Additions to the Genus Rhytidhysteron in Hysteriaceae Author(s): Kasun M. Thambugala , Kevin D. Hyde , Prapassorn D. Eungwanichayapant , Andrea I. Romero & Zuo-Yi Liu Source: Cryptogamie, Mycologie, 37(1):99-116. Published By: Association des Amis des Cryptogames https://doi.org/10.7872/crym/v37.iss1.2016.99 URL: http://www.bioone.org/doi/full/10.7872/crym/v37.iss1.2016.99 BioOne (www.bioone.org) is a nonprofit, online aggregation of core research in the biological, ecological, and environmental sciences. BioOne provides a sustainable online platform for over 170 journals and books published by nonprofit societies, associations, museums, institutions, and presses. Your use of this PDF, the BioOne Web site, and all posted and associated content indicates your acceptance of BioOne’s Terms of Use, available at www.bioone.org/page/terms_of_use. Usage of BioOne content is strictly limited to personal, educational, and non- commercial use. Commercial inquiries or rights and permissions requests should be directed to the individual publisher as copyright holder. BioOne sees sustainable scholarly publishing as an inherently collaborative enterprise connecting authors, nonprofit publishers, academic institutions, research libraries, and research funders in the common goal of maximizing access to critical research. Cryptogamie, Mycologie, 2016, 37 (1): 99-116 © 2016 Adac. Tous droits réservés $GGLWLRQVWRWKHJHQXVRhytidhysteronLQ+\VWHULDFHDH Kasun M. THAMBUGALA a,b,c, Kevin D. HYDEb,c,d, Prapassorn D. EUNGWANICHAYAPANT c, Andrea I. ROMERO e & Zuo-Yi LIU a* aGuizhou Key Laboratory of Agricultural Biotechnology, Guizhou Academy of Agricultural Sciences, Guiyang, Guizhou 550006, People’s Republic of China bCenter of Excellence in Fungal Research, Chiang Rai 57100, Thailand cSchool of Science, Mae Fah Luang University, Chiang Rai 57100, Thailand dKey Laboratory for Plant Diversity and Biogeography of East Asia, Kunming Institute of Botany, Chinese Academy of Science, Kunming 650201, Yunnan, China ePrhideb-Conicet, Deptomento Cs. Biológicas,Facultad de Cs. Exactas y Naturales (UBA), Ciudad Universitaria,Pabellón II, 4to Piso,1428 Buenos Aires, Argentina $EVWUDFW– Rhytidhysteron (Hysteriaceae) species are widely distributed as saprobes or weak pathogens on a wide range of woody plants. In this study, several Rhytidhysteron collections were made in northern Thailand and multi-gene phylogenetic analyses were used to resolve the phylogenetic boundaries of species. Two novel species, R. thailandicum and R. neorufulum are introduced, based on morphological traits and multi-gene phylogeny. The genus is revised with a key to species. +\VWHULDOHV3K\ORJHQ\QHZVSHFLHVPXOWLJHQHSK\ORJHQ\7KDLODQG ,1752'8&7,21 Hysteriaceous fungi are primarily lignicolous or corticolous and are saprobes on wide host range in temperate and tropical regions (Boehm et al., 2009a, b; Murillo et al., 2009; Hyde et al., 2013; de Almeida et al/LXet al., 2015; Yacharoen et al., 2015). The family Hysteriaceae was introduced by Chevallier (1826) as “Hysterineae” and this family has been treated with different genera by various authors (Zogg 1962; von Arx & Müller 1975; Kirk et al., 2001; Eriksson /XPEVFKDQG+XKQGRUI 5HFHQWPROHFXODUSK\ORJHQHWLFDQDO\VHVSODFHG Hysteriaceae in Hysteriales, Pleosporomycetidae (Boehm et al., 2009a, b; Hyde et al., 2013; Wijayawardene et al., 2014). Hyde et al. (2013) and Wijayawardene et al. (2014) accepted 13 genera including Actidiographium /DU1 9DVVLOMHYD Coniosporium /LQN Gloniella Sacc., Gloniopsis De Not., Hysterium Pers., Hysterobrevium(%RHKP &/6FKRFKHysterocarina H. Zogg, Hysteropycnis Hilitzer, Oedohysterium(%RHKP &/6FKRFKOstreichnion Duby, Psiloglonium * Corresponding author: [email protected] doi/10.7872/crym/v37.iss1.2016.99 100 K. M. Thambugala et al. Höhn. Rhytidhysteron Speg. and Sphaeronaema Fr in the family, while de Almeida et al., (2014) introduced a new genus Hysterodifractum. Hysteriaceae is characterized E\ FDUERQDFHRXV LPPHUVHG WR HUXPSHQW WR HQWLUHO\ VXSHU¿FLDO K\VWHURWKHFLD distinctly navicular in outline, bearing a pronounced, longitudinal slit, running the length of the long axis, bitunicate asci and hyaline to pigmented, 1 to multi-septate, or muriform ascospores (Boehm et al., 2009a, b; Hyde et al., 2013; de Almeida et al., 2014). Rhytidhysteron was introduced by Spegazzini (1881) to accommodate R. brasiliense Speg. and R. viride Speg., without a type species being designated. Subsequently, Clements & Shear (1931) designated R. brasiliense as the type species (Samuels & Müller 1979; Yacharoen et al /DWHUVHYHUDOVSHFLHVZHUHDGGHG to the genus based on morphology (Spegazzini 1921; Sharma & Rawla 1985; Barr 1990; Magnes 1997) and currently there are 17 epithets listed in Index Fungorum 9DULRXVDXWKRUVKDYHFODVVL¿HGRhytidhysteron within the family Patellariaceae %DUU.XWRUJD +DZNVZRUWK(ULNVVRQ/XPEVFKDQG+XKQGRUI 2010). However, recent multi-gene phylogenetic studies have shown that Rhytidhysteron should be placed in Hysteriaceae (Boehm et al., 2009a, b, de Almeida et al., 2014; Wijayawardene et al., 2014). The genus Rhytidhysteron is characterized by closed and navicular ascomata, later opening by a longitudinal slit to become irregularly apothecioid at maturity and heavily pigmented, and with thick-walled ascospores (Boehm et al., 2009b). Aposphaeria-like’ or diplodia-like’ asexual morphs have been reported for Rhytidhysteron (Samuels & Müller 1979). In this study, we made several Rhytidhysteron collections on dead woody branches in northern Thailand. We used morphological characters and multi-gene molecular analyses to resolve species in the genus. 0$7(5,$/6$1'0(7+2'6 Collection of samples, isolation and morphological examination Fresh specimens were collected from dead wood in northern Thailand and samples were grown on potato dextrose agar (PDA). Isolates were derived via single spore isolation following the protocols of Chomnunti et al., (2014). Germinating spores were transferred to PDA and incubated at 25°C in the dark. Cultural characteristics, such as mycelium colour, shape, texture and growth rate were determined. Type and voucher specimens are deposited in the herbarium of Mae Fah /XDQJ8QLYHUVLW\ 0)/8 &KLDQJ5DLDQG1HZ=HDODQG)XQJDO 3ODQW'LVHDVH &ROOHFWLRQ 3'' ZKLOH FXOWXUHV DUH GHSRVLWHG DW WKH 0DH )DK /XDQJ 8QLYHUVLW\ &XOWXUH &ROOHFWLRQ 0)/8&& ZLWK GXSOLFDWHV LQ ,QWHUQDWLRQDO &ROOHFWLRQ RI 0LFURRUJDQLVPVIURP3ODQWV ,&03 /DQGFDUH5HVHDUFK1HZ=HDODQG Specimens were observed and examined with a Motic SMZ 168 stereomicroscope. Micro-morphological characters of the taxon were examined with D 1LNRQ (&/,36( L FRPSRXQG PLFURVFRSH DQG LPDJHV ZHUH FDSWXUHG XVLQJ D 1LNRQ(&/,36(LFRPSRXQGPLFURVFRSHZLWKD&DQRQ(26'GLJLWDOFDPHUD Observations and photographs were made from material mounted in water and Indian ink was added to water mounts to show the presence of gelatinous sheaths around the ascospores. Measurements were made with the Tarosoft (R) Image Frame :RUN DQG LPDJHV XVHG IRU ¿JXUHV ZHUH SURFHVVHG ZLWK $GREH 3KRWRVKRS &6 Additions to the genus Rhytidhysteron in Hysteriaceae 101 Extended version 10.0 software. Faces of fungi numbers and Index Fungorum numbers are provided as detailed in Jayasiri et al. (2015) and Index Fungorum (2016). '1$H[WUDFWLRQ3&5DPSOL¿FDWLRQDQGVHTXHQFLQJ Fresh fungal mycelium was grown on PDA at 25°C for 21 days. Extraction of genomic DNA from mycelia and sequencing of PCR products were carried out following the method of Thambugala et al. (2015). Four partial gene portions were DPSOL¿HGLQWKLVVWXG\LQFOXGLQJ/525DQG/5 9LOJDO\VDQG+HVWHU IRUWKH QXFOHDU ULERVRPDO ODUJH VXEXQLW /68 ,76 DQG ,76 :KLWH et al., 1990) for the internal transcribed spacer (ITS), EF1-983F and EF1-2218R (Carbone and Kohn IRU WUDQVODWLRQ HORQJDWLRQ IDFWRU DOSKD ()Į DQG 16 DQG 16 :KLWH et al IRUWKHQXFOHDUULERVRPDOVPDOOVXEXQLW 668 7KHDPSOL¿FDWLRQVZHUH × SHUIRUPHG LQ ȝ/ RI 3&5 PL[WXUHV FRQWDLQLQJ ȝ/ GG+22 ȝ/ PCR 0DVWHU0L[ 7,$1*(1&R&KLQD ȝ/RI'1$WHPSODWHȝ/RIHDFKSULPHU ȝ0 7KHDPSOL¿FDWLRQUHDFWLRQVZHUHSHUIRUPHGDQGDQDO\]HGDVGHVFULEHGE\ Thambugala et al., (2015). Phylogenetic analyses The phylogenies of the new strains were determined using two analyses: 7KH ¿UVW SK\ORJHQHWLF DQDO\VLV ZDV FDUULHG RXW EDVHG RQ D FRPELQHG GDWD VHW RI /68 668 DQG ()Į VHTXHQFH GDWD RI LVRODWHV EHORQJLQJ WR Hysteriaceae (Table 1) in order to show the placement of the genus Rhytidhysteron in the family, with Delitschia winteri (CBS 225.62) as the outgroup taxon. The second phylogenetic DQDO\VLVEDVHGRQDFRPELQHGGDWDVHWRI/68668()ĮDQG,76VHTXHQFHGDWD were done only for the species in the genus Rhytidhysteron, using 19 isolates and Gloniopsis praelonga (CBS 112415) as the outgroup taxon. All newly generated sequences are deposited in GenBank. Isolates and GenBank accession numbers used in these analyses are listed in Table 1. SeqMan v. 7.0.0 (DNASTAR, Madison, WI) was used to compute consensus sequences. The sequence data were aligned and combined using Bioedit (Hall 1999) and MEGA 5.0 (Tamura et al., 2011). Alignments were checked and manual improved manually where necessary. The maximum likelihood analysis was SHUIRUPHGXVLQJ5$[0/Y%ODFN%R[VRIWZDUH 6WDPDWDNLVet al., 2008). The general time reversible model (GTR) including estimation of invariable sites was applied with a discrete gamma distribution with four rate classes. The best scoring trees were selected for the each analysis and illustrated with MEGA 5.0 and with Adobe Illustrator CS3. 5(68/76 Phylogenetic analyses 7KH ¿UVW DQDO\VLV ZDV EDVHG RQ D FRPELQHG GDWD VHW RI /68 668 DQG ()ĮVHTXHQFHGDWDEHORQJLQJWR+\VWHULDFHDHDQGWKHEHVWVFRULQJ5$[0/WUHHLV