Anthracnose of <Emphasis Type="Italic">Polygonatum falcatum</Emphasis> caused by <Emphasis Type="Italic">Colletotrichum dematium</Emphasis>

Severe spotting, blight and drop of leaves caused by Colletotrichum dematium were found on potted plants of Polygonatum falcatum, a liliaceous ornamental, in open fields in Kagawa Prefecture, Japan, in May 2001. This new disease was named anthracnose of P. falcatum. Keisuke Tomioka, Jouji Moriwaki, Toyozo Sato contributed equally to this work. The fungal isolate and its nucleotide sequence data obtained in this study were deposited in Genebank, National Institute of Agrobiological Sciences and the DDBJ/EMBL/GenBank databases under accessions MAFF239500 and AB334523, respectively.     

Anthracnose of salt-wort (<Emphasis Type="Italic">Salsola komarovii</Emphasis>) caused by <Emphasis Type="Italic">Colletotrichum truncatum</Emphasis>

In July 2006, black rot was observed on the leaves of 4-leaf-stage seedlings of salt-wort (Salsola komarovii) in Yamagata Prefecture, Japan. We isolated two single-conidial isolates from the diseased leaves. Although colony appearance of the isolates was different from that of each other, both isolates were identified as Colletotrichum truncatum by morphology and molecular similarity. After inoculation of healthy salt-wort plants with the isolates, the isolates were reisolated from symptomatic plants. We thus propose a new disease, anthracnose of salt-wort.     

Anthracnose of <Emphasis Type="Italic">Enkianthus campanulatus</Emphasis> and <Emphasis Type="Italic">Rhynchosia acuminatifolia</Emphasis> caused by <Emphasis Type="Italic">Colletotrichum gloeosporioides</Emphasis> (new occurrence)

In 2003–2004, anthracnoses of Enkianthus campanulatus and Rhynchosia acuminatifolia were found for the first time in Kanagawa Prefecture and Tokyo in Japan. These pathogens were identified as Colletotrichum gloeosporioides based on their pathogenicity, morphology and ribosomal DNA spacer sequences. Results were presented at the annual meeting of The Phytopathological Society of Japan in 2004.     

First report of celery stunt anthracnose caused by <Emphasis Type="Italic">Colletotrichum simmondsii</Emphasis> in Japan

A new disease was found in Japan, on celery (Apium graveolens var. dulce) having severe chlorotic leaf spot, stunt, and dwarf with leaf curl. A spore suspension from the fungus isolated from affected plants induced identical symptoms 14 days after plants were sprayed. Identification and molecular characterization showed that the causal agent is Colletotrichum simmondsii. This report is the first of stunt anthracnose on celery caused by C. simmondsii. We propose the name “stunt anthracnose” for the new disease. Colletotrichum acutatum sensu lato, as reference pathogen of celery anthracnose, should be changed to C. fioriniae based on morphological and molecular characteristics.     

Anthracnose of snapdragon caused by <Emphasis Type="Italic">Colletotrichum destructivum</Emphasis>

Severe spotting and blight of leaves caused by Colletotrichum destructivum were found on snapdragon (Antirrhinum majus L.), a scrophulariaceous ornamental, in open fields in Shizuoka Prefecture, Japan, from June through September 2004. The fungus is added to the group of the pathogens causing anthracnose of snapdragon.     

First report of blueberry red ringspot disease caused by <Emphasis Type="Italic">Blueberry</Emphasis><Emphasis Type="Italic">red</Emphasis><Emphasis Type="Italic">ringspot</Emphasis><Emphasis Type="Italic">virus</Emphasis> in Japan

Virus-like symptoms—red ringspots on stems and leaves, circular blotches or pale spots on fruit—were found on commercial highbush blueberry (Vaccinium corymbosum) cultivars Blueray, Weymouth, Duke and Sierra in Japan. In PCR testing, single DNA fragments were amplified from total nucleic acid samples of the diseased blueberry bushes using primers specific to Blueberry red ringspot virus (BRRV). Sequencing analysis of the amplified products revealed 95.7–97.7% nucleotide sequence identity with the BRRV genome. This paper is the first report of blueberry red ringspot disease caused by BRRV in Japan. The nucleotide sequence data reported in this paper are available in the GenBank/EMBL/DDBJ database as accessions AB469884 to AB469893 for BRRV isolates from Japan.     

Anthracnose of three-leaf akebia (<Emphasis Type="Italic">Akebia trifoliata</Emphasis> Koidzumi) caused by <Emphasis Type="Italic">Colletotrichum acutatum</Emphasis>

In October 2001, anthracnose caused by Colletotrichum acutatum Simmonds ex Simmonds was found on three-leaf akebia (Akebia trifoliata) in Saitama, Japan. This is the first report of anthracnose on three-leaf akebia caused by C. acutatum.     

<Emphasis Type="Italic">Colletotrichum destructivum from</Emphasis> cowpea infecting <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis> and its identity to <Emphasis Type="Italic">C. higginsianum</Emphasis>

Colletotrichum isolates isolated from cowpea in the Hangzhou area of China were identified as C. destructivum based on morphological characteristics, pathogenicity tests, sequence analysis of the internal transcribed spacer (ITS)1, 5.8S RNA gene and ITS2 regions of ribosomal DNA and the infection process. The ability of the C. destructivum isolates to infect Arabidopsis thaliana was investigated under laboratory conditions and showed a two-phase hemibiotrophic infection process. In addition, the sequences of the rDNA ITS region of C. destructivum isolates from cowpea were identical with 100% similarity to that of isolates of C. higginsianum originating from cruciferous plants. This article presents new evidence in support of C. higginsianum as a synonym of C. destructivum.     

Fusarium rot of hyacinth caused by <Emphasis Type="Italic">Gibberella zeae</Emphasis> (anamorph: <Emphasis Type="Italic">Fusarium</Emphasis><Emphasis Type="Italic"> graminearum</Emphasis>)

Severe rot of leaves, peduncles and flowers caused by Gibberella zeae (anamorph: Fusarium graminearum) was found on potted plants of hyacinth (Hyacinthus orientalis), a liliaceous ornamental, in greenhouses in Kagawa Prefecture, Japan, in January 2001. This disease was named “Fusarium rot of hyacinth” as a new disease because only the anamorph, F. graminearum, was identified on the diseased host plant. The authors contributed equally to this work. The fungal isolate and its nucleotide sequence data obtained in this study were deposited in the Genebank, National Institute of Agrobiological Sciences and the DDBJ/EMBL/GenBank databases under the accession numbers MAFF239499 and AB366161, respectively.     

Cobweb disease on <Emphasis Type="Italic">Agaricus bisporus</Emphasis> caused by <Emphasis Type="Italic">Cladobotryum mycophilum</Emphasis> in Korea

This report is the first of cobweb disease on Agaricus bisporus in Korea. Cobweb on both fruit bodies and casing soils were observed on several mushroom farms in Gyeongbuk Province, Korea. Classical and molecular characterization indicated that the causal agent is Cladobotryum mycophilum. The isolated fungus was used to inoculate fruiting bodies of A. bisporus and caused the same symptoms.     

Pathogenicity and characterization of <Emphasis Type="Italic">Colletotrichum lentis</Emphasis>: A causal agent of anthracnose in common vetch (<Emphasis Type="Italic">Vicia sativa</Emphasis>)

A series of studies were carried out on Colletotrichum lentis which had been been identified in 2015 based largely on the distinctive shape of conidia and ITS sequences, and which has been causing severe anthracnose disease symptoms on common vetch plants (Vicia sativa) in Gansu Province in the northwest region of China. A key focus of the present studies was to determine how vetch crops become infected. The addition of residues from harvested common vetch crops to land being prepared as a seedbed was shown to result in the highest levels of disease severity indicating that this management practice was the most likely way for crops to become severely infected. Seed transmission was unlikely to be the cause of severe outbreaks as less than 5% of seeds harvested from severely infected plants carried C. lentis. To verify that the species causing the severe outbreaks of anthracnose disease of vetch crops was C. lentis, sequence analysis of the ITS, TUB2, ACT, HIS3 and GAPDH genes was conducted. C. lentis isolates from common vetch and lentil (Lens culinaris) formed a distinctive group among Colletotrichum species, including those species that infect other forage and field crops. The unique shape of conidia of C. lentis, straight with only one end curved, was confirmed as being reliable for rapid identification of disease outbreaks caused by this damaging fungal pathogen.     

<Emphasis Type="Italic">Agrobacterium tumefaciens</Emphasis>-mediated transformation for random insertional mutagenesis in <Emphasis Type="Italic">Colletotrichum lagenarium</Emphasis>

Random insertional mutagenesis using a marker DNA fragment is an effective method for identifying fungal genes relevant to morphogenesis, metabolism, and so on. Agrobacterium tumefaciens-mediated transformation (AtMT) has long been used as a tool for the genetic modification of a wide range of plant species. Recent study has indicated that A. tumefaciens could transfer T-DNA not only to plant cells but also to fungal cells. In this study, AtMT was applied to Colletotrichum lagenarium for random insertional mutagenesis. We constructed a binary vector pBIG2RHPH2 carrying a hygromycin-resistant gene cassette between the right and left borders of T-DNA. Optimal co-cultivation of C. lagenarium wild-type 104-T with pBIG2RHPH2-introduced A. tumefaciens C58C1 led to the production of 150–300 hygromycin-resistant transformants per 10⁶ conidia. Southern blot analysis revealed that T-DNA was mainly integrated at a single site in the genome and at different sites in transformants. The T-DNA inserts showed small truncations of either end, but the hygromycin-resistant gene cassette inside the T-DNA was generally intact. The mode of T-DNA insertion described above resulted in highly efficient gene recovery from the transformants by thermal asymmetrical interlaced-polymerase chain reaction. The fungal genomic DNA segments flanking T-DNA were identified from five of eight mutants that had defective melanin biosynthesis. The sequence from one of the segments was identical to that of the melanin biosynthesis gene PKS1 of C. lagenarium, which we previously characterized. These results strongly support our notion that AtMT is a possible tool for tagging genes relevant to pathogenicity in the plant pathogenic fungus C. lagenarium.     

White rust of <Emphasis Type="Italic">Ipomoea</Emphasis> caused by <Emphasis Type="Italic">Albugo ipomoeae-panduratae</Emphasis> and <Emphasis Type="Italic">A. ipomoeae-hardwickii</Emphasis> and their host specificity

In some areas of Japan, yellow spots with white pustules on leaves, stems, petioles, peduncles and calyces were found on Ipomoea nil, I. triloba, I. lacunosa and I. hederacea var. integriuscula. We demonstrated that the diseases on I. nil, I. triloba and I. lacunosa were caused by host-specific strains of Albugo ipomoeae-panduratae and defined three forma speciales of the fungus, respectively, for the three Ipomoea species: “f. sp. nile”, “f. sp. trilobae” and “f. sp. lacunosae”. Because the diseases were new to Japan, we coined the Japanese name “shirosabi-byo”, which means white rust. We also showed that the disease on I. hederacea var. integriuscula was caused by A. ipomoeae-hardwickii. We named this new disease “white rust (shirosabi-byo in Japanese)”.     

Pythium rot of figmarigold (<Emphasis Type="Italic">Lampranthus spectabile</Emphasis>) caused by <Emphasis Type="Italic">Pythium aphanidermatum</Emphasis>

A new leaf rot disease was found on the leaves of figmarigold (Lampranthus spectabile). The causal organism, identified as Pythium aphanidermatum was found to cause the same symptoms after artificial inoculation and was then reisolated from the inoculated plants. We propose to name the disease Pythium rot of figmarigold.     

First report of leaf spot caused by <Emphasis Type="Italic">Alternaria alternata</Emphasis> on <Emphasis Type="Italic">Drimia maritima</Emphasis>

Drimia maritima (squill) is a historically important medicinal plant. During the spring of 2016, small, yellow leaf spots, which became brown and finally necrotic, were observed on squill plants in Kohgiluyeh and Boyer-Ahmad Provinces in Iran. A fungus was consistently isolated from infected leaves and identified as Alternaria alternata based on morphological and phylogenetic analyses. Pathogenicity tests confirmed A. alternata to be the causal agent of the newly observed leaf spot disease. This is the first report of leaf spot on D. maritima caused by A. alternata in the world.     

Characterization of <Emphasis Type="Italic">Inago1</Emphasis> and <Emphasis Type="Italic">Inago2</Emphasis> retrotransposons in <Emphasis Type="Italic">Magnaporthe oryzae</Emphasis>

From the genome of a Japanese field isolate of the rice blast fungus, Magnaporthe oryzae, we newly identified Inago1 and Inago2 LTR retrotransposons. Both elements were found to be Ty3/gypsy-like elements whose copies were dispersed within the genome of Magnaporthe spp. isolates infecting rice and other monocot plants. Southern hybridization patterns of nine re-isolates derived from conidia of the strain Ina168 produced after a methyl viologen treatment were not changed, indicating that the insertion pattern of Inago elements is relatively stable.     

Epidemiology of apple bitter rot caused by <Emphasis Type="Italic">Colletotrichum acutatum sensu lato</Emphasis>

Apple bitter rot caused by Colletotrichum acutatum sensu lato results in fruit decay before and after harvest. We investigated the epidemiology of the disease in terms of conidial formation and dispersal as well as the change in susceptibility of fruits in Iwate, Japan. Conidia of C. acutatum were detected in rainwater collected from inside the tree canopy from May to August with peaks in production in mid-May to early June and mid- to late July. The first peak corresponded to the most conidia being produced on fruit scars, but the second peak was due to conidiation on mummified fruitlets and peduncles collected in July. Inoculation experiments revealed that fruits were susceptible to the pathogen between 20 and 90 days after petal fall and that immature fruits infected as early as 20 days after petal fall frequently developed lesions on the lower fruit half as growth progressed. These results suggest that C. acutatum sporulates on infested fruit scars to infect immature fruits, resulting in bitter rot and that the fungus also colonizes mummified fruitlets and peduncles, contributing to survival of the pathogen on fruit scars. Thus, infested fruit scars represent the primary source of inoculum.     

Pathogenicity of <Emphasis Type="Italic">Beauveria bassiana</Emphasis> and <Emphasis Type="Italic">Metarhizium anisopliae</Emphasis> against <Emphasis Type="Italic">Galleria mellonella</Emphasis>

The greater wax moth Galleria mellonella L. (Lepidoptera: Pyralidae) is occasionally found in beehives and is a major pest of stored wax. Entomopathogenic fungi have recently received attention as possible biocontrol elements for certain insect pests. In this study, 90 isolates of Beauveria bassiana and 15 isolates of Metarhizium anisopliae were screened for proteases and lipases production. The results showed significant variations in the enzymatic action between the isolates. In the bioassay, the selected isolates evinced high virulence against the 4th instar of the G. mellonella larvae. The isolates BbaAUMC3076, BbaAUMC3263 and ManAUMC3085 realized 100% mortality at concentrations of 5.5 × 10⁶ conidia ml⁻¹, 5.86 × 10⁵ conidia ml⁻¹, and 4.8 × 10⁶ conidia ml⁻¹, respectively. Strong enzymatic activities in vitro did not necessarily indicate high virulence against the tested insect pest. The cuticle of the infected larvae became dark and black-spotted, indicating direct attack of fungus on the defense system of the insects. The LC₅₀ values were 1.43 × 10³, 1.04 × 10⁵ and 5.06 × 10⁴ for Bba3263AUMC, Bba3076AUMC and Man3085AUMC, respectively, and their slopes were determined by computerized probit analysis program as 0.738 ± 0.008, 0.635 ± 0.007 and 1.120 ± 0.024, respectively.