Journal of General Plant Pathology最新文献

Infection of Zizania latifolia by the basidiomycete Ustilago esculenta enlarges the stem tissue to produce edible galls called "makomotake" in Japan. The development of stem galls in Z. latifolia may be influenced by the colonization level of U. esculenta at the seedling stage. To evaluate this possibility, it is necessary to establish a method to investigate the relationship between fungal levels in seedlings and the production of makomotake from the same seedlings in a non-destructive manner. To achieve this goal, in this study, we attempted to develop a real-time loop-mediated isothermal amplification (LAMP) method with toothpick sampling for U. esculenta detection. An U. esculenta gene-specific LAMP primer set designed for real-time LAMP showed sufficient detection sensitivity for fungal genomic DNA, which is only about 10 times lower than that of quantitative PCR. Real-time LAMP reactions following toothpick sampling detected U. esculenta efficiently in the culm part of Z. latifolia seedlings that eventually produced makomotake, but did not detect in those that failed to develop stem galls. Our data show that the combination of real-time LAMP and toothpick sampling is a simple and useful method for estimating U. esculenta levels in Z. latifolia seedlings and for studying the subsequent development of stem galls in the same individuals.

Berkeleyomyces rouxiae, a soil-borne fungal pathogen, causes black root rot on various plant species including carrot, okra, tobacco, lettuce, and other important crops. Here we analyzed the entire genome sequence of a Japanese isolate of B. rouxiae and discovered 25 microsatellite sequences. These sequences were amplified from the genomic DNA of several Japanese isolates by PCR and sequenced, revealing that the microsatellite sequences were polymorphic among isolates. These markers are useful for phylogenetic analysis, epidemiological population analysis, identification of specific isolates, and other genetic investigation of B. rouxiae.

Huanglongbing (HLB) is the most devastating disease of citrus. Although no citrus species are known to be immune to HLB, some (e.g., Persian lime [Citrus latifolia]) have a high level of tolerance. Differentially expressed genes in the WRKY transcription factor (TF) family have been identified in another tolerant citrus, suggesting an association with such tolerance. Therefore, here we searched for the conserved WRKY domain of WRKY family members in the transcriptome of C. latifolia infected with HLB, characterized the identified ClWRKY genes, and determined which were differentially expressed in leaves with HLB symptoms; 177 transcripts with the WRKY domain were identified, and 32 ClWRKY genes were characterized for conserved motifs, gene ontology, and coexpression networks among the 32 ClWRKY genes. These genes were thus shown to be involved in response to external stimuli and biotic and abiotic stresses. Four differentially expressed ClWRKY genes (ClWRKY20, 23, 47, and 65) were identified in the transcriptome and validated by qRT-PCR. The possible roles of these genes in the tolerance of C. latifolia to HLB are discussed based on the function of homologs of these genes in other plant species. These WRKY genes could contribute to the genetic improvement of susceptible citrus to manage HLB.

In the inducible chlorosis model tobacco, i-hpHSP90C, the silencing of HSP90C activated both salicylic acid (SA)- and cell death-related gene expression and sporadic cell death, resulting in severe chlorosis. In this model plant, we found a transient SA accumulation to a significantly high level at 8 h after induction of HSP90C silencing and consistent upregulation of CBP60-type transcription factors and some SA biosynthetic genes. Exogenous treatment of the model plant with SA alone did not induce chlorosis. The introgression of a gene encoding SA-degrading enzyme, nahG^A430V, into tobacco plants with functional N′ tobamovirus resistance gene partially compromised their resistance to tomato mosaic virus but without a clear reduction in SA levels. Expression of nahG^A430V stochastically alleviated chlorosis and, subsequently, sporadic cell death upon induction of HSP90C silencing. We applied tenoxicam, a potent inhibitor of the NPR1-dependent SA signaling pathway in Arabidopsis, and found that it alleviated chlorosis in i-hpHSP90C, which accompanied a reduced expression of a CBP60-type transcription factor. However, the expression of PR1a, a well-characterized SA signal marker, was not suppressed by tenoxicam in the i-hpHSP90 plants with alleviated chlorosis. The findings collectively suggest that the plant immunity-related response, including SA production, could have a role in increasing the severity of chlorosis, although the underlying mechanisms remain to be elucidated.

It is still difficult to manipulate the nucleotide-binding site-leucine-rich repeat (NB-LRR) class disease resistance genes because of their large multigenic family. Here, we report the successful application of CRISPR/Cas9 to resurrect a functional allele from a susceptible allele of the N′ tobamovirus resistance gene. The susceptible alleles of N′ from some Nicotiana tabacum cultivars (Nt-n′) have a partial duplication of the N′ coding sequence upstream of a complete coding sequence of the N′ gene, which likely abolished the N′-mediated resistance. We first established a transgenic tobacco line expressing Cas9 under the control of a chemically inducible promoter. The plant line was retransformed with a construct expressing a guide RNA targeting the sequences common to the duplicated partial sequence in the upstream and the complete sequence in the downstream. The T₀ transformants had different ratios of the sequences devoid of the duplicated partial sequence. Sequencing proved that some of them had sequences identical to that of the functional N′ gene, suggesting the successful resurrection of the functional N′ gene. The resurrected allele, N′^-R, was inherited by a few T₁ progenies and subsequent generations with the least mutation at the target site under Cas9-uninduced conditions. The plants homozygous for N′^-R showed resistance to a tobamovirus, indicating that the resurrected N′^-R allele is functional.

Theobroma cacao is primarily cultivated for its seeds. In the Philippines, where cacao yields are decreased by pod rots and vascular streak dieback (VSD)-like symptoms, the fungus Lasiodiplodia theobromae (Botryospaheriaceae) is the only reported species of Lasiodiplodia reported to cause disease in cacao. Here to identify and determine the pathogenicity of Lasiodiplodia isolates from cacao trees, we isolated Lasiodiplodia from from a leaf with blight (isolate PH22-080), stem with vascular streak (PH22-060), pod rot (PH22-120), an asymptomatic stem (PH22-014), and leaf litter (PH22-007). All strains were pathogenic on wounded leaves and stems and infected pods even without wounds. On the basis of molecular analysis of combined sequence data for ITS, rpb2, tef1-α and tub2 and morphological characteristics, the strains were identified as L. pseudotheobromae (PH22-080), L. theobromae (PH22-120), and Lasiodiplodia sp. (PH22-060), which were confirmed to be pathogenic on cacao leaves, stems, and pods. L. theobromae (PH22-007) and L. hormozganensis (PH22-014) showed potential to cause disease in cacao but further investigation is needed.

Anthracnose caused by Colletotrichum spp. is a serious threat to many crops, and C. siamense has become increasingly prevalent worldwide. However, the mycelial growth and virulence of many of these species, especially below 26 °C, has largely remained unknown. Here, mycelial growth of five representative Colletotrichum strains (three C. siamense [Cs], one C. fructicola [Cf] and one C. gloeosporioides s.s. [Cg]) from diseased strawberry was measured at 10, 14, 18, 22 and 26 °C. Their virulence on Fragaria vesca and highly susceptible F. × ananassa cv. Benihoppe was tested at 14 °C and 22 °C. The strains did not differ much in mycelial growth between 10 and 26 °C, although growth of the Cs and Cg strains was somewhat faster than that of Cf. However, the strains varied greatly in their virulence on the two hosts. Strain Cs GQHZJ19, which grew fastest at 22 °C, was also the most virulent on both hosts at 14 °C and 22 °C. But growth rate of Cs was only positively correlated with virulence on susceptible cv. Benihoppe after 3 days at 22 °C. Based on the virulence analysis of the five Colletotrichum strains on hosts that vary in susceptibility at distinct temperatures, host genotype might play a decisive role in disease severity at an early stage, while temperature might influence the host–Colletotrichum spp. interaction at a later stage. More Colletotrichum strains (especially C. siamense) need to be tested to determine the mycelial growth and virulence in a wider temperature range and thus we can effectively manage disease.

Wheat stem rust caused by the fungus Puccinia graminis f. sp. tritici is one of the most destructive diseases. Here, we collected 43 stem rust isolates from six provinces of Iran during 2020–2022 and assessed variability in their virulence on 20 North American differential lines carrying one of the stem rust resistance genes. More than 90% of isolates were virulent on lines with gene Sr5, Sr9d, Sr9g, Sr30, Sr36, Sr38, SrTmp and SrMcN and the least virulent on those with Sr31 (16.2%) and Sr24 (0%). The majority of isolates belonged to races TKTTF, TTTTF, and TTTTK. Races TTTTK, TKTTK and PTTTK are new in Iran.

Anthracnose caused by Colletotrichum species is a serious disease of loquat (Eriobotrya japonica), but the species of Colletotrichum that are associated with the disease in Japan have not been adequately surveyed. In this study, 46 isolates were isolated from diseased leaves and fruits of loquat in Kagawa and Tokushima prefectures in 2017 and 2018. On the basis of morphological features and molecular approaches, seven Colletotrichum species were identified: C. fioriniae, C. nymphaeae, C. karsti, C. aenigma, C. fructicola, C. gloeosporioides s. s., and C. siamense. The most prevalent species was C. karsti (43.5% of total species), followed by C. fioriniae (21.7%) and C. gloeosporioides s. s. (13.0%). All identified species were confirmed to cause loquat anthracnose by fulfilling Koch’s postulates. The loquat isolates of C. fioriniae, C. fructicola, and C. siamense also caused symptoms on all tested plants of kiwifruit, satsuma mandarin, persimmon, and peach. This report is the first on C. karsti and C. aenigma associated with loquat anthracnose and the first record of C. fructicola in Japan.

Rice false smut has become a serious threat to rice production. Durable chlamydospores of the fungus are the primary inoculum in paddy soil. Mixing converter slag into paddy soil can decrease disease incidence, but the mechanism is unknown. Here we first confirmed the suppressive effect of converter slag mixed into a paddy field: 3t/ha suppressed disease incidence by 75.6%. We then tested a converter slag solution (CSS), neutralized CSS (NCSS), and converter-slag-amended soil extract (CSSE), all at 3t slag/ha. In the CSS test, chlamydospores rapidly became transparent, then gradually collapsed, but NCSS slowed chlamydospore collapse. More germ tubes were abnormal in the CSS treatment than in the NCSS. The percentages of transparent, collapsed, and normally germinating chlamydospores in the CSSE treatment were intermediate to those obtained with CSS and NCSS. Hyphal invasion into rice roots in CSSE was also suppressed compared to that in control. These results suggest chlamydospores gradually collapse when exposed to cations and the pH is neutralized by soil, thus reducing hyphal production and infection of rice roots, contributing to disease suppression in fields.