is 91.8%. Bakanae disease, or "foolish seedling", is a seed-borne disease of rice (Oryza sativa L.). Infected plants can yield empty panicles or perish, resulting in a loss of grain yield. The disease occurs most frequently when contaminated seeds are used. Once the seeds are contaminated, the pathogen Fusarium fujikuroi spreads in the field. Therefore, infected plants must be screened at early developmental stages. This work proposes an approach to nondestructively distinguish infected and healthy seedlings at the age of 3weeks using machine vision. Seeds of the rice cultivars Tainan 11 and Toyonishiki were inoculated with a conidial suspension of F. fujikuroi. The seedling were cultivated in an incubator for 3weeks. The images of infected and control seedlings were acquired using flatbed scanners to quantify their morphological and color traits. Support vector machine (SVM) classifiers were developed for distinguishing the infected and healthy seedlings. A genetic algorithm was used for selecting essential traits and optimal model parameters for the SVM classifiers. The proposed approach distinguished infected and healthy seedlings with an accuracy of 87.9% and a positive predictive value of 91.8%.