First report of Seimatosporium vitifusiforme causing trunk disease in Chilean grapevines (Vitis vinifera).

Grapevine is one of the most important fruit crops in Chile and trunk diseases reduce the productivity, quality, and longevity of the vineyards. A survey was conducted in ancient (> 50 years) vineyards of Cauquenes (35°57´14´´S 72°17´07´´W) and Itata valleys (36°38´13´´S 72°30´57´´W), located in the central area of Chile, during 2019. Trunks and cordons showing dieback and dark brown to black wood discoloration were collected from 50 to 200-year-old plants of six cultivars: País, Moscatel, Torontel Amarilla, Carignan, Aliatica and Aligote. The bark was removed and 0.5-cm sections were cut from the edges of necrotic wood lesions. Subsequently, pieces were surface disinfected using 10% v/v sodium hypochlorite bleach (4.9% chlorine), plated on acidified quarter-strength potato dextrose agar (APDA) (25% PDA, acidified with 0.1% v/v 85% lactic acid) and incubated at 25°C, for 14 to 28 days. Hyphal tips were excised and transferred to PDA to obtain pure cultures. Along with the conidiomata and conidia produced, growth rate, color and shape of the colonies on PDA, after 7 and 14 days of incubation at 25°C (n=17), were recorded. DNA was extracted from pure cultures of three isolates on PDA: HMV3, HMV64 and HMV81. The internal transcribed spacer region and partial ß-tubulin genes were amplified, using ITS1/ITS4 (White et al. 1990) and bt2A/bt2B (Glass & Donaldson 1995) primers, respectively. Sequences were subjected to NCBI BLAST search and compared to the published sequences. Isolated colonies were whitish to light-brown, cottony with a smooth margin (n=37). Their mycelium grew 1.9 cm after 7-days and 3.2 cm after 14-days of incubation on PDA, at 25°C. Colonies produced black globose pycnidia and curved, slightly-pigmentated, three-septated conidia 22.3-(29.8)-32.2 x 3.9-(4.8)-5.3 µm (n=30), with apical and basal flexuous appendages 4.3-(12.7)-21.5 µm (n=20). When compared to type sequences of Seimatosporium vitifusiforme (Lawrence et al. 2018), ITS and ßtub sequences identity of these isolates were 99 to 100% identical. To produce uniform healthy plants for pathogenicity tests, Petit Syrah canes (1-year old) were rooted in tap water amended with 500 ppm of indole-butyric acid, for 30 days. Plants were inoculated with 0.5-cm diameter mycelial plugs of actively growing colonies of the isolates HMV3, HMV64 and HMV81 (GenBank accessions no. MW026664, MW048518; MW026665, MW048519, and MW026666, MW048520, respectively). Sterile agar plugs were used for controls. Five plants per pathogen isolate were incubated at 25°C, in a humid chamber, for 25 days, and seven additional plants per isolate were incubated in aerated tap water, for 55 days. After the incubation period, the bark was removed and the lesions were measured. Dark necrotic lesions identical to the original observations were reproduced, both in the high humidity chamber (6% length) and water (10% length). There were no differences in lesion length among the isolates (P < 0.05). Control vines remained asymptomatic. To fulfill Koch´s postulates, isolations were made from symptomatic vines and compared to the ones used for inoculation, and found to be identical. Seimatosporium vitifusiforme was previously reported as a pathogen of Vitis vinifera in California, USA (Lawrence et al. 2018). Consequently, this is the second report of this fungus as a grapevine pathogen and the first one affecting Latin-American grapevines.

Sporocadaceae, a family of coelomycetous fungi with appendage-bearing conidia.

Species of Sporocadaceae are endophytic, plant pathogenic or saprobic, and associated with a wide range of host plants. Recent molecular studies that have attempted to address familial and generic boundaries of fungi belonging to Sporocadaceae were based on a limited number of samples and DNA loci. The taxonomy of this group of fungi is therefore still not fully resolved. The aim of the present study is to provide a natural classification for the Sporocadaceae based on multi-locus phylogenetic analyses, using LSU, ITS, tef-1α, tub2 and rpb2 loci, in combination with morphological data. A total of 30 well-supported monophyletic clades in Sporocadaceae are recognised, representing 23 known and seven new genera. Typifications are proposed for the type species of five genera (Diploceras, Discosia, Monochaetia, Sporocadus and Truncatella) to stabilise the application of these names. Furthermore, Neotruncatella and Dyrithiopsis are synonymised under Hymenopleella, and the generic circumscriptions of Diploceras, Disaeta, Hymenopleella, Monochaetia, Morinia, Pseudopestalotiopsis, Sarcostroma, Seimatosporium, Synnemapestaloides and Truncatella are emended. A total of 51 new species, one nomina nova and 15 combinations are introduced.

Microsphaerol and seimatorone: two new compounds isolated from the endophytic fungi, Microsphaeropsis sp. and Seimatosporium sp.

A new polychlorinated triphenyl diether named microsphaerol (1), has been isolated from the endophtic fungus Microsphaeropsis sp. An intensive phytochemical investigation of the endophytic fungus Seimatosporium sp., led to the isolation of a new naphthalene derivative named seimatorone (2) and eight known compounds, i.e., 1-(2,6-dihydroxyphenyl)-3-hydroxybutan-1-one (3), 1-(2,6-dihydroxyphenyl)butan-1-one (4), 1-(2-hydroxy-6-methoxyphenyl)butan-1-one (5), 5-hydroxy-2-methyl-4H-chromen-4-one (6), 2,3-dihydro-5-hydroxy-2-methyl-4H-chromen-4-one (7), 8-methoxynaphthalen-1-ol (8), nodulisporins A and B (9 and 10, resp.), and daldinol (11). The structures of 1 and 2 were elucidated by detailed spectroscopic analysis including (1) H- and (13) C-NMR, COSY, HMQC, HMBC, and HR-EI-MS, while the structures of the known compounds were deduced from comparison of their spectral data with those in the literature. Preliminary studies revealed that microsphaerol (1) showed good antibacterial activities against B. Megaterium and E. coli, and good antilagal and antifungal activities against C. fusca, M. violaceum, respectively. On the other hand, seimatorone (2) exhibited moderate antibacterial, antialgal, and antifungal activities.

Seimisochromenes A and B: two new dihydroisochromenes from the endophytic fungus, Seimatosporium sp.

Two new dihydroisochromenes, named seimisochromenes A and B (1 and 2), were isolated from an endophytic fungus, Seimatosporium sp. The structures of seimisochromenes A and B have been determined from 1D ((1)H and (13)C NMR spectra) and 2D (COSY, HMQC, HMBC, and NOESY) NMR experiments.

Wild Rosa Endophyte M7SB41-Mediated Host Plant's Powdery Mildew Resistance.

Our previous studies indicated that endophyte M7SB41 (Seimatosporium sp.) can significantly enhance host plants powdery mildew (PM) resistance. To recover the mechanisms, differentially expressed genes (DEGs) were compared between E+ (endophte-inoculated) and E- (endophyte-free) plants by transcriptomics. A total of 4094, 1200 and 2319 DEGs between E+ and E- were identified at 0, 24, and 72 h after plants had been infected with PM pathogen Golovinomyces cichoracearum, respectively. Gene expression pattern analysis displayed a considerable difference and temporality in response to PM stress between the two groups. Transcriptional profiling analysis revealed that M7SB41 induced plant resistance to PM through Ca2+ signaling, salicylic acid (SA) signaling, and the phenylpropanoid biosynthesis pathway. In particular, we investigated the role and the timing of the SA and jasmonic acid (JA)-regulated defensive pathways. Both transcriptomes and pot experiments showed that SA-signaling may play a prominent role in PM resistance conferred by M7SB41. Additionally, the colonization of M7SB41 could effectively increase the activities and the expression of defense-related enzymes under PM pathogen stress. Meanwhile, our study revealed reliable candidate genes from TGA (TGACG motif-binding factor), WRKY, and pathogenesis-related genes related to M7SB41-mediate resistance. These findings offer a novel insight into the mechanisms of endophytes in activating plant defense responses.

Grapevine Trunk Diseases in Greece: Disease Incidence and Fungi Involved in Discrete Geographical Zones and Varieties.

A three-year survey was conducted to estimate the incidence of grapevine trunk diseases (GTDs) in Greece and identify fungi associated with the disease complex. In total, 310 vineyards in different geographical regions in northern, central, and southern Greece were surveyed, and 533 fungal strains were isolated from diseased vines. Morphological, physiological and molecular (5.8S rRNA gene-ITS sequencing) analyses revealed that isolates belonged to 35 distinct fungal genera, including well-known (e.g., Botryosphaeria sp., Diaporthe spp., Eutypa sp., Diplodia sp., Fomitiporia sp., Phaeoacremonium spp., Phaeomoniella sp.) and lesser-known (e.g., Neosetophoma sp., Seimatosporium sp., Didymosphaeria sp., Kalmusia sp.) grapevine wood inhabitants. The GTDs-inducing population structure differed significantly among the discrete geographical zones. Phaeomoniella chlamydospora (26.62%, n = 70), Diaporthe spp. (18.25%, n = 48) and F. mediterranea (10.27%, n = 27) were the most prevalent in Heraklion, whereas D. seriata, Alternaria spp., P. chlamydospora and Fusarium spp. were predominant in Nemea (central Greece). In Amyntaio and Kavala (northern Greece), D. seriata was the most frequently isolated species (>50% frequency). Multi-genes (rDNA-ITS, LSU, tef1-α, tub2, act) sequencing of selected isolates, followed by pathogenicity tests, revealed that Neosetophoma italica, Seimatosporium vitis, Didymosphaeria variabile and Kalmusia variispora caused wood infection, with the former being the most virulent. To the best of our knowledge, this is the first report of N. italica associated with GTDs worldwide. This is also the first record of K. variispora, S. vitis and D. variabile associated with wood infection of grapevine in Greece. The potential associations of disease indices with vine age, cultivar, GTD-associated population structure and the prevailing meteorological conditions in different viticultural zones in Greece are presented and discussed.

A Novel Seimatosporium and Other Sporocadaceae Species Associated with Grapevine Trunk Diseases in Cyprus.

Besides well-known grapevine trunk disease (GTD)-related pathogens, there is an increased interest in wood-colonizing fungi that infect grapevines. During 2017-2018, a survey was conducted in Cyprus and wood samples were collected from vines exhibiting typical GTD symptoms. Based on morphological and multilocus phylogenetic analyses (ITS, LSU, bt2, tef1-a), four species in the Sporocadaceae family were described and typified; two in the genus of Seimatosporium: Seim. cyprium sp. nov. and Seim. vitis-viniferae and two in Sporocadus: Spo. kurdistanicus and Spo. rosigena. The teleomorph of Seim. cyprium sp. nov. was also described. Pathogenicity trials with representative isolates of each species were performed on woody stems of two-year-old potted grapevines for 12 months under field conditions. All isolates were pathogenic, causing dark brown to black vascular discoloration, extending upward and downward from the inoculation point. Sporocadus isolates were significantly more aggressive than Seimatosporium with lesion lengths ranging from 9.24 to 6.90 and 4.13 to 4.00 cm, respectively. Successful re-isolations were also evident for all species and isolates. Seim. cyprium sp. nov. is a newly described species, while Spo. kurdistanicus and Spo. rosigena are reported for the first time in Europe on Vitis vinifera, suggesting the potential role of Sporocadaceae in the GTDs complex.

Reassessing Vermisporium (Amphisphaeriaceae), a genus of foliar pathogens of eucalypts.

The genus Vermisporium presently accommodates 13 species, 11 of which are associated with leaf spots of eucalypts in the Southern Hemisphere. Vermisporium is chiefly distinguished from Seimatosporium (Amphisphaeriaceae) on the basis of a short exogenous basal appendage, and the absence of a recognisable apical appendage. Due to the increasing importance of these species in native forests, and confusion pertaining to their taxonomy, a revision of the genus was undertaken based on fresh collections and dried herbarium specimens. Results from DNA sequence data analyses of the nrDNA-ITS and 28S nrRNA genes for species of Vermisporium indicated the genus to be a synonym of Seimatosporium. New combinations are introduced in Seimatosporium for several species: S. acutum, S. biseptatum, S. brevicentrum, S. obtusum, S. orbiculare, S. verrucisporum and S. walkeri. An updated key to all species occurring on eucalypts is also provided.