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. 2024 Mar 8;103:1–24. doi: 10.3897/mycokeys.103.116153

Molecular phylogeny and morphology reveal two new entomopathogenic species of Ophiocordyceps (Ophiocordycipitaceae, Hypocreales) parasitic on termites from China

Qi Fan 1,2, Tao Yang 2,3, Hui Li 2, Xue-Mei Wang 2,3, He-Fa Liao 2,3, Pei-Hong Shen 1, Zhu-Liang Yang 2, Wen-Bo Zeng 4,✉, Yuan-Bing Wang 2,✉
PMCID: PMC10943269  PMID: 38495949

Abstract

Two new termite-pathogenic species, Ophiocordycepsglobiperitheciata and O.longistipes, are described from Yunnan Province, China. Six-locus (ITS, nrSSU, nrLSU, tef-1α, rpb1 and rpb2) phylogenetic analyses in combination with morphological observations were employed to characterize these two species. Phylogenetically, O.globiperitheciata is most closely related to Hirsutellacryptosclerotium and O.communis, whereas O.longistipes shares a sister relationship with O.fusiformis. However, O.globiperitheciata differs from H.cryptosclerotium by parasitizing Blattodea and producing clavate, unbifurcated stromata. Ophiocordycepsglobiperitheciata is distinguished from O.communis by multiple stromata, shorter asci and ascospores. Ophiocordycepslongistipes differs from O.fusiformis in producing larger stromata, perithecia, asci and ascospores, as well as smaller citriform or oval conidia. Morphological descriptions of the two new species and a dichotomous key to the 19 termite-pathogenic Ophiocordyceps species are presented.

Key words: New species, morphology, Ophiocordyceps , phylogeny, termites

Introduction

Invertebrate-associated fungi are intriguing and diverse, widely distributed around the world (Araújo et al. 2018; Luangsa-ard et al. 2018; Haelewaters and Kasson 2020; Wilson et al. 2021; Santamaria et al. 2023). There are two typical relationships between fungi and invertebrates. One is mutualism. Mutualism is reciprocally positive interactions between pairs of species (Bronstein 2009). For example, Termitomyces Heim (Lyophyllaceae, Agaricomycetes) can decompose plants to provide food for termites; in return, termites shelter Termitomyces from external threats (Da Costa et al. 2019). The other is parasitism. Parasitism is the interaction between two species where one party (the parasite) benefits, while the other party (the host) suffers harm (Roper et al. 2019). As exemplified by species of Cordyceps Fr. sensu lato (s. l.), fungi parasitize invertebrates and eventually kill them. Invertebrate-pathogenic fungi are considered as the most well-known parasitic fungi (Araújo et al. 2018; Araújo et al. 2021; Wilson et al. 2021). They are ubiquitous inhabitants of forests worldwide, especially in tropical and subtropical regions. Invertebrate-pathogenic fungi are highly virulent and are known to have significant effects on host populations (Evans 1974). Cordyceps s. l. represents the most abundant and diverse group among invertebrate-pathogenic fungi (Araújo et al. 2021). Representatives of this group can colonize hosts in more than 10 invertebrate orders (Sanjuan et al. 2015; Araújo and Hughes 2016). They spread primarily through their hosts, evolving extensively in their morphologies and parasitic strategies. (Araújo and Hughes 2016). According to the current status of Cordyceps s. l. taxonomy, it belongs to four families: Clavicipitaceae, Cordycipitaceae, Ophiocordycipitaceae and Polycephalomycetaceae (Sung et al. 2007a; Xiao et al. 2023). Among them, the genus Ophiocordyceps Petch (Ophiocordycipitaceae) has received significant attention for its unique interactions with hosts and medical values (Zou et al. 2017; Araújo et al. 2018; Luangsa-ard et al. 2018; Khonsanit et al. 2019; Wang et al. 2021a; Zou et al. 2022; Tang et al. 2023a).

Ophiocordyceps is the largest genus in the family Ophiocordycipitaceae (Araújo et al. 2018; Luangsa-ard et al. 2018). The genus was established by Petch based on the type species O.blattae Petch (Petch 1931). In recent years, an increasing number of species have been described in Ophiocordyceps, with approximately 410 accepted species names to date (http://www.indexfungorum.org/names/Names.asp) (Sung et al. 2007a; Sanjuan et al. 2015; Spatafora et al. 2015; Araújo et al. 2018; Evans et al. 2018; Luangsa-ard et al. 2018; Wijayawardene et al. 2018; Khonsanit et al. 2019; Mongkolsamrit et al. 2023; Tang et al. 2023a, b).

The majority of species in Ophiocordyceps exhibit clavate, entirely, or partially darkly pigmented stromata or synnemata, especially those species with a hirsutella-like anamorph, while some species possess brightly colored stromata with hymenostilbe-like anamorph. The stromata are mostly wiry, tough, leathery, and flexible. Perithecia are superficial to pseudo-immersed to fully immersed, and are vertically or obliquely inserted in the stromata. Asci are usually cylindrical with thickened apex and contain eight ascospores. Ascospores are typically cylindrical or clavate, multiseptate, either disarticulating into secondary spores or remaining whole after discharge (Sung et al. 2007a; Quandt et al. 2014; Luangsa-ard et al. 2018). Species in Ophiocordyceps mainly attack insects of Coleoptera, Diptera, Hemiptera, Hymenoptera, Lepidoptera, Odonata, and Orthoptera. Generally, they can attack all stages (larva, pupa, nymph, and adult) of the insects, with the majority of targets being larvae of Coleoptera and Lepidoptera living in wood or buried in soil (Sung et al. 2007a; Shrestha et al. 2016). Among species of Ophiocordyceps, only 17 species attack termites (Tasanathai et al. 2019; Araújo et al. 2021; Tang et al. 2022; Tasanathai et al. 2022; Xu et al. 2022).

Termites (Termitidae, Blattodea) are typically eusocial soil-dwelling insects, widely distributed around the world, especially in tropical and subtropical regions (Pearce 1997). Most termites are considered pests, causing significant impacts on forest ecosystems, and agricultural and forestry crops, with subterranean termites being particularly destructive (Rust and Su 2012; Scharf 2015). Some species of termite-pathogenic Ophiocordyceps have been regarded as potential biological agents to control termite populations (Rath 2000).

During surveys of invertebrate-pathogenic fungi in Yunnan Province, China, several specimens attacking termites were collected. Morphological and molecular evidence indicates that they belong to two different taxa distinct from previously described species. This study aims to introduce these two new species and discuss their evolutionary placement among related species.

Materials and methods

Collection and isolation

Stromata emerging above fallen leaves were found in subtropical evergreen broad-leaved forests of Ruili City and Jinghong City, Yunnan Province, China. Specimens were documented and photographed in the field using a Canon 90D digital camera, and then each was placed in a sterilized 50 mL plastic centrifugal tube. All samples were stored in a cooler with ice packs until they were taken to the laboratory. Pure cultures were obtained on potato dextrose agar (PDA) with the composition of 200 g/L potato, 20 g/L dextrose, and 20 g/L agar, following the method previously presented (Wang et al. 2020). Subsequently, pure cultures were transferred to PDA slants and stored at the Kunming Institute of Botany Culture Collection (KUNCC), Chinese Academy of Sciences. Dried specimens were deposited in the Cryptogamic Herbarium of the Kunming Institute of Botany, Chinese Academy of Sciences (KUN-HKAS).

Morphological observations

The newly collected specimens were macroscopically examined with the Canon 750D camera and Olympus SZ60 stereo microscope. The characteristics of stromata (size, texture, shape, and color) were recorded. For the observation of teleomorph, perithecia were removed from the stromata and mounted on a glass slide with either 3% potassium hydroxide (KOH) (w/v) or 0.04% lactophenol cotton blue stain solution (w/v). Subsequently, the sizes and shapes of the perithecia, asci, and ascospores were measured under Olympus BX53 microscope. For each species, at least two specimens are measured, and each characteristic is measured at least 15 times repeatedly. The characteristics of pure cultures (size, texture, and color) were photographed using a Canon 750D camera after six weeks of culturing in an incubator at 25 °C. For the morphological description of anamorph, microscope slide cultures were prepared using the previous described method (Wang et al. 2020). Conidiogenous structures and conidia were measured and photographed using an Olympus BX53 microscope.

DNA extraction, amplification and sequencing

Genomic DNA was extracted from fresh mycelia cultured for three weeks using Ezup Column Fungi Genomic DNA Extraction Kit (Sangon Bio Co., Ltd., Shanghai, China), following the manufacturer’s protocol. Polymerase chain reactions (PCRs) were used to amplify genetic markers using the following primer pairs: nrSSU-COF/nrSSU-COR for the nuclear ribosomal small subunits (nrSSU) (Wang et al. 2015), LR0R/LR5 for the nuclear ribosomal large subunits (nrLSU) (Vilgalys and Hester 1990; Hopple 1994), ITS5/ITS4 for the internal transcribed spacer (ITS) (White et al. 1990), EF1α-EF/EF1α-ER for the translation elongation factor 1α (tef-1α) (Bischoff et al. 2006; Sung et al. 2007b), RPB1-5F/RPB1-5R for the largest subunits of RNA polymerase II (rpb1) (Bischoff et al. 2006), and RPB2-5F/RPB2-7cR for the second largest subunits of RNA polymerase II (rpb2) (Bischoff et al. 2006; Sung et al. 2007b).

Each 25 µL-PCR reaction contained 12.5 µL of 2× Taq PCR Master Mix (Tiangen Biotech Co., Ltd., Beijing, China), 9.5 µL of RNase-Free water (Sangon Bio Co., Ltd., Shanghai, China), 1 µL of each forward and reverse primer (10 µmol/L), 1 µL of DNA template (500 ng/µL). PCR reactions were placed in a LongGene T20 multi-block thermal cycler (Hangzhou LongGene Scientific Instruments Co., Ltd., Hangzhou, China) under the following conditions: For ITS, (1) 3 min at 95 °C, (2) 36 cycles of denaturation at 94 °C for 30 sec, annealing at 55 °C for 50 sec and extension at 72 °C for 1 min, (3) extension at 72 °C for 5 min and 12 °C soak. For nrSSU, (1) 4 min at 95 °C, (2) 22 cycles of denaturation at 94 °C for 1 min, annealing at 51 °C for 1 min and extension at 72 °C for 90 sec, followed by (3) 12 cycles of denaturation at 94 °C for 1 min, annealing at 50 °C for 1 min and extension at 72 °C for 95 sec, (4) extension at 72 °C for 10 min and 12 °C soak. For nrLSU, (1) 4 min at 95 °C, (2) 36 cycles of denaturation at 94 °C for 1 min, annealing at 50 °C for 1 min and extension at 72 °C for 2 min, (3) extension at 72 °C for 10 min and 12 °C soak. For tef-1α, (1) 3 min at 95 °C, (2) 36 cycles of denaturation at 94 °C for 30 sec, annealing at 50 °C for 30 sec and extension at 72 °C for 1 min, (3) extension at 72 °C for 10 min and 12 °C soak. For rpb1, (1) 4 min at 95 °C, (2) 36 cycles of denaturation at 94 °C for 40 sec, annealing at 50 °C for 40 sec and extension at 72 °C for 90 sec, (3) extension at 72 °C for 10 min and 12 °C soak. For rpb2, (1) 3 min at 95 °C, (2) 36 cycles of denaturation at 94 °C for 45 s, annealing at 58 °C for 45 s and extension at 72 °C for 90 s, (3) extension at 72 °C for 10 min and 12 °C soak. Standard DNA markers (Sangon Bio Co., Ltd., Shanghai, China) of known size and weight were used to quantify the PCR products. PCR products were purified using the DiaSpin PCR Product Purification Kit (Sangon Bio Co., Ltd., Shanghai, China), following the manufacturer’s instructions. Purified PCR products were sent to Sangon Bio Co., Ltd., (Kunming, China) for Sanger sequencing. The newly generated sequences were checked using MEGA v. 7.0 (Kumar et al. 2016). Consensus sequences were obtained using SeqMan of the Lasergene software package v. 14.1 (DNAstar, Madison, Wisconsin, USA) and deposited in NCBI GenBank (https://www.ncbi.nlm.nih.gov/genbank).

Sequencing alignments and phylogenetic analyses

We generated sequences for six loci from five specimens (Table 1). These were complemented with sequences of 125 related samples downloaded from NCBI GenBank based on BLAST searches and recent publications on Ophiocordycipitaceae (Tang et al. 2022; Xu et al. 2022). Tolypocladiuminflatum Gams OSC 71235 and T.ophioglossoides (J.F. Gmel.) Quandt et al. CBS 100239 were selected as the outgroup. The sequence datasets were aligned using MAFFT v. 7, and alignments were manually corrected in MEGA v. 7.0 (Katoh and Standley 2013; Kumar et al. 2016). Ambiguously aligned sites were manually eliminated and gaps were regarded as missing data. ModelFinder (Kalyaanamoorthy et al. 2017) was used to select the best-fit nucleotide substitution models for Maximum Likelihood (ML) and Bayesian Inference (BI) analyses under the Akaike Information Criterion (AIC). The optimized models for each locus partition are presented in Table 2. Partitioned ML and BI analyses were performed on the concatenated data set. The BI analysis was conducted using the MrBayes v. 3.2 (Ronquist et al. 2012). Four simultaneous Markov chains were run for 2,000,000 generations with a sub-sampling frequency every 100 generations. A burn-in of the first 25% of the total run was discarded. ML analysis was conducted using IQ-TREE v. 2.1.3 (Nguyen et al. 2015) under partitioned models (Chernomor et al. 2016) with 1000 ultrafast bootstrap (Hoang et al. 2018). Trees were visualized with its Maximum-Likelihood bootstrap proportions (ML-BS) and Bayesian posterior probability (BI-PP) in FigTree v. 1.4.4 and edited with Adobe Illustrator CS6.0.

Table 1.

Voucher information and GenBank accession numbers for the sequences included in this study.

Species Voucher information GenBank accession no. Reference
ITS nrSSU nrLSU tef-1α rpb1 rpb2
Hirsutellasatumaensis ARSEF 996 – KM652082 KM652125 KM652008 KM652047 – Simmons et al. 2015
H.cf.haptospora ARSEF 2228 KM652166 KM652075 KM652118 KM652001 KM652041 – Simmons et al. 2015
H.citriformis ARSEF 1446 KM652154 KM652065 KM652106 KM651990 KM652031 – Simmons et al. 2015
H.cryptosclerotium ARSEF 4517 KM652157 KM652066 KM652109 KM651992 KM652032 – Simmons et al. 2015
H.fusiformis ARSEF 5474 – KM652067 KM652110 KM651993 KM652033 – Simmons et al. 2015
H.gigantea ARSEF 30 – – JX566977 JX566980 KM652034 – Simmons et al. 2015
H.guyana ARSEF 878 – KM652068 KM652111 KM651994 KM652035 – Simmons et al. 2015
H.haptospora ARSEF 2226 KM652159 – – KM651995 KM652036 – Simmons et al. 2015
H.illustris ARSEF 5539 KM652160 KM652069 KM652112 KM651996 KM652037 – Simmons et al. 2015
H.kirchneri ARSEF 5551 – KM652070 KM652113 KM651997 – – Simmons et al. 2015
H.lecaniicola ARSEF 8888 KM652162 KM652071 KM652114 KM651998 KM652038 – Simmons et al. 2015
H.liboensis ARSEF 9603 KM652163 KM652072 KM652115 – – – Simmons et al. 2015
H.necatrix ARSEF 5549 KM652164 KM652073 KM652116 KM651999 KM652039 – Simmons et al. 2015
H.nodulosa ARSEF 5473 KM652165 KM652074 KM652117 KM652000 KM652040 – Simmons et al. 2015
H.radiata ARSEF 1369 – KM652076 KM652119 KM652002 KM652042 – Simmons et al. 2015
H.repens nom. inval. ARSEF 2348 KM652167 KM652077 KM652120 KM652003 – – Simmons et al. 2015
H.rhossiliensis ARSEF 3747 KM652168 KM652080 KM652123 KM652006 KM652045 – Simmons et al. 2015
H.strigosa ARSEF 2197 KM652174 KM652085 KM652129 KM652012 KM652050 – Simmons et al. 2015
H.subulata ARSEF 2227 KM652176 KM652086 KM652130 KM652013 KM652051 – Simmons et al. 2015
H.thompsonii ARSEF 257 KM652182 – KM652139 KM652019 KM652056 – Simmons et al. 2015
ARSEF 414 KM652184 – KM652143 KM652021 KM652059 – Simmons et al. 2015
H.thompsoniivar.vina ARSEF 254 – KM652101 KM652149 KM652028 KM652062 – Simmons et al. 2015
H.versicolor ARSEF 1037 – KM652102 KM652150 KM652029 KM652063 – Simmons et al. 2015
Ophiocordycepsacicularis OSC 110988 – EF468951 EF468804 EF468745 EF468853 – Sung et al. 2007a
O.agriotidis ARSEF 5692 JN049819 DQ522540 DQ518754 DQ522322 DQ522368 DQ522418 Spatafora et al. 2007
O.annulata CEM 303 – KJ878915 KJ878881 KJ878962 KJ878995 – Quandt et al. 2014
O.appendiculata NBRC 106960 JN943326 JN941728 JN941413 AB968577 JN992462 AB968539 Schoch et al. 2012
O.arborescens NBRC 105891 AB968398 AB968386 AB968414 AB968572 – AB968534 Ban et al. 2015
O.asiana MY11878 MW285719 – MW280213 MW292448 MW296049 – Khao-ngam et al. 2021
O.asiatica BCC 30516 MH754722 – MH753675 MK284263 MK214105 MK214091 Tasanathai et al. 2019
BCC 86435 MH754723 – MH753676 – MK214106 MK214092 Tasanathai et al. 2019
O.barnesii BCC 28560 – EU408776 – – EU408773 EU418599 Luangsa-ard et al. 2010
O.bidoupensis YFCC 8793 – OM304638 – OK556894 OK556898 OK556900 Zou et al. 2022
O.brunneinigra BCC 69032 – – MF614654 MF614638 MF614668 MF614681 Luangsa-ard et al. 2018
O.brunneiperitheciata BCC 66167 – – MF614659 MF614644 – MF614684 Luangsa-ard et al. 2018
O.brunneipunctata OSC 128576 – DQ522542 DQ518756 DQ522324 DQ522369 DQ522420 Spatafora et al. 2007
O.brunneirubra BCC 14384 MH754736 – MH753690 GU797121 MK751465 MK751468 Tasanathai et al. 2019
O.campes BCC 36938 MT783955 – MT118175 MT118167 MT118183 MT118188 Tasanathai et al. 2020
O.communis BCC 1842 MH754726 – MH753680 MK284266 MK214110 MK214096 Tasanathai et al. 2019
BCC 1874 MH754725 – MH753679 MK284267 MK214109 MK214095 Tasanathai et al. 2019
BCC 2754 MH754727 – MH753681 MK284268 MK214111 MK214097 Tasanathai et al. 2019
O.cossidarum MFLU 17-0752 – MF398186 MF398187 MF928403 MF928404 – Xiao et al. 2019
O.crinalis GDGM 17327 – KF226253 KF226254 KF226256 KF226255 – Wang et al. 2014
O.dipterigena OSC 151911 – KJ878919 KJ878886 KJ878966 KJ879000 – Quandt et al. 2014
O.elongata OSC 110989 – – EF468808 EF468748 EF468856 – Sung et al. 2007a
O.flavida BCC 84256 – – MT512655 MT533482 MT533476 – Mongkolsamrit et al. 2021
O.formosana TNM F13893 – KJ878908 – KJ878956 KJ878988 KJ878943 Quandt et al. 2014
O.furcatosubulata YFCC 902 – MT774214 MT774221 MT774242 MT774228 MT774235 Wang et al. 2021b
O.fusiformis BCC 93025 MZ676743 – MZ675422 MZ707849 MZ707855 MZ707805 Tasanathai et al. 2022
BCC 93026 MZ676744 – MZ675423 MZ707850 MZ707856 MZ707806 Tasanathai et al. 2022
O.geometridicola BCC 35947 – – MF614647 MF614631 MF614664 MF614678 Luangsa-ard et al. 2018
O.globiceps MFLU 18-0661 MH725816 MH725812 MH725830 MH727388 – – Xiao et al. 2019
O.globiperitheciata HKAS 126130 OR015963 OR082950 OR015968 OR030532 OR119834 – This study
HKAS 126131 OR015964 OR082951 OR015969 OR030533 OR119835 – This study
O.globosa BCC 93023 MZ676740 MZ675419 MZ707846 MZ707861 – Tasanathai et al. 2022
O.halabalaensis MY5151 GU723763 KM655826 – GU797110 – – Luangsa-ard et al. 2011
O.hydrangea YFCC 8832 – OM304636 OM304640 OM831277 OM831280 OM831283 Zou et al. 2022
O.irangiensis BCC 82795 MH028142 – – MH028186 MH028164 MH028174 Khonsanit et al. 2019
O.isopterae MY12376 MZ676741 – MZ675420 MZ707847 MZ707859 MZ707803 Tasanathai et al. 2022
BCC 93042 MZ676742 – MZ675421 MZ707848 MZ707804 Tasanathai et al. 2022
O.karstii MFLU 15-3884 – KU854952 – KU854945 KU854943 – Li et al. 2016
O.khokpasiensis BCC 48071 MH754728 – MH753682 MK284269 MK214112 – Tasanathai et al. 2019
BCC 48072 MH754729 – MH753683 MK284270 MK214113 – Tasanathai et al. 2019
BCC 1764 MH754730 –– MH753684 MK284271 MK214114 MK214098 Tasanathai et al. 2019
BCC 81464 MK632043 MK632128 MK632103 MK632077 MK632170 MK632159 Tasanathai et al. 2019
O.kimflemingiae SC09B – KX713631 KX713620 KX713698 KX713724 – Araújo et al. 2018
O.konnoana EFCC 7315 – EF468959 – EF468753 EF468861 EF468916 Sung et al. 2007a
O.longissima EFCC 6814 – – EF468817 EF468757 EF468865 – Sung et al. 2007a
NBRC 106965 AB968406 AB968392 AB968420 AB968584 – AB968546 Ban et al. 2015
O.longistipes KUNCC 5224 OR015962 OR082949 OR015967 OR030530 OR062224 OR113082 This study
HKAS 126186 OR015960 OR082947 OR015966 OR030531 OR062225 This study
HKAS 126187 OR015961 OR082948 OR015965 OR030529 OR062223 This study
O.longistromata BCC 44497 MT783956 – MT118178 MT118170 – MT118191 Tasanathai et al. 2020
O.macroacicularis NBRC 100685 AB968400 AB968388 AB968416 AB968574 – AB968536 Ban et al. 2015
O.megacuculla BCC 82984 – – MH028162 MH028192 – MH028181 Khonsanit et al. 2019
O.mosingtoensis BCC 30904 MH754732 – MH753686 MK284273 MK214115 MK214100 Tasanathai et al. 2019
O.mosingtoensis BCC 36921 MH754731 – MH753685 MK284272 MK214116 MK214099 Tasanathai et al. 2019
O.multiperitheciata BCC 22861 – – MF614656 MF614640 MF614670 MF614683 Luangsa-ard et al. 2018
O.myrmecophila CEM 1710 – – KJ878894 KJ878974 KJ879008 – Quandt et al. 2014
O.nigrella EFCC 9247 JN049853 EF468963 EF468818 EF468758 EF468866 EF468920 Sung et al. 2007a
O.nutans OSC 110994 – DQ522549 DQ518763 DQ522333 DQ522378 – Spatafora et al. 2007
O.ovatospora YHH 2206001 OP295105 OP295110 OP295113 OP313801 OP313803 OP313805 Tang et al. 2022
YFCC 22069184 OP295106 OP295111 OP295114 OP313802 OP313804 – Tang et al. 2022
O.pauciovoperitheciata TBRC 8096 – – MF614649 MF614636 MF614665 MF614672 Luangsa-ard et al. 2018
O.phuwiangensis BCC 85351 MT783958 – – MT118174 MT118187 MT118195 Tasanathai et al. 2020
BCC 86208 – – MT118180 MT118172 MT118185 MT118193 Tasanathai et al. 2020
O.pruinosa NHJ 12994 – EU369106 EU369041 EU369024 EU369063 EU369084 Johnson et al. 2009
O.pseudoacicularis BCC 53843 – – MF614646 MF614630 MF614661 MF614677 Luangsa-ard et al. 2018
O.pseudocommunis NHJ 12581 – EF468973 EF468831 EF468775 – EF468930 Luangsa-ard et al. 2018
NHJ 12582 EF468975 EF468830 EF468771 – EF468926 Luangsa-ard et al. 2018
O.pseudocommunis BCC 16757 MH754733 – MH753687 MK284274 MK214117 MK214101 Tasanathai et al. 2019
O.pseudolloydii MFLUCC 15-0689 MF351725 – – MF372758 MF372761 – Xiao et al. 2017
O.pseudorhizoidea BCC 48879 MH754720 – MH753673 MK284261 MK214104 MK214089 Tasanathai et al. 2019
BCC 86431 MH754721 – MH753674 MK284262 MK751469 MK214090 Tasanathai et al. 2019
NHJ 12522 JN049857 – EF468825 EF468764 EF468873 EF468923 Tasanathai et al. 2019
NHJ 12529 – – EF468824 EF468765 EF468872 EF468922 Tasanathai et al. 2019
O.puluongensis YFCC 6442 – MT141118 MT270528 MT270520 MT270523 MT270526 Xu et al. 2022
YFCC 6443 – MT141119 MT270529 MT270521 MT270524 MT270527 Xu et al. 2022
YHH 16017 – – MT270530 MT270522 MT270525 – Xu et al. 2022
O.pulvinata TNS-F 30044 AB721302 GU904208 – GU904209 GU904210 – Kepler et al. 2011
O.radiciformis BCC 93036 MZ676746 – MZ675425 MZ707852 MZ707857 MZ707808 Tasanathai et al. 2022
BCC 93035 MZ676747 – MZ675426 MZ707853 MZ707858 MZ707809 Tasanathai et al. 2022
O.ramosissimum GZUHHN8 KJ028007 KJ028012 – KJ028014 KJ028017 – Wen et al. 2014
O.ravenelii OSC 110995 – DQ522550 DQ518764 DQ522334 DQ522379 DQ522430 Spatafora et al. 2007
O.rhizoidea NHJ 12522 JN049857 EF468970 EF468825 EF468764 EF468873 EF468923 Tasanathai et al. 2019
NHJ 12529 – EF468969 EF468824 EF468765 EF468872 EF468922 Tasanathai et al. 2019
O.robertsii KEW 27083 AJ309335 – EF468826 EF468766 – – Sung et al. 2007a
O.rubiginosiperitheciata NBRC 106966 JN943344 JN941704 JN941437 AB968582 JN992438 AB968544 Kepler et al. 2012
O.salganeicola Mori01 – MT741705 MT741719 MT759575 MT759578 MT759580 Araújo et al. 2021
Mori02 – MT741704 MT741718 MT759572 MT759579 MT759581 Araújo et al. 2021
O.satoi J7 – KX713653 KX713599 KX713683 KX713711 – Araújo et al. 2018
O.sinensis ARSEF 6282 KM652173 KM652083 KM652126 KM652009 KM652048 – Simmons et al. 2015
EFCC 7287 JN049854 EF468971 EF468827 EF468767 EF468874 EF468924 Kepler et al. 2012
O.sobolifera NBRC 106967 AB968409 AB968395 AB968422 AB968590 – – Ban et al. 2015
O.spataforae NHJ 12525 – EF469125 EF469078 EF469063 EF469092 EF469111 Sung et al. 2007a
OSC 128575 JN049845 EF469126 EF469079 EF469064 EF469093 EF469110 Sung et al. 2007a
O.sphecocephala NBRC 101752 JN943351 JN941696 JN941445 AB968591 JN992430 AB968552 Schoch et al. 2012
O.spicatus MFLU 18-0164 MK863254 MK863047 MK863054 MK860192 – – Zha et al. 2021
O.stylophora OSC 111000 JN049828 DQ522552 DQ518766 DQ522337 DQ522382 DQ522433 Spatafora et al. 2007
OSC 110999 – EF468982 EF468837 – EF468882 EF468931 Sung et al. 2007a
O.termiticola BCC 1920 MH754724 – MH753678 MK284265 MK214108 MK214094 Tasanathai et al. 2019
BCC 1770 GU723780 – MH753677 MK284264 MK214107 MK214093 Tasanathai et al. 2019
BCC 93002 – – MZ675427 MZ707854 MZ707862 MZ707810 Tasanathai et al. 2019
O.thanathonensis MFLU 16-2909 MF850376 – MF850377 MF872613 MF872615 – Xiao et al. 2017
O.tricentri NBRC 106968 AB968410 AB968393 AB968423 AB968593 – AB968554 Ban et al. 2015
O.unilateralis OSC 128574 – DQ522554 DQ518768 DQ522339 DQ522385 DQ522436 Spatafora et al. 2007
O.unituberculata YFCC HU1301 – KY923214 – KY923216 KY923218 KY923220 Wang et al. 2018
O.xuefengensis GZUHHN13 KC631804 KC631785 – KC631790 KC631795 – Wen et al. 2013
GZUH2012HN13 KC631801 KC631787 – KC631792 KC631797 – Wen et al. 2013
trichospora CBS 109876 – AF543766 AF543790 AF543779 AY489669 DQ522457 Sung et al. 2007a
Tolypocladiuminflatum OSC 71235 JN049844 EF469124 EF469077 EF469061 EF469090 EF469108 Sung et al. 2007b
T.ophioglossoides CBS 100239 – KJ878910 KJ878874 KJ878958 KJ878990 KJ878944 Quandt et al. 2014
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Note: Newly-generated sequences are shown in bold.

Table 2.

Results of the best-ftting model for maximum likelihood (ML) and Bayesian inference (BI) for six loci partitions.

Gene name ML BI
ITS GTR+F+I+G4 GTR+F+I+G4
nrSSU TNe+I+G4 SYM+I+G4
nrLSU TIM+F+I+G4 GTR+F+I+G4
tef-1α TIM2+F+I+G4 GTR+F+I+G4
rpb1 TIM+F+I+G4 GTR+F+I+G4
rpb2 TIM3+F+I+G4 GTR+F+I+G4
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Results

Phylogenetic analyses

The combined dataset of six loci was composed of 5021 bp (585 bp for ITS, 903 bp for nrLSU, 1037 bp for nrSSU, 859 bp for tef-1α, 664 bp for rpb1, and 973 bp for rpb2). Phylogenetic trees inferred from ML and BI analyses exhibited nearly consistent overall topologies and recognized four statistically well-supported clades within Ophiocordyceps, namely Hirsutella Pat, O.sphecocephala (Klotzsch ex Berk.) Sung et al., O.sobolifera (Hill ex Watson) Sung et al., and O.ravenelii (Berk. & M.A. Curtis) Sung et al. clades (Fig. 1). Among them, the Hirsutella clade includes six distinct subclades, namely H.citriformis Speare, H.thompsonii Fisher, H.nodulosa Petch, H.guyana Minter & Brady, H.sinensis (Berk.) Sung et al., and the Hirsutella ant pathogen subclades. As revealed from phylogenetic analyses, all specimens collected in this study were placed in the H.thompsonii subclade. Three samples (HKAS 126185, HKAS 126186, and HKAS 126187), newly described as O.longistipes, were clustered closely with O.fusiformis Tasan et al. However, the phylogenetic evidence indicated that these three samples formed a monophyletic clade in Ophiocordyceps, with high statistical support (ML-BS/BI-PP=100/1). The other two samples (HKAS 126130 and HKAS 126131), newly described as O.globiperitheciata, clustered together and formed a separate clade, distinguishing from other species in Ophiocordyceps with moderate bootstrap support (ML-BS/BI-PP=84/0.99). Therefore, the phylogenetic data supported the recognition of O.longistipes and O.globiperitheciata as distinct species in Ophiocordyceps.

Figure 1.

Figure 1.

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Phylogenetic tree based on the combined dataset of nrSSU, nrLSU, tef-1α, rpb1, rpb2, and ITS showing the relationship of two new species on termites from China with other Ophiocordyceps species. Values at the nodes before and after the backslash are BI posterior probabilities (BI-PP greater than 0.60) and ML bootstrap proportions (ML-BP greater than 70%), respectively. New species described in this paper are shown in bold red.

Taxonomy

. Ophiocordyceps longistipes

Y.B. Wang, T. Yang, Q. Fan & Zhu L. Yang sp. nov.

CBABB789-0E59-5460-8EFB-97C7F203A012

Index Fungorum: IF901029

Fig. 2

Figure 2.

Figure 2.

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OphiocordycepslongistipesA, B stromata of fungus arising from termites C fertile part D perithecia E colony on PDA (obverse and reverse) F, G ascospores H, I asci J–L conidiogenous cells and conidia. Scale bars: 1 cm (A, B, E); 2 mm (C); 100 µm (D); 20 µm (F–L).

Etymology.

Referring to the long stipe of stromata.

Type.

Holotype: China, Yunnan Province, Ruili City, 26°1.07'N, 97°51.33'E, alt. 1140 m, on a termite buried in soil, 2 July 2022, Tao Yang (holotype HKAS 126185, ex-type culture KUNCC 5224). Ex-type sequences (ITS: OR015962, nrLSU: OR015967, nrSSU: OR082949, tef-1α: OR030530, rpb1: OR062224, rpb2: OR113082).

Description.

Stromata arising from the back of termites buried in soil, solitary, unbranched, cylindrical, flexible, leathery, 17–24 cm long, 0.5–1.0 mm wide, grayish white to yellowish brown. Fertile parts cylindrical, yellowish brown, 3–5.5 cm long, generating toward the upper part of stromata, covered by a spinous surface, with a sterile tip of 11–28 × 0.5–1.0 mm. Perithecia superficial, pale yellow at early stage, brown at maturity, pyramidal to oval, densely distributed in the upper of stromata, arranged in a disordered manner, 390–420 × 295–350 µm. Asci 8-spored, filiform, hyaline, 160–195 × 4.5–6.5 µm, with hemispheric apical cap. Ascospores whole, hyaline, filiform, tapering at both ends, 70–85 × 3.5–4.5 µm, multiseptate, septa 4.5–13.8 μm long.

Anamorph.

hirsutella-like. Colonies on PDA growing very slowly, reaching 3–3.5 cm diam after six weeks at 25 °C, felty, irregularly convex, cream, reverse pale brown to dark brown. Hyphae hyaline, branched, septate, smooth-walled, 2–3 µm wide. Conidiogenous cells arising from aerial mycelia, monophialidic or rarely polyphialidic, on hyphae laterally or terminally, hyaline, smooth, flask-shaped, 29–60 μm long, with a swollen base, 4–4.5 μm wide, tapering sharply into a thin neck, 0.5–0.8 μm wide. Conidia borne directly on the tip of phialides, hyaline, one-celled, solitary, smooth-walled, citriform or oval, 7–10 × 4.5–7 µm, with a mucous sheath.

Additional specimens examined.

China, Yunnan Province, Ruili City, 26°1.07'N, 97°51.33'E, alt. 1140 m, on a termite buried in soil, 2 July 2022, Tao Yang (HKAS 126186), sequences (ITS: OR015960, nrLSU: OR015966, nrSSU: OR082947, tef-1α: OR030531, rpb1: OR062225). Ibid., (HKAS 126187), sequences (ITS: OR015961, nrLSU: OR015965, nrSSU: OR082948, tef-1α: OR030529, rpb1: OR062223).

Habitat and ecology.

Parasitic on termites buried in soil of the subtropical evergreen broad-leaved forests, emerging from fallen leaves on the forest floor.

Known distribution.

Ruili City, Yunnan Province, China.

Notes.

Ophiocordycepslongistipes is characterized by solitary stromata, superficial and pyramidal to oval perithecia, filiform asci, and filiform ascospores, hirsutella-like anamorph with monophialidic or rarely polyphialidic, flask-shaped conidiogenous cells, and citriform or oval conidia embedded in a mucous sheath. Phylogenetically, all specimens of O.longistipes are clustered in the H.thompsonii subclade of Hirsutella lineages and form a monophyletic clade, which is placed sister to O.fusiformis with maximum support (Fig. 1). However, O.longistipes exhibits significant morphological differences from O.fusiformis in its both teleomorph and anamorph. For the teleomorph, O.longistipes produce longer stromata of 17–24 cm (up to 6 cm long for O.fusiformis), larger perithecia of 390–420 × 295–350 µm (300–360 × 180–270 µm for O.fusiformis). For the anamorph, O.longistipes possess both monophialidic and polyphialidic conidiogenous cells, but O.fusiformis is only monophialidic. Moreover, O.longistipes produces oval conidia, while O.fusiformis produces narrower fusiform conidia (Table 3).

Table 3.

Morphological comparison between Ophiocordyceps species parasitic on termites.

Species Host Stromata (cm) Perithecia (µm) Asci (µm) Ascospore (µm) Conidiogenous cells (µm) Conidia (µm) Reference
O.asiatica Termites Solitary, simple, filiform, orange brown, up to 15 long Superficial, globose to subglobose, 240–320 × 180–260 Filiform, 92.5–175 × 5–6.3 Filiform, septate, whole, 80–132.5 × 1–2 Monophialidic or rarely polyphialidic, 15–20 × 2–3 Fusiform, 7–9 × 2–3 (Tasanathai et al. 2019)
O.bispora Termites Multiple (20–30), simple or branched, clavate Immersed, globose, 300–375 × 375 Clavate, 162–163 × 58–61 Elliptical closely appressed, septate, 95–105 × 34–35.4 (Blackwell and Gilbertson 1984; Ochiel et al. 1997)
O.brunneirubra Termites Solitary, simple or branched, narrowly clavate, orange brown to red brown, 9.5 long Immersed, ovoid, 300–400 × 130–200 Cylindrical, 155–225 × 4.5–8 Filiform, septate, whole, 156.5–197.5 × 2–3 Monophialidic, 32–50 × 2–3 Fusiform, 12–17 × 2–4 (Tasanathai et al. 2019)
O.communis Termites Solitary, simple, filiform, base whitish-grey, upper part yellow-brown, 5–13 long Superficial, 285–675 × 195–390 Filiform, 215–250 × 15 Filiform, septate, whole, 100–180 × 5–6 Monophialidic or rarely polyphialidic, 10–14 × 2.7–3.3 Almond-shaped, 7–9 × 2.5–3 (Sung et al. 2007a)
O.fusiformis Termite Solitary, simple, cylindrical, brown, up to 6 long Superficial, ovoid, 300–360 × 180–270 Cylindrical, 141–227 × 7–15 Cylindrical, septate, whole, 36–78 × 5–6.5 Monophialidic, 9–24 × 2–4 Fusiform, 6–18 × 2–4 (Tasanathai et al. 2022)
O.globosa Termites Solitary, simple, cylindrical, brown, up to 8 long Pseudo-immersed, ovoid, 190–245 × 120–190 Filiform, 100–157 × 7–13 Filiform, septate, whole, 58–118 × 2–3 Monophialidic or polyphialidic, 9–15 × 3–5 Globose, 2–4 (Tasanathai et al. 2022)
O.globiperitheciata Termites Multiple (2–5), unbifurcated, clavate, base brown, tip gray, 8–15 long Superficial, subglobose, 240–295 × 215–280 Filiform, 135–170 × 8.5–13.5 Filiform, septate, whole, 85–110 × 3.5–4.5 This study
O.isopterae Termites Solitary, simple, cylindrical, brown, up to 10 long Superficial, ovoid, 270–320 × 140–180 Filiform, 81–137 × 5–9 Filiform, septate, whole, 55–78 × 2–2.5 Monophialidic, 14–28 × 2–4 Fusiform, 6–11 × 1.5–3 (Tasanathai et al. 2022)
O.khokpasiensis Termites Solitary, simple, cylindrical, brown, 16 long Pseudo-immersed, subglobose, 200–250 × 120–200 Filiform, 62.5–125 × 4–5 Filiform, whole, 46–90 × 2–3 Monophialidic or polyphialidic, 15–28 × 3–5 Globose to oval, 4–6 × 2.5–4 (Khonsanit et al. 2019)
O.koningsbergeri Termites Solitary, filiform, gray-white, 8–10 long Immersed, 450 × 90 Cylindrica, 180–200 × 4–5 Filiform, whole, 150 × 1 (Penzig and Saccardo 1904)
O.longistipes Termites Solitary, unbifurcated, cylindrical, grayish white to yellowish brown, 17–24 long Superficial, pyramidal to oval, 390–420 × 295–350 Filiform, 160–195 × 4.5–6.5 Filiform, septate, whole, 70–85 × 3.5–4.5 Monophialidic or rarely polyphialidic, on hyphae laterally or terminally, 29–60 long, with a swollen base, 4–4.5 wide, tapering sharply into a thin neck, 0.5–0.8 wide. Citriform or oval, 7–10 × 4.5–7 This study
O.mosingtoensis Termites Solitary, simple, cylindrical, brown to grey, 11 long Pseudo-immersed, ovoid, 400–500 × 200–300 Filiform, 187.5–287.5 × 4.5–7.5 Filiform, septate, whole, 230–315 × 1.5–3 Monophialidic, 10–17 × 2–3 Oval, 3–5 × 2–3 (Tasanathai et al. 2019)
O.octospora Termites Multiple, clavate, white to pale tan, 0.2–0.3 long Immersed, subglobose to ovoid, 180–220 × 200 Clavate, about 250 × 60 Cylindrical, septate, 40–70 × 15–30 (Blackwell and Gilbertson 1981)
O.ovatospora Termites Solitary, simple, cylindrical or clavate, light-yellow, up to 13 long Pseudo-immersed, ovoid to pyriform, 110–140 × 80–110 Filiform, 110–125 × 5–7 Filiform, septate, whole, 110–130 × 1–2 Monophialidic or rarely polyphialidic, 15–35 × 3–6 Oval, 3–5 × 3–4 (Tang et al. 2022)
O.pseudocommunis Termites Solitary, simple, cylindrical, brown, 21 long Superficial, Subglobose, 520–600 × 360–440 Filiform, 160–165 × 14–17 Filiform,septate, whole, 107.5–147.5 × 6–7.5 Arising from hyphae laterally or terminally Fusiform, septate (2–3), 13–27 × 3–5 (Tasanathai et al. 2019)
O.pseudorhizoidea Termites Solitary, simple, filiform, light brown, up to 21 long Superficial, ovoid, 280–390 × 160–220 Cylindrical, 120–150 × 5–7 Filiform, whole, 65–82.5 × 2–3 Monophialidic, 9–21 × 2–4 Fusiform, 5–10 × 1–2 (Tasanathai et al. 2019)
O.puluongensis Termites Solitary, simple or branched, filiform, pale orange to red brown, 7.1–13.3 long Superficial, subglobose, 181.8–251.0 × 123.7–205.4 Fliform, 74.3–138.5 × 4.6–6.5 Filiform, septate, whole, 67.0–124.5 × 1.5–2.5 Monophialidic or rarely polyphialidic, 7.9–21.2 × 1.7–5.0 Fusiform or citriform, 2.8–6.1 × 1.9–3.4 (Xu et al. 2022)
O.radiciformis Termites Solitary, simple, cylindrical, brown, up to 11 long Superficial, ovoid, 330–460 × 200–320 Cylindrical, 140–296 × 6–10 Filiform septate, whole, 154–215 × 2–3 6–15 × 2–5 Fusiform, 5–7 × 2–3 (Tasanathai et al. 2022)
O.termiticola Termites Solitary, simple, filiform, yellow brown, up to 14 long Pseudo-immersed, globose to subglobose, 200–280 × 150–250 Filiform 62.5–110 × 4–6 Filiform, whole, 85 × 2 Monophialidic to polyphialidic, 7–11 × 2.5–4 Globose, 2.5–3.5 (Tasanathai et al. 2019)
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. Ophiocordyceps globiperitheciata

Y.B. Wang, T. Yang, Q. Fan & Zhu L. Yang sp. nov.

825A1CAC-16CC-5671-BD8C-95405C4B9186

Index Fungorum: IF901030

Fig. 3

Figure 3.

Figure 3.

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OphiocordycepsglobiperitheciataA stromata of fungus arising from termites B sterile tip and fertile part C fertile part D perithecia E–G asci H–J ascospores. Scale bars: 1 cm (A); 2 mm (B); 500 µm (C); 50 µm (D); 20 µm (E–J).

Etymology.

Referring to the shape of perithecia, with “globi” meaning globose.

Type.

Holotype: China, Yunnan Province, Jinghong City, Puwen Town, 22°26.35'N, 101°1.32'E, alt. 970 m, on a termite buried in soil, 28 Sep. 2022, Tao Yang (HKAS 126130). Holotype sequences (ITS: OR015963, nrLSU: OR015968, nrSSU: OR082950, tef-1α: OR030532, rpb1: OR119834).

Description.

Stromata arising from the termite buried in soil, multiple (2–5), clavate, unbranched, flexible, leathery, 8–15 cm long, 1–1.5 mm wide, tapering from base to tip, base brown, tip gray. Fertile parts cylindrical, pale brown, generating toward the upper part of stromata, covered by a spinous surface, with a sterile tip. Perithecia superficial, pale brown to brown, subglobose, aggregating loosely at the upper of stromata, arranged in a disordered manner, 240–295 × 215–280 µm. Asci 8-spored, filiform, hyaline, 135–170 × 8.5–13.5 µm, with a hemispheric apical cap. Ascospores whole, hyaline, tapering at both ends, filiform, 85–110 × 3.5–4.5 µm, multiseptate, septa 11–14.5 μm long. Anamorph not detected.

Additional specimens examined.

China, Yunnan Province, Jinghong City, Puwen Town, 22°26.35'N, 101°1.32'E, alt. 970 m, on a termite buried in soil, 28 Sep. 2022, Tao Yang (HKAS 126131). Sequences (ITS: OR015964, nrLSU: OR015969, nrSSU: OR082951, tef-1α: OR030533, rpb1: OR119835).

Habitat and ecology.

Parasitic on termites buried in soil of tropical evergreen broad-leaved forests, emerging from fallen leaves on the forest floor.

Known distribution.

Puwen Town, Jinghong City, Yunnan Province, China.

Notes.

Ophiocordycepsglobiperitheciata is characterized by multiple and unbranched stromata, superficial and subglobose perithecia, and filiform asci and ascospores. Phylogenetically, O.globiperitheciata forms a separate clade from other Ophiocordyceps species in the H.thompsonii subclade with moderate bootstrap support (Fig. 1). It is closed to H.cryptosclerotium Fern. et al. and O.communis Hywel-Jones & Samson. However, it differs from H.cryptosclerotium in parasitizing Blattodea (H.cryptosclerotium parasitic on Hemiptera), producing multiple clavate stromata (H.cryptosclerotium stroma absence). Ophiocordycepsglobiperitheciata is distinguished from O.communis by multiple and thicker stromata, shorter asci of 135–170 µm (215–250 µm for O.communis) and ascospores of 85–110 µm (100–180 µm for O.communis) (Table 3).

Key to species of Ophiocordyceps parasitic on termites

1 Stromata multiple 2
– Stromata solitary 4
2 Perithecia superficial O.globiperitheciata
– Perithecia immersed 3
3 Perithecia subglobose to ovoid O.octospora
– Perithecia globose O.bispora
4 Perithecia nonsuperficial 5
– Perithecia superficial 11
5 Perithecia immersed 6
– Perithecia pseudo-immersed 7
6 Stromata orange brown to red brown O.brunneirubra
– Stromata gray-white O.koningsbergeri
7 Only monophialidic O.mosingtoensis
– Possessing polyphialidic 8
8 Large asci (100–160 µm long) O.globosa
– Small asci (60–130 µm long) 9
9 Large ascospores (> 100 µm long) O.ovatospora
– Small ascospores (< 100 µm long) 10
10 Conidia globose O.termiticola
– Conidia globose to oval O.khokpasiensis
11 Stromata sometimes branched O.puluongensis
– Stromata unbranched 12
12 Long stromata (≥ 15 cm long) 13
– Short stromata (< 15 cm long) 16
13 Conidia have septa O.pseudocommunis
– Conidia have no septa 14
14 Short stromata (< 16 cm long) O.asiatica
– Long stromata (> 16 cm long) 15
15 Long conidiogenous cells (> 25 µm long) O.longistipes
– Short conidiogenous cells (< 25 µm long) O.pseudorhizoidea
16 Conidia almond-shaped O.communis
– Conidia fusiform 17
17 Short asci (< 140 µm long) O.isopterae
– Long asci (≥ 140 µm long) 18
18 Long stromata (> 6 cm long) O.radiciformis
– Short stromata (≤6 cm long) O.fusiformis
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Discussion

Thus far, only 17 species of Ophiocordyceps parasitic on termites were described, mainly clustered in the H.thompsonii subclade (Tasanathai et al. 2019; Tasanathai et al. 2022). These species are: O.asiatica Tasanathai et al., O.bispora (Stifler) G.H. Sung et al., O.brunneirubra Tasanathai et al., O.communis Hywel-Jones & Samson, O.fusiformis Tasanathai et al., O.globosa Tasanathai et al., O.isopterae Tasanathai et al., O.khokpasiensis Tasanathai et al., O.koningsbergeri (Penz. & Sacc.) G.H. Sung et al., O.mosingtoensis Tasanathai et al., O.octospora (M. Blackw. & Gilb.) G.H. Sung et al., O.ovatospora H. Yu et al., O.pseudocommunis Tasanathai et al., O.pseudorhizoidea Tasanathai et al., O.puluongensis H. Yu et al., O.radiciformis Tasanathai et al., and O.termiticola Tasanathai et al. Most the termite-pathogenic Ophiocordyceps species are found in tropical and subtropical regions, which may be related to the higher diversity of both Ophiocordyceps fungi and their termite hosts in these climatic zones (Sung et al. 2007a; Tasanathai et al. 2019; Cerezer et al. 2020; Araújo et al. 2021; Wilson et al. 2021; Tang et al. 2022; Tasanathai et al. 2022; Xu et al. 2022).

Phylogenetically, almost all Ophiocordyceps species parasitic on termites are placed in the H.thompsonii subclade, except for O.brunneirubra. Termite-pathogenic species exhibit significant morphological variation overall. Among these species, the length of stromata ranges from extremely short to very long, the existence pattern of perithecia from superficial to pseudo-immersed to immersed, and the size of perithecia ranges from about 100 to 600 µm (Tasanathai et al. 2019; Araújo et al. 2021; Tasanathai et al. 2022; Xu et al. 2022). However, some of these species exhibit minimal interspecific morphological variation, making it challenging to distinguish them only through morphological studies. Therefore, the use of molecular systematics is necessary to accurately identify these species. For example, O.asiatica and O.puluongensis, as well as O.khokpasiensis and O.termiticola, share similar morphological characteristics. Ophiocordycepsasiatica and O.puluongensis produce subglobose superficial perithecia, similar asci, ascospores, conidiogenous cells, and conidia (Tasanathai et al. 2019; Xu et al. 2022). Ophiocordycepskhokpasiensis and O.termiticola possess similar colored and shaped stromata, pseudo-immersed perithecia, and similar asci, ascospores, and conidiogenous cells (Tasanathai et al. 2019). Although these species are morphologically indistinguishable, phylogenetic analyses support them as separate taxa.

It’s worth noting that the hosts of these termite-pathogenic Ophiocordyceps species are usually buried underground, typically 5 to 15 cm below the ground, which may be relevant to the subterranean living habits of the host termites (Martelossi et al. 2023). However, this can pose a challenge for species identification, as hosts are often lost due to separation from fragile stromata during excavation (Tasanathai et al. 2022).

Termites are notorious pests known for damaging wood, cultivated plants, buildings, pastures, forests, and even non-cellulosic materials like cables, causing annual economic losses amounting to tens of billions of dollars. Subterranean termites are responsible for about 80% of the total damage (Rust and Su 2012; Scharf 2015; Oi 2022). Therefore, the control of termites has become the focus of attention in various industries. Previously, many chlorinated hydrocarbon insecticides were used for termite control, but they were banned due to their irreversible environmental impact and negative effects on crop production. Consequently, environmentally friendly and sustainable control measures for termites are urgently needed. Entomopathogenic fungi may represent a potent solution (Afzal et al. 2019; Tasanathai et al. 2019; Oi 2022; Moon et al. 2023). These fungi, with strong infectivity, can continuously spread spores in the field to control pests and are considered environmentally non-polluting, so they have significant advantages in pest control (Shimazu et al. 1995; Meyling and Eilenberg 2007). Most members of H.thompsonii subclade have been found to obligately parasitize termites, they may have a regulatory effect on natural termite populations. Particularly, O.bispora, for which field investigations have revealed a high infection rate against termites, and laboratory experiments have also shown that it can effectively kill termites (Blackwell and Gilbertson 1984; Suh et al. 1998). Although laboratory experiments have not been conducted with O.longistipes, field observations have found that termites infected by this fungus often appear in groups. This may indicate that it has strong lethality against termites and possesses the potential to become a biological control agent for termites.

Supplementary Material

XML Treatment for Ophiocordyceps longistipes
XML Treatment for Ophiocordyceps globiperitheciata

Acknowledgements

The authors gratefully acknowledge Mr. Maolin Yan, Mr. Shouhua Cun, Mr. Haijun Yin, and Ms. Zhaolin Yang of the Tongbiguan Provincial Nature Reserve in Yunnan for their invaluable assistance and support during the sample collection process.

Citation

Fan Q, Yang T, Li H, Wang X-M, Liao H-F, Shen P-H, Yang Z-L, Zeng W-B, Wang Y-B (2024) Molecular phylogeny and morphology reveal two new entomopathogenic species of Ophiocordyceps (Ophiocordycipitaceae, Hypocreales) parasitic on termites from China. MycoKeys 103: 1–24. https://doi.org/10.3897/mycokeys.103.116153

Funding Statement

This work was financially supported by the Science and Technology Planning Project of Yunnan Province (202207AB110016, 202001BA070001-078), the High Level Talent Introduction Plan, Kunming Institute of Botany, CAS (E16N61), and the Innovation Project of Guangxi Graduate Education (YCBZ2022028).

Contributor Information

Wen-Bo Zeng, Email: zengwenboherb@163.com.

Yuan-Bing Wang, Email: wangyuanbing@mail.kib.ac.cn.

Additional information

Conflict of interest

The authors have declared that no competing interests exist.

Ethical statement

No ethical statement was reported.

Funding

This work was financially supported by the Science and Technology Planning Project of Yunnan Province (202207AB110016, 202001BA070001-078), the High Level Talent Introduction Plan, Kunming Institute of Botany, CAS (E16N61), and the Innovation Project of Guangxi Graduate Education (YCBZ2022028).

Author contributions

All authors contributed to the study conception and design. Material preparation, data collection and analysis were performed by Xue-Mei Wang, He-Fa Liao, Qi Fan and Tao Yang. The first draft of the manuscript was written by Qi Fan and Tao Yang. Pei-Hong Shen, Zhu-Liang Yang, Wen-Bo Zeng, and Yuan-Bing Wang reviewed and revised the manuscript. All authors commented on previous versions of the manuscript.

Author ORCIDs

Qi Fan https://orcid.org/0000-0003-3168-0347

Tao Yang https://orcid.org/0009-0000-8579-1574

Hui Li https://orcid.org/0000-0003-4255-8312

Xue-Mei Wang https://orcid.org/0009-0001-0171-4924

He-Fa Liao https://orcid.org/0009-0006-7287-1384

Pei-Hong Shen https://orcid.org/0000-0003-0980-9562

Zhu-Liang Yang https://orcid.org/0000-0001-9745-8453

Wen-Bo Zeng https://orcid.org/0009-0000-0251-5890

Yuan-Bing Wang https://orcid.org/0000-0002-3305-9418

Data availability

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

References

  1. Afzal M, Farman M, Rasib KZ, Qureshi NA. (2019) Biocidal action of silver oak (Grevillearobusta) leaf extract on the termite Heterotermesindicola wasmann (Blattodea: Rhinotermitidae). International Biodeterioration & Biodegradation 139: 1–10. 10.1016/j.ibiod.2019.02.001 [DOI] [Google Scholar]
  2. Araújo JPM, Hughes DP. (2016) Diversity of entomopathogenic fungi: Which groups conquered the insect body? Advances in Genetics 94: 1–39. 10.1016/bs.adgen.2016.01.001 [DOI] [PubMed]
  3. Araújo JPM, Evans HC, Kepler R, Hughes DP. (2018) Zombie-ant fungi across continents: 15 new species and new combinations within Ophiocordyceps. I. Myrmecophilous hirsutelloid species. Studies in Mycology 90(1): 119–160. 10.1016/j.simyco.2017.12.002 [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Araújo JPM, Moriguchi MG, Uchiyama S, Kinjo N, Matsuura Y. (2021) Ophiocordycepssalganeicola, a parasite of social cockroaches in Japan and insights into the evolution of other closely-related Blattodea-associated lineages. IMA Fungus 12(1): 1–17. 10.1186/s43008-020-00053-9 [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Ban S, Sakane T, Nakagiri A. (2015) Three new species of Ophiocordyceps and overview of anamorph types in the genus and the family Ophiocordyceptaceae. Mycological Progress 14(1): 1–12. 10.1007/s11557-014-1017-8 [DOI] [Google Scholar]
  6. Bischoff JF, Rehner SA, Humber RA. (2006) Metarhiziumfrgidum sp. nov.: A cryptic species of M.anisopliae and a member of the M.flavoviride complex. Mycologia 98(5): 737–745. 10.1080/15572536.2006.11832645 [DOI] [PubMed] [Google Scholar]
  7. Blackwell M, Gilbertson RL. (1981) Cordycepioideusoctosporus, a termite suspected pathogen from Jalisco, Mexico. Mycologia 73(2): 358–362. 10.1080/00275514.1981.12021355 [DOI] [Google Scholar]
  8. Blackwell M, Gilbertson RL. (1984) New information on Cordycepioideusbisporus and Cordycepioideusoctosporus. Mycologia 76(4): 763–765. 10.1080/00275514.1984.12023912 [DOI] [Google Scholar]
  9. Bronstein JL. (2009) The evolution of facilitation and mutualism. Journal of Ecology 97(6): 1160–1170. 10.1111/j.1365-2745.2009.01566.x [DOI] [Google Scholar]
  10. Cerezer FO, de Azevedo RA, Nascimento MAS, Franklin E, de Morais JW, de Sales Dambros C. (2020) Latitudinal gradient of termite diversity indicates higher diversification and narrower thermal niches in the tropics. Global Ecology and Biogeography 29(11): 1967–1977. 10.1111/geb.13167 [DOI] [Google Scholar]
  11. Chernomor O, Von Haeseler A, Minh BQ. (2016) Terrace aware data structure for phylogenomic inference from supermatrices. Systematic Biology 65(6): 997–1008. 10.1093/sysbio/syw037 [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Da Costa RR, Vreeburg SME, Shik JZ, Aanen DK, Poulsen M. (2019) Can interaction specificity in the fungus-farming termite symbiosis be explained by nutritional requirements of the fungal crop? Fungal Ecology 38: 54–61. 10.1016/j.funeco.2018.08.009 [DOI]
  13. Evans HC. (1974) Natural control of arthropods, with special reference to ants (Formicidae), by fungi in the tropical high forest of Ghana. Journal of Applied Ecology 11(1): 37–49. 10.2307/2402003 [DOI] [Google Scholar]
  14. Evans HC, Araújo JPM, Halfeld VR, Hughes DP. (2018) Epitypification and re-description of the zombie-ant fungus, Ophiocordycepsunilateralis (Ophiocordycipitaceae). Fungal Systematics and Evolution 1(1): 13–22. 10.3114/fuse.2018.01.02 [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Haelewaters D, Kasson MT. (2020) Animal-associated fungi. Mycologia 112(6): 1045–1047. 10.1080/00275514.2020.1841469 [DOI] [PubMed] [Google Scholar]
  16. Hoang DT, Chernomor O, Von Haeseler A, Minh BQ, Vinh LS. (2018) UFBoot2: Improving the ultrafast bootstrap approximation. Molecular Biology and Evolution 35(2): 518–522. 10.1093/molbev/msx281 [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Hopple JS. (1994) Phylogenetic investigations in the genus Coprinus based on morphological and molecular characters. PhD dissertation, Duke University.
  18. Johnson D, Sung GH, Hywel-Jones NL, Luangsa-ard JJ, Bischoff JF, Kepler RM, Spatafora JW. (2009) Systematics and evolution of the genus Torrubiella (Hypocreales, Ascomycota). Mycological Research 113(3): 279–289. 10.1016/j.mycres.2008.09.008 [DOI] [PubMed] [Google Scholar]
  19. Kalyaanamoorthy S, Bui Quang M, Wong TKF, von Haeseler A, Jermiin LS. (2017) ModelFinder: Fast model selection for accurate phylogenetic estimates. Nature Methods 14(6): 587–589. 10.1038/nmeth.4285 [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. 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. 10.1093/molbev/mst010 [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Kepler RM, Kaitsu Y, Tanaka E, Shimano S, Spatafora JW. (2011) Ophiocordycepspulvinata sp. nov., a pathogen of ants with a reduced stroma. Mycoscience 52(1): 39–47. 10.1007/S10267-010-0072-5 [DOI] [Google Scholar]
  22. Kepler RM, Sung GH, Ban S, Nakagiri A, Chen MJ, Huang B, Li Z, Spatafora JW. (2012) New teleomorph combinations in the entomopathogenic genus Metacordyceps. Mycologia 104(1): 182–197. 10.3852/11-070 [DOI] [PubMed] [Google Scholar]
  23. Khao-ngam S, Mongkolsamrit S, Rungjindamai N, Noisripoom W, Pooissarakul W, Duangthisan J, Himaman W, Luangsa-ard JJ. (2021) Ophiocordycepsasiana and Ophiocordycepstessaratomidarum (Ophiocordycipitaceae, Hypocreales), two new species on stink bugs from Thailand. Mycological Progress 20(3): 341–353. 10.1007/s11557-021-01684-x [DOI] [Google Scholar]
  24. Khonsanit A, Luangsa-ard JJ, Thanakitpipattana D, Kobmoo N, Piasai O. (2019) Cryptic species within Ophiocordycepsmyrmecophila complex on formicine ants from Thailand. Mycological Progress 18(1–2): 147–161. 10.1007/s11557-018-1412-7 [DOI] [Google Scholar]
  25. 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. 10.1093/molbev/msw054 [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Li GJ, Hyde KD, Zhao RL, Hongsanan S, Abdel-Aziz FA, Abdel-Wahab MA, Alvarado P, Alves-Silva G, Ammirati JF, Ariyawansa HA, Baghela A, Bahkali AH, Beug M, Bhat DJ, Bojantchev D, Boonpratuang T, Bulgakov TS, Camporesi E, Boro MC, Ceska O, Chakraborty D, Chen JJ, Chethana KWT, Chomnunti P, Consiglio G, Cui BK, Dai DQ, Dai YC, Daranagama DA, Das K, Dayarathne MC, De Crop E, De Oliveira RJV, Fragoso de Souza CA, de Souza JI, Dentinger BTM, Dissanayake AJ, Doilom M, Drechsler-Santos ER, Ghobad-Nejhad M, Gilmore SP, Goes-Neto A, Gorczak M, Haitjema CH, Hapuarachchi KK, Hashimoto A, He MQ, Henske JK, Hirayama K, Iribarren MJ, Jayasiri SC, Jayawardena RS, Jeon SJ, Jernimo GH, Jesus AL, Jones EBG, Kang JC, Karunarathna SC, Kirk PM, Konta S, Kuhnert E, Langer E, Lee HS, Lee HB, Li WJ, Li XH, Liimatainen K, Lima DX, Lin CG, Liu JK, Liu XZ, Liu ZY, Luangsa-ard JJ, Luecking R, Lumbsch HT, Lumyong S, Leano EM, Marano AV, Matsumura M, McKenzie EHC, Mongkolsamrit S, Mortimer PE, Thi Thuong Thuong N, Niskanen T, Norphanphoun C, O’Malley MA, Parnmen S, Pawlowska J, Perera RH, Phookamsak R, Phukhamsakda C, Pires-Zottarelli CLA, Raspe O, Reck MA, Rocha SCO, de Santiago ALCMA, Senanayake IC, Setti L, Shang QJ, Singh SK, Sir EB, Solomon KV, Song J, Srikitikulchai P, Stadler M, Suetrong S, Takahashi H, Takahashi T, Tanaka K, Tang LP, Thambugala KM, Thanakitpipattana D, Theodorou MK, Thongbai B, Thummarukcharoen T, Tian Q, Tibpromma S, Verbeken A, Vizzini A, Vlasak J, Voigt K, Wanasinghe DN, Wang Y, Weerakoon G, Wen HA, Wen TC, Wijayawardene NN, Wongkanoun S, Wrzosek M, Xiao YP, Xu JC, Yan JY, Yang J, Yang SD, Hu Y, Zhang JF, Zhao J, Zhou LW, Persoh D, Phillips AJL, Maharachchikumbura SSN. (2016) Fungal diversity notes 253–366: Taxonomic and phylogenetic contributions to fungal taxa. Fungal Diversity 78(1): 1–237. 10.1007/s13225-016-0366-9 [DOI] [Google Scholar]
  27. Luangsa-ard JJ, Ridkaew R, Mongkolsamrit S, Tasanathai K, Hywel-Jones NL. (2010) Ophiocordycepsbarnesii and its relationship to other melolonthid pathogens with dark stromata. Fungal Biology 114(9): 739–745. 10.1016/j.funbio.2010.06.007 [DOI] [PubMed] [Google Scholar]
  28. Luangsa-ard JJ, Ridkaew R, Tasanathai K, Thanakitpipattana D, Hywel-Jones N. (2011) Ophiocordycepshalabalaensis: A new species of Ophiocordyceps pathogenic to Camponotusgigas in Hala Bala Wildlife Sanctuary, Southern Thailand. Fungal Biology 115(7): 608–614. 10.1016/j.funbio.2011.03.002 [DOI] [PubMed] [Google Scholar]
  29. Luangsa-ard JJ, Tasanathai K, Thanakitpipattana D, Khonsanit A, Stadler M. (2018) Novel and interesting Ophiocordyceps spp. (Ophiocordycipitaceae, Hypocreales) with superficial perithecia from Thailand. Studies in Mycology 89(1): 125–142. 10.1016/j.simyco.2018.02.001 [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Martelossi J, Forni G, Iannello M, Savojardo C, Martelli PL, Casadio R, Mantovani B, Luchetti A, Rota-Stabelli O. (2023) Wood feeding and social living: Draft genome of the subterranean termite Reticulitermeslucifugus (Blattodea; Termitoidae). Insect Molecular Biology 32(2): 118–131. 10.1111/imb.12818 [DOI] [PubMed] [Google Scholar]
  31. Meyling NV, Eilenberg J. (2007) Ecology of the entomopathogenic fungi Beauveriabassiana and Metarhiziumanisopliae in temperate agroecosystems: Potential for conservation biological control. Biological Control 43(2): 145–155. 10.1016/j.biocontrol.2007.07.007 [DOI] [Google Scholar]
  32. Mongkolsamrit S, Noisripoom W, Pumiputikul S, Boonlarppradab C, Samson RA, Stadler M, Becker K, Luangsa-ard JJ. (2021) Ophiocordycepsflavida sp. nov. (Ophiocordycipitaceae), a new species from Thailand associated with Pseudogibellulaformicarum (Cordycipitaceae), and their bioactive secondary metabolites. Mycological Progress 20(4): 477–492. 10.1007/s11557-021-01683-y [DOI] [Google Scholar]
  33. Mongkolsamrit S, Noisripoom W, Hasin S, Sinchu P, Jangsantear P, Luangsa-ard JJ. (2023) Multi-gene phylogeny and morphology of Ophiocordycepslaotii sp. nov. and a new record of O.buquetii (Ophiocordycipitaceae, Hypocreales) on ants from Thailand. Mycological Progress 22(1): 1–5. 10.1007/s11557-022-01855-4 [DOI] [Google Scholar]
  34. Moon JH, Ajuna HB, Won SJ, Choub V, Choi SI, Yun JY, Hwang WJ, Park SW, Ahn YS. (2023) Entomopathogenic potential of Bacillusvelezensis CE 100 for the biological control of termite damage in wooden architectural buildings of korean cultural heritage. International Journal of Molecular Sciences 24(9): e8189. 10.3390/ijms24098189 [DOI] [PMC free article] [PubMed]
  35. Nguyen LT, Schmidt HA, Von Haeseler A, Minh BQ. (2015) IQ-TREE: A fast and effective stochastic algorithm for estimating maximum-likelihood phylogenies. Molecular Biology and Evolution 32(1): 268–274. 10.1093/molbev/msu300 [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Ochiel GS, Evans HC, Eilenberg J. (1997) Cordycepioideus, a pathogen of termites in Kenya. The Mycologist 1(11): 7–9. 10.1016/S0269-915X(97)80059-6 [DOI] [Google Scholar]
  37. Oi F. (2022) A review of the evolution of termite control: A continuum of alternatives to termiticides in the United States with emphasis on efficacy testing requirements for product registration. Insects 13(1): 1–50. 10.3390/insects13010050 [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Pearce MJ. (1997) Termites: Biology and Pest Management. CAB International, Wallingford, 172 pp. 10.1079/9780851991306.0000 [DOI] [Google Scholar]
  39. Penzig O, Saccardo PA. (1904) Icones Fungorum Javanicorum. Buchhandlung and Druckerei EJ Brill, Leiden, 55–57. 10.5962/bhl.title.17234 [DOI]
  40. Petch T. (1931) Notes on entomogenous fungi. Transactions of the British Mycological Society 16(1): 55–75. 10.1016/S0007-1536(31)80006-3 [DOI] [Google Scholar]
  41. Quandt CA, Kepler RM, Gams W, Araújo JPM, Ban S, Evans HC, Hughes D, Humber R, Hywel-Jones N, Li Z, Luangsa-ard JJ, Rehner SA, Sanjuan T, Sato H, Shrestha B, Sung GH, Yao YJ, Zare R, Spatafora JW. (2014) Phylogenetic-based nomenclatural proposals for Ophiocordycipitaceae (Hypocreales) with new combinations in Tolypocladium. IMA Fungus 5(1): 121–134. 10.5598/imafungus.2014.05.01.12 [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Rath AC. (2000) The use of entomopathogenic fungi for control of termites. Biocontrol Science and Technology 10(5): 563–581. 10.1080/095831500750016370 [DOI] [Google Scholar]
  43. 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. 10.1093/sysbio/sys029 [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Roper C, Castro C, Ingel B. (2019) Xylellafastidiosa: Bacterial parasitism with hallmarks of commensalism. Current Opinion in Plant Biology 50: 140–147. 10.1016/j.pbi.2019.05.005 [DOI] [PubMed] [Google Scholar]
  45. Rust MK, Su NY. (2012) Managing social insects of urban importance. Annual Review of Entomology 57(1): 355–375. 10.1146/annurev-ento-120710-100634 [DOI] [PubMed] [Google Scholar]
  46. Sanjuan TI, Franco-Molano AE, Kepler RM, Spatafora JW, Tabima J, Vasco-Palacios AM, Restrepo S. (2015) Five new species of entomopathogenic fungi from the Amazon and evolution of neotropical Ophiocordyceps. Fungal Biology 119(10): 901–916. 10.1016/j.funbio.2015.06.010 [DOI] [PubMed] [Google Scholar]
  47. Santamaria B, Verbeken A, Haelewaters D. (2023) Mycophagy: A Global Review of Interactions between Invertebrates and Fungi. Journal of Fungi 9(2): e163. 10.3390/jof9020163 [DOI] [PMC free article] [PubMed]
  48. Scharf ME. (2015) Termites as targets and models for biotechnology. Annual Review of Entomology 60(1): 77–102. 10.1146/annurev-ento-010814-020902 [DOI] [PubMed] [Google Scholar]
  49. Schoch CL, Seifert KA, Huhndorf S, Robert V, Spouge JL, Levesque CA, Chen W, Bolchacova E, Voigt K, Crous PW, Miller AN, Wingfield MJ, Aime MC, An KD, Bai FY, Barreto RW, Begerow D, Bergeron MJ, Blackwell M, Boekhout T, Bogale M, Boonyuen N, Burgaz AR, Buyck B, Cai L, Cai Q, Cardinali G, Chaverri P, Coppins BJ, Crespo A, Cubas P, Cummings C, Damm U, de Beer ZW, de Hoog GS, Del-Prado R, Dentinger B, Dieguez-Uribeondo J, Divakar PK, Douglas B, Duenas M, Duong TA, Eberhardt U, Edwards JE, Elshahed MS, Fliegerova K, Furtado M, Garcia MA, Ge ZW, Griffith GW, Griffiths K, Groenewald JZ, Groenewald M, Grube M, Gryzenhout M, Guo LD, Hagen F, Hambleton S, Hamelin RC, Hansen K, Harrold P, Heller G, Herrera G, Hirayama K, Hirooka Y, Ho HM, Hoffmann K, Hofstetter V, Hognabba F, Hollingsworth PM, Hong SB, Hosaka K, Houbraken J, Hughes K, Huhtinen S, Hyde KD, James T, Johnson EM, Johnson JE, Johnston PR, Jones EB, Kelly LJ, Kirk PM, Knapp DG, Koljalg U, Kovacs GM, Kurtzman CP, Landvik S, Leavitt SD, Liggenstoffer AS, Liimatainen K, Lombard L, Luangsa-ard JJ, Lumbsch HT, Maganti H, Maharachchikumbura SS, Martin MP, May TW, McTaggart AR, Methven AS, Meyer W, Moncalvo JM, Mongkolsamrit S, Nagy LG, Nilsson RH, Niskanen T, Nyilasi I, Okada G, Okane I, Olariaga I, Otte J, Papp T, Park D, Petkovits T, Pino-Bodas R, Quaedvlieg W, Raja HA, Redecker D, Rintoul TL, Ruibal C, Sarmiento-Ramirez JM, Schmitt I, Schussler A, Shearer C, Sotome K, Stefani FOP, Stenroos S, Stielow B, Stockinger H, Suetrong S, Suh SO, Sung GH, Suzuki M, Tanaka K, Tedersoo L, Telleria MT, Tretter E, Untereiner WA, Urbina H, Vagvolgyi C, Vialle A, Vu TD, Walther G, Wang QM, Wang Y, Weir BS, Weiss M, White MM, Xu J, Yahr R, Yang ZL, Yurkov A, Zamora JC, Zhang N, Zhuang WY, Schindel D, Fungal Barcoding C. (2012) Nuclear ribosomal internal transcribed spacer (ITS) region as a universal DNA barcode marker for Fungi. Proceedings of the National Academy of Sciences of the United States of America 109(16): 6241–6246. 10.1073/pnas.1117018109 [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Shimazu M, Tsuchiya D, Sato H, Kushida T. (1995) Microbial control of Monochamusalternatus Hope (Coleoptera: Cerambycidae) by application of nonwoven fabric strips with Beauveriabassiana (Deuteromycotina: Hyphomycetes) on infested tree trunks. Applied Entomology and Zoology 30(1): 207–213. 10.1303/aez.30.207 [DOI] [Google Scholar]
  51. Shrestha B, Tanaka E, Hyun MW, Han JG, Kim CS, Jo JW, Han SK, Oh J, Sung GH. (2016) Coleopteran and lepidopteran hosts of the entomopathogenic genus Cordyceps sensu lato. Journal of Mycology 2016: 1–14. 10.1155/2016/7648219 [DOI] [Google Scholar]
  52. Simmons DR, Kepler RM, Rehner SA, Groden E. (2015) Phylogeny of Hirsutella species (Ophiocordycipitaceae) from the USA: Remedying the paucity of Hirsutella sequence data. IMA Fungus 6(2): 345–356. 10.5598/imafungus.2015.06.02.06 [DOI] [PMC free article] [PubMed] [Google Scholar]
  53. Spatafora JW, Sung GH, Sung JM, Hywel-Jones NL, White Jr JF. (2007) Phylogenetic evidence for an animal pathogen origin of ergot and the grass endophytes. Molecular Ecology 16(8): 1701–1711. 10.1111/j.1365-294X.2007.03225.x [DOI] [PubMed] [Google Scholar]
  54. Spatafora JW, Quandt CA, Kepler RM, Sung GH, Shrestha B, Hywel-Jones NL, Luangsa-ard JJ. (2015) New 1F1N species combinations in Ophiocordycipitaceae (Hypocreales). IMA Fungus 6(2): 357–362. 10.5598/imafungus.2015.06.02.07 [DOI] [PMC free article] [PubMed] [Google Scholar]
  55. Suh SO, Spatafora JW, Ochiel GRS, Evans HC, Blackwell M. (1998) Molecular phylogenetic study of a termite pathogen Cordycepioideusbisporus. Mycologia 90(4): 611–617. 10.1080/00275514.1998.12026950 [DOI] [Google Scholar]
  56. Sung GH, Hywel-Jones NL, Sung JM, Luangsa-ard JJ, Shrestha B, Spatafora JW. (2007a) Phylogenetic classification of Cordyceps and the clavicipitaceous fungi. Studies in Mycology 57: 5–59. 10.3114/sim.2007.57.01 [DOI] [PMC free article] [PubMed] [Google Scholar]
  57. Sung GH, Sung JM, Hywel-Jones NL, Spatafora JW. (2007b) A multi-gene phylogeny of Clavicipitaceae (Ascomycota, Fungi): Identification of localized incongruence using a combinational bootstrap approach. Molecular Phylogenetics and Evolution 44(3): 1204–1223. 10.1016/j.ympev.2007.03.011 [DOI] [PubMed] [Google Scholar]
  58. Tang DX, Zhu JY, Luo LY, Hou DH, Wang ZQ, Yang SD, Yu H. (2022) Ophiocordycepsovatospora sp. nov. (Ophiocordycipitaceae, Hypocreales), pathogenic on termites from China. Phytotaxa 574(1): 105–117. 10.11646/phytotaxa.574.1.8 [DOI] [Google Scholar]
  59. Tang DX, Huang O, Zou WQ, Wang YB, Wang Y, Dong QY, Sun T, Yang G, Yu H. (2023a) Six new species of zombie-ant fungi from Yunnan in China. IMA Fungus 14(1): 1–9. 10.1186/s43008-023-00114-9 [DOI] [PMC free article] [PubMed] [Google Scholar]
  60. Tang DX, Xu ZH, Wang Y, Wang YB, Tran NL, Yu H. (2023b) Multigene phylogeny and morphology reveal two novel zombie-ant fungi in Ophiocordyceps (Ophiocordycipitaceae, Hypocreales). Mycological Progress 22(4): 1–22. 10.1007/s11557-023-01874-9 [DOI] [Google Scholar]
  61. Tasanathai K, Noisripoom W, Chaitika T, Khonsanit A, Hasin S, Luangsa-ard JJ. (2019) Phylogenetic and morphological classification of Ophiocordyceps species on termites from Thailand. MycoKeys 56: 101–129. 10.3897/mycokeys.56.37636 [DOI] [PMC free article] [PubMed] [Google Scholar]
  62. Tasanathai K, Thanakitpipattana D, Himaman W, Phommavong K, Dengkhhamounh N, Luangsa-ard JJ. (2020) Three new Ophiocordyceps species in the Ophiocordycepspseudoacicularis species complex on Lepidoptera larvae in Southeast Asia. Mycological Progress 19(10): 1043–1056. 10.1007/s11557-020-01611-6 [DOI] [Google Scholar]
  63. Tasanathai K, Khonsanit A, Noisripoom W, Kobmoo N, Luangsa-ard JJ. (2022) Hidden species behind Ophiocordyceps (Ophiocordycipitaceae, Hypocreales) on termites: Four new species from Thailand. Mycological Progress 21(10): 86–102. 10.1007/s11557-022-01837-6 [DOI] [Google Scholar]
  64. 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. 10.1128/jb.172.8.4238-4246.1990 [DOI] [PMC free article] [PubMed] [Google Scholar]
  65. Wang L, Li HH, Chen YQ, Zhang WM, Qu LH. (2014) Polycephalomyceslianzhouensis sp. nov., a new species, co-occurs with Ophiocordycepscrinalis. Mycological Progress 13(4): 1089–1096. 10.1007/s11557-014-0996-9 [DOI] [Google Scholar]
  66. Wang YB, Yu H, Dai YD, Wu CK, Zeng WB, Yuan F, Liang ZQ. (2015) Polycephalomycesagaricus, a new hyperparasite of Ophiocordyceps sp. infecting melolonthid larvae in southwestern China. Mycological Progress 14(9): 1–70. 10.1007/s11557-015-1090-7 [DOI] [Google Scholar]
  67. Wang YB, Thi Tra N, Dai YD, Yu H, Zeng WB, Wu CK. (2018) Molecular phylogeny and morphology of Ophiocordycepsunituberculata sp. nov. (Ophiocordycipitaceae), a pathogen of caterpillars (Noctuidae, Lepidoptera) from Yunnan, China. Mycological Progress 17(6): 745–753. 10.1007/s11557-017-1370-5 [DOI] [Google Scholar]
  68. Wang YB, Wang Y, Fan Q, Duan DE, Zhang GD, Dai RQ, Dai YD, Zeng WB, Chen ZH, Li DD, Tang DX, Xu ZH, Sun T, Nguyen TT, Tran NL, Dao VM, Zhang CM, Huang LD, Liu YJ, Zhang XM, Yang DR, Sanjuan T, Liu XZ, Yang ZL, Yu H. (2020) Multigene phylogeny of the family Cordycipitaceae (Hypocreales): New taxa and the new systematic position of the Chinese cordycipitoid fungus Paecilomyceshepiali. Fungal Diversity 103(1): 1–46. 10.1007/s13225-020-00457-3 [DOI] [Google Scholar]
  69. Wang Y, Dai YD, Yang ZL, Guo R, Wang YB, Yang ZL, Ding L, Yu H. (2021a) Morphological and molecular phylogenetic data of the Chinese medicinal fungus Cordycepsliangshanensis reveal its new systematic position in the family Ophiocordycipitaceae. Mycobiology 49(4): 297–307. 10.1080/12298093.2021.1923388 [DOI] [PMC free article] [PubMed] [Google Scholar]
  70. Wang Y, Wu HJ, Tran NL, Zhang GD, Souvanhnachit S, Wang YB, Yu H. (2021b) Ophiocordycepsfurcatosubulata, a new entomopathogenic fungus parasitizing beetle larvae (Coleoptera: Elateridae). Phytotaxa 482(3): 268–278. 10.11646/phytotaxa.482.3.5 [DOI] [Google Scholar]
  71. Wen TC, Zhu RC, Kang JC, Huang MH, Tan DB, Ariyawansha H, Hyde KD, Liu H. (2013) Ophiocordycepsxuefengensis sp. nov. from larvae of Phassusnodus (Hepialidae) in Hunan Province, southern China. Phytotaxa 123(1): 41–50. 10.11646/phytotaxa.123.1.2 [DOI] [Google Scholar]
  72. Wen TC, Xiao YP, Li WJ, Kang JC, Hyde KD. (2014) Systematic analyses of Ophiocordycepsramosissimum sp. nov., a new species from a larvae of Hepialidae in China. Phytotaxa 161(3): 227–234. 10.11646/phytotaxa.161.3.6 [DOI] [Google Scholar]
  73. 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, 315–322. 10.1016/B978-0-12-372180-8.50042-1 [DOI]
  74. Wijayawardene NN, Hyde KD, Lumbsch HT, Liu JK, Maharachchikumbura SSN, Ekanayaka AH, Tian Q, Phookamsak R. (2018) Outline of Ascomycota: 2017. Fungal Diversity 88(1): 167–263. 10.1007/s13225-018-0394-8 [DOI] [Google Scholar]
  75. Wilson M, Barden P, Ware J. (2021) A review of ectoparasitic fungi associated with termites. Annals of the Entomological Society of America 114(4): 373–396. 10.1093/aesa/saab001 [DOI] [Google Scholar]
  76. Xiao YP, Wen TC, Hongsanan S, Sun JZ, Hyde KD. (2017) Introducing Ophiocordycepsthanathonensis, a new species of entomogenous fungi on ants, and a reference specimen for O.pseudolloydii. Phytotaxa 328(2): 115–126. 10.11646/phytotaxa.328.2.2 [DOI] [Google Scholar]
  77. Xiao YP, Hongsanan S, Hyde KD, Brooks S, Xie N, Long FY, Wen TC. (2019) Two new entomopathogenic species of Ophiocordyceps in Thailand. MycoKeys 47: 53–74. 10.3897/mycokeys.47.29898 [DOI] [PMC free article] [PubMed] [Google Scholar]
  78. Xiao YP, Wang YB, Hyde KD, Eleni G, Sun JZ, Yang Y, Meng J, Yu H, Wen TC. (2023) Polycephalomycetaceae, a new family of clavicipitoid fungi segregates from Ophiocordycipitaceae. Fungal Diversity 120(1): 1–76. 10.1007/s13225-023-00517-4 [DOI] [Google Scholar]
  79. Xu ZH, Tran NL, Wang Y, Zhang GD, Dao VM, Nguyen TT, Wang YB, Yu H. (2022) Phylogeny and morphology of Ophiocordycepspuluongensis sp. nov. (Ophiocordycipitaceae, Hypocreales), a new fungal pathogen on termites from Vietnam. Journal of Invertebrate Pathology 192: e107771. 10.1016/j.jip.2022.107771 [DOI] [PubMed]
  80. Zha LS, Kryukov VY, Ding JH, Jeewon R, Chomnunti P. (2021) Novel taxa and species diversity of Cordyceps sensu lato (Hypocreales, Ascomycota) developing on wireworms (Elateroidea and Tenebrionoidea, Coleoptera). MycoKeys 78: 79–117. 10.3897/mycokeys.78.61836 [DOI] [PMC free article] [PubMed] [Google Scholar]
  81. Zou J, Wu L, He ZM, Zhang P, Chen ZH. (2017) Determination of the main nucleosides and nucleobases in natural and cultured Ophiocordycepsxuefengensis. Molecules 22(9): e1530. 10.3390/molecules22091530 [DOI] [PMC free article] [PubMed]
  82. Zou WQ, Tang DX, Xu ZH, Huang O, Wang YB, Tran NL, Yu H. (2022) Multigene phylogeny and morphology reveal Ophiocordycepshydrangea sp. nov. and Ophiocordycepsbidoupensis sp. nov. (Ophiocordycipitaceae). MycoKeys 92: 109–130. 10.3897/mycokeys.92.86160 [DOI] [PMC free article] [PubMed] [Google Scholar]

Associated Data

This section collects any data citations, data availability statements, or supplementary materials included in this article.

Supplementary Materials

XML Treatment for Ophiocordyceps longistipes
mycokeys.103.116153-treatment1.xml (48.6KB, xml)
XML Treatment for Ophiocordyceps globiperitheciata
mycokeys.103.116153-treatment2.xml (45.8KB, xml)

Data Availability Statement

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

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