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a -e. Nectria peziza subsp. reyesiana (PAD S00015: herbarium Saccardo, n. 1609, holotype). a. Original herbarium specimen; b -c. perithecia on natural substrate; d -e. asci and ascospores in cotton blue. -f-j. Nectria illudens (PAD S00012: herbarium Saccardo, neotype). f. Original herbarium specimen; g. perithecia on natural substrate; h -j. ascospores in cotton blue. -Scale bars: b, g = 500 μm; c = 150 μm; d -e, h -j = 10 μm. -Photos: a -e by N. Forin; f-j by S. Nigris.
Source publication
Specimens of Nectria spp. and Nectriella rufofusca were obtained from the fungarium of Pier Andrea Saccardo, and investigated via a morphological and molecular approach based on MiSeq technology. ITS1 and ITS2 sequences were successfully obtained from 24 specimens identified as ‘ Nectria ’ sensu Saccardo (including 20 types) and from the type speci...
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Citations
... Previous studies have reported that Clonostachys rosea was associated with avocado fruit rot in Puebla, Mexico [24]. Clonostachys ambigua and C. pallens were identified on bark (unknown host) from Indonesia [25]. Clonostachys species, such as C. rosea, are well-known biological control agents for various plant pathogens [26]. ...
During an investigation of micro-fungi in soil, two fungal isolates belonging to the phylum Ascomycota, namely KNUF-20-NI011 and KNUF-20-NI006, were collected from Gyeongbuk Province and Dokdo Island in Korea and identified as Clonostachys divergens and Chrysosporium merdarium, respectively. The fungal isolates were confirmed through molecular phylogenetic analyses of the internal transcribed spacer regions, 28S rDNA large subunit, and β-tubulin sequences. Cultural and morphological characteristics were observed and determined using different media. These species were identified based on phylogenetic relationships along with their cultural and morphological characteristics. To our knowledge, this is the first report on Clonostachys divergens and Chrysosporium merdarium in Korea.
... Unculturable taxa represent a significant portion of the Kingdom Fungi, potentially representing missing lineages that can be analyzed from ecological samples using HTS. In addition, HTS technologies are proving valuable in analyzing DNA from older herbarium specimens that lack DNA sequences from the PCR/Sanger sequencing approach [52]. ...
Citation: Wijayawardene, N.N.; Boonyuen, N.; Ranaweera, C.B.; de Zoysa, H.K.S.; Padmathilake, R.E.; Nifla, F.; Dai, D.-Q.; Liu, Y.; Suwannarach, N.; Kumla, J.; et al. OMICS and Other Advanced Technologies in Mycological Applications. J. Fungi 2023, 9, 688. Abstract: Fungi play many roles in different ecosystems. The precise identification of fungi is important in different aspects. Historically, they were identified based on morphological characteristics, but technological advancements such as polymerase chain reaction (PCR) and DNA sequencing now enable more accurate identification and taxonomy, and higher-level classifications. However, some species, referred to as "dark taxa", lack distinct physical features that makes their identification challenging. High-throughput sequencing and metagenomics of environmental samples provide a solution to identifying new lineages of fungi. This paper discusses different approaches to tax-onomy, including PCR amplification and sequencing of rDNA, multi-loci phylogenetic analyses, and the importance of various omics (large-scale molecular) techniques for understanding fungal applications. The use of proteomics, transcriptomics, metatranscriptomics, metabolomics, and in-teractomics provides a comprehensive understanding of fungi. These advanced technologies are critical for expanding the knowledge of the Kingdom of Fungi, including its impact on food safety and security, edible mushrooms foodomics, fungal secondary metabolites, mycotoxin-producing fungi, and biomedical and therapeutic applications, including antifungal drugs and drug resistance, and fungal omics data for novel drug development. The paper also highlights the importance of exploring fungi from extreme environments and understudied areas to identify novel lineages in the fungal dark taxa.
... = 5.3 × 3.5 μm). A morphological comparison with C. vacuolata and C. pallens is difficult, because C. palllens was only described based on a sexual morph (Forin et al. 2020), while we observed only asexual morph characters in culture although working with the same strain. Schroers (2001) ...
Clonostachys ( Bionectriaceae , Hypocreales ) species are common soil-borne fungi, endophytes, epiphytes, and saprotrophs. Sexual morphs of Clonostachys spp. were placed in the genus Bionectria , which was further segregated into the six subgenera Astromata , Bionectria , Epiphloea , Myronectria , Uniparietina , and Zebrinella . However, with the end of dual nomenclature, Clonostachys became the single depository for sexual and asexual morph-typified species. Species of Clonostachys are typically characterised by penicillate, sporodochial, and, in many cases, dimorphic conidiophores (primary and secondary conidiophores). Primary conidiophores are mononematous, either verticillium-like or narrowly penicillate. The secondary conidiophores generally form imbricate conidial chains that can collapse to slimy masses, particularly on sporodochia. In the present study, we investigated the species diversity within a collection of 420 strains of Clonostachys from the culture collection of, and personal collections at, the Westerdijk Fungal Biodiversity Institute in Utrecht, the Netherlands. Strains were analysed based on their morphological characters and molecular phylogeny. The latter used DNA sequence data of the nuclear ribosomal internal transcribed spacer regions and intervening 5.8S nrDNA (ITS) and partial 28S large subunit (LSU) nrDNA and partial protein encoding genes including the RNA polymerase II second largest subunit ( RPB2 ), translation elongation factor 1-alpha ( TEF1 ) and β-tubulin ( TUB2 ). Based on these results, the subgenera Astromata , Bionectria , Myronectria and Zebrinella are supported within Clonostachys . Furthermore, the genus Sesquicillium is resurrected to accommodate the former subgenera Epiphloea and Uniparietina . The close relationship of Clonostachys and Sesquicillium is strongly supported as both are inferred phylogenetically as sister-genera. New taxa include 24 new species and 10 new combinations. Recognition of Sesquicillium distinguishes species typically forming a reduced perithecial stroma superficially on plant tissue from species in Clonostachys often forming well-developed, through bark erumpent stromata. The patterns of observed perithecial wall anatomies, perithecial wall and stroma interfaces, and asexual morph diversifications described in a previously compiled monograph are used for interpreting ancestral state reconstructions. It is inferred that the common ancestor of Clonostachys and Sesquicillium may have formed perithecia superficially on leaves, possessed a perithecial wall consisting of a single region, and formed intercalary phialides in penicilli of conidiophores. Character interpretation may also allow hypothesising that diversification of morphs occurred then in the two genera independently and that the frequently stroma-linked Clonostachys morphs evolved together with the occupation of woody host niches and mycoparasitism.
... The phylogenetic overview of Fusicolla based on multilocus sequence analyses showed that the genus is monophyletic [2]. The present phylogeny, including the newly added taxa, inferred from sequences of the acl1, ITS, LSU, rpb2 and tub2 regions, resulted in a similar tree topology to that demonstrated in the previous studies [8,9,46,47]. The result indicated that the four Chinese strains (CGMCC 3.24907, 3.24908, 3.24909 and 3.24910) grouped with the known species of Fusicolla (BIBP/MLBP = 1.0/96%), which confirmed their taxonomic placements. ...
... Among the known species of Fusicolla, F. aquaeductuum, F. betae, F. bharatavarshae [2,5,8,10,34,46,47,49], as well as the newly described species. Large-scale surveys covering different ecosystems and substrates in unexplored regions will further improve our knowledge of the species diversity of the genus and establish connections between the sexual and asexual stages of Fusicolla species, which will permit a better understanding of the whole fungus. ...
To explore the species diversity of the genus Fusicolla, specimens from Henan, Hubei and Jiangsu Provinces in China are examined, and three undescribed taxa are encountered. The morphological characteristics and DNA sequence analyses of the combined acl1, ITS, LSU, rpb2 and tub2 regions support their placement in Fusicolla and their recognition as new species. Fusicolla aeria sp. nov. is distinguished by the formation of abundant aerial mycelia on PDA, falcate, (1–)3-septate macroconidia 16–35 × 1.5–2.8 μm and subcylindrical, aseptate microconidia 7.5–13 × 0.8–1.1 μm. Fusicolla coralloidea sp. nov. has a coralloid colony on PDA, falcate, 2–5-septate macroconidia 38–70 × 2–4.5 μm and rod-shaped to ellipisoidal, aseptate microconidia 2–7 × 1–1.9 μm. Fusicolla filiformis sp. nov. is characterized by filiform, 2–6-septate macroconidia 28–58 × 1.5–2.3 μm and lacking microconidia. Morphological differences between these novel species and their close relatives are compared in detail. The previously recorded species of the genus in China are listed and a key to these taxa is provided.
... Notes: Clonostachys leucaenae is morphologically and phylogenetically closely related to Clonostachys pallens (PAD S00004) (97% MLBT, Fig. 7). Ascomata and asci dimensions of C. leucaenae are similar to C. pallens (Forin et al. 2020). Clonostachys leucaenae is morphologically distinguishable from C. pallens by its warted ascomatal surface and smaller ascospores than the latter species (12-17 vs. 16.2-18.2 ...
... Clonostachys leucaenae is morphologically distinguishable from C. pallens by its warted ascomatal surface and smaller ascospores than the latter species (12-17 vs. 16.2-18.2 µm) (Forin et al. 2020). There are 25 nucleotides differences (5.1%, 4 gaps) between C. leucaenae (MFLUCC 20-0008) and C. pallens (PAD S00004) ITS sequences. ...
... 10.5-15.2 μm) (Forin et al. 2020). Clonostachys spinulosa is morphologically distinguishable from C. eriocamporesiana by its distinctly warted ascomatal surface, larger ascomata (250-400 × 250-385 μm vs. 160-260 × 160-285 μm) and larger asci than the latter species (69-80 × 8.6-11 μm vs. 58-66 × 7.3-10.3 ...
This paper provides outlines for Bionectriaceae, Calcarisporiaceae, Hypocreaceae, Nectriaceae, Tilachlidiaceae, Ijuhyaceae, Stromatonectriaceae and Xanthonectriaceae with taxonomic treatments. We provide up-to-date DNA sequence-based phylogenies including combined gene analysis of ITS, LSU, rpb2, tef1 and tub2 for Hypocreales and accept 17 families. Three new families and 12 new species are introduced with descriptions and illustrations, while 13 new records and one new species combination are provided. Here we mainly detail the taxonomy of Bionectriaceae, Hypocreaceae, Nectriaceae and Tilachlidiaceae, Ijuhyaceae fam. nov., Stromatonectriaceae fam. nov. and Xanthonectriaceae fam. nov. are introduced in this study based on phenotypic and molecular analyses. For each family we provide a list of accepted genera, the taxonomic history, morphological descriptions, taxonomic placement based on DNA sequence data and illustrate the type genus. Representatives of each family are illustrated based on the type herbarium material or fresh specimens where available, or provide relevant references. Notes on ecological and economic importance of the families are also given.
... About 100 names have been created under the genus Clonostachys (www.indexfungorum. org (accessed on 1 July 2022)), among which 65 species are commonly accepted [2,3,[7][8][9][10][11][12][13][14][15][16][17][18][19][20]. Twenty-four species are known from China [2,7,21]. ...
Three new species of Clonostachys are introduced based on specimens collected from China. Clonostachys chongqingensis sp. nov. is distinguished by pale yellow to pale orange-yellow perithecia with a very low papilla, clavate to subcylindrical asci possessing ellipsoidal to elongate-ellipsoidal spinulose ascospores 13–16 × 4.5–5.5 μm; it has acremonium- to verticillium-like conidiophores and ellipsoidal to rod-shaped conidia. Clonostachys leptoderma sp. nov. has pinkish-white subglobose to globose perithecia on a well-developed stroma and with a thin perithecial wall, clavate to subcylindrical asci with ellipsoidal to elongate-ellipsoidal spinulose ascospores 7.5–11 × 2.5–3.5 μm; it produces verticillium-like conidiophores and ellipsoidal to subellipsoidal conidia. Clonostachys oligospora sp. nov. features solitary to gregarious perithecia with a papilla, clavate asci containing 6–8 smooth-walled ascospores 9–17 × 3–5.5 μm; it forms verticillium-like conidiophores and sparse, subfusiform conidia. The morphological characteristics and phylogenetic analyses of combined nuclear ribosomal DNA ITS1-5.8S-ITS2 and beta-tubulin sequences support their placement in Clonostachys and their classification as new to science. Distinctions between the novel taxa and their close relatives are compared herein.
... The internal transcribed spacer (ITS) region was amplified by PCR using the primers ITS1 and ITS4; PCR products were sequenced in an ABI 3500 DNA sequencer. A phylogenetic tree of Clonostachys was constructed with sequences from ex-type cultures or reference strains used in other taxonomic studies (Moreira et al. 2016;Forin et al. 2020;Hyde et al. 2020) and sequences from GenBank database. Maximum Likelihood (ML) analyses were done as described by Montalva et al. 2018. ...
... The anamorphic genus Clonostachys (Bionectriaceae, Hypocreales), described by Corda in 1839, includes a high diversity with more than 50 known species throughout the world (Schroers 2001;Forin et al. 2020;Hyde et al. 2020). These fungi are commonly found in decaying plants in soils, and rarely specimens were isolated from water, air or insects (Schroers 2001;Chang et al. 2021). ...
Aedes aegypti (Linnaeus, 1762) is an important vector of arboviruses in the tropics and subtropics. New control strategies based on natural enemies such as entomopathogenic fungi are of utmost importance, and the present study reports the first isolation of Clonostachys spp. (Hypocreales: Bionectriaceae) from mosquitoes and their activity against A. aegypti. Entomopathogenic fungi were surveyed in central Brazil using A. aegypti larvae as sentinels and, also, a CDC light trap. Clonostachys eriocamporesii R.H. Perera & K.D. Hyde, 2020 (IP 440) and Clonostachys byssicola Schroers, 2001 (IP 461) were identified by sequence analysis of the nuclear ribosomal internal transcribed spacer gene, and tested against eggs, larvae, and adults. Both strains were highly active against A. aegypti third instar larvae, with mortalities ≥ 80% at 107 conidia/mL after 5 days but distinctly less active against eggs and adults. This is the first report of both C. eriocamporesii and C. byssicola as naturally occurring pathogens affecting mosquitoes, and IP 440 appears to be a promising control agent against aquatic stages of A. aegypti.
... Field trips for macrofungi should therefore pay more attention to isolation of living cultures, although preserved specimens remain important for taxonomical studies. In addition, innovative methodologies in DNA extraction and genome sequencing and assembly should be developed to bring huge amounts of old specimens in global fungaria to the genomics era (Forin et al. 2018(Forin et al. , 2020. More importantly, the accuracy of functional annotation for fungal genomes urgently needs to be improved (Mohanta and Al-Harrasi 2021). ...
Fungal taxonomy is a fundamental discipline that aims to recognise all fungi and their kinships. Approximately 5% of a practical estimate of 2.2–3.8 million species globally are currently known, and consequently the Fungal Tree of Life (FTOL) is very incompletely reconstructed. With the advances of new technologies, mycology is marching into the interdisciplinary and globalisation era. To make fungal taxonomy relevant, innovative sampling methods and phylogenomics analyses should be performed to reconstruct a much more comprehensive FTOL. In association with this densely sampled FTOL, multiomics will reveal what drives fungal species diversification and how fungal traits evolve to adapt to various environments, while metagenomics will facilitate the understanding and protection of the ecological functions of fungi. A coordinated approach to pursuing these research agendas that includes conceiving of and costing a mission to describe all the fungi on the planet will unlock potential of fungi to support sustainable development of our society.
... У разі роботи з недозрілими, пошкодженими або аберантними плодовими тілами цих грибів, є велика вірогідність хибного визначення. Рішенням у таких ситуаціях може бути залучення сучасних молекулярно-генетичних методів досліджень (Kotlaba, Zehnálek, 2018;Forin et al., 2020;Pärtel et al., 2021). ...
The article provides results of our revision of three herbarium specimens of the genus Tulostoma collected in Ukraine and deposited in the Mycological Herbarium of V.N. Karazin Kharkiv National University, CWU (Myc). Previously, they have been identified as Tulostoma kotlabae, T. niveum, and T. subsquamosum and reported as newly recorded species in Ukraine. For the examined specimens, we analyzed the nucleotide sequences of the ITS region of ribosomal DNA. For one specimen, the large subunit (LSU) of rDNA was also obtained. Based on the nucleotide sequencing results, one specimen (GB00121) was re-identified as Tulostoma melanocyclum and the other two specimens (GB00125 and GB00129) – as T. simulans. The latter species is reported in Ukraine for the first time. Given that genus Tulostoma has been subject to a number of recent taxonomic changes, it would be reasonable to make a revision of other herbarium specimens from Ukraine using molecular methods.
... Hence, old material would be the only option to obtain DNA, but it is difficult to obtain cultures from these old taxa since the viability of spores is lower. Thus, direct extracting DNA from old material has been suggested in some studies [79,81,82]. ...
... However, due to degradation, fragmentation, and the deamination, the DNA quality and quantity can be low, reducing the efficacy of PCR [187]. Forin et al. [79,81] used specimens of Nectria and Peziza from P. A. Saccardo's mycological collection to extract DNA and revisit the taxonomic status of particular taxa. In both studies, Forin et al. [79,81] used high-throughput sequencing to overcome the DNA fragmentation and exogenous DNA contaminants. ...
... Forin et al. [79,81] used specimens of Nectria and Peziza from P. A. Saccardo's mycological collection to extract DNA and revisit the taxonomic status of particular taxa. In both studies, Forin et al. [79,81] used high-throughput sequencing to overcome the DNA fragmentation and exogenous DNA contaminants. Daru et al. [188] is an interesting study which showed that old specimens harbored unrevealed endophytic species. ...
Culture techniques are vital in both traditional and modern fungal taxonomy. Establishing sexual–asexual links and synanamorphs, extracting DNA and secondary metabolites are mainly based on cultures. However, it is widely accepted that a large number of species are not sporulating in nature while others cannot be cultured. Recent ecological studies based on culture-independent methods revealed these unculturable taxa, i.e., dark taxa. Recent fungal diversity estimation studies suggested that environmental sequencing plays a vital role in discovering missing species. However, Sanger sequencing is still the main approach in determining DNA sequences in culturable species. In this paper, we summarize culture-based and culture-independent methods in the study of ascomycetous taxa. High-throughput sequencing of leaf endophytes, leaf litter fungi and fungi in aquatic environments is important to determine dark taxa. Nevertheless, currently, naming dark taxa is not recognized by the ICN, thus provisional naming of them is essential as suggested by several studies.
