MycoKeys 83: | 23-1 44 (202 | ) sae A peer-reviewed open-access journal doi: 10.3897/mycokeys.83.68582 RESEARCH ARTICLE . 03 MycoKkeys https://mycokeys.pensoft. net Launched to accelerate biodiversity research Three new species of Rhytidhysteron (Dothideomycetes, Ascomycota) from Mexico Aurora Cobos-Villagran', Ricardo Valenzuela', César Hernandez-Rodriguez’, Rosa Paulina Calvillo-Medina’, Lourdes Villa-Tanaca’, Luz Elena Mateo-Cid?, Abigail Pérez-Valdespino’, César Ramiro Martinez-Gonzalez®, Tania Raymundo! | Instituto Politécnico Nacional, Escuela Nacional de Ciencias Biolégicas, Laboratorio de Micologia, Prolongacién de Carpio y Plan de Ayala s/n, Mexico City 11340, Mexico 2 Instituto Politécnico Nacional, Escuela Nacional de Ciencias Biolégicas, Laboratorio Experimental de Bacterias y Levaduras, Prolongacién de Carpio y Plan de Ayala s/n, Mexico City 11340, Mexico 3 Instituto Politécnico Nacional, Escuela Nacional de Ciencias Biolégicas, Laboratorio de Ficologta, Prolongacién de Carpio y Plan de Ayala s/n, Mexico City 11340, Mexico 4 Instituto Politécnico Na- cional, Escuela Nacional de Ciencias Biolégicas, Laboratorio de Ingenieria Genética, Prolongacién de Carpio y Plan de Ayala s/n, Mexico City 11340, Mexico 5 Facultad de Quimica, Universidad Auténoma de Querétaro, Cerro de las Campanas s/n, Querétaro 76010, Mexico 6 Universidad Auténoma Chapingo, Departamento de Fitotecnia, Instituto de Horticultura, Km 38.5 Carretera Federal México-Texcoco, Texcoco, Estado de México 56230, Mexico Corresponding author: Tania Raymundo (traymundoo@ipn.mx) Academic editor: Cecile Gueidan | Received 12 May 2021 | Accepted 9 August 2021 | Published 14 September 2021 Citation: Cobos-Villagran A, Valenzuela R, Hernandez-Rodriguez C, Calvillo-Medina RP, Villa-Tanaca L, Mateo- Cid LE, Pérez-Valdespino A, Martinez-Gonzdlez CR, Raymundo T (2021) Three new species of Rhytidhysteron (Dothideomycetes, Ascomycota) from Mexico. MycoKeys 83: 123-144. https://doi.org/10.3897/mycokeys.83.68582 Abstract The genus Rhytidhysteron is characterised by forming navicular to apothecial hysterothecia, exposing the green, yellow, orange, red, vinaceous or black colours of the hymenium which generally releases pigments in the presence of KOH. The exciple is smooth or striated, the asci bitunicate and ascospores have 1—5 transverse septa. To date, twenty-six Rhytidhysteron species have been described from the Tropics. The pre- sent study aims to describe three new species in the Neotropics of Mexico based on molecular methods and morphological features. Illustrations and a taxonomic key are provided for all known species of this genus. Rhytidhysteron cozumelense from the Isla Cozumel Biosphere Reserve, R. esperanzae from the Sierra Juarez, Oaxaca and R. mesophilum from the Sierra Madre Oriental, Hidalgo are described as new species. With the present study, the number of species of R/hytidhysteron known from Mexico is now increased to eight. Keywords Hysteriaceae, Hysteriales, Neotropic, phylogeny, taxonomy Copyright Aurora Cobos-Villagrdn et al. 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. 124 Aurora Cobos-Villagran et al. / MycoKeys 83: 123-144 (2021) Introduction The genus Rhytidhysteron was described by Spegazzini (1881) and has been shown to belong to the Hysteriaceae (Boehm et al. 2009a, 2009b; Wijayawardene et al. 2020). The genus is characterised by forming hysterothecia, with lenticular or ir- regular, striated, or smooth openings; epithecium of various colours; excipulum composed of 1-2 layers of cells of angularis texture or globose texture. Rhytid- hysteron presents dense hamathecium, composed of branched pseudo-paraphyses, enclosed in a gelatinous matrix; octosporic, bitunicate, cylindrical asci; 1-3 septa ascospores, constricted in the central septum, reddish-brown to brown (Spegazzini 1881; Samuels and Miiller 1979; Kutorga and Hawksworth 1997; Boehm et al. 2009b; Thambugala et al. 2016). The distribution of the genus is Pantropical. It has been reported as an endophytic fungus (Rashmi et al. 2019) and causes mycosis in humans (Chowdhary et al. 2008; Mishra et al. 2014; Mahajan et al. 2014; Chander et al. 2016). The species with the largest distribution is Rhytidhysteron rufulum. It has been de- scribed from various places, with slight morphological differences depending on where it was found. R. rufulum have hysterothecia 1500-2000 um long, ascospores of (19— )26—36(—43) um and the colour of the red epithecium in Melzer’s Reagent changes to bright orange (Samuels and Miiller 1979). According to Kutorga and Hawksworth (1997), the length of the hysterothecia ranges from 2500-4000 um, ascospores from (22—)25-—35(—39) pm and has dark brown to reddish epithecium in potassium hydrox- ide (KOH) which changes to pale greenish-brown or from red wine to intense pink. On the other hand, in the description made by Almeida et al. (2014), the size of the ascomata ranges from 800-2500 um, ascospores from 21—32 um and has black or red epithecium without extractable KOH pigment. The specimens from Thailand have ascomata from 900-2350 pm, ascospores from 28-36 um and black or red epithe- cium are not reported to have a reaction with any reagent (Thambugala et al. 2016). Finally, Cobos-Villagran et al. (2020), for the Mexican specimens, report ascomata of 1000-3000 um, ascospores of 22.4—30.4 um and orange-reddish, yellow or black epithecia changing to magenta in reaction with KOH. These morphological varia- tions within R. rufulum have caused confusion in various fungal collections around the world and, as a result, they have been grouped into a complex of species (Boehm et al. 2009b; Murillo et al. 2009; Yacharoen et al. 2015; Doilom et al. 2016; Thambugala et al. 2016; Soto-Medina and Liicking 2017). Twenty-six species are known worldwide according to the Fungorum Index (2021) and, in the last two years, it has had greater relevance, since at least seven species have been described. In the present work, morphological and molecular analyses of distinct specimens of Rhytidhysteron obtained from different locations in Mexico were performed. Phylogenetic relationships were inferred based on internal transcribed spacer (ITS), nuclear large subunit ribosomal DNA (LSU) and elonga- tion factor l-alpha (tefl). Additionally, a dichotomous key is provided with all the species described so far. Three new species of Rhytidhysteron from Mexico 125 Materials and methods Study zone The specimens have been found from three different sites: one from Cozumel Island Biosphere Reserve, Quintana Roo, which is located between coordinates 20°35'20" and 20°17'16" north latitude (N) and —86°43'55" and —87°00'07" west longitude (W). The climate, according to the Képpen system, modified by Garcia (1981), is of the AmW (I) type, warm humid with abundant rain in summer. ‘The average annual temperature is 25.5 °C. Average annual rainfall is 1570 mm (INEGI 2013; Garcta- Martinez et al. 2021). The type of vegetation present in the town of San Gervasio is tropical dry forest, at 0 m above sea level. The second specimen from La Esperanza, Santiago Comaltepec, Chinantla was collected from the Sierra de Juarez in the State of Oaxaca, between coordinates 17°32' and 17°44’ north latitude (N) and —96°16' and —96°36' west longitude (W); altitude between 100 and 3200 m a.s.l. La Esperanza presents different types of climates, the main ones, according to the Képpen system, modified by Garcia (1981), are temperate humid with abundant rain in summer, C (m) and semi-warm humid with rain all year round. The temperature range is 10—26 °C. The range of precipitation is 800-4000 mm (INEGI 2008). ‘The type of vegetation present in the town of La Esperanza is tropical cloud forest, at 1600 m a.s.l. The last of the specimens is from Laguna de Atezca, Molango de Escamilla, which is located in the Sierra Madre Oriental in the State of Hidalgo, between the coordinates 20°42' and 20°59' of north latitude (N) and —98°41' and —98°53' of West longitude (W), altitude between 300 and 2200 m a.s.l. The Laguna de Atezca presents different types of climates, the main ones, according to the K6ppen system, modified by Garcia (1981), are semi-warm humid with rain throughout the year, ACf and temperate hu- mid with abundant rain in summer, C (m). The average annual temperature is 17 °C. Average annual rainfall is 1438 mm (INEGI 2009). The type of vegetation present in the town of Laguna de Atezca is tropical cloud forest, at 1281 ma.s.l. Morphological study The specimens were obtained by searching for dry or fallen branches in each of the lo- calities. The material was examined following traditional techniques in mycology (Ci- fuentes et al. 1986). Photographs were taken using a digital camera (Nikon, D7000, Tokyo, Japan) with an 85 mm macro lens (Nikon, Tokyo, Japan). The fresh collected specimens were used to obtain morphological data such as the colour of the epithe- cium, growth habit and habitat. Ascomata were measured by a stereomicroscope (Zeiss 475002, Jena, Germany). Cross sections were made in the middle part of the ascomata and mounted on temporary slides in 70% alcohol and 10% KOH. Sections were ob- served under an optical microscope (Zeiss K-7, Jena, Germany) for the measurement of the characters of taxonomic importance. 126 Aurora Cobos-Villagran et al. / MycoKeys 83: 123-144 (2021) DNA extraction, amplification and sequencing The DNA of each specimen of Rhytidhysteron spp. was obtained using the cetyl- trimethylammonium bromide (CTAB) method, according to Doyle and Doyle (1987). Three molecular markers were used, the ribosomal large subunit (LSU), the internal transcribed spacer rDNA-ITS1 5.88 rDNA-ITS2 (ITS) and trans- lation elongation translation factor l-« (tef7). The primers used for LSU were LOROf and LR5r (Vilgalys and Hester 1990), for ITS, these were ITS1f and ITS4r (White et al. 1990; Schoch et al. 2012) and tef? EF1-B-F1 and EF1-B-R (Wu et al. 2014). DNA amplifications were performed in a GeneAmp PCR System 9700 thermal cycler (Thermo Fisher Scientific), following recommendations by White et al. (1990) for ITS, Vilgalys and Hester (1990) for LSU and Wu et al. (2014) for tefl. The PCR products were verified by agarose gel electrophoresis. The gels were run for 1 h at 95 V cm*in 1.5% agarose and 1x TAE buffer (Tris Acetate-EDTA). The products were then dyed with GelRed (Biotium, USA) and viewed in a tran- silluminator (Infinity 300 Vilber, Loumat, Germany). Finally, the products were purified using the ExoSap Kit (Affymetrix, USA) according to the manufacturer’s instructions and were prepared for the sequencing reaction using the BigDye Ter- minator Cycle Sequencing Kit v. 3. 1 (Applied BioSystems). Sequencing was car- ried out in a genetic analyser (Sanger sequencing) by Macrogen Inc. (Seoul, Korea). The sequences of both strains of each sample were analysed, edited and assembled using BioEdit v. 1.0.5 (Hall 1999) to create consensus sequences. The consensus sequences were compared with those in the GenBank database of the National Center for Biotechnology Information (NCBI) using the BLASTN 2.2.19 tool (Zhang et al. 2000). Phylogenetic analyses In order to study phylogenetic relationships, our newly produced sequences of six individuals of Rhytidhysteron were added to reference sequences of ITS, LSU and tefl (Table 1) deposited in the NCBI database (http://www.ncbi.nlm.nih.gov/ genbank/). Each gene region was independently aligned using the online version of MAFFT v7 (Katoh et al. 2002, 2017; Katoh and Standley 2013). Alignments were reviewed in PhyDE (Miller et al. 2005), followed by minor manual adjustments to ensure character homology between taxa. The matrices were formed for ITS by 28 taxa (667 characters), for LSU by 31 taxa (875 characters); while the tef7 con- sisted of 24 taxa (896 characters). Gloniopsis calami was used as the outgroup. The aligned matrices were concatenated into a single matrix (31 taxa, 2438 characters). Five partitioning schemes were established: one for the ITS, one for the LSU, and three to represent the three codon positions of the tef7 gene region, which were es- tablished using the option to minimize the stop codons with Mesquite v3.2 (Mad- dison and Maddison 2017). The best evolutionary model for alignment was sought using PartitionFinder (Lanfear et al. 2014, 2017; Frandsen et al. 2015). Phylogeny Three new species of Rhytidhysteron from Mexico 127 Table |. Species names, strain numbers, isolation source, locality and GenBank accession numbers for the taxa used in this phylogenetic analysis. Sequences generated for this study are in bold. Species Rhytidhysteron bruguierae R. bruguierae R. bruguierae R. bruguierae R. bruguierae* R. camporesii R. chromolanae R. cozumelense R. cozumelense erioi esperanzae esperanzae hysterinum aR RR BD hongheense R. hongheense R. magnoliae* os) mangrovei * mesophilum mesophilum mexicanum* mexicanum neorufulum * neorufulum neorufulum opuntiae rufulum tectonae thailandicum PRR R RRR RDVDDAAR PD thailandicum Gloniopsis calami* *Ex-type strains. thailandicum* Isolate No. MFLUCC 17-1502 MFLUCC 17-1509 MFLUCC 17-1511 MFLUCC 17-1515 MFLU 18-0571 KUN-HKAS 104277 MFLUCC 17-1516 A. Cobos-Villagran 951 T. Raymundo 7321 MEFLU 16-0584 T. Raymundo 6579 R. Valenzuela 17206 EB 0351 KUMCC 20-0222 HKAS112348 MFLUCC 18-0719 MFLU 18-1894 A. Trejo 74 A. Cobos-Villagran 1800 RV17107.1 RV17107.2 MFLUCC 13-0216 MFLUCC 13-0221 MFLUCC 17-2236 GKM 1190 MFLUCC 14-0577 MFLUCC 13-0710 MFLUCC 14-0503 MEFLU 19-2373 MFLUCC 13-0051 MFLUCC 15-0739 LSU MN632453.1 MN632455.1 MN632454.1 MN632452.1 NG_068292.1 MN429072.1 MN632456.1 MW9394459 MW9394460 MN429071.1 MW9394457 MW9394458 GU397350.1 MW264193.1 MW541820.1 MN989384.1 NG_067868.1 MW9394461 MW939462 MT626026 MT626027 NG_059649.1 KU377567.1 MH063266.1 GQ221892.1 KU377565.1 KU764698.1 NG_059648.1 MN989429.1 MN509434.1 NG_059715.1 ITS MN632458.1 MN632460.1 MN632459.1 MN632457.1 MN429069.1 MN632461.1 MZ056797 MZ056798 MN429068.1 MZA77203 MZA77204 MW264214.1 MW541824.1 NR_170019.1 NR_165548.1 MZ056799 MZ056800 MT626028 MT626029 NR_164242.1 KU377562.1 MH062956.1 KU377560.1 KU144936.1 NR_164241.1 MN989428.1 MN509433.1 KX669036.1 tefl MN635662.1 MN635661.1 MNO077056.1 MN442087.1 MN635663.1 MZ457338 MZ457339 MN442086.1 MZ457336 MZ457337 GU397340.1 MW256815.1 MW556132.1 MN997309.1 MK450030.1 MZA457340 MZ457341 KU510400.1 GU397341.1 KU510399.1 KU497490.1 MN989431.1 MN509435.1 KX671965.1 Source and Locality Dead stems of Chromolaena odorata, Thailand Dead stems of Chromolaena odorata, Thailand Dead stems of Chromolaena odorata, Thailand Dead stems of Chromolaena odorata, Thailand Submerged branches of Bruguiera sp. Thailand Dead stems, China Dead stems of Chromolaena odorata, Thailand Dead twigs of Tabebuia rosea, Mexico Dead twigs of Tabebuia rosea, Mexico Dead stems, Thailand Dead stems Mexico Dead stems Mexico Dead branches, France Dead twigs of Dodonaea, China Dead twigs of Dodonaea, China Dead twigs of Magnolia grandiflora, China Dead twigs of mangrove, Thailand Dead stems, México Dead stems, México Dead wood, Mexico Dead wood, Mexico Dead wood, Thailand Dead wood, Thailand Dead wood, Thailand Kenya Woody litter, Thailand Dead branches, India Dead wood, Thailand Dead wood, Thailand Dead wood, Thailand Unknown was performed with Bayesian inference using MrBayes v3.2.6 x64 (Huelsenbeck and Ronquist 2001). The information block for the matrix includes two inde- pendent runs of the MC3 chains using 10 million generations (standard deviation <0.1). The convergence of the chains was displayed in Tracer v1 (Rambaut et al. 2014). The highest credibility phylogram of the clades recovered with TreeAnnota- tor v. 1.8 (Bouckaert et al. 2014) was chosen with a 25% burn-in. 128 Aurora Cobos-Villagran et al. / MycoKeys 83: 123-144 (2021) Results Phylogenetic analysis The ITS, LSU and ¢ef/ sequences obtained from Rhytidhysteron cozumelense, Rhytidhysteron esperanzae and Rhytidhysteron mesophilum were deposited in GenBank (Table 1). In the Bayesian analysis, the standard deviation between the chains stabilized at 0.001 after 10 million generations, indicating that MC3 reached a stationary phase. To confirm that the sample size was sufficient, the parameter file was examined in Tracer 1.6 (Rambaut et al. 2014): all parameters had an estimated sample size of over 1,500. The posterior probabili- ties (PP) obtained were estimated by generating a strict consensus tree in MrBayes. Bayes- ian inference analysis recovered well-supported clades (PP = 1) of the three species Rhytid- hysteron cozumelense, Rhytidhysteron esperanzae and Rhytidhysteron mesophilum (Figure 1). Taxonomy Rhytidhysteron cozumelense Cobos-Villagran, R. Valenz., Hdz-Rdz., Calvillo- Medina & Raymundo sp. nov MycoBank No: 839084 Fig. 2 Diagnosis. Differs from Rhytidhysteron rufulum in its host (Bignoniaceae), size of asco- mata (2.5—3.5 x 1.1-1.5 x 1.0-1.9 mm), asci (182-191 x 12-13 ym) and its reaction with KOH being faster (one to five seconds). Type. Holotype: Mexico. Quintana Roo, Cozumel Municipality, San Gervasio Chen-tuk archaeological zone, 20°29'50"N, —86°50'39"W, 0 m as.l., 21 January 2018, A. Cobos-Villagran 951 (ENCB), on Tabebuia rosea DC. (Bignoniaceae), Gen- Bank: LSU MW9394459, ITS MZ056797, tefl MZ457338. Description. Ascomata hysterothecial to apothecial 2.5—3.5 mm long, 1.1—-1.5 mm wide, (0.8)1.0-1.9 mm high, erumpent, solitary, boat-shaped hysterothecia, subglo- bose, elongated, compressed in the apex, with conspicuous longitudinal groove or cleft and becoming lenticular when mature or exposed to moisture, black, carbonaceous when dry. Margin involute, smooth to perpendicularly slightly striated, black. Exciple integrated in two layers, the first carbonaceous, glabrous, 45—100 pm thick, wide at the base, composed of pseudoparenchymal cells of textura prismatica (iso-radiating cells), thick-walled, the second composed of cells hyaline, thin-walled. Pseudoparaphyses up to 2.5 um wide, filamentous, capitate, hyaline, septate, enclosed in a gelatinous matrix, strongly anastomosed above the asci. Epithecium reddish brown (8F7) when fresh, black in old specimens or when dry, becoming greyish magenta (13B5) in the presence of 10% KOH. Asci 182-191 x 12-13 um, bitunicate, cylindrical, hyaline, uniseriate, octosporic, thick-walled, with a sinuous base. Ascospores 26—29(-31) x 9-11 (-13) um, (x= 28 x 10.2 um, n = 30), ellipsoidal to fusiform, rounded at both ends, dark brown in colour with three transverse septa, with a thick and smooth wall. Three new species of Rhytidhysteron from Mexico 129 1 R. neorufulum 221 R. neorufulum 216 0.41 R. neorufulum 236 0.75 Rhytidhysteron magnoliae 18-0719 1 Rhytidhysteron rufulum 14-0577 Rhytidhysteron tectonae 13-0710 i Rhytidhysteron mesophilum 1800 Rhytidhysteron mesophilum 74 1 Rhytidhysteron mexicanum RV17107.1 1 Rhytidhysteron mexicanum RV17107.2 1 Rhytidhysteron esperanzae 17206 Rhytidhysteron esperanzae 6579 1 Rhytidhysteron camporesii 104277 1 Rhytidhysteron chromolanae 17-1516 0.86 Rhytidhysteron hongheense 112348 Rhytidhysteron hongheense 20-0222 1 Rhytidhysteron cozumelense 7321 Rhytidhysteron cozumelense 951 1 Rhytidhysteron mangrovei 18-1894 1 Rhytidhysteron opuntiae 1190 Rhytidhysteron hysterinum 0351 0.91 Rhytidhysteron bruguierae 17-1511 0.93 Rhytidhysteron bruguierae 17-1502 Rhytidhysteron bruguierae 17-1509 Rhytidhysteron bruguierae 18-0571 Rhytidhysteron bruguierae 17-1515 | Rhytidhysteron erioi 16-0584 Rhytidhysteron thailandicum 19-2373 1 Rhytidhysteron thailandicum 14-0503 Rhytidhysteron thailandicum 13-0051 0.91 0.99 Gloniopsis calami 15-0739 0.002 Figure |. Phylogenetic relationships within the genus R/ytidhysteron based on a Bayesian analysis of a combined dataset of ITS, LSU and tef7 sequence data. Gloniopsis calami 150739 was used as the out- group. The posterior probabilities for each clade are shown above the branches. The new species Rhytidhys- teron cozumelense, Rhytidhysteron esperanzae and Rhytidhysteron mesophilum are shown in bold. Distribution. Known from a single local Island in the Cozumel Biosphere Re- serve, Mexico. Ecology. Dead twigs of Tabebuia rosea DC. (Bignoniaceae). Etymology. The epithet refers to the Island in the Cozumel Biosphere Reserve where the species was found. Specimens examined. Mexico, Quintana Roo, Cozumel Municipality, San Gervasio Chen-tuk archaeological zone, 20°29'54"N, —86°50'43"W, 13 m as.l., 21 January 2018, T. Raymundo 7321, R. Valenzuela 17985 (ENCB); 17 June 2018, A. Cobos-Villagran 1838 (ENCB). Notes. Rhytidhysteron cozumelense is characterised by black ascomata with a red- dish brown epithecium and a smooth to slightly striated margin that, in reaction with 10% KOH, changes to greyish magenta. R. mesophilum has a similar reaction in KOH, but with several tones of green in the hysterothecia, a reddish orange to orange red 130 Aurora Cobos-Villagran et al. / MycoKeys 83: 123-144 (2021) Figure 2. Rhytidhysteron cozumelense (Holotype, A. Cobos-Villagran 951) A appearance of ascomata hystero- 10 pm “4 thecial and apothecial on host B ascomata apothecial close-up, striated margin and black epithecium C=F mi- croscopical features stained with alcohol (70%) and KOH (10%) reagent C ascomata apothecial cross-section with alcohol (70%) D exciple of iso-radiating cells (textura prismatica), close-up E asci F ascospores. Three new species of Rhytidhysteron from Mexico 131 epithecium and a perpendicularly striate with irregular slits and yellowish green pruina in margin. R. rufulum has a magenta reaction in KOH and strongly striated margin. Tabebuia rosea is reported as a new host for a Rhytidhysteron species. Rhytidhysteron esperanzae Cobos-Villagran, R.Valenz. & Raymundo sp. nov Mycobank No: 839086 Fig. 3 Diagnosis. Different from most Rhytidhysteron species by having greyish-green asco- mata with greenish-grey to yellow epithecium in the presence of KOH, and large and wide ascospores (45-47 x 17-19 um). Type. Holotype: Mexico. Oaxaca, Sierra de Juarez, Chinantla, Santiago Co- maltepec Municipality, La Esperanza, Carretera Oaxaca-Tuxtepec Km 51, 17°37'55"N, —96°22'01"W, 1600 m a.s.l., 23 May 2017, T. Raymundo 6579 (ENCB). GenBank: LSU MW9394457, ITS MZ056795. Etymology. The epithet refers to the locality “La Esperanza” where the species was found. Description. Ascomata hysterothecial to apothecial, (2—)3-4.5 mm long, (1.2- )2-3 mm wide, (1—)1.7—2.4 mm high, superficial, solitary, rarely gregarious, boat- shaped hysterothecia, elongated, straight or flexuous, with sharp ends, opening in a discoid shape when ripe or with humidity, exposing the hymenium, taking the apothecial shape of 3-4 mm in diameter, brown (6D7), dull-green (30E4) to black. Margin involute, perpendicularly striate, greyish green (30C4) to dull green (30D4). Exciple integrated in two layers, the first carbonaceous, glabrous, 60-220 um wide, thinning in the apical part, the middle part and the base are thicker, composed of pseudoparenchymal cells of textura globulosa-angularis (isodiametric cells), 11 x 10 um, thick-walled, 3 um wide, the second slightly pigmented to hyaline, thin- walled. Pseudoparaphyses up to 4 um wide, filamentous, capitate, apical part wider, straight, hyaline, with a septum, enclosed in a gelatinous matrix, strongly anastomo- sed above the asci. Epithecium dark green (30F4) to black, becoming yellow (2A7) in the presence of 10% KOH. Asci (250—)265-270 x (18—)19-—20 um, bitunicate, cylindrical, rounded apex, hyaline, uniseriate, octosporic, thick-walled, with a short pedicel. Ascospores of (42—)45—47(-49) x (15—)17—19(—23) um, (x= 45 x 17.2 um, n = 30), ellipsoidal to spindle-shaped, rounded or pointed at both ends, reddish-brown to brown when mature, with three transverse septa, constricted at the septa, thick- walled and smooth. Distribution. Known from a single locality in a forest in La Esperanza, Mexico. Ecology. Dead stems and twigs in tropical cloud forest dominated by Oreomunnea mexicana Standl. J.-F. Leroy (Juglandaceae). Specimens examined. Mexico. Oaxaca. Sierra de Juarez, Santiago Comaltep- ec Municipality, La Esperanza, Carretera Oaxaca-Tuxtepec Km 51, 17°37'55"N, —96°22'01"W, 1600 m a.s.l., 22 May, 2017, R. Valenzuela 17206 (ENCB); 23 May 132 Aurora Cobos-Villagran et al. / MycoKeys 83: 123-144 (2021) - 0pm 50 um Figure 3. Rhytidhysteron esperanzae (Holotype, T. Raymundo 6579) A appearance of ascomata apothe- cial on host B ascomata apothecial close-up, greyish-green to dull green and striated margin and dark green to black epithecitum C-F microscopical features stained with alcohol (70%) and KOH (10%) reagent C ascomata apothecial cross-section with alcohol (70%) D exciple of isodiametric cells (textura globulosa-angularis), close-up E asci F ascospores. Three new species of Rhytidhysteron from Mexico 133 2017, A. Cobos-Villagran 498 (ENCB); 25 May 2017, E. Campero 3 (ENCB), 30 April 2018, A. Cobos-Villagran 1119 (ENCB), A. Gay AG30041814 (ENCB). Notes. khytidhysteron esperanzae, is characterised by a brown, dull-green to black exciple and dark green to black epithecium that, in reaction with 10% KOH, changes to yellow colouration. This colouration with KOH is very different than those of of R. rufulum and R. neorufulum which are magenta and violet, respectively. R. esperanzae have larger ascospores than R. rufulum (22.4—30.4 x 8-9.6 um) and R. mexicanum (34-40 x 10-12 pm). Ecologically, this new species grows in a tropical cloud forest dominated by Oreomunnea mexicana Standl. J.-F. Leroy (Juglandaceae). Rhytidhysteron mesophilum Cobos-Villagran, R. Valenz., Hdz.-Rdz., Calvillo- Medina & Raymundo sp. nov. Mycobank No: 839097 Fig. 4 Diagnosis. Differs from Rhytidhysteron rufulum by its green-yellowish pruina on the mar- gins, size of asci (267—282 x 15.5—16 um) and larger ascospores (40-44 x 12-14 um). Type. Molango de Escamilla Municipality, Laguna Atezca, 20°48'32"N, —98°44'52"W, 1281 m asl, 01 June 2018, A. Trejo 74 (ENCB). GenBank: LSU MW9394461, ITS MZ056799. Etymology. ‘The epithet refers to the type of vegetation (mountain mesophilic for- est) it was collected from. Description. Ascomata hysterothecial to apothecial, 2.5-4 mm _ long, 1.0-1.5 mm wide, 1.4—-1.7 mm high, superficial or erumpent, gregarious, rarely solitary, with small hysterothecial ascomata, ellipsoid to oblong and black when young, then boat-shaped hysterothecia, with some constriction in the middle part, flexuous, open in apothecioid ascomata, dark green (30F3-—4), dull green (30E3—4), greyish green (30E6-—7), deep green (30D-E8) to yellowish green (30B-C8) when mature, forming small ascomata within disc in old specimens. Margin involute, perpendicularly striate, marks are not roughness, rather irregular slits, with yellow- ish green (30B-C8) pruina. Exciple integrated in two layers, the first carbonaceous, glabrous, green yellowish, 62.5—75 um thick, in the middle part widening more (112.5-125 um), composed of pseudoparenchymal cells of textura prismatica (iso- radiating cells), the second composed of cells hyaline, thin-walled. Pseudoparaphy- ses 2.0—2.5 um up to 3.0 um wide, filamentous, capitate, hyaline, without septa, branched towards the apex, enclosed in a gelatinous matrix, strongly anastomosed above the asci. Epithecium reddish orange (7B8) to orange red (8A8), becoming greyish magenta (13D6) in the presence of 10% KOH. Asci 267-282 x 15.5- 16 um, bitunicate, cylindrical, hyaline, uniseriate, octosporic, thick-walled, with a sinuous base. Ascospores (38—)40—44(—46) x 12-14 um, (x= 44.2 x 13.6, n = 30), ellipsoidal to oblong, light brown in colour, with three transverse septa, constricted at the septa, with a thick and smooth wall. 134 Aurora Cobos-Villagran et al. / MycoKeys 83: 123-144 (2021) Figure 4. Rhytidhysteron mesophilum (Holotype, A. Trejo 74) A appearance of ascomata hysterothecial on host B ascomata hysterothecial close-up, striated margin with yellowish green pruina and reddish orange to orange red epithecium C=F microscopical features stained with alcohol (70%) and KOH (10%) rea- gent C ascomata hysterothecial cross-section with alcohol (70%) D exciple of iso-radiating cells (textura prismatica), close up E asci F ascospores. Three new species of Rhytidhysteron from Mexico 135 Distribution. Known from a single locality in Laguna de Atezca, Molango de Escamilla, Hidalgo, Mexico. Ecology. Dead stems in tropical cloud forest. Specimens examined. Mexico. Hidalgo, Molango de Escamilla Municipality, La- guna Atezca, 20°48'32"N, —98°44'52"W, 1281 ma.s.l., 01 June 2018; C. Herrera 40 (ENCB), A. Cobos-Villagran 1800 (ENCB). Notes. Rhytidhysteron mesophilum is characterised by a dark green, dull green, greyish green, deep green to yellowish green hysterothecium, forming small ascomata within disc in old specimens. This fungus could be confused with R. esperanzae because both are found in tropical cloud forest (mesophilic forests) and have similar ascospores. However, R. mesophi- lum is distinguished by a reddish orange to orange red epithecium, while in R. esperanzae, the epithecium is dark green to black. R. mesophilum also resembles R. columbiense by the presence of a yellowish green (30B-C8) pruina in the margin, but the ascospores are larger (38-52 x 13-18 um) and the epithecium is brown to dark brown in the second species. A dichotomous key is presented with the species of R/ytidhysteron accepted by Index Fungorum (2021), including the three new species proposed in this work. ‘The key includes the recently described R. mexicanum Cobos-Villagran, Raymundo, Calvillo-Medina & R. Valenz and R. hongheense Wanas. It should be noted that R. fuscum (Ellis & Everh.) J.L. Bezerra & Kimbr. and R. minor (Cooke) A. Pande are not considered because the first belongs to the genus Jryblidiella and the second is a nom. inval., because the basionym was not indicated and bibliographic reference omitted (Art. 41.5, see Art. 41.7, Melbourne). Key to the known species of Rhytidhysteron 1 ASCOS POtes Su TIT IbOr Ml tere oe ee tees eee ome EER a Sa 2 Ascosporestransversely- septate 1—5 septa’. .ivsec.o.0s n.d ee 3 2 Ascospores with 3—5 transverse and 1-3 longitudinal septa, 20-25 x 7.5— 10 um, epithecium brown-red, on Cylindropuntia fulgida; type: USA........... Rta asics pecs ase Bath sas Cee de ton RUN a ae R. opuntiae (J.G. Br.) M.E. Barr - Ascospores with 3 transverse septa mainly and rarely with 3 transverse septa and 1 longitudinal septum, 20-33 x 9-13 pm, epithecium reddish orange, on Dodonaea viscosa, type: China ......ceeeeeseeeeseeeeeeee R. hongheense Wanas. 3 ASCOS POLES — SEPALS at stra 2 a8. 7 Mess t 28s ass Oe testa, Fons teehee MeN don ls Soles 4 — PASCOSPOLESEO—9) SCDUEAUG sseinstasg- reeset tae Ronee sericea. nach etal tntar etn tas inee eedalacailn 5 4 Epithecium ferruginous brown, ascospores 22-32 x 10-16 um, on Buxus sempervirens, Diospyros spp. or Ilex spp.; type: France oo. esceseeseeseeeeseceeeeees < Rn ria Ree helene merle, re 27 R. hysterinum (Dufour) Samuels & E. Mill. = Epithecium orange, ascospores 24.8—29(—31) x 8.8—10(—11.2) um, on Acacia RO] eercinigu repens < [cl Ohir aanarm nantes in ile Rasa Rlatlni hn ieys. Arie Ran Et iret in ceeds R. neohysterinum Cobos-Villagran, Hdz.-Rdz., R. Valenz. & Raymundo 5 Five septa in mature ascospores, 30-46 x 12—20 um, epithecium yellowish orange, on Pinus spp.; type: Finland ........... R. dissimile (P. Karst.) Magnes = Mhceetseptacit. matuitcrascospOkesen 4.0102 ule tant Pesan Some Ae ies Lu! f 136 co | Aurora Cobos-Villagran et al. / MycoKeys 83: 123-144 (2021) Ascospores 12—15 x 5—6 um, exciple brownish green, epithecium brown, on monocotyledonous; type: Sri Lanka...... R. beccarianum (Ces.) Bat. & Valle AxSGOs poresiloneer thant LOMA tesa ceteucussiuniceutersine coneeenesrucgdivndecedea teetdaticette 7 HASCOSPOLeS:DeLWceIl GMOIGO- Mind OTM ds.chchsbiutouites ohesedonseiis ohabecurastunenns 8 PSSCOSPOLCS GIVE CHAC HAP OO TIE, gust aay cusgundtrtdracieusnuettobenewekers costar terenectona 22 Ascomata with exciple and/or margin in several tones of green ........ eee 9 Ascomata with exciple and margin reddish brown to black... 10 Ascomata with exciple and margin vivid green, perpendicularly striate, as- cospores 20-30 x 7—9 um, constricted at the central septum, on angiosperm; Cyqpes Braz. fs Mesenteric in Bes te neta Stee Sess ds og tae vl aes R. viride Speg. Ascomata dark brown to black with yellowish green on the margin, smooth, not striate, ascospores 23-28 x 8-11 um, slightly constricted at the central septum, on Chromolaena odorata; type: Thailand ........eeeeeeseeseceneeeeeeeeeeeees SSeS Ah eid Bel APs J oeh AB acesh R. chromolaenae Mapook & K.D. Hyde Epithecium with yellow, orange, red or green colour in some development Epithecium brown to black in young and mature specimens.............0.0 18 Epithecium yellowish green, margin perpendicularly striate, ascospores 20.3— 30.4 x 7.6-10.1 um, on Prosopis jungiflora; type: USA.....R. prosopidis Peck Epithecium with yellow, orange or red COlOUL oie eeeeeeeseeseeseeeseeeeeeees 12 Ascomata hysterotecial, epithecium yellow, margin smooth, ascospores (19- )28—29(—31) x (8-)10-12(-13) um constricted at the central septum, on Tectona grandis; type: Thailand............... R. tectonae Doilom & K.D. Hyde HNSC@ mata a pO LMC eialeN.2.78. Peet eee a ety Bsus teen 13 Epithecium with red tones in young or mature speciMeNs ........... cece 14 Epithecium with orange tones in young or mature specimens..............00+ 16 Epithecium vivid red or cinnabar red, ascospores 19.0—24.7 x 7.6-11.4 um, constricted atthe septa; ionwQuercustspe: type: dindia® Me ly eevee eshte Hoenete sesie setae: R. quercinum (B.G. Desai & V.N. Pathak) M.P. Sharma & Rawla Eprtheciummr dar ktred £0: D babes is troche et Arata Hae ben cubs ux eetewnet ss ihak tae 15 Growing on mangrove tree, epithecium dark red to dark brown, ascospores 21— 28 x 7.5—8.5 um; type: Thailand ......R. mangrovei Vin. Kumar & K.D. Hyde Growing mainly on Fabaceae, not on mangroves, epithecium orange red, red, dark red to black, 22.4-30.4 x 8-9.6 um, type: Puerto Rico... eee eeeeeees prescient deel tended eae ca Dee tain die Oe eect R. rufulum (Spreng.) Speg. Ascospores 28—30 x 10—12 um, on angiosperm, type: Paraguay... Bh ha MAA ea Mia WEEN, oh OS ou Buta Medel ae EL R. discolor (Speg.) Speg. scospotes Siria lle rst: 2.8: pitt akar.soees eae ceseh-See eRe Be ee Saas 17 Ascospores 6.2—9 um broad, on Bruguiera sp. and Chromolaena odorata; Ey pes nailer ea Pc N hve Ae ae coda ea et R. bruguierae Dayarathne Ascospores 9-11 um, on angiosperm; type: Thailand... eeeeeeeeeeeeeee seats aS SeraRE ae ovat Soa tata levees ec heaieoteeecndethe R. erioi Ekanayaka & K.D. Hyde I Vesea te Wioraig ofc) sc HOE LEMAR eA We CCR ter ROD ra ys BEN Eee eet ey 19 Marois @otlintO=s WO MU STR AGE acc. mlucinictaer Se ed aa nds rcmnutons we tsonp sents 20 19 22 ZS 26 27, 28 Three new species of Rhytidhysteron from Mexico 137 Ascospores 25—27 um broad, on angiosperm; type: Australia... eee Joss ter ciec eatin Nid 75h Pitt a Rot 9 ot oo rope a ae re ar R. scortechinii Sacc. & Berl. Ascospores 28—30(—32) um broad, on Magnolia grandiflora; type: China...... Ul AIRMEN Ae NaS We R. magnoliae N.I. de Silva, Lumyong S & K.D. Hyde Ascomata apothecial, ascospores 26—29(—31) x 9-11 (—13) um, on Tabebuia rosea DC.; type: Mexico......... R. cozumelense Cobos-Villagran, R.Valenz., Hdz-Rdz., Calvillo-Medina & Raymundo ENSCOMMAtA: MY STCROLE Clan tne tox denen tui catch oats oe deo tnetectieaed texttedtsteanetaions 21 Ascospores 25-28 x 9-11 um, hamathecium release magenta pigment in KOE ontanciose pers ty per G Hina: cci3 sce bs sca snoncatncentlsatantag ecee ieee ati 1 ARR SUR cE AOL RSS R. camporesii Ekanayaka & K.D. Hyde Ascospores 20—28(-31) x 7.5-12 pm, hamathecium do not release pigment in’ KOE son-angioseper; type: Vnailand: 5:2. coteoe tea eet ec eetdeseasarseent dees Leh assist adel seid tats nate eam bats R. thailandicum Thambugala & K.D. Hyde PXSCOSPO LES 3 04 Or HIN ease dices eteeesons retobt aenoonee tacks T¥ireinced tee teaircomecemeeoeaties 23 Ascosporesiloneer than:-40: Wins icsrcotesirenreavigendethive sony erensskincatee macs iteatenren 26 Margin perpendicularly striate, epithecium yellowish green to pistachio green when fresh, light green to pale green when dry, 34-40 x 10-12 um, on angio- Spetimy aby pes VieKheOn 2 tuer. Att saves Namie vedas wecnlrcoeneatuneadted aashs cues terete case. b R. mexicanum Cobos-Villagran, Raymundo, Calvillo-Medina & R. Valenz. Margin smooth, epithecium yellow, reddish orange or black... 24 Epithecium yellow, orange to reddish orange, ascospores 27—34 x (6.5—)7— 10.6:(=12.5) um; ottangiospermnnty pe: gl Nata nd sia). eutii estas oioeoaenesbouphden’ Us swt Seb sccdes skeen sen em coset R. neorufulum Thambugala & K.D. Hyde Epithecivpit black: See see es oie aha or Se Ee. 25 Ascospores 10—12 um broad, constricted at the central septum, on angio- SPERMS. ty Pew PatAOUAy sc: fe. cetieasalbes Seckasel ee secaseraaetielr R. guaraniticum Speg. Ascospores 13—14 um broad, constricted at the septa, on Scutia indica; type: Vivace: Aa sae AoC gst fed R. indicum (Anahosur) M.P. Sharma & K.S. Thind Exciple black, epithecium black, ascospores 40-45 x 15—20 um, on angio- BOE Ebi ty IOS GAT Mek chestnaceds bate ntean shade secon tte weds maians tees R. brasiliense Speg. Exeiple-or margin wath ereen tones: j.4 Sie ca tas cueires dost vecteandon tens dorceth ae Sateen. 27 Exciple and margin dark green, dull green, greyish green, deep green to yel- lowish green when mature, epithecium reddish orange to orange red, as- cospores (38—)40—44(—46) x 12-14 um, on angiosperm; type: Mexico......... R. mesophilum Cobos-Villagran, R. Valenz., Hdz.-Rdz., Calvillo-Medina & Raymundo Exciple brawn, lat kabrowar to! black ..ccn.2:82.4. ue hbase Be ac eee Se 28 Margin with a yellowish-green pruina, epithecium brown to dark brown, as- cospores 38-52 x 13—18 um, on angisoperm; type: Colombia... Posies Patents sal aan dthactne geist ee oct eoeutue R. columbiense Soto-Medina & Liicking Margin greyish green to dull green, epithecium dark green (30F4) to black, ascospores (42—)45—47(—49) x (15—)17—19(—23) um, on angiosperm; type: Mexico...........066- R. esperanzae Cobos-Villagran, R.Valenz. & Raymundo 138 Aurora Cobos-Villagran et al. / MycoKeys 83: 123-144 (2021) Discussion The genus khytidhysteron is a highly diverse group with a mainly Pantropical distribu- tion (Samuels and Miiller 1979). The morphological characteristics that have, so far, helped in the segregation of the species are: shape and border of the hysterothecium, ornamentation of the exciple, colour and reaction of the epithecium, and size of the ascospores which only, in some cases, have helped delimiting species, as in the case of Rhytidhysteron columbiense Soto-Medina & Liicking and R. neohysterinum Cobos- Villagran, Hern-Rodr., R. Valenz. & Raymundo. Therefore, species in which the size of spores overlap, have been clarified by mo- lecular methods and the use of molecular markers, such as ITS, LSU, elongation factor 1 alpha (TEF1), amongst others. For example, in the case of R. rufulum, catalogued as a species complex based on morphology, the fungal barcodes have been helpful in describing different species that are morphologically similar (Boehm et al. 2009b; Murillo et al. 2009; Yacharoen et al. 2015; Doilom et al. 2016; Thambugala et al. 2016; Soto-Medina and Liicking 2017). In recent years, part of the taxonomy has been resolved using collections from different countries around the globe. For example, in Thailand, R. neorufulum and R. thailandicum were described in the work of Thambu- gala et al. 2016. In the same year, Doilom et al. (2016) described R. tectone on Tectona grandis L. (Verbenaceae) also from Thailand. In recent years, eight new species were described: Kumar et al. (2019) described R. mangrovei Vinit & K.D. Hyde, isolated from dead mangrove branches; Dayarathne et al. (2020) described R. bruguierae Dayarathne, also isolated from mangrove branches Bruguiera Lam. (Rhizophoraceae); Hyde et al. (2020) described R. camporesii Eka- nayaka & K.D. Hyde and R. erioi Ekanayaka & K.D. Hyde; Mapook et al. (2020) described R. chromolaenae Mapook & K.D. Hyde, isolated from branches of Chromo- laena odorata (L.) King & Robinson (Asteraceae); Wanasinghe et al. (2021) described R. hongheense Wanas. isolated from dead twigs of Dodonaea Mill. (Sapindaceae); and in Mexico, Cobos-Villagran et al. (2020) described R. neohysterinum Cobos-Villagran, Hdez.-Rdz., R. Valenz. & Raymundo and Cobos-Villagran et al. (2021) R. mexica- num Cobos-Villagran, Raymundo, Calvillo-Medina & R. Valenz. With this new study, three more species have been described from Mexico. In the present study, we observed that R. cozumelense is phylogenetically close to R. hongheense, R. camporesii and R. chromolaenae. The four species are similar in terms of ascospore size in the range of 23-30 x 8-13 um and have a margin smooth to slight- ly striate. R. hongheense has slightly longer ascospores (20-33 x 9-13 um). However, they have ascomata of contrasting sizes. R. chromolaenae forms smaller navicular hys- terothecia, 750-885 um diam., with orange epithecium, turning purple in KOH and is described from Chiang Rai Province, Thailand (Mapook et al. 2020). R. camporesii has hysterothecial ascomata of 800-1100 pm long with black epithecium that changes to magenta in KOH and it is described from Yunnan Province, China (Hyde et al. 2020). Finally, Rk. hongheense has ascomata hystherothecial 1200-2000 pm long with reddish- orange epithecium and it is described from Honghe County, Yunnan Province, China (Wanasinghe et al. 2021). R. cozumelense produces longer ascomata, hysterothecial to Three new species of Rhytidhysteron from Mexico 139 apothecial, 2500 to 3500 um long with reddish brown to black epithecium that chang- es to greyish magenta in KOH and it grows on Tabebuia rosea DC. (Bignoniaceae). R. esperanzae is phylogenetically close to R. mexicanum, both species described from Mexico presenting similar hysterothecial to apothecial ascomata, sizes of 2000-— 4500 x 1200-2500 um and a perpendicularly striate margin. However, they differ by the colour of the ascomata and the epithecium: in R. esperanzae, the ascomata is brown, the exciple dull-green to black, and the epithecium dark green to black, with a yellow reaction in KOH. In contrast, in R. mexicanum, the exciple is completely black and the epithecium yellowish green to pistachio green when fresh, light green, pale green to lemon yellow when dry, becoming ocher to yellow gold in KOH. Another difference is the size of the ascospores which are longer and wider in R. esperanzae: they are (42—)45-47(-49) x (15—)17—19(—23) um, while in R. mexicanum, they are 34-40 (—44) x 10-12(—15) um (Cobos-Villagran et al. 2021). On the other hand, Rk. mesophilum is characterised by navicular hysterothecia, stri- ated margin with green-yellowish pruina, reddish orange to orange red epithecium that changes to greyish magenta in KOH, and long ascospores. It is related phylogenetically to R. tectonae and R. rufulum. However, it is morphologically different, including in the size and colour of the hysterothecium, colour of the epithecium, colouration in the reaction with 10% KOH and the size of asci and ascospores. The hysterothecia of R. tectonae are 1225-3365 um long, with a smooth margin, yellow epithecium without reaction in KOH, ascospores (19—)28—29(—31) x (8—)10-—12(—13) pm and the species grows on Tectona grandis L., in Chiang Rai, Thailand (Doilom et al. 2016). In R. ru- fulum, the size of the ascomata ranges from 1500-2000 um long, the exciple is black, the epithecium brown, orange, or reddish, changing to magenta in KOH, and the as- cospores are 21—32(—39) x 8—9.6 um (Kutorga and Hawksworth 1997; Almeida et al. 2014; Thambugala et al. 2016; Cobos-Villagran et al. 2020). In contrast, the hystero- thecia of R. mesophilum are 2500-4000 um long, the epithecium orange, changing to greyish magenta in KOH, and the ascospores (38—)40—44(—46) x 12-14 um, therefore much longer and wider. In Mexico, the tropical dry forest is the best represented vegetation with four k/yz- idhysteron species: R. cozumelense, R. neorufulum, R. rufulum and R. neohysterinum. This is followed by the xerophilous scrub with R. thailandicum, R. rufulum and R. neohys- terinum, and only R. mexicanum in Quercus forest. Finally, in this study, we describe R. esperanzae and R. mesophilum in a tropical cloud forest, which is a vulnerable ecosystem and therefore these species are in danger of extinction. With the present study, the num- ber of khytidhysteron species known from Mexico reaches a total of eight and together with Thailand, they form the countries with the most species diversity of the genus. Acknowledgements Dr. T. Raymundo and Dr. R. Valenzuela gratefully acknowledge the financial support received from CONACYT and IPN of Project 252934, and Projects SIP-20210315 and SIP-20210661, respectively. Dr. C. Hernandez Rodriguez, Dr. L. Villa Tanaca, 140 Aurora Cobos-Villagran et al. / MycoKeys 83: 123-144 (2021) Dr. L. E. Mateo Cid and Dr. A. Pérez Valdespino thank IPN for financial support for their research of the Projects SIP 20200782, SIP-20210508, SIP-20210885 and SIP- 20210609, respectively. Dr. Calvillo Medina thanks CONACYT Postdoctoral scholar- ship (005352). Cobos Villagran thanks Posgrado en Biociencias, Escuela Nacional de Ciencias Bioldgicas, IPN. The authors gratefully acknowledge the Sistema Nacional de Investigadores (CONACYT) and COFAA (IPN). References Almeida DAC, Gusmao LFP, Miller AN (2014) Brazilian Semi-Arid Ascomycetes I: new and interesting records of hysteriaceous ascomycetes. Mycosphere 5(2): 379-391. https://doi. org/10.5943/mycosphere/5/2/11 Boehm EWA, Mugambi GK, Miller AN, Huhndorf SM, Marincowitz S, Spatafora JW, Schoch CL (2009a) A molecular phylogenetic reappraisal of the Hysteriaceae, Mytilinidiaceae and Gloniaceae (Pleosporomycetidae, Dothideomycetes) with keys to world species. 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