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Neoerysiphe leontopodii sp. nov., a new powdery mildew from China
2023 Volume 64 Issue 6 Pages 150-155
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2023 Volume 64 Issue 6 Pages 150-155
A powdery mildew was found on Leontopodium leontopodioides (Asteraceae) in China. Phylogenetic analyses using a combination of internal transcribed spacer and 28S rDNA sequences showed that this species, which clusters as sister to Neoerysiphe joerstadii, is allied to N. galii, N. geranii, and N. nevoi. This species differs from the closely allied N. joerstadii in the number and size of asci (3-10 asci, 55-75 × 20-40 µm versus 16-32 asci, 40-60 × 20-30 µm). This species is morphologically very similar to N. gnaphalii, but clearly differs from this species in having larger chasmothecia and colorless appendages. Therefore, the powdery mildew on L. leontopodioides is described as N. leontopodii sp. nov.
Leontopodium leontopodioides (Willd.) Beauv. (Asteraceae), a perennial herb, is widely distributed in northeast, northwest, north and southwest China, especially in the Qinghai Tibet Plateau (Chen et al., 2011). Leontopodium leontopodioides is commonly used in the traditional Chinese medicine for the treatment of albuminuria, nephritis, urinary tract infection, hematuria, vaginitis, bronchopneumonia, gastric ulcer and traumatic bleeding (Li et al., 2012; Zhang et al., 2018; Zhao et al., 2019). Its main chemical components include phenylpropanoids, flavonoids, sesquiterpene, phytosterols, etc. (Pan, 2009). In addition, it plays an important role in the succession of alpine meadows (Zhang et al., 2019).
Leontopodium leontopodioides belongs to Asteraceae subtribe Gnaphalinae (Chen et al., 2011). Two powdery mildew species of Neoerysiphe have been described on host species of subtribe Gnaphalinae. Neoerysiphe joerstadii V. P. Heluta & S. Takam. parasitizes on the Phagnalon rupestre and is distributed in the Golan Heights of Israel in the Mediterranean region (Heluta et al., 2010; Braun & Cook, 2012). Neoerysiphe joerstadii has larger chasmothecia and a larger number of asci compared to N. leontopodii. The second species, N. gnaphalii U. Braun, parasitizes on Omalotheca norvegica and is distributed in northern Europe, including Finland, Norway, and Sweden (Braun & Cook, 2012). Its morphology is similar to N. galeopsidis, and appendages are geniculate-sinuous to spirally twisted, usually unbranched (Braun & Cook, 2012).
In late Aug of 2012, we found chasmothecia on the surface of leaves of L. leontopodioides in the Inner Mongolia Autonomous Region, China. The leaves of L. leontopodioides are characterized by having a dense white tomentum, so that the asexual morph of this species is difficult to detect. Morphological and molecular analyses revealed that this morph represents an undescribed species belonging to the genus Neoerysiphe. Neoerysiphe leontopodii sp. nov is proposed in this study for this fungus, including a detailed morphological description and illustration.
So far, only a single powdery mildew sample on L. leontopodioides has been collected in Sumushan Forest Park, Xinghe County, Ulanqab City, Inner Mongolia Autonomous Region, China. The collection is deposited in the Mycological Herbarium of Chifeng University (CFSZ). Conidiophores, conidia, and chasmothecia were stripped from the leaf surface with a clean needle, mounted on a microscope slide, and examined in lactic acid using a light microscope (ZEISS scopeA1, Germany) with phase contrast at 10 ×, 20 ×, and 40 × objectives. Thirty measurements for all structures of taxonomical value were performed.
2.2. Molecular phylogenyGenomic DNA was extracted from mycelia and chasmothecia using the chelex-100 method (Walsh et al., 1991) as previously described (Hirata & Takamatsu, 1996). PCR and DNA sequencing were conducted according to the procedures described in Liu et al. (2020), except for the primers that were used. PM2/PM10 and PM3/LSU2 primer sets were used to amplify 5'-half of the internal transcribed spacer (ITS) region and 3'-half of ITS and 5'-end of the 28S rDNA gene (including domains D1 and D2), respectively (Bradshaw & Tobin, 2020; Scholin et al., 1994). The newly obtained sequences in this study were deposited in GenBank with the accession number OR269994. The sequences were aligned with closely related sequences of other Neoerysiphe species retrieved from GenBank using MUSCLE implemented in the MEGA7 program (Kumar et al., 2016). Alignments were further manually refined and deposited in TreeBASE (https://www.treebase.org/) under the accession number 30579. Phylogenetic trees were obtained from the data using the maximum-parsimony (MP) method in PAUP4.0 (Swofford, 2002) with heuristic search option using the tree-bisection-reconstruction (TBR) algorithm with 100 random sequence additions to find the global optimum tree. All sites were treated as unordered and unweight with gaps treated as missing data. The strength of internal branches of resulting trees was tested with bootstrap analysis (1000 replications) (Felsenstein, 1985). Bootstrap (BS) values of 50% or higher are indicated in the tree (Fig. 3). Tree scores, including tree length, consistency index (CI), retention index (RI) and rescaled consistency index (RC) were also calculated.
Neoerysiphe leontopodii T.Z. Liu & L. Liu, sp. nov. Figs. 1, 2.
MycoBank no.: MB849490.
Diagnosis: Differs from Neoerysiphe gnaphalii in having larger chasmothecia (vs. smaller chasmothecia, 85-140 µm diam in N. gnaphalii) and colorless appendages (vs. appendages yellowish brown to moderate dark brown in N. gnaphalii).
Type: CHINA, Sumushan Forest Park, Xinghe County, Ulanqab City, Inner Mongolia Autonomous Region, on Leontopodium leontopodioides (Asteraceae), 21 Aug 2012, Tie-Zhi Liu, Zhen-Hua Jia and De-Nan Su (holotype, CFSZ 5854).
Gene sequences ex-holotype: OR269994
Etymology: The specific epithet “leontopodii” is derived from the host genus name.
Description: Mycelium on leaves, amphigenous, and stems, white, in irregular patches, sometimes confluent or covering the entire surface of the leaves, evanescent to persistent; hyphal cells 5-10 µm wide, smooth to slightly rough, hyaline; hyphal appressoria and secondary hyphae not observed; conidiophores arising mostly centrally from the upper surface of the hyphal mother cell, erect, about 85-175 µm long, foot-cells 50-100 × 10-15 µm, cylindrical, straight to mostly somewhat flexuous at the base, followed by 1-2 shorter cell, forming catenescent conidia; conidia ellipsoid, doliiform-cylindrical, subcylindrical, 20-35(-37.5) × 10-15(-17.5) µm, with a longitudinal surface wrinkling pattern. Chasmothecia scattered to gregarious, sometimes immersed in the pannose tomentum of the host, 120-180 µm diam, dark brown, depressed globose; peridium cells irregularly polygonal or daedaleoid, 8-25 µm diam; appendages numerous, about 10-40, arising from the lower half, mycelioid, unbranched, rarely irregularly branched, rarely stiff, sometimes poorly developed, sinuous or geniculate, length 20-150 μm, 0.2-1 times as long as the diameter of the chasmothecium, mostly shorter than the chasmothecial diam., 2.5-8 μm wide, thin-walled, walls thick below, becoming gradually thinner towards the tip, smooth or slightly rough, 0-3-septate, hyaline; asci 3-10, ellipsoid, obovoid, with irregular outline, 55-75 × 20-40 µm, sessile or short-stalked, content often yellowish, but always immature in the current season; ascospores not developed before overwintering.
Host range and distribution: On Leontopodium leontopodioides. Inner Mongolia Autonomous Region, China, only known from the type locality.
3.2. Molecular phylogenyA phylogenetic tree has been constructed, using a data set of ITS and 28S rDNA sequences, to clarify the phylogenetic affinity of N. leontopodii. These sequences were aligned with 51 sequences from closely related species reported in Bradshaw et al. (2022). Two sequences of Microidium phyllanthi-reticulati were used as outgroup based on Bradshaw and Tobin (2020). The data-set consisted of 54 sequences. Of the 1200 characters, 28 characters were variable and 228 characters were informative for parsimony analysis. A total of 100 equally parsimonious trees were constructed by the MP analysis. Tree topologies were almost consistent among the trees, except for branching orders of the terminal groups and branch lengths. One of the trees with the highest likelihood value is shown in Fig. 3. Neoerysiphe leontopodii forms an independent cluster with N. joerstadii with high BS support (98%).
The ITS sequence of Neoerysiphe leontopodii clusters close to a corresponding sequence of N. joerstadii (AB498976), with a similarity of 98.04%. The analysis of the 28S rDNA sequence yielded a maximum similarity of 98.48% to both N. galeopsidis (ON415299), N. galii (ON415305) and N. chelones (ON415293). Based on the phylogenetic tree constructed by ITS and 28S rDNA sequences, N. leontopodii is sister to N. joerstadii, with N. galii, N. nevoi, and N. geranii as further allied species. The hosts of N. leontopodii and N. joerstadii both belong to subtribe Gnaphalinae, and in the phylogenetic tree, they cluster together, indicating that these two species may come from a common ancestor, reflecting co-evolution with the host plants. Hosts of N. galii belong to the genera Cruciata, Galium, and Phuopsis (Rubiaceae). The taxonomic-phylogenetic placement of the latter species has been proven by several authors (Takamatsu et al., 2008, 2009). Based on overwintering experiments of this species, it was demonstrated that the asci and ascospores within chasmothecia mature in the next year (Braun & Cook, 2012). Neoerysiphe nevoi is parasitic on hosts of the tribe Cichorieae of the Asteraceae. The host range of this species is still insufficiently known and needs further research. Some of the collections reported as “Erysiphe cumminsiana”, “E. galeopsidis”, “E. galii”, and “E. cichoracearum” on Asteraceae might belong to N. nevoi and need to be microscopically (and/or phylogenetically) re-examined (Amano, 1986; Braun, 1987; Gorter, 1987). Neoerysiphe geranii was introduced as a new species by Nomura (1997), and its phylogenetic position was later clarified by Heluta et al. (2010). Originally reported from Japan, this species has later been confirmed for China (Liu, 2010; Liu et al., 2006), India (Braun & Paul, 2009), Russia and Ukraine (Heluta et al., 2010), the United Kingdom, and New Zealand (Braun & Cook, 2012), but it might have a much wider distribution. Neoerysiphe leontopodii differs from N. joerstadii in having 3-10 asci, 55-75 × 20-40 µm (vs., 16-32 asci, 40-60 × 20-30 µm). Neoerysiphe leontopodii is morphologically very close to N. gnaphalii, but differs in having larger chasmothecia, 120-180 μm diam, whereas the latter species has smaller chasmothecia, 85-140 μm diam. Furthermore, the appendages of the new species are colorless, in contrast to yellowish brown to moderately dark brown in N. gnaphalii (Braun & Cook, 2012). Unfortunately, N. gnaphalii has not yet been sequenced, so that the phylogenetic relationship between N. gnaphalii and N. leontopodii can currently not be elucidated. In general, it is recommendable to base new species at least on two specimens and sequences. However, in the present case it was not possible to find a second collection. The texture of the leaf surface of the host, Leontopodium leontopodioides, with a dense white hairiness, strongly impedes the tracing of this powdery mildew, so that the discovery of this powdery happened rather by chance.
The authors declare no conflicts of interest. All the experiments undertaken in this study comply with the current laws of the country where they were performed.
This research was supported by the National Natural Science Foundation of China (No. 31760004, No. 31970019, No. U21A20177) and First-class disciplines construction project of Chifeng University (No. CFXYYLXKB 202102). The authors sincerely acknowledge Prof. Uwe Braun for reading the full text, for critical comments and for corrections.
