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Research Article
Identification and fungicide sensitivity of Microdochium chrysopogonis (Ascomycota, Amphisphaeriaceae), a new species causing tar spot of Chrysopogon zizanioides in southern China
expand article infoXiang Lu‡, Mengxian Mai‡, Wenhui Tan‡, Muyan Zhang‡, Jie Xie‡, Yi Lu‡, Xue Li Niu‡, Wu Zhang‡
‡ Lingnan Normal University, Zhanjiang, China

Corresponding author: Wu Zhang ( ldzw1987@163.com )

Academic editor: Chayanard Phukhamsakda
© 2023 Xiang Lu, Mengxian Mai, Wenhui Tan, Muyan Zhang, Jie Xie, Yi Lu, Xue Li Niu, Wu Zhang.
This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Citation: Lu X, Mai M, Tan W, Zhang M, Xie J, Lu Y, Niu XL, Zhang W (2023) Identification and fungicide sensitivity of Microdochium chrysopogonis (Ascomycota, Amphisphaeriaceae), a new species causing tar spot of Chrysopogon zizanioides in southern China. MycoKeys 100: 205-232. https://doi.org/10.3897/mycokeys.100.112128
Open Access

Abstract

Vetiver grass (Chrysopogon zizanioides) has received extensive attention in recent years due to its diverse applications in soil and water conservation, heavy metal remediation, as well as essential oil and phenolic acids extraction. In 2019, the emergence of tar spot disease on C. zizanioides was documented in Zhanjiang, Guangdong Province, China. Initially, the disease manifested as black ascomata embedded within leaf tissue, either scattered or clustered on leaf surfaces. Subsequently, these ascomata became surrounded by fisheye lesions, characterised by brown, elliptical, necrotic haloes, which eventually coalesced, resulting in leaf withering. Koch’s postulates demonstrated that the fungus isolated from these lesions was the causal agent. Microscopic examination showed that the pathogen morphologically belonged to Microdochium. The phylogenetic tree inferred from the combined ITS, LSU, tub2 and rpb2 sequences revealed the three isolates including GDMCC 3.683, LNU-196 and LNU-197 to be a novel species of Microdochium. Combining the results of phylogenetic, pathogenicity and morphological analyses, we propose a new species named M. chrysopogonis as the causal agent of C. zizanioides in southern China. The optimum growth temperature for M. chrysopogonis was determined to be 30 °C. The in vitro fungicide sensitivity of M. chrysopogonis was determined using a mycelial growth assay. Four demethylation-inhibiting (DMI) fungicides, including difenoconazole, flusilazole, propiconazole and tebuconazole and one methyl benzimidazole carbamate (MBC) fungicide, carbendazim, were effective against M. chrysopogonis, with mean 50% effective concentration (EC50) values of 0.077, 0.011, 0.004, 0.024 and 0.007 μg/ml, respectively. These findings provide essential references for the precise diagnosis and effective management of M. chrysopogonis.

Key words

fungicide sensitivity, multilocus phylogeny, new taxon, pathogenicity, tar spot

Introduction

Vetiver (Chrysopogon zizanioides) is one of the main grasses in tropical and subtropical areas (Maurya et al. 2023). With a large root system that penetrates deep into the soil, C. zizanioides is strongly tolerant to adverse environments, such as drought, salinity and heavy metals. Recently, vetiver has been widely utilised in various applications, such as land restoration, soil and water conservation and phytoremediation of heavy metal-contaminated soils (Chen et al. 2021). Moreover, the essential oil and phenolic acids extracted from vetiver root possess significant aromatic and biological properties, giving it an important role in perfumery, the food industry and medicine (David et al. 2019; Moon et al. 2020).

To our knowledge, four diseases on C. zizanioides have been reported, namely, leaf blight caused by Curvularia trifolii in India (Babu et al. 2021), root and basal stem rot caused by Gaeumannomyces graminicola (Lin et al. 2019), leaf spot caused by Phoma herbarum (Zhang et al. 2017) and leaf streak caused by Stenocarpella chrysopogonis (Jia et al. 2023).

Microdochium species were originally introduced with the type species, M. phragmitis, identified on the leaves of Phragmites australis in Germany (Sydow 1924). Presently, there are 49 species included in this genus. However, only a subset of these species can induce diseases, primarily affecting grasses and cereals. For example, M. albescens (also referred to as Monographella albescens) typically induces leaf scald and grain discoloration in rice, leading to a global reduction in rice yield (Araujo et al. 2016; Dirchwolf et al. 2023). M. bolleyi is recognised for inducing root necrosis and basal rot in creeping bent grass in Korea, as well as causing root rot on triticale in Kazakhstan (Hong et al. 2008; Alkan et al. 2021). M. nivale and M. majus frequently result in the occurrence of pink snow mould or Fusarium Patch on wheat, barley and turf grass in cold to temperate regions (Ren et al. 2015; Abdelhalim et al. 2020). M. opuntiae leads to brown spotting on Opuntia (Braun 1995). M. poae triggers leaf blight disease in turf-grasses, such as Poa pratensis and Agrostis stolonifera (Liang et al. 2019). M. paspali is recognised for its ability to cause leaf blight in seashore paspalum (Paspalum vaginatum) (Zhang et al. 2015). M. panattonianum has the potential to induce anthracnose in lettuce (Galea et al. 1986). M. sorghi is accountable for the formation of zonate leaf spots and decay on sorghum species (Stewart et al. 2019).

The application of fungicides has always been an effective approach for disease control. In recent decades, demethylation-inhibiting (DMI) fungicides have emerged as a significant and extensive group of fungicides, exhibiting notable efficacy in the control of diseases caused by the Microdochium genus. Notably, compounds such as prochloraz, difenoconazole, propiconazole, metconazole, myclobutanil, tebuconazole and triticonazole have shown substantial antifungal efficacy against M. panattonianum, M. majus and M. nivale (Wicks et al. 1994; Debieu et al. 2000; Glynn et al. 2008; Mao et al. 2023). Additionally, fungicide subgroups, including phenylpyrrole (PP) fungicides, such as fludioxonil, dicarboximides, such as iprodione and quinone outside inhibitors (QoIs), such as trifloxystrobin, have demonstrated noteworthy efficacy in the management of diseases induced by M. nivale (Glynn et al. 2008; Koch et al. 2015; Aamlid et al. 2017). Therefore, to promote effective control against tar spot of C. zizanioides, it is necessary to determine the sensitivity of the pathogen to fungicides.

The main objectives of this study were to identify the pathogenic fungi causing tar spot of C. zizanioides in southern China on the basis of morphological characteristics and multigene sequence analysis; to determine the pathogenicity to C. zizanioides; and to determine the inhibitory effect of fungicides against mycelial growth of the pathogen.

Materials and methods

Sample collection and fungal isolation

Leaves exhibiting symptoms of tar spot on C. zizanioides were collected in fields of the Grass Research Station of Lingnan Normal University (LNU), Zhanjiang, Guangdong, China. Leaf segments (0.5 × 0.5 cm) from the transition zone from diseased to healthy tissue were cut and surface-sterilised for 30 s with 75% ethanol and 2% sodium hypochlorite (NaClO) for 1 min, rinsed with distilled water 3 times, dried on sterile filter paper and placed on 2% potato dextrose agar (PDA) (Crous et al. 2021b). Additionally, ascomata developing on the surface of diseased tissue were gently scraped using a sterile scalpel. Subsequently, a small number of ascospores were transferred and evenly spread on to the surface of a water agar (WA) plate. Hyphal tips originating from leaf tissue fragments and single germinating conidia were transferred on to PDA medium. They were then incubated at 30 °C in darkness (Polizzi et al. 2009). After a period of 7 days, the isolates were transferred on to PDA slants and preserved at 4 °C in the culture collection of Lingnan Normal University. Additionally, they were deposited in the Guangdong Microbial Culture Collection Center (GDMCC) in Guangzhou, China. The holotype specimen was preserved in the Herbarium of the Chinese Academy of Forestry (CAF) in Beijing, China.

Morphological characterisation

Colonies were subcultured on 2% malt extract agar (MEA) and oatmeal agar (OA) at 30 °C for 10 days in the dark (Crous et al. 2009). Colony colour was characterised using Rayner’s Mycological Color Chart (Rayner 1970) and colony diameters were measured after incubation for 10 days at 30 °C in the dark. Morphological characters of ascomata, asci, ascospores, sporodochia, hyphae, conidiomata, conidiophores, conidiogenous cells and conidia were determined in sterile water using an Olympus BX53 compound microscope (Tokyo, Japan), equipped with cellSens Dimension software (version 1.17).

DNA extraction, PCR amplification and sequencing

Fungal genomic DNA was extracted from mycelia grown on PDA medium after 10 days using the ENZA Fungal DNA Miniprep Kit (Omega Bio-tek, Doraville, Norcross, GA, U.S.A.), according to the protocol of manufacturer. Four loci, including internal transcribed spacer (ITS) rDNA region, large subunit ribosomal acid (LSU) rDNA region, RNA polymerase II second largest subunit gene (rpb2) and part of the beta-tubulin gene (tub2), were amplified by the following primer pairs: ITS1 and ITS4 for ITS (White et al. 1990), LR0R and LR5 for LSU (Vilgalys and Hester 1990), RPB150F (Jewell and Hsiang 2013) and fRPB2-7Cr (Liu et al. 1999) and Btub526F and Btub1332R (Jewell and Hsiang 2013). The polymerase chain reaction (PCR) conditions were as follows: 94 °C for 5 min; 94 °C for 30 s, annealing temperature for 45 s and 72 °C for 1 min, 35 cycles; and a final extension step at 72 °C for 10 min. The annealing temperature for ITS and LSU was 54 °C, for tub2 was 55 °C and for rpb2 was 57 °C. PCR products were sequenced by Sangon Biotech Co., Ltd. (Shanghai, China). Sequences were edited and assembled using DNAMAN version 5.2.2 and deposited in the NCBI GenBank nucleotide database (Table 1).

Table 1.
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Strains included in the phylogenetic analyses with collection details and GenBank accession numbers.

Species Voucher Country GenBank Accession Number
LSU ITS tub2 rpb2
Microdochium albescens CBS 290.79 Ivory Coast KP858950 KP859014 KP859078 KP859123
CBS 291.79 Ivory Coast KP858932 KP858996 KP859059 KP859105
CBS 243.83 Unknown country KP858930 KP858994 KP859057 KP859103
M. bolleyi CBS 172.63 Germany MH869857 MH858255 – –
CBS 540.92 Syria KP858946 KP859010 KP859073 KP859119
Kaz_Mb01 Kazakhstan – MW301448 – –
Kaz_Mb02 Kazakhstan – MW301449 – –
CBS 137.64 Netherlands MH870023 MH858394 – –
CPC 25994 Canada KP858954 KP859018 KP859074 KP859127
CBS 102891 Germany MH874405 – – –
CBS 618.72 Germany MH872294 MH860598 – –
M. chrysanthemoides CGMCC 3.17929T China KU746736 KU746690 KU746781 –
CGMCC 3.17930 China KU746735 KU746689 KU746782 –
M. chuxiongense YFCC 8794T China OK586160 OK586161 OK556901 OK584019
M. citrinidiscum CBS 109067T Peru KP858939 KP859003 KP859066 KP859112
M. colombiense CBS 624.94T Colombia KP858935 KP858999 KP859062 KP859108
M. dawsoniorum BRIP 65649T Australia ON394569 MK966337 – –
BRIP 67439 Australia OM333563 MN492650 – ON624208
M. fisheri CBS 242.90T UK KP858951 KP859015 KP859079 KP859124
NFCCI 4083 India KY777594 KY777595 – –
C30 ITI Sri Lanka – MT875317 – –
M. graminearum CGMCC 3.23525T China OP104016 OP103966 OP236029 OP236026
CGMCC 3.23524 China OP104015 OP103965 OP242835 OP236026
M. hainanense SAUCC210781T China OM959323 OM956295 OM981146 OM981153
SAUCC210782 China OM959324 OM956296 OM981147 OM981154
M. indocalami SAUCC1016T China MT199878 MT199884 MT435653 MT510550
M. insulare BRIP 75114a Australia OQ892168 OQ917075 – OQ889560
M. lycopodinum CBS 146.68 The Netherlands KP858929 KP858993 KP859056 KP859102
CBS 109397 Germany KP858940 KP859004 KP859067 KP859113
CBS 109398 Germany KP858941 KP859005 KP859068 KP859114
CBS 109399 Germany KP858942 KP859006 KP859069 KP859115
CBS 125585T Austria KP858952 KP859016 KP859080 KP859125
M. maculosum COAD 3358T Brazil OK966953 OK966954 – OL310501
M. majus CBS 741.79 Germany KP858937 KP859001 KP859064 KP859110
10099 France – JX280597 JX280563 JX280560
10098 France – – JX280564 JX280561
99027 Canada – JX280583 JX280566 –
200107 Norway – KT736191 KT736253 KT736287
M. miscanthi SAUCC211092T China OM957532 OM956214 OM981141 OM981148
SAUCC211093 China OM957533 OM956215 OM981142 OM981149
SAUCC211094 China OM957534 OM956216 OM981143 OM981150
M. musae CBS 143500T Malaysia MH107942 MH107895 MH108041 MH108003
CBS 143499 Malaysia MH107941 MH107894 MH108040 –
M. musae CBS 111018 Costa Rica – AY293061 – –
CPC 11240 Mauritius MH107944 MH107897 MH108043 –
CPC 16258 Mexico MH107945 MH107898 MH108044 –
CPC 11234 Mauritius MH107943 MH107896 MH108042 –
CPC 32681 Malaysia MH107946 MH107899 – –
M. neoqueenslandicum CBS445.95 The Netherlands KP858933 KP858997 KP859060 KP859106
CBS108926T New Zealand KP858938 KP859002 KP859065 KP859111
M. nivale CBS 116205T UK KP858944 KP859008 KP859071 KP859117
200114 Norway – KT736185 – KT736279
200119 Norway – KT736199 KT736240 KT736263
200120 Norway – KT736210 KT736221 KT736273
200566 Norway – KT736220 KT736224 –
201050 Norway – KT736217 KT736236 KT736257
M. novae-zelandiae CBS 143847 New Zealand – LT990655 LT990608 LT990641
CPC 29693 New Zealand – LT990656 LT990609 LT990642
M. paspali CBS 138620T China – KJ569509 KJ569514 –
CBS 138621 China – KJ569510 KJ569515 –
CBS 138622 China – KJ569511 KJ569516 –
M. phragmitis CBS 285.71T Poland KP858949 KP859013 KP859077 KP859122
CBS 423.78 Germany KP858948 KP859012 KP859076 KP859121
M. poae CGMCC3.19170T China – MH740898 MH740914 MH740906
LC12115 China – MH740901 MH740917 MH740909
LC12116 China – MH740902 MH740918 MH740910
LC12117 China – MH740903 MH740919 MH740911
LC12118 China – MH740897 MH740913 MH740905
LC12119 China – MH740899 MH740915 MH740907
LC12120 China – MH740904 MH740920 MH740912
LC12121 China – MH740900 MH740916 MH740908
M. ratticaudae BRIP 68298T Australia MW481666 MW481661 – MW626890
M. rhopalostylidis CBS 145125T New Zealand MK442532 MK442592 MK442735 MK442667
M. salmonicolor NC14-294 South Korea MK836108 MK836110 – –
M. seminicola CBS 122706 Switzerland KP858943 KP859007 KP859070 KP859116
CBS 122707 Switzerland KP858947 KP859011 KP859081 KP859120
CBS 139951T Switzerland KP858974 KP859038 KP859101 KP859147
KAS1516 Canada KP858961 KP859025 KP859088 KP859134
KAS3574 Switzerland KP858973 KP859037 KP859100 KP859146
KAS3158 Canada KP858970 KP859034 KP859097 KP859143
KAS1527 Canada KP858966 KP859030 KP859093 KP859139
KAS1473 Canada KP858955 KP859019 KP859082 KP859128
M. shilinense CGMCC 3.23531T China OP104022 OP103972 OP242834 –
M. sinense SAUCC211097T China OM959225 OM956289 OM981144 OM981151
SAUCC211098 China OM959226 OM956290 OM981145 OM981152
M. sorghi CBS 691.96 Cuba KP858936 KP859000 KP859063 KP859109
Microdochium sp. SAUCC1017 China MT199879 MT199885 MT435654 –
M. tainanense CBS 269.76T Taiwan KP858945 KP859009 KP859072 KP859118
CBS 270.76 Taiwan KP858931 KP858995 KP859058 KP859104
M. trichocladiopsis CBS 623.77T Unknown country KP858934 KP858998 KP859061 KP859107
M. triticicola RR 241 UK – AJ748691 – –
M. chrysopogonis GDMCC 3.683 China MT988024 MT988022 MW002441 MW002444
LNU-196 China MT988023 MT988020 MW002442 MW002445
LNU-197 China MT988025 MT988021 MW002443 MW002446
M. yunnanense SAUCC1018 China MT199880 MT199886 MT435655 –
SAUCC1015 China MT199877 MT199883 MT435652 MT510549
SAUCC1012 China MT199876 MT199882 – MT510548
SAUCC1011T China MT199875 MT199881 MT435650 MT510547
Thamnomyces dendroidea CBS 123578 France KY610467 FN428831 KY624313 KY624232

Note: “T” denotes ex-type strain. Newly-generated sequences are indicated in bold. “-” means no data available in GenBank.

Phylogenetic analyses

The sequences of the strains from C. zizanioides and those of Microdochium species, as well as the outgroup Idriella lunata obtained from NCBI GenBank, were aligned with MAFFT version 7 using the default settings. Manual adjustments were made to optimise the alignment in MEGA version 7.0 (Katoh and Standley 2013; Kumar et al. 2016). To elucidate the taxonomic phylogenetic relationships, single and concatenated ITS, LSU, rpb2 and tub2 sequence alignments were subjected to analysis by applying Bayesian Inference (BI) using MrBayes version 3.2.5 and Maximum Likelihood (ML) using RAxML on the CIPRES portal (www.phylo.org) (Swofford 2002; Crous et al. 2006; Ronquist et al. 2012). For BI analysis, the best evolutionary model was determined through the utilisation of MrModelTest version 2.2 (Nylander 2004). Subsequently, in MrBayes v. 3.2.5, the Markov Chain Monte Carlo 180 (MCMC) algorithm was used to generate phylogenetic trees. The first 25% of saved trees were discarded as the burn-in phase. Posterior probabilities (PPs) were determined from the remaining trees prior to calculation of the 50% majority rule consensus trees; PP values exceeding 0.90 were considered significant. The ML analyses were performed by using RAxML-HPC BlackBox version 8.2.6, based on 1000 bootstrap replicates. A general time reversible (GTR) model was applied with gamma-distributed rate variation. Bootstrap values (BSs) equal to or higher than 70% were regarded as significant. The phylogenetic tree was viewed in FigTree version 1.4.4 (Rambaut 2018) and edited by Adobe Illustrator CC2018.

Pathogenicity test

Three isolates of M. chrysopogonis (GDMCC 3.683, LNU-196 and LNU-197) were used to conduct the pathogenicity test. C. zizanioides plants were cultivated within a greenhouse, utilising plastic pots containing field-collected soil from the location where the plants had been established. The isolates were cultured on PDA for 2 weeks at 30 °C in the dark to collect conidia.

For the detached leaf assay, 1-cm wide leaves were harvested from 2-month-old plants cultivated in a greenhouse, washed under running tap water, surface disinfected with 70% ethanol for 1 min, rinsed with sterile water for 30 seconds and finally air-dried on sterilised filter paper. The conidial suspension was adjusted to a concentration of 1 × 106 conidia/ml in sterile distilled water. An equivalent volume of sterile distilled water was used as a control. Leaf blades were then wounded with a sterilised pin and each leaf was sprayed with 2 ml of conidial suspension. All inoculated and control leaves were placed in a moist chamber at 25 °C with 100% relative humidity (RH) under cool fluorescent light with a 12-h photoperiod. After seven days, the disease incidence was assessed and calculated as the percentage of leaves with leaf tar spot symptoms. Each treatment consisted of five replicates and the experiment was conducted three times.

For the attached leaf assay, the leaf blades of healthy leaves were also pin-pricked and the conidial suspension was adjusted to a concentration of 2 × 106 conidia/ml in sterile distilled water. An equivalent volume of sterile distilled water was used as a control. In each treatment, five plants were included, with each plant being sprayed with approximately 20 ml of inoculum. All sprayed and control plants were incubated in a plastic container in a greenhouse at 25 ± 2 °C under cool fluorescent light with a 12-h photoperiod. For the first 3 days, the plastic container was covered with transparent polyethylene bags to maintain a high humidity. The disease incidence was assessed 10 days post inoculation and calculated as the percentage of plants displaying tar spot symptoms. Each treatment had three replicates and the pathogenicity test was repeated twice.

To fulfil Koch’s postulates, symptomatic leaf tissues were subjected to surface sterilisation as described above. Subsequently, these tissues were plated on to PDA medium to enable the re-isolation of the fungi. These isolates were identified, based on comparison of the cultures with those of the original strains. Furthermore, the identifications were confirmed by sequencing of the isolates.

Effect of temperature on mycelial growth rate

Mycelial growth rates of M. chrysopogonis isolates were assessed across various temperatures. Mycelial plugs with a diameter of 5 mm were excised using a sterile hole puncher from the periphery of 10-day-old PDA cultures. Subsequently, they were translocated to the central area of 90 mm PDA Petri dishes. The cultures were subjected to incubation across a temperature range of 5, 10, 15, 20, 25, 30, 35, 40 and 45 °C. Four replicate plates per isolate were prepared for each temperature. The plates were enveloped using Parafilm (Bemis Company, Neenah, WI, U.S.A.) and then positioned within plastic containers prior to their placement in incubators. The colony diameter was measured along two mutually perpendicular axes and the mean of these two measurements was documented as the radial colony diameter. Following a 10-day duration, mycelial growth rates were determined, based on colony diameter and subsequently quantified in millimetres per day. Each treatment was replicated four times.

Fungicide sensitivity

To determine possible control measures for this pathogen in the field, six groups including nine fungicides were tested for their ability to inhibit the growth of M. chrysopogonis in vitro. Fungicide sensitivity assays were conducted, based on methods developed by Yin et al. (2019). The commercial formulations of fungicides were serially diluted using sterilised distilled water. These diluted solutions were added to autoclaved PDA medium that had been cooled to 55 °C to obtain the desired concentrations in micrograms per millilitre (Table 2). Isolates were cultured on PDA plates at 30 °C for 10 days in darkness to supply inoculum. Mycelial discs (5 mm in diameter) from the periphery of colonies actively growing on PDA were positioned at the centre of the fungicide-amended plates and unamended (control) plates. The plates were then incubated at a temperature of 30 °C in darkness for a duration of 2 weeks. Subsequently, the diameter of each colony was measured along two perpendicular axes and the mean diameter was recalibrated by deducting the diameter of the original plug utilised for inoculation. The effective concentration for 50% mycelial growth inhibition (EC50) was estimated by performing a regression analysis of the percentage of mycelial growth inhibition against the log10 of fungicide concentrations. Each treatment was replicated four times.

Table 2.
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List of the fungicides used in this study.

Active ingredient Chemical family Trade name FRAC code Concentration (µg/ml)
Pyrimethanil anilino-pyrimidines Syngenta 9 0, 0.01, 0.1, 1, 10, 100
Difenoconazole triazoles Syngenta 3 0, 0.01, 0.1, 1, 10, 100
Fludioxonil phenylpyrroles Syngenta 12 0, 1, 10, 30, 100, 300
Iprodione dicarboximides Syngenta 2 0, 1, 10, 30, 100, 300
Flusilazole triazoles Syngenta 3 0, 0.001, 0.01, 0.1, 1, 10
Propiconazole triazoles BASF 3 0, 0.0016, 0.008, 0.04, 0.2, 1
Carbendazim benzimidazoles Syngenta 1 0, 0.0016, 0.008, 0.04, 0.2, 1
Metalaxyl acylalanines BASF 4 0, 10, 30, 100, 300, 1000
Tebuconazole triazoles Bayer 3 0, 0.0016, 0.008, 0.04, 0.2, 1

Statistical analysis

The dataset was tested for variance homogeneity using the Levene test. If the variances were equal, an analysis of variance (ANOVA) followed by a least significant difference (LSD) test was conducted. In cases where the variances were unequal, the Dunnett T3 test was applied. All statistical analyses were carried out using IBM SPSS version 20.0 (SPSS Inc., Chicago, IL, U.S.A.). The significance threshold for detecting treatment disparities was set at P < 0.05.

Results

Disease symptoms and isolation of the pathogen

From 2019 to 2022, a previously unknown disease of vetiver grass occurred during late spring and early autumn at the Grass Research Station of Lingnan Normal University (LNU) in Guangdong Province, China. Symptoms consistently appeared on 85% of C. zizanioides grown under field conditions. The initial symptoms appeared as small and scattered punctate spots (< 1 cm) embedded within the leaf tissue. Gradually, these spots clustered on leaf surfaces. Subsequently, brown, elliptical, fish-eye necrotic haloes emerged, encircling the lesion spots and aligning parallel to the leaf veins (Fig. 1A–C). As these necrotic haloes coalesced, the leaf underwent chlorosis and wilting, eventually leading to blighting of the entire plant. Ascomata were visible on the diseased leaf surfaces.

Figure 1. 

Disease symptoms and morphological characters of Microdochium chrysopogonis on infected leaf tissue (CAF 800053) A–C tar spot symptoms of Chrysopogon zizanioides from natural infection in the field D appearance of immersed ascomata on infected leaves E ascomata in longitudinal section F–H asci I–N ascospores. Scale bars: 100 μm (D, E); 10 μm (F–H).

A total of 67 isolates were obtained on PDA. As the colony morphology of the isolates was consistent, three representative isolates (GDMCC 3.683, LNU-196 and LNU-197), one from each field, were selected for further studies.

Phylogeny

Based on a Megablast search on NCBI’s GenBank nucleotide database, the closest hits for the ITS sequence of strain GDMCC 3.683 were M. dawsoniorum sequences with 98% identity (538/551, MK966337; 532/543, MN492650) and a Microdochium sp. sequence with 97% identity (545/562, FJ536210). The closest hits for LSU sequence of this strain were M. dawsoniorum sequences with 99% identity (868/871, OM333563; 864/867, ON394569) and a M. yunnanense sequence with 99% identity (875/882, MT199880). The closest hits for its rpb2 sequence were M. tainanense sequences with 85% identity (711/841, KP859118 and KP859104) and a M. neoqueenslandicum sequence with 83% identity (698/842, KP859106). The closest hits for tub2 sequence were a M. tainanense sequence with 95% identity (661/697, KP859058), a M. neoqueenslandicum sequence with 95% identity (665/703, KP859060) and a M. colombiense sequence with 95% identity (658/695, KP859062). Therefore, molecular analyses with all available Microdochium species were performed. The alignment of each single locus and concatenated sequence dataset of ITS, LSU, rpb2 and tub2 were used to confirm species resolution in Microdochium.

There were in total 99 aligned sequences, including the outgroup, Thamnomyces dendroidea. A total of 3,033 characters (547 bp from the ITS, 843 bp from LSU, 848 bp from tub2 and 795 bp from rpb2) were included in the phylogenetic analyses. RAxML analysis of the combined dataset yielded a best scoring tree with a final ML optimisation likelihood value of -21, 329.537402 (ln). The matrix had 1,096 distinct alignment patterns with 27.41% undetermined characters or gaps. The tree length was 3.410120. Estimated base frequencies were: A = 0.234382, C = 0.267827, G = 0.258835, T = 0.238956; substitution rates were AC = 1.101009, AG = 4.781387, AT = 1.240884, CG = 0.955029, CT = 6.933148 and GT = 1.000000; gamma distribution shape parameter α = 0.152657. Based on the results of MrModelTest, the SYM + I + gamma for ITS, GTR + I + gamma for LSU and rpb2 and HKY + I + gamma model for tub2 were selected as the best fit models for Bayesian analyses. A total of 47,402 trees were generated by BI, amongst which 11,851 trees were discarded as the burn-in phase and the remaining 35,551 trees were used to calculate the posterior probabilities (PPs). The BI consensus tree confirmed the tree topology obtained with ML. The well-supported clade (1/100) formed by the three strains from C. zizanioides clustered with high support (1/100) with M. dawsoniorum (0.92/97), which was sister to one single-strain clade representing M. ratticaude. This clade clustered with high support (0.92/93) with the clade formed by M. albescens, M. seminicola, M. graminearum, M. shilinense, M. insulare, M. paspali, M. citrinidiscum, M. sorghi, M. tainanense and M. trichocladiopsis strains. The M. neoqueenslandicum clade (1/100) was basal to this clade (Fig. 2). Single gene-based phylogenies are presented in the Suppl. material 1. Nonetheless, these individual gene trees did not yield a conclusive taxonomic classification for the new species, in contrast to the comprehensive resolution achieved through the concatenated sequence analysis.

Figure 2. 

Phylogenetic tree inferred from a Maximum Likelihood analysis, based on a combined alignment of ITS, LSU, tub2 and rpb2 sequences from 99 isolates of Microdochium sp. Bootstrap support values obtained with ML above 70% and Bayesian (BI) posterior probability values above 0.90 are shown at the nodes (BI/ML). The tree was rooted to Thamnomyces dendroidea CBS 123578. Numbers of ex-type strains are emphasised with an asterisk and species are delimited with shaded blocks. Isolates of M. chrysopogonis are indicated with lighter text.

Taxonomy

Based on multilocus phylogenetic analyses, the three strains isolated from C. zizanioides represent a previously unknown species within the genus Microdochium that is closely related to M. dawsoniorum and M. ratticaudae. Morphological data placed the new species in the genus Microdochium. This species is characterised below.

Microdochium chrysopogonis W. Zhang & X. Lu, sp. nov.

MycoBank No: 845624
Figs 1, 3

Etymology

Name refers to Chrysopogon, the host genus from which this fungus was collected.

Description

Sexual morph on infected leaf tissue of the host plant (CAF 800054). Ascomata perithecial, 300–350 μm diam., solitary or in groups, immersed, pale brown to black, subglobose to oval, uniloculate, non-ostiolate. Paraphyses filiform, hyaline, straight or curved, apically free. Asci 50–60 × 10–18, x¯ = 55 × 13 μm (n = 50), hyaline, fasciculate, unitunicate, oblong to narrowly clavate, fusiform, 8 biseriate spores with a short stipe. Ascospores clavate, hyaline, guttulate, 20–22 × 8–11.5, x¯ = 21 × 9 μm (n = 50), aseptate, smooth. Sporodochia salmon-pink, slimy. Conidiophores reduced to conidiogenous cells. Conidiogenous cells with percurrent proliferation, hyaline, smooth, aseptate, ampulliform or obpyriform, 10–23 × 8–11.5, x¯ = 17 × 9.5 μm (n = 50). Conidia fusiform, lunate, curved, solitary, guttulate, variable in length, 0–1-septate, 18–72 × 2–3.5, x¯ = 38.5 × 3 μm (n = 50), apex rounded, base usually flattened. Chlamydospores not observed. Vegetative hyphae on PDA (GDMCC 3.683) superficial and immersed, septate, branched, hyaline, smooth, 1–5.5 μm wide.

Culture characteristics

Colonies on PDA reaching 4.0–4.5 cm within seven days in the dark at 30 °C, flat, white cottony aerial mycelium, dense, saffron rounded sporodochia produced after 3 weeks; reverse saffron. On MEA, sparse white cottony aerial mycelium, orange rounded sporodochia produced; reverse salmon-pink. On OA, periphery with white scarce cottony aerial mycelium, concentric rings of orange rounded sporodochia produced; reverse orange.

Type

China, Guangdong Province, Zhanjiang City, field of the Grass Research Station of Lingnan Normal University (LNU), from a leaf of vetiver grass (Chrysopogon zizanioides) with leaf tar spot disease, September 2019, W. Zhang & X. Lu, holotype CAF 800054, ex-type living strain GDMCC 3.683.

Additional materials examined

China, Guangdong Province, Zhanjiang City, field of the Grass Research Station of Lingnan Normal University (LNU), from a leaf of vetiver grass (C. zizanioides) with leaf tar spot disease, September 2019, W. Zhang & X. Lu, strain LNU-196; China, Guangdong Province, Zhanjiang City, field of the Grass Research Station of Lingnan Normal University (LNU), from a leaf of vetiver grass (C. zizanioides) with leaf tar spot disease, September 2019, W. Zhang & X. Lu, strain LNU-197.

Notes

A multilocus phylogenetic analysis of the ITS, LSU, tub2 and rpb2 loci placed three strains of M. chrysopogonis in a distinct and monophyletic clade (1/100) sister to M. dawsoniorum and M. ratticaudae. Notably, M. chrysopogonis has longer conidia (18–72 × 2–3.5 μm) than M. ratticaudae (7–11 × 1.5–2.5 μm) and wider conidia than M. dawsoniorum (25–75 × 1–2 μm). Furthermore, the conidia of M. chrysopogonis are guttulate and 0–1-septate, while those of M. dawsoniorum are 0–3-septate and those of M. ratticaudae are aseptate. The conidiogenous cells of M. chrysopogonis appear as percurrent, ampulliform or obpyriform, whereas those of M. ratticaudae are indistinct from the hyphae and those of M. dawsoniorum are cylindrical to irregular and flexuous. Additionally, the conidiogenous cells of M. chrysopogonis (10–23 × 8–11.5 μm) are wider than those of M. ratticaudae (20–30 × 1–2 μm) (Table 3). Differences are also evident in the sexual morph of these three species. In particular, the sexual morph is not observed in M. dawsoniorum. Ascomata size varies, with that of M. ratticaudae (100–160 μm) being smaller than that of M. chrysopogonis (300–350 μm). Ascospores of M. ratticaudae (14–24 × 4–7 μm) are fusoid to navicular, while those of M. chrysopogonis are clavate, guttulate and wider (20–22 × 8–11.5 μm). In addition, M. ratticaudae features abundant, pale to olivaceous brown, subglobose or cylindrical chlamydospores, while these are not observed in M. chrysopogonis (Crous et al. 2020, 2021a; Table 3). Consequently, based on both morphological characteristics and phylogenetic analyses, all three isolates of M. chrysopogonis were proposed as a new species.

Figure 3. 

Microdochium chrysopogonis (from ex-type: GDMCC 3.683) A colonies after 7 days on PDA B colonies after 7 days on MEA C colonies after 7 days on OA D colony overview of the sporodochia on PDA in culture after incubation for three weeks E aggregated conidiophores F conidiophores with conidiogenous cells G, H conidia. Scale bars: 20 μm (B).

Table 3.
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Morphological characters of Microdochium chrysopogonis and its related species.

Taxa M. albescens M. citrinidiscum M. neoqueenslandicum M. paspali M. seminicola M. trichocladiopsis M. tainanense M. sorghi M. dawsoniorum M. ratticaudae M. graminearum M. shilinense M. chrysopogonis
Asexual morph Conidia Shape falcate, slightly to strongly curved, apex pointed cylindrical, clavate, obovoid lunate, allantoid, curved falcate, apex pointed cylindrical to fusiform, straight or curved oblong, fusiform to obovoid, straight or curved lunate filiform, narrowly acicular fusiform, obclavate flexuous to falcate, sometimes with a geniculation, acute at the tip, narrow at the base fusoid, falcate, acute at the apex and narrowed at the base n/a n/a fusiform, lunate, curved, guttulate
Size (μm) 11–16 × 3.5–4.5 7–31 × 2–3 4–9 × 1.5–3 7–20.5 × 2.5–4.5 19–54 × 3–4.5 6–18 × 2–3.5 10–15 × 2–3 20–90 × 1.5–4.5 25–75 × 1–2 7–11 × 1.5–2.5 n/a n/a 18–72 × 2–3.5
Septa 0–1(–3) 0–3 0(–1) 0–3 (0–)3(–5) 0(–1) 0–1 1–7(–10) 0–3 aseptate n/a n/a 0–1
Conidiogenous cells Shape subcylindrical, doliiform to obpyriform denticulate, cylindrical ampulliform, lageniform to subcylindrical ampulliform, lageniform to cylindrical ampulliform to lageniform cylindrical to clavate, straight but often curved at the tip sympodial, apical, cylindrical or ampulliform with conspicuous rhachides sympodial, ovoid, ampulliform to obclavate cylindrical to irregular, flexuous, narrowed towards the tip indistinct from hyphae, terminal, solitary. n/a n/a ampulliform or obpyriform
Size (μm) 6–15 × 1.5–4 11–29 × 1.5–2 4.5–10 × 2–3.5 6.5–15.5 × 2.5–4 7–9.5 × 3–4 4–37 × 2–3 3–10 × 1–3 5–13 × 3–4 20–30 × 1–2 n/a n/a 16.3–22.4 × 4.1–5.7 10–23 × 8–11.5
Sexual morph Chlamydospores Shape n/a n/a n/a n/a n/a present n/a n/a n/a subglobose or cylindrical n/a n/a n/a
Perithecia Size (μm) 150–180 × 90–120 n/a n/a n/a 110–149 n/a n/a n/a n/a 100–160 n/a n/a 300–350
Asci Size (μm) 40–85 × 8–12 n/a n/a n/a 41–66 × 7.6–11 n/a n/a n/a n/a 50–75 × 10–14 55–77.5 × 9.5–15. 50–76 × 7–10 50–60 × 10–18
Ascospores Size (μm) 14–23 × 3.5–4.5 n/a n/a n/a 12–22 × 3–4.5 n/a n/a n/a n/a 14–24 × 4–7 16.5–24 × 4–5.5 14–18 × 3–5.5 20–22 × 8–11.5
Septa 1–3(–5) n/a n/a n/a 0–3 n/a n/a n/a n/a aseptate 0–3 0–3 aseptate
References Hernández-Restrepo et al. (2016) Hernández-Restrepo et al. (2016) Hernández-Restrepo et al. (2016) Zhang et al. (2015) (Continued on next page) Hernández-Restrepo et al. (2016) Hernández-Restrepo et al. (2016) De Hoog & Hermanides-Nijhof (1977) Braun (1995) Crous et al. (2020)
(Continued on next page)		 Crous et al. (2021a) Gao et al. (2022) Gao et al. (2022) This study

Note: “n/a” means not provided in the literature.

Pathogenicity test

The symptoms observed on leaves of C. zizanioides after inoculation with the representative isolate GDMCC 3.683 were similar to those observed in the field. No symptoms were observed on the leaves of the negative controls (Fig. 4). The average disease incidence of detached leaves that were wounded and sprayed with the isolates GDMCC 3.683, LNU-196 and LNU-197 was 93.3%, 80.0% and 93.3%, respectively. The average disease incidence of whole plants after spraying with the same isolates was 76.7%, 73.3% and 73.3%, respectively (Fig. 5). Koch’s postulates were fulfilled by successful re-isolation of the fungal strains from all leaf spot tissues inoculated with the three isolates. The morphology and DNA sequences of the isolates re-isolated from the inoculated tissues were consistent with those of the strains used for inoculations.

Figure 4. 

Tar spot symptoms of Chrysopogon zizanioides 7 days after spraying on detached leaves (A, B) and 10 days after spraying on leaves attached to whole plants (C, D) with Microdochium chrysopogonis isolate GDMCC 3.683 (B, D) and sterilised water (A, C).

Figure 5. 

Disease incidence of tar spot symptoms on Chrysopogon zizanioides for leaves 7 days after spraying detached leaves (A) and for whole plants 10 days after spraying leaves attached to potted plants (B), respectively, with Microdochium chrysopogonis isolates GDMCC 3.683, LNU-196 and LNU-197. Values are shown as the means, with the error bars representing the standard error. For each pathogen, columns with the same letter indicate means that are not significantly different according to a least significant difference (LSD) test (P < 0.05).

Effect of temperature on mycelial growth

The mycelial growth of M. chrysopogonis was significantly affected by temperature (P < 0.01). All three isolates of M. chrysopogonis grew between 10 and 40 °C, with maximum growth observed at 30 °C (Fig. 6). No isolates grew at 5 or 45 °C after 3 days. The highest average mycelial growth rate was observed at 30 °C (26.5 ± 2.0 mm/day), followed by 25 °C (20.1 ± 4.7 mm/day).

Figure 6. 

Colony growth rate of three isolates, GDMCC 3.683, LNU-196 and LNU-197, of Microdochium chrysopogonis from Chrysopogon zizanioides under different temperatures. Error bars represent the standard error.

Fungicide sensitivity

The EC50 values of various fungicides were analysed for their effectiveness against M. chrysopogonis isolates. A total of 17 isolates of M. chrysopogonis were collected from diseased leaves spanning the period from 2019 to 2022.

The frequency distribution showed that difenoconazole, fludioxonil, flusilazole, carbendazim and iprodione exhibited distributions resembling normal curves, while pyrimethanil, propiconazole, metalaxyl and tebuconazole displayed unimodal curves (Fig. 7). EC50 values for the inhibition of 17 M. chrysopogonis isolates, based on mycelial radial growth, varied across fungicide treatments (P < 0.05) (Table 4). Amongst the tested fungicides, flusilazole had the lowest EC50 values, with a notably concentrated response range of 0.001 to 0.007 µg/ml and an average of 0.004 µg/ml. Tebuconazole closely followed with a slightly narrower range, exhibiting values ranging from 0.002 to 0.009 µg/ml and an average of 0.007 µg/ml. Furthermore, there was no significant difference between flusilazole and tebuconazole. Propiconazole displayed EC50 values spanning from 0.006 to 0.016 µg/ml, with an average of 0.011 µg/ml, while those of carbendazim ranged from 0.008 to 0.031 µg/ml, with an average of 0.024 µg/ml. In contrast, those of difenoconazole exhibited a broader range, varying from 0.013 to 0.127 µg/ml, with a mean value of 0.077 µg/ml, while those of pyrimethanil ranged from 0.054 to 0.605 µg/ml, with an average of 0.411 µg/ml. Those of iprodione, on the other hand, spanned from 15.018 to 260.335 µg/ml, with an average of 193.031 µg/ml. Metalaxyl exhibited the highest EC50 value, displaying the widest range amongst all fungicides, extending from 302.785 to 1056.896 µg/ml with an average of 892.677 µg/ml. Overall, these findings indicate varying degrees of sensitivity to different fungicides amongst M. chrysopogonis isolates. These variations in sensitivity could be essential considerations for designing effective fungicide application strategies against vetiver leaf tar spot disease.

Figure 7. 

Frequency distribution of the 50% effective concentration (EC50) values of six groups including nine fungicides for Microdochium chrysopogonis isolates, based on mycelial growth from 2019 to 2022.

Table 4.
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In vitro sensitivity ranges and mean 50% effective concentration (EC50) values for the inhibition of Microdochium chrysopogonis.

Fungicide EC50 (μg/ml)
Lowest Highest Mean ± SE
Difenoconazole 0.013 0.127 0.077 ± 0.039e
Pyrimethanil 0.054 0.605 0.411 ± 0.180d
Fludioxonil 0.014 6.128 4.525 ± 1.626c
Iprodione 15.018 260.335 193.031 ± 99.462b
Flusilazole 0.001 0.007 0.004 ± 0.003h
Propiconazole 0.006 0.016 0.011 ± 0.003g
Carbendazim 0.008 0.031 0.024 ± 0.009f
Metalaxyl 302.785 1056.896 892.677 ± 236.145a
Tebuconazole 0.002 0.009 0.007 ± 0.002h

Note: The letters indicate the comparison amongst the different fungicide treatments. Means followed by the same letter do not differ according to a post hoc Dunnett T3 test (p < 0.05).

The inhibition of mycelial growth revealed that all nine fungicides exhibited a reduction in fungal growth in vitro when compared to plates without amendments. The effectiveness of these fungicides in diminishing the mycelial growth of the isolates was contingent upon both the specific chemical compound and its concentration. Four DMI fungicides, namely, difenoconazole, propiconazole, flusilazole and tebuconazole and one MBC fungicide, carbendazim, displayed strong activity against M. chrysopogonis growth at concentrations below 1 µg/ml, specifically at concentrations of 1, 0.2, 0.1, 0.2 and 0.2 µg/ml, respectively (Fig. 8). However, M. chrysopogonis showed a tendency to exhibit better growth in the presence of pyrimethanil, fludioxonil, iprodione and metalaxyl, with mycelial growth being completely inhibited at concentrations exceeding 100 µg/ml (Fig. 8).

Figure 8. 

Effect of fungicides on the mycelial growth of Microdochium chrysopogonis. Values are shown as the means, with the error bars representing the standard error.

Discussion

In a survey of disease on C. zizanioides in Guangdong Province, China, from 2019 to 2022, tar spot was the predominant leaf spot disease. Isolation, morphological features, multilocus phylogenetic analysis and pathogenicity tests confirmed that a new Microdochium species, M. chrysopogonis was the causal agent. To effectively control the disease, the sensitivity of M. chrysopogonis to six groups of fungicides, including nine fungicides was determined. Results indicated that four DMI fungicides, namely difenoconazole, propiconazole, flusilazole and tebuconazole and one MBC fungicide, carbendazim, were highly effective against the new species.

The morphology of the new species is introduced along with its sexual and asexual morphological features, which are consistent with the following of Microdochium: pale brown to black, subglobose to oval, uniloculate, perithecial ascomata; hyaline, fasciculate, unitunicate, oblong to narrowly clavate, eight biseriate spores with short stipe asci, from which hyaline, clavate, smooth ascospores arise. Conidiophores reduced to hyaline, smooth, aseptate, percurrent, ampulliform or obpyriform, conidiogenous cells, from which hyaline, 0–1-septate, fusiform, lunate conidia with the apex rounded and base flattened usually arise (Figs 1, 3) (Hernández-Restrepo et al. 2016). The concatenated ITS, LSU, tub2 and rpb2 sequences were able to identify species in Microdochium and proved to be suitable barcoding markers in the process of species resolution (Hernández-Restrepo et al. 2016). Phylogenetic analysis indicated that M. chrysopogonis formed a distinct well-supported clade (1/100) and was closely related to M. dawsoniorum and M. ratticaudae (Fig. 2). Nevertheless, the classification of the new species in the genus Microdochium is well supported by morphology, based on sexual and asexual morphs, which are different from those of M. dawsoniorum and M. ratticaudae.

Temperature is a major factor affecting plant disease epidemics. In recent years, tar spot disease of C. zizanioides has become increasingly prevalent in Guangdong Province, China, especially in hot and rainy summers. Thus, the effect of temperature on the growth rate of M. chrysopogonis in vitro was evaluated in this study. There were no significant differences in the minimum and optimum growth temperatures amongst the three isolates and the optimum growth temperature was 30 °C (Fig. 6). Research revealed that the highest growth rate of M. paspali occurred at 25–28 °C and M. majus, M. seminicola and M. nivale strains in Russia and Europe grew optimally at 20–25 °C (Doohan et al. 2003; Gagkaeva et al. 2020), while M. nivale from Slovakia grew better at temperatures below 20 °C (Hudec and Muchová 2010). Thus, the optimum growth temperature varies amongst Microdochium species.

A previous study showed that P. herbarum could initially induce leaf spots and blight on vetiver grass, causing round or irregular dark brown spots, which are similar to the symptoms on M. chrysopogonis (Zhang et al. 2017). However, the symptoms on M. chrysopogonis were different from those on P. herbarum in the later period. Specifically, P. herbarum caused fusiform or irregular with reddish-brown margins on the host plant, whereas M. chrysopogonis caused fish-eye necrotic haloes surrounding the spot lesions on leaves. Additionally, the disease incidences were different. P. herbarum affected 26% to 42% of plants, while M. chrysopogonis showed a 100% disease incidence. Given the high disease incidence associated with M. chrysopogonis and its induction of leaf spots on vetiver grass, as well as the identification of this new species, it is imperative to conduct future studies addressing the host spectrum, epidemic conditions, biological characteristics and distribution patterns of M. chrysopogonis.

The effectiveness of biofungicides, such as bacterial seed treatments using Pseudomonas and Pantoea in controlling diseases caused by Microdochium, has been established (Johansson et al. 2003). However, there remains substantial reliance on registered chemical fungicides. Currently, research on fungicide sensitivity within Microdochium mainly focuses on three species: M. panattonianum, M. majus and M. nivale. Six groups of fungicides, namely, MBCs, DMIs, QoIs, SDHIs, PPs, and dicarboximides, have been shown to have significant inhibitory activity (Kaneko and Ishii 2009; Aamlid et al. 2017, 2018; Matušinsky et al. 2017; Gagkaeva et al. 2022). In this study, consistent with previous findings, four DMI fungicides (difenoconazole, propiconazole, flusilazole and tebuconazole) and one MBC fungicide (carbendazim) exhibited significant inhibitory effects on the growth of M. chrysopogonis, with mean EC50 values of 0.077, 0.011, 0.004, 0.024 and 0.007 μg/ml, respectively (Table 4). However, dicarboximides (iprodione), which were effective against snow mould and Microdochium patch caused by M. nivale on turf-grass in previous studies (Gourlie and Hsiang 2017), showed ineffectiveness in this study, with a mean EC50 value of 193.031 μg/ml. Additionally, while the PP fungicide fludioxonil demonstrated good antifungal activity against M. majus in other research (Mao et al. 2023), the isolates in this study displayed only moderate sensitivity to fludioxonil, with an EC50 value of 4.525 μg/ml and complete inhibition of mycelial growth required concentrations exceeding 100 µg/ml (Table 4, Fig. 8). These variations in fungicide sensitivity could be attributed to genetic structural changes, introducing bias in chemical control efficacy (Matušinsky et al. 2019). Furthermore, the response of the same pathogen to fungicides can vary amongst regions. For example, the DMI fungicides, tebuconazole and metconazole, were reported to be ineffective against M. nivale in the Czech Republic and France (Ioos et al. 2005; Matušinsky et al. 2019). Similarly, M. nivale exhibited sensitivity to SDHI fungicides, including pydiflumetofen, fluxapyroxad and penthiopyrad, in vitro, but these fungicides were ineffective in providing acceptable control under field conditions in the USA (Hockemeyer and Koch 2022). These differences may be attributed to variations in environmental factors, such as temperature and humidity, as well as diverse biological characteristics, including epidemiology, fungicide sensitivity and aggressive nature of the pathogen (Abdelhalim et al. 2020). Overall, this study offers valuable insights into fungicide application strategies for effectively managing the disease. Further research is needed to analyse the influences of environmental variables and conduct field trials to validate the effects of DMI fungicides, ultimately enhancing the ability to successfully manage vetiver leaf tar spot disease.

Acknowledgements

We thank the Herbarium of the Chinese Academy of Forestry for helping with the preservation of plant specimens.

Additional information

Conflict of interest

The authors have declared that no competing interests exist.

Ethical statement

No ethical statement was reported.

Funding

This research was financially supported by the Guangdong Basic and Applied Basic Research Foundation (2020A1515110167), School-level Talents Project of Lingnan Normal University (ZL 2034), Natural Science Foundation of Guangdong Province, China (2023A1515011676) and Key Scientific Research Platform and Project of Guangdong Education Department (2021KCXTD054)

Author contributions

Xiang Lu, Xue-Li Niu and Wu Zhang carried out the investigation and sampling. Xiang Lu and Wu Zhang conducted the morphological and phylogenetic analysis. Xiang Lu and Wu Zhang carried out the pathogenicity test. Xiang Lu, Meng-Xian Mai, Wen-Hui Tan, Mu-Yan Zhang, Jie Xie and Yi Lu undertook the fungicide sensitivity experiment. Xiang Lu wrote, edited and reviewed the manuscript. Xiang Lu and Wu Zhang reviewed the manuscript and provided funding. All authors have read and agreed to the published version of the manuscript.

Author ORCIDs

Xiang Lu https://orcid.org/0000-0001-9582-1319

Mengxian Mai https://orcid.org/0009-0001-3824-2895

Wenhui Tan https://orcid.org/0009-0008-6054-2174

Muyan Zhang https://orcid.org/0009-0005-8880-9780

Jie Xie https://orcid.org/0009-0002-4200-0140

Yi Lu https://orcid.org/0009-0003-0848-2390

Data availability

All of the data that support the findings of this study are available in the main text or Supplementary Information.

References

  • Aamlid TS, Niemeläinen O, Paaske K, Widmark D, Ruuttunen P, Kedonperä A (2018) Evaluation of a petroleum-derived spray oil for control of Microdochium patch and turfgrass spring performance on Nordic golf greens. Agronomy Journal 110(6): 2189–2197. https://doi.org/10.2134/agronj2018.07.0475
  • Abdelhalim M, Brurberg MB, Hofgaard IS, Rognli OA, Tronsmo AM (2020) Pathogenicity, host specificity and genetic diversity in Norwegian isolates of Microdochium nivale and Microdochium majus. European Journal of Plant Pathology 156(3): 885–895. https://doi.org/10.1007/s10658-020-01939-5
  • Alkan M, Özer G, İmren M, Özdemir F, Morgounov A, Dababat AA (2021) First report of Fusarium culmorum and Microdochium bolleyi causing root rot on Triticale in Kazakhstan. Plant Disease 105(7): е2015. https://doi.org/10.1094/PDIS-12-20-2659-PDN
  • Babu S, Rameshkumar S, Prakash M (2021) Vetiver-a Blessing to Coastal Ecosystem for an Integral Prosperity and Ecological Stability. Coastal Agriculture and Climate Change. CRC Press, Florida, 13 pp. https://doi.org/10.1201/9781003245285-11
  • Braun U (1995) A Monograph of Cercosporella, Ramularia and Allied Genera (Phytopathogenic Hyphomycetes). IHW Verlag, Eching, 2 pp.
  • Crous PW, Slippers B, Wingfield MJ, Rheeder J, Marasas WFO, Philips AJL, Alves A, Burgess T, Barber P, Groenewald JZ (2006) Phylogenetic lineages in the Botryosphaeriaceae. Studies in Mycology 55: 235–253. https://doi.org/10.3114/sim.55.1.235
  • Crous PW, Verkley GJM, Groenewald JZ, Samson RA (2009) Fungal Biodiversity. American Phytopathological Society, St.Paul, 269 pp.
  • Crous PW, Wingfield MJ, Chooi Y-H, Gilchrist CLM, Lacey E, Pitt JI, Roets F, Swart WJ, Cano-Lira JF, Valenzuela-Lopez N, Hubka V, Shivas RG, Stchigel AM, Holdom DG, Jurjević Ž, Kachalkin AV, Lebel T, Lock C, Martín MP, Tan YP, Tomashevskaya MA, Vitelli JS, Baseia IG, Bhatt VK, Brandrud TE, De Souza JT, Dima B, Lacey HJ, Lombard L, Johnston PR, Morte A, Papp V, Rodríguez A, Rodríguez-Andrade E, Semwal KC, Tegart L, Abad ZG, Akulov A, Alvarado P, Alves A, Andrade JP, Arenas F, Asenjo C, Ballarà J, Barrett MD, Berná LM, Berraf-Tebbal A, Bianchinotti MV, Bransgrove K, Burgess TI, Carmo FS, Chávez R, Čmoková A, Dearnaley JDW, Santiago ALCMA, Freitas-Neto JF, Denman S, Douglas B, Dovana F, Eichmeier A, Esteve-Raventós F, Farid A, Fedosova AG, Ferisin G, Ferreira RJ, Ferrer A, Figueiredo CN, Figueiredo YF, Reinoso-Fuentealba CG, Garrido-Benavent I, Cañete-Gibas CF, Gil-Durán C, Glushakova AM, Gonçalves MFM, González M, Gorczak M, Gorton C, Guard FE, Guarnizo AL, Guarro J, Gutiérrez M, Hamal P, Hien LT, Hocking AD, Houbraken J, Hunter GC, Inácio CA, Jourdan M, Kapitonov VI, Kelly L, Khanh TN, Kisło K, Kiss L, Kiyashko A, Kolařík M, Kruse J, Kubátová A, Kučera V, Kučerová I, Kušan I, Lee HB, Levicán G, Lewis A, Liem NV, Liimatainen K, Lim HJ, Lyons MN, Maciá-Vicente JG, Magaña-Dueñas V, Mahiques R, Malysheva EF, Marbach PAS, Marinho P, Matočec N, McTaggart AR, Mešić A, Morin L, Muñoz-Mohedano JM, Navarro-Ródenas A, Nicolli CP, Oliveira RL, Otsing E, Ovrebo CL, Pankratov TA, Paños A, Paz-Conde A, Pérez-Sierra A, Phosri C, Pintos Á, Pošta A, Prencipe S, Rubio E, Saitta A, Sales LS, Sanhueza L, Shuttleworth LA, Smith J, Smith ME, Spadaro D, Spetik M, Sochor M, Sochorová Z, Sousa JO, Suwannasai N, Tedersoo L, Thanh HM, Thao LD, Tkalčec Z, Vaghefi N, Venzhik AS, Verbeken A, Vizzini A, Voyron S, Wainhouse M, Whalley AJS, Wrzosek M, Zapata M, Zeil-Rolfe I, Groenewald JZ (2020) Fungal Planet description sheets: 1042–1111. Persoonia 44(1): 301–459. https://doi.org/10.3767/persoonia.2020.44.11
  • Crous PW, Cowan DA, Maggs-Kölling G, Yilmaz N, Thangavel R, Wingfield MJ, Noordeloos ME, Dima B, Brandrud TE, Jansen GM, Morozova OV, Vila J, Shivas RG, Tan YP, Bishop-Hurley S, Lacey E, Marney TS, Larsson E, Le Floch G, Lombard L, Nodet P, Hubka V, Alvarado P, Berraf-Tebbal A, Reyes JD, Delgado G, Eichmeier A, Jordal JB, Kachalkin AV, Kubátová A, Maciá-Vicente JG, Malysheva EF, Papp V, Rajeshkumar KC, Sharma A, Spetik M, Szabóová D, Tomashevskaya MA, Abad JA, Abad ZG, Alexandrova AV, Anand G, Arenas F, Ashtekar N, Balashov S, Bañares Á, Baroncelli R, Bera I, Biketova AYu, Blomquist CL, Boekhout T, Boertmann D, Bulyonkova TM, Burgess TI, Carnegie AJ, Cobo-Diaz JF, Corriol G, Cunnington JH, da Cruz MO, Damm U, Davoodian N, de A Santiago ALCM, Dearnaley J, de Freitas LWS, Dhileepan K, Dimitrov R, Di Piazza S, Fatima S, Fuljer F, Galera H, Ghosh A, Giraldo A, Glushakova AM, Gorczak M, Gouliamova DE, Gramaje D, Groenewald M, Gunsch CK, Gutiérrez A, Holdom D, Houbraken J, Ismailov AB, Istel Ł, Iturriaga T, Jeppson M, Jurjević Ž, Kalinina LB, Kapitonov VI, Kautmanová I, Khalid AN, Kiran M, Kiss L, Kovács Á, Kurose D, Kušan I, Lad S, Læssøe T, Lee HB, Luangsa-ard JJ, Lynch M, Mahamedi AE, Malysheva VF, Mateos A, Matočec N, Mešić A, Miller AN, Mongkolsamrit S, Moreno G, Morte A, Mostowfizadeh-Ghalamfarsa R, Naseer A, Navarro-Ródenas A, Nguyen TTT, Noisripoom W, Ntandu JE, Nuytinck J, Ostrý V, Pankratov TA, Pawłowska J, Pecenka J, Pham THG, Polhorský A, Pošta A, Raudabaugh DB, Reschke K, Rodríguez A, Romero M, Rooney-Latham S, Roux J, Sandoval-Denis M, Smith MTh, Steinrucken TV, Svetasheva TY, Tkalčec Z, van der Linde EJ, Vegte M, Vauras J, Verbeken A, Visagie CM, Vitelli JS, Volobuev SV, Weill A, Wrzosek M, Zmitrovich IV, Zvyagina EA, Groenewald JZ (2021a) Fungal Planet description sheets: 1182–1283. Persoonia 46: 313–528. https://doi.org/10.3767/persoonia.2021.46.11
  • Crous PW, Osieck ER, Jurjevi Ž, Boers J, Van Iperen AL, Starink-Willemse M, Dima B, Balashov S, Bulgakov TS, Johnston PR, Morozova OV, Pinruan U, Sommai S, Alvarado P, Decock CA, Lebel T, McMullan-Fisher S, Moreno G, Shivas RG, Zhao L, Abdollahzadeh J, Abrinbana M, Ageev DV, Akhmetova G, Alexandrova AV, Altés A, Amaral AGG, Angelini C, Antonín V, Arenas F, Asselman P, Badali F, Baghela A, Bañares A, Barreto RW, Baseia IG, Bellanger J-M, Berraf-Tebbal A, Biketova AYu, Bukharova NV, Burgess TI, Cabero J, Câmara MPS, Cano-Lira JF, Ceryngier P, Chávez R, Cowan DA, de Lima AF, Oliveira RL, Denman S, Dang QN, Dovana F, Duarte IG, Eichmeier A, Erhard A, Esteve-Raventós F, Fellin A, Ferisin G, Ferreira RJ, Ferrer A, Finy P, Gaya E, Geering ADW, Gil-Durán C, Glässnerová K, Glushakova AM, Gramaje D, Guard FE, Guarnizo AL, Haelewaters D, Halling RE, Hill R, Hirooka Y, Hubka V, Iliushin VA, Ivanova DD, Ivanushkina NE, Jangsantear P, Justo A, Kachalkin AV, Kato S, Khamsuntorn P, Kirtsideli IY, Knapp DG, Kochkina GA, Koukol O, Kovács GM, Kruse J, Kumar TKA, Kušan I, Læssøe T, Larsson E, Lebeuf R, Levicán G, Loizides M, Marinho P, Luangsa-ard JJ, Lukina EG, Magaña-Dueñas V, Maggs-Kölling G, Malysheva EF, Malysheva VF, Martín B, Martín MP, Matočec N, McTaggart AR, Mehrabi-Koushki M, Mešić A, Miller AN, Mironova P, Moreau P-A, Morte A, Müller K, Nagy LG, Nanu S, Navarro-Ródenas A, Nel WJ, Nguyen TH, Nóbrega TF, Noordeloos ME, Olariaga I, Overton BE, Ozerskaya SM, Palani P, Pancorbo F, Papp V, Pawłowska J, Pham TQ, Phosri C, Popov ES, Portugal A, Pošta A, Reschke K, Reul M, Ricci GM, Rodríguez A, Romanowski J, Ruchikachorn N, Saar I, Safi A, Sakolrak B, Salzmann F, Sandoval-Denis M, Sangwichein E, Sanhueza L, Sato T, Sastoque A, Senn-Irlet B, Shibata A, Siepe K, Somrithipol S, Spetik M, Sridhar P, Stchigel AM, Stuskova K, Suwannasai N, Tan YP, Thangavel R, Tiago I, Tiwari S, Tkalčec Z, Tomashevskaya MA, Tonegawa C, Tran HX, Tran NT, Trovão J, Trubitsyn VE, Van Wyk J, Vieira WAS, Vila J, Visagie CM, Vizzini A, Volobuev SV, Vu DT, Wangsawat N, Yaguchi T, Ercole E, Ferreira BW, de Souza AP, Vieira BS, Groenewald JZ (2021b) Fungal Planet description sheets: 1284–1382. Persoonia 47: 178–374. https://doi.org/10.3767/persoonia.2021.47.06
  • David A, Wang F, Sun X, Li H, Lin J, Li P, Deng G (2019) Chemical composition, antioxidant, and antimicrobial activities of Vetiveria zizanioides (L.) nash essential oil extracted by carbon dioxide expanded ethanol. Molecules (Basel, Switzerland) 24(10): 1897. https://doi.org/10.3390/molecules24101897
  • Debieu D, Bach J, Arnold A, Brousset S, Gredt M, Taton M, Rahier A, Malosse C, Leroux P (2000) Inhibition of ergosterol biosynthesis by morpholine, piperidine, and spiroketalamine fungicides in Microdochium nivale: Effect on sterol composition and sterol Δ8 → Δ7-isomerase activity. Pesticide Biochemistry and Physiology 67(2): 85–94. https://doi.org/10.1006/pest.2000.2485
  • Dirchwolf PM, Moschini RC, Gutiérrez SA, Carmona MA (2023) Modelling of the effects of environmental factors on rice grain discoloration incidence in Corrientes province, Argentina. Journal of Phytopathology 171(1): 12–22. https://doi.org/10.1111/jph.13150
  • Doohan FM, Brennan J, Cooke BM (2003) Epidemiology of Mycotoxin Producing Fungi. Springer, Dordrecht, 129 pp.
  • Gagkaeva T, Orina A, Gavrilova O, Usoltseva M, Crouch JA, Normann K, Entwistle K, Torp T, Espevig T (2022) In vitro fungicide sensitivity of Clarireedia, Fusarium, and Microdochium isolates from grasses. International Turfgrass Society Research Journal 14(1): 972–980. https://doi.org/10.1002/its2.139
  • Glynn NC, Hare MC, Edwards SG (2008) Fungicide seed treatment efficacy against Microdochium nivale and M. majus in vitro and in vivo. Pest Management Science 64(8): 793–799. https://doi.org/10.1002/ps.1558
  • Gourlie R, Hsiang T (2017) Resistance to dicarboximide fungicides in a Canadian population of Microdochium nivale. International Turfgrass Society Research Journal 13(1): 133–138. https://doi.org/10.2134/itsrj2016.06.0497
  • Hernández-Restrepo M, Groenewald JZ, Crous PW (2016) Taxonomic and phylogenetic re-evaluation of Microdochium, Monographella and Idriella. Persoonia 36(1): 57–82. https://doi.org/10.3767/003158516X688676
  • Hockemeyer K, Koch PL (2022) Field evaluations and in vitro sensitivity of Microdochium nivale to succinate dehydrogenase (SDHI) fungicides. International Turfgrass Society Research Journal 14(1): 951–957. https://doi.org/10.1002/its2.48
  • Hong SK, Kim WG, Choi HW, Lee SY (2008) Identification of Microdochium bolleyi associated with basal rot of creeping bent grass in Korea. Mycobiology 36(2): 77–80. https://doi.org/10.4489/MYCO.2008.36.2.077
  • Hudec K, Muchová D (2010) Influence of temperature and species origin on Fusarium spp. and Microdochium nivale pathogenicity to wheat seedlings. Plant Protection Science 46(2): 59–65. https://doi.org/10.17221/12/2009-PPS
  • Ioos R, Belhadj A, Menez M, Faure A (2005) The effects of fungicides on Fusarium spp. and Microdochium nivale and their associated trichothecene mycotoxins in French naturally-infected cereal grains. Crop Protection 24(10): 894–902. https://doi.org/10.1016/j.cropro.2005.01.014
  • Jia X, Zheng X, Damm U, Lu X, Niu XL, Feng Y, Zhang W (2023 [June 04]) Stenocarpella chrysopogonis, a new species causing leaf streak disease of Chrysopogon zizanioides in southern China. Plant Disease PDIS-02-23-0383-SR. https://doi.org/10.1094/PDIS-02-23-0383-SR
  • Johansson PM, Johnsson L, Gerhardson B (2003) Suppression of wheat-seedling diseases caused by Fusarium culmorum and Microdochium nivale using bacterial seed treatment. Plant Pathology 52(2): 219–227. https://doi.org/10.1046/j.1365-3059.2003.00815.x
  • Kaneko I, Ishii H (2009) Effect of azoxystrobin on activities of antioxidant enzymes and alternative oxidase in wheat head blight pathogens Fusarium graminearum and Microdochium nivale. Journal of General Plant Pathology 75(5): 388–398. https://doi.org/10.1007/s10327-009-0178-9
  • Katoh K, Standley DM (2013) MAFFT multiple sequence alignment software version 7: Improvements in performance and usability. Molecular Biology and Evolution 30(4): 772–780. https://doi.org/10.1093/molbev/mst010
  • Koch PL, Stier JC, Kerns JP (2015) Snow cover has variable effects on persistence of fungicides and their suppression of microdochium patch on amenity turfgrass. Plant Pathology 64(6): 1417–1428. https://doi.org/10.1111/ppa.12379
  • Kumar S, Stecher G, Tamura K (2016) MEGA7: Molecular evolutionary genetics analysis version 7.0 for bigger datasets. Molecular Biology and Evolution 33(7): 1870–1874. https://doi.org/10.1093/molbev/msw054
  • Lin JM, Liu JX, Niu XL, Zhang W (2019) Root and basal stem rot caused by Gaeumannomyces graminicola on Vetiveria zizanioides in southern China. Plant Disease 103(8): e2128. https://doi.org/10.1094/PDIS-12-18-2161-PDN
  • Mao Y, Wu J, Song W, Zhao B, Zhao H, Cai Y, Wang J-X, Zhou M, Duan Y (2023) Occurrence and chemical control strategy of wheat brown foot rot caused by Microdochium majus. Plant Disease 107(11): 3523–3530. https://doi.org/10.1094/PDIS-02-23-0392-RE
  • Matušinsky P, Svačinová I, Jonavičienė A, Tvarůžek L (2017) Long-term dynamics of causative agents of stem base diseases in winter wheat and reaction of Czech Oculimacula spp. and Microdochium spp. populations to prochloraz. European Journal of Plant Pathology 148(1): 199–206. https://doi.org/10.1007/s10658-016-1082-8
  • Matušinsky P, Svobodová LL, Svačinová I, Havis N, Hess M, Tvarůžek L (2019) Population genetic structure of Microdochium majus and Microdochium nivale associated with foot rot of cereals in the Czech Republic and adaptation to penthiopyrad. European Journal of Plant Pathology 155(1): 1–12. https://doi.org/10.1007/s10658-019-01737-8
  • Maurya R, Yadav P, Krishna R, Kulkarni P, Rastogi S, Mohapatra S, Srivastava S, Akhtar MQ, Shukla AK, Chauhan HS, Verma RK, Chanotiya CS, Shasany AK (2023) Transcriptome and metabolome analysis of sesquiterpene diversity in Indian vetiver (Chrysopogon zizanioides L. Roberty). Industrial Crops and Products 200: e116798. https://doi.org/10.1016/j.indcrop.2023.116798
  • Moon UR, Durge AA, Kumar M (2020) Value addition of khus (Vetiveria zizanioides L. Nash) root extracts by elicitor and heat treatment. Industrial Crops and Products 144: e112037. https://doi.org/10.1016/j.indcrop.2019.112037
  • Nylander JAA (2004) MrModeltest 2.0. MrModeltest Version 2. Uppsala University, Uppsala.
  • Polizzi G, Aiello D, Vitale A, Giuffrida F, Groenewald JZ, Crous PW (2009) First report of shoot blight, canker, and gummosis caused by Neoscytalidium dimidiatum on citrus in Italy. Plant Disease 93(11): 1215–1215. https://doi.org/10.1094/PDIS-93-11-1215A
  • Rambaut A (2018) FigTree v.1.4.4. University of Edinburgh, Edinburgh.
  • Rayner RW (1970) A Mycological Colour Chart. Commonwealth Mycological Institute & British Mycological Society, Kew, 34 pp.
  • Ren R, Yang X, Ray RV (2015) Comparative aggressiveness of Microdochium nivale and M. majus and evaluation of screening methods for Fusarium seedling blight resistance in wheat cultivars. European Journal of Plant Pathology 141(2): 281–294. https://doi.org/10.1007/s10658-014-0541-3
  • Ronquist F, Teslenko M, van der Mark P, Ayres DL, Darling A, Höhna S, Larget B, Liu L, Suchard MA, Huelsenbeck JP (2012) MrBayes 3.2: Efficient bayesian phylogenetic inference and model choice across a large model space. Systematic Biology 61(3): 539–542. https://doi.org/10.1093/sysbio/sys029
  • Stewart S, Rodríguez M, Mattos N, Abreo E, Stewart S, Rodríguez M, Mattos N, Abreo E (2019) First report of zonate leaf spot in sorghum caused by Microdochium sorghi in Uruguay. Agrociencia 23(2): 94–98. https://doi.org/10.31285/AGRO.23.74
  • Swofford D (2002) PAUP*. phylogenetic analysis using parsimony (*and other methods). version 4.0b10. Sinauer Associates, Sunderland.
  • Sydow H (1924) Mycotheca germanica. Fasc. XLII–XLV (No. 20512250). Annales Mycologici 22: 257–268.
  • Vilgalys R, Hester M (1990) Rapid genetic identification and mapping of enzymatically amplified ribosomal DNA from several Cryptococcus species. Journal of Bacteriology 172(8): 4238–4246. https://doi.org/10.1128/jb.172.8.4238-4246.1990
  • White TJ, Bruns T, Lee S, Taylor J (1990) Amplification and Direct Sequencing of Fungal Ribosomal RNA Genes for Phylogenetics. PCR Protocols: a Guide to Methods and Applications. Academic Press, San Diego, 482 pp. https://doi.org/10.1016/B978-0-12-372180-8.50042-1
  • Wicks TJ, Hall B, Pezzaniti P (1994) Fungicidal control of anthracnose (Microdochium panattonianum) on lettuce. Australian Journal of Experimental Agriculture 34(2): 277–283. https://doi.org/10.1071/EA9940277
  • Yin LF, Du SF, Chaisiri C, Cheewangkoon R, Luo CX (2019) Phylogenetic analysis and fungicide baseline sensitivities of Monilia mumecola in China. Plant Disease 103(9): 2231–2236. https://doi.org/10.1094/PDIS-11-18-1953-RE
  • Zhang W, Nan Z, Tian P, Hu M, Gao Z, Li M, Liu G (2015) Microdochium paspali, a new species causing seashore paspalum disease in southern China. Mycologia 107(1): 80–89. https://doi.org/10.3852/14-119

Supplementary materials

Supplementary material 1 

Phylogenetic tree inferred from a maximum likelihood analysis based on a combined alignment of ITS sequences of 97 isolates of the Microdochium sp.

Xiang Lu, Mengxian Mai, Wenhui Tan, Muyan Zhang, Jie Xie, Yi Lu, Xue Li Niu, Wu Zhang

Data type: tif

Explanation note: Bootstrap support values obtained with ML above 70% and Bayesian (BI) posterior probability values above 0.90 are shown at the nodes (BI/ML). The designated outgroup taxa are Thamnomyces dendroidea CBS 123578. Numbers of ex-type strains are emphasized with an asterisk and species are delimited with shaded blocks. Isolates of M. chrysopogonis are indicated with lighter text.

This dataset is made available under the Open Database License (http://opendatacommons.org/licenses/odbl/1.0/). The Open Database License (ODbL) is a license agreement intended to allow users to freely share, modify, and use this Dataset while maintaining this same freedom for others, provided that the original source and author(s) are credited.
Download file (28.70 MB)
Supplementary material 2 

Phylogenetic tree inferred from a maximum likelihood analysis based on a combined alignment of LSU sequences of 72 isolates of the Microdochium sp.

Xiang Lu, Mengxian Mai, Wenhui Tan, Muyan Zhang, Jie Xie, Yi Lu, Xue Li Niu, Wu Zhang

Data type: tif

Explanation note: Bootstrap support values obtained with ML above 70% and Bayesian (BI) posterior probability values above 0.90 are shown at the nodes (BI/ML). The designated outgroup taxa are Thamnomyces dendroidea CBS 123578. Numbers of ex-type strains are emphasized with an asterisk and species are delimited with shaded blocks. Isolates of M. chrysopogonis are indicated with lighter text.

This dataset is made available under the Open Database License (http://opendatacommons.org/licenses/odbl/1.0/). The Open Database License (ODbL) is a license agreement intended to allow users to freely share, modify, and use this Dataset while maintaining this same freedom for others, provided that the original source and author(s) are credited.
Download file (28.25 MB)
Supplementary material 3 

Phylogenetic tree inferred from a maximum likelihood analysis based on a combined alignment of rpb2 sequences of 71 isolates of the Microdochium sp.

Xiang Lu, Mengxian Mai, Wenhui Tan, Muyan Zhang, Jie Xie, Yi Lu, Xue Li Niu, Wu Zhang

Data type: tif

Explanation note: Bootstrap support values obtained with ML above 70% and Bayesian (BI) posterior probability values above 0.90 are shown at the nodes (BI/ML). The designated outgroup taxa are Thamnomyces dendroidea CBS 123578. Numbers of ex-type strains are emphasized with an asterisk and species are delimited with shaded blocks. Isolates of M. chrysopogonis are indicated with lighter text.

This dataset is made available under the Open Database License (http://opendatacommons.org/licenses/odbl/1.0/). The Open Database License (ODbL) is a license agreement intended to allow users to freely share, modify, and use this Dataset while maintaining this same freedom for others, provided that the original source and author(s) are credited.
Download file (20.65 MB)
Supplementary material 4 

Phylogenetic tree inferred from a maximum likelihood analysis based on a combined alignment of tub2 sequences of 80 isolates of the Microdochium sp.

Xiang Lu, Mengxian Mai, Wenhui Tan, Muyan Zhang, Jie Xie, Yi Lu, Xue Li Niu, Wu Zhang

Data type: tif

Explanation note: Bootstrap support values obtained with ML above 70% and Bayesian (BI) posterior probability values above 0.90 are shown at the nodes (BI/ML). The designated outgroup taxa are Thamnomyces dendroidea CBS 123578. Numbers of ex-type strains are emphasized with an asterisk and species are delimited with shaded blocks. Isolates of M. chrysopogonis are indicated with lighter text.

This dataset is made available under the Open Database License (http://opendatacommons.org/licenses/odbl/1.0/). The Open Database License (ODbL) is a license agreement intended to allow users to freely share, modify, and use this Dataset while maintaining this same freedom for others, provided that the original source and author(s) are credited.
Download file (21.81 MB)
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