Neovaginatispora mangiferae Tennakoon, M.S. Calabon, E.B.G. Jones & K.D. Hyde, sp. nov.

Index Fungorum number: IF 559843; MycoBank number: MB 559843; Facesoffungi number: FoF 12720; Fig. 1

Etymology – Name reflects the host Mangifera indica, from which the holotype was collected.

Holotype – MFLU 18–i069

Saprobic on decaying stem of Mangifera indica. Sexual morph: Ascomata 430–500 μm high, 320–350 μm diam., scattered, semi-immersed to immersed, papilla erumpent through host surface, coriaceous to carbonaceous, dark brown to black, globose to subglobose, ostiolate. Ostiole crest-like, variable in shape, central, periphysate, broadly papillate, with an irregular pore-like opening. Peridium 25–40 μm wide, thick, composing two layers of irregular cells arranged in a textura angularis, outer layer with darker, light brown to brown flattened cells, inner layer comprising several layers of hyaline cells. Hamathecium 1.5–3 μm wide, massive, long cylindrical cellular, anastomosed, cellular pseudoparaphyses, hyaline, septate with small guttules. Asci 60–80×7–9 μm (x̅=60.9×7.2 μm, n=20), 8-spored, bitunicate, cylindrical to cylindric-clavate, short pedicellate with furcate to obtuse ends, apically rounded with an indistinct ocular chamber. Ascospores 13–17×3.0–5.5 μm (x̅=15.1×3.7 μm, n=25), biseriate, overlapping, fusiform to sunbfusoid, strongly constricted at the median septum, straight or slightly curved, 1-septate at the center, enlarged near the septum at the upper cell, hyaline, guttulate, smoothwalled, mostly with 4 guttules, with a helmet-shaped to subcylindrical mucilaginous sheath at each end when immature, invisible at maturity, 2.1–5.6 μm long, 2.2–4.4 μm wide. Asexual morph: Not observed.

Culture characteristics – Conidia germinating on potato dextrose agar (PDA) within 24 h. Germ tubes produced from the apical cell of conidia. Colonies growing on PDA, reaching 25–30 mm in 2 weeks at 25 °C. Mycelia superficial, medium dense, irregular, flat, slightly raised, surface smooth with crenate edge, fluffy to velvety with smooth aspects, zonate with different sector yellowish-brown to moss brown at the margin brownish-grey at the center; from below, light moss brown at margin, dark gray at the middle, brown at the center, no pigmentation and sporulation.

Material examined – Taiwan, Chiayi, Fanlu Township area, Dahu village, dead stems of Mangifera indica (Anacardiaceae), 5 August 2017, D.S. Tennakoon, DTW 018C (18–0069, holotype), ex-type living culture, MFLUCC 17–2652.

GenBank numbers – MG931027(LSU), MG931030 (SSU), MG931033 (ITS).

Notes – The isolate MFLUCC 17–2652 has formerly identified as Neovaginatispora fuckelii by Tennakoon et al. (2018) but in our phylogenetic analysis, it clustered with Neovaginatispora sp. MFLUCC 11–0577, and forms a separate subclade with N. fuckelii and N. clematidis with 100% MP, 1.00 BYPP support (Fig. 2). The pairwise nucleotide comparison of LSU and ITS of N. mangiferae MFLUCC 17–2652 show 2 bp (0/24%, 838 bp) and 1 bp (0.23%, 442 bp) differences with Neovaginatispora sp. MFLUCC 11–0577, respectively, with no differences on SSU sequence data. For this reason, we named the unidentified Neovaginatispora species (strain MFLUCC 11–0577) as another strain of N. mangiferae. In pairwise nucleotide comparisons of Neovaginatispora mangiferae MFLUCC 17–2652 with N. fuckelii MFLUCC 17–1334, there is a nucleotide difference of 6.94% (30/432 bp) in ITS, and 2.71% (22/813 bp) in LSU genes. A pairwise nucleotide comparisons of LSU, SSU, and ITS sequence data of N. mangiferae MFLUCC 17–2652 and N. clematidis reveals 22 bp (2.72%, 810 bp), 6 bp (0.64%, 938 bp), and 27 bp (7.03%, 384 bp) differences, respectively. Neovaginatispora mangiferae differs from N. fuckelii KT 634 by having larger ascomata (430–500 × 320–350 μm vs 150–180 × 200–250 μm) and thicker peridium (25–40 μm vs 15–25 μm), and periphyses in ostiole. The asci and ascospores are similar in size but ascospores of N. mangiferae is fusiform with acute ends but N. fuckelii has a fusiform ascospores with obtuse ends (Thambugala et al. 2015; Tennakoon et al. 2018; Bao et al. 2019). Neovaginatispora mangiferae has larger ascomata (430–500 × 320–350 μm vs 145–250 × 108–160 μm), shorter asci (60–80 × 7–9 μm vs. 53–105 × 9–12 µm), and smaller ascospores (13–17 × 3.0–5.5 μm vs. 16–19 × 5–7 µm) compared to N. clematidis (Phukhamsakda et al. 2020).

Figure 1– Neovaginatispora mangiferae (MFLU 18–0069, holotype). a Appearance of ascomata in host substrate. b Vertical section of ascoma. c Peridium. d Pseudoparaphyses. e Ostiole with numerous periphysis. f, g Bitunicate asci. h–k Ascospores in different stages of maturity. Scale bars: b=100 µm, c, d, h–k=5 µm, e, f, g=20 µm

Figure 2 – Phylogram generated from maximum likelihood analysis based on combined LSU, SSU, ITS, tef1, and rpb2 sequence data representing Lophiostomataceae (Pleosporales). Eighty-one strains are included in the combined analyses which comprised 3726 characters (741 characters for LSU, 973 characters for SSU, 526 characters for ITS, 1001 characters for tef1, and 1011 characters for rpb2) after alignment. Angustimassarina acerina (MFLUCC 14–0505) and Angustimassarina populi (MFLUCC 13–0034) in Amorosiaceae (Pleosporales) were used as the outgroup taxa. Bootstrap support values for ML≥75% are given above the nodes (left side) and BYPP≥0.95 are given above the nodes (right side). Ex-type strains are in bold and newly generated sequences are in blue