Two new green-spored species of Trichoderma (Sordariomycetes, Ascomycota) and their phylogenetic positions Author Zhu, Zhao-Xiang Author Xu, Hao-Xiang Author Zhuang, Wen-Ying Author Li, Yu text MycoKeys 2017 26 61 75 http://dx.doi.org/10.3897/mycokeys.26.14919 journal article http://dx.doi.org/10.3897/mycokeys.26.14919 1314-4049-26-61 Trichoderma zonatum Z.X. Zhu, W.Y. Zhuang & Y. Li sp. nov. Figure 3 Diagnosis. Characterized by pulvinate, pale yellow to light brown stromata with densely disposed dark green to black ostioles, long asci (93-112 x 5.8-6.6 μm ), monomorphic and subglobose ascospores (4.2-5 x 4-4.7 μm ), simple trichoderma-like conidiophores, green, (sub)globose to pyriform conidia (2.8-3.8 x 2.3-2.8 μm ). Figure 3. The new species Trichoderma zonatum , holotype (HMJAU 34820). a-o Sexual state. a-g Dry stromata on nature substrate h Mature stroma after rehydration i Perithecium in section j Cortical and subcortical tissue in section k Subperithecial tissue in section l Stroma base in section. m-o Ascus with part-ascospores. p-aa Asexual state. p-r Cultures after 7 d at 25 °C (pPDA, qCMD, rSNA). s-y Conidiophores and phialides (SNA, 7 d). z, aa Conidia (SNA, 7 d). Scales bars: 2 mm (a); 1 mm (b, g); 500 μm ( c-f , h); 20 μm ( i-l , s, x, y); 5 μm ( m-o , z, aa); 20 mm ( p-r ); 10 μm ( t-w ). Type. CHINA. Henan: Xinyang City, Jigong mountain. 31°49'2"N , 114°04'16"E , 1500 m alt., on bark, 16 Jul 2015, B. Zhang 220 (HMJAU 34820, holotype), Ex-type culture CGMCC 3.18758. Description. Stromata generally solitary, scattered, gregarious, or aggregated in small groups, broadly attached, pulvinate to somewhat flattened, outline circular or with lobed margin, (0.5 -)1.0-2.5(- 3) mm diam (n = 20), (0.3 -)0.5- 0.8 mm high (n = 20). Surface flat, smooth, with slight perithecial protuberances, pale yellow to light brown, not changing colour in KOH, ostiolar openings obvious due to the green ascospores. In section stroma cortical tissue of textura angularis, 13-28 μm thick, not changing colour in 3% KOH, cells yellow, thin-walled, 6 -12(- 17) x 5 -10(- 13) μm (n = 40); subcortical tissue of textura angularis, cells hyaline, thin-walled, 4-10 x 5-8 μm (n = 40); subperithecial tissue of textura epidermoidea, cells hyaline, thin-walled, 10-22 x 8-17 μm (n = 40); tissue at the base of textura intricata, hyphae hyaline, thin-walled, (2.5 -)3.5-6(- 8) μm (n = 40) wide. Perithecia subglobose or flask-shaped, crowded, 178-216 x120- 165 μm (n = 40); peridium yellow in lactic acid, not changing colour in 3% KOH, (8 -)10-14(- 17) μm thick at the sides, (12 -)14-21(- 27) μm at the base (n = 40). Ostioles conical or cylindrical, 51-70 μm high, 31-54 μm wide at the apex (n = 40). Asci cylindrical, 93-112 x 5.8 -6.6(- 7) μm , with a stipe (13 -)18- 23 μm long (n = 60). Part-ascospores green, turning brown in KOH, distinctly verrucose, cells monomorphic, subglobose, also slightly ovoid, 4.2-5 x 4-4.7 μm (n = 100), l/w 1.0-1.1. On CMD colony radius after 72 h 30-43 mm at 20 °C, 32-46 mm at 25 °C, 17-34 mm at 30 °C, no growth at 35 °C. Colony hyaline, circular, loose, forming obvious zonate, covering the plate after 5-7 d at 25 °C. Aerial hyphae radially arranged. Conidiation at 25 °C noted after 3 d, first effuse, soon followed by formation of granules or pustules, particularly along the margin, spreading from the centre across the entire plate. No distinct odor, no diffusing pigment observed. On PDA after 72 h 38-48 mm at 20 °C, 55-62 mm at 25 °C, 28-30 mm at 30 °C, no growth at 35 °C; mycelium covering the plate after 8 d at 25°C. Colony circular, conspicuously dense, becoming zonate with broad, slightly downy zones and narrow, well-defined, convex, white to green farinose zones. Aerial hyphae numerous, mostly short, becoming fertile from the centre. Conidiation at 25 °C starting after 2 d, green after 4 d, first simple, mostly on short aerial hyphae concentrated in the centre and in denser zones, later abundant in pustules. Autolytic activity lacking or inconspicuous, no coilings seen. No diffusing pigment, no distinct odour noted. On SNA after 72 h 12-14 mm at 20 °C, 18-20 mm at 25 °C, 15-17 mm at 30 °C, no growth at 35 °C; mycelium covering the plate after 8-9 d at 25°C. Colony hyaline, thin, loose, irregularly lobed, not zonate. Aerial hyphae inconspicuous. Autolytic activity moderate. Conidiophores visible after 4 d, trichoderma-like, with 2 -3(- 4) whorls arising from the main axis. Phialides solitary or divergent in whorls of 2-3, mostly asymmetrically arranged, lageniform, (5 -)7-11(- 14) x 2 -3(- 4) μm , l/w 1.7 -2.8(- 4) (n = 60). Conidia green, (sub)globose to pyriform, smooth, (2.5 -)2.8- 3.8 x 2.3-2.8 μm , l/w (1.0 -)1.1-1.3(- 1.5) (n = 70). No chlamydospores formed. No distinct odor, no diffusing pigment observed. Habitat and distribution. On the surface of rotten wood in humid forests of south central and east China. Etymology. The specific epithet refers to the zonate colony on PDA. Other specimens examined. CHINA. Fujian: Quanzhou City, Qingyuan mountain. 24°55'53"N, 118°36'31"E, 200 m alt., on bark, 6 Aug 2015, Z.X. Zhu 225, HMJAU 34825, Ex-type culture CGMCC 3.18759. Remarks. Phylogenetic analyses based on RPB2 and TEF1-α indicated that Trichoderma zonatum belongs to the Chlorosporum clade, previously consisting of eight species, T. sinuosum , T. cremeum , T. surrotundum , T. chlorosporum , T. thelephoricola , T. rosulatum , T. cremeoides and T. costaricense . Phylogenetically, T. zonatum is most related to T. rosulatum and T. costaricense , but T. rosulatum is clearly distinguishable by dimorphic ascospores, gliocladium -like conidiophores, production of abundant chlamydospores and rosulate colony on CMD ( Zhu and Zhuang 2015 ); T. costaricense produces dimorphic and larger ascospores (5.5-6.0 x 5.2-5.7 μm ), verticillium-like conidiophores and ellipsoidal to cylindrical conidia ( Chaverri and Samuels 2003 , Zhu and Zhuang 2015 ). Species of the Chlorosporum clade usually produce pale yellow or pale green, semi-translucent stromata, globose to subglobose ascospores and gliocladium -like or verticillium-like conidiophores ( Chaverri and Samuels 2003 , Zhu and Zhuang 2015 ). Trichoderma zonatum is characterized by pulvinate, pale yellow to light brown stromata with densely disposed dark green to black ostioles, monomorphic ascospores, simple trichoderma-like conidiophores, green, (sub)globose to pyriform conidia. Morphologically, stromata of T. zonatum are not typical of the Chlorosporum clade and differ from all other species by relatively larger and non-transparent. It is most similar to T. chromospermum in gross stromata morphology, while the latter fungus clearly differs by much shorter asci [(78 -)85-90(- 102) μm ], gliocladium -like conidiophores and ellipsoidal to cylindrical conidia ( Chaverri and Samuels 2003 , Zhu and Zhuang 2015 ).