Phytotaxa 270 (2): 089–102 http://www.mapress.com/j/pt/ Copyright © 2016 Magnolia Press ISSN 1179-3155 (print edition) Article PHYTOTAXA ISSN 1179-3163 (online edition) http://dx.doi.org/10.11646/phytotaxa.270.2.2 Lamproconiaceae fam. nov. to accommodate Lamproconium desmazieri CHADA NORPHANPHOUN1,2,3, SINANG HONGSANAN3, MINGKWAN DOILOM2,3, DARBHE J. BHAT6,7, TING-CHI WEN1*, INDUNIL C. SENANAYAKE2,3,4, TIMUR S. BULGAKOV5 & KEVIN D. HYDE1,2,3,4,7 1 The Engineering Research Center of Southwest Bio-Pharmaceutical Resources, Ministry of Education, Guizhou University, Guiyang, 550025, Guizhou Province, China 2 School of Science, Mae Fah Luang University, Chiang Rai, 57100, Thailand 3 Center of Excellence in Fungal Research, Mae Fah Luang University, Chiang Rai, 57100 Thailand 4 World Agroforestry Centre, East and Central Asia, 132 Lanhei Road, Kunming 650201, China 5 Academy of Biology and Biotechnology, Southern Federal University, Rostov-on-Don 344090, Rostov region, Russia 6 Formerly Department of Botany, Goa University, Goa, India 7 No. 128/1–J, Azad Housing Society, Curca, Goa Velha, India *email:tingchiwen@yahoo.com Abstract The genus Lamproconium comprises species that are endophytes, saprobes and pathogens on a wide variety of plants. This genus is currently placed in Diaporthales genera incertae sedis. Fresh specimens of Lamproconium were collected in Russia and studied to provide morphological and phylogenetic data. Phylogenetic analyses of single spore isolates generated from maximum likelihood, maximum parsimony and Bayesian inference analyses using combined ITS and LSU sequence data, place L. desmazieri in the order Diaporthales. Melanconis desmazieri is synonymized under Lamproconium desmazieri, and Lamproconiaceae is introduced as a new family to accommodate L. desmazieri and Hercospora tiliae, based on morphology and phylogenetic analyses. Key words: Foliar pathogens, genera incertae sedis, morphological data, phylogenetic analyses, synonym Introduction Lamproconium (Grove) Grove was introduced as a monotypic genus by Grove (1937) to accommodate L. desmazieri (Berk. & Broome) Grove, and was placed in genera incertae sedis in the order Diaporthales by Cannon & Minter (2014). This genus is apparently a saprobe with an endophytic life style, or vice versa. Some authors, however, have considered a few species in this genus to be plant pathogens (Grove 1937, Cannon & Minter 2014, Sutton 1980). The type species of Lamproconium, originally described as Discella desmazieri Berk. & Broome, was collected from twigs of lime (Citrus aurantiifolia) in the UK (Berkeley & Broome 1850). Grove (1918) transferred the species to Melanconium in the subgenus Lamproconium as Melanconium desmazieri (Berk. & Broome) Sacc. Subsequently, Grove (1937) found that M. desmazieri differed from the type species of Melanconium (M. atrum Link) in having bluish to glistening or dark blue, but not brownish black, 1-septate conidia and accordingly re-circumscribed the species. Melanconium desmazieri differed from species of Discella by the absence of a true peridium, whereas Discella species have a proliferous stratum at the base. Grove (1937) therefore considered Discella desmazieri and Melanconium desmazieri as conspecific and introduced a new genus Lamproconium to accommodate this taxon (Grove 1937, Sutton 1980). Melanconium has been reported as the asexual morph of Melanconis Tul. & C. Tul. (Sutton 1980). The genus Melanconis belongs in the family Melanconidaceae in Diaporthales, Sordariomycetes (Maharachchikumbura et al. 2015, 2016). Castlebury et al. (2002) used LSU sequence data from M. stilbostoma (Fr.) Tul. & C. Tul., the type species of Melanconis, plus M. alni Tul. & C. Tul. and M. marginalis (Peck) Wehm. in a phylogenetic analysis and showed that Melanconis desmazieri Petr. clustered with Hercospora tiliae Tul. & C. Tul. However, the group clustered distantly from the family Melanconidaceae. Thus, Castlebury et al. (2002) placed Melanconis desmazieri and Hercospora tiliae in Melanconis sensu lato as genera incertae sedis in Diaporthales. This taxonomic treatment was followed by Voglmayr et al. (2012), Voglmayr & Jaklitsch (2014) and Maharachchikumbura et al. (2015, 2016). Accepted by Eric McKenzie: 25 Jul. 2016; published: 17 Aug. 2016 Licensed under a Creative Commons Attribution License http://creativecommons.org/licenses/by/3.0 89 Hercospora tiliae differs from Melanconis desmazieri in having ellipsoidal to cylindric-ellipsoidal, aseptate, hyaline conidia, while the sexual morph has ellipsoidal to cylindrical, hyaline, 1-septate ascospores (Tulasne & Tulasne 1863, Petrak 1938, Sutton 1980). In M. desmazieri conidia are oblong-fusoid, straight, at first acute, and later becoming obtuse at one or both ends (Petrak 1938, Castlebury et al. 2002, Voglmayr et al. 2012, Voglmayr & Jaklitsch 2014). In this study, we provide morphological and phylogenetic data for Lamproconium based on five specimens collected in Russia. Furthermore, Melanconis desmazieri is synonymized under Lamproconium desmazieri based on morphological characters and multi-locus phylogenetic analyses. These taxa form a distinct lineage in Diaporthales and therefore we introduce the family Lamproconiaceae to accommodate them. Material and methods Sample collection and examination of specimens The samples were collected from dead branches of Tilia cordata Mill. in the Rostov region, Russia, in May 2014. The specimens were returned to the laboratory in small paper bags, examined, identified and described following Norphanphoun et al. (2015). Micro-morphological characters were studied using a Motic SMZ 168 dissecting microscope for fungal fruiting bodies. Hand sections of the fruiting structures were mounted in water and examined for morphological details. Fungi were also examined using a Nikon Ni compound microscope and photographed with a Canon EOS 600D digital camera fitted to the microscope. Photo-plates were made by using Adobe Photoshop CS6 Extended version 13.0 × 64 (Adobe Systems, USA), while Tarosoft (R) Image Frame Work program v. 0.9.7 was used for measurements. The contents inside the conidiomata, which comprised conidiophores, conidiogenous cells, conidia and paraphyses, were removed with a sterile needle and soaked in sterile water in a glass container prior to examination. Cultures were obtained by single spore isolation as described in Chomnunti et al. (2014). Spore germination was observed and photographed using a Nikon Ni compound microscope fitted with Canon EOS 600D digital camera. Geminated spores were transferred aseptically to fresh malt extract agar (MEA) and incubated at room temperature (18−25°C). Colony characters were observed and measured after one week and also one month. The Herbarium specimens are deposited in the Mae Fah Luang University Herbarium, Chiang Rai, Thailand (MFLU) and duplicated in New Zealand Fungarium (PDD). Living cultures are deposited at Mae Fah Luang University Culture Collection (MFLUCC) and Kunming Culture Collection (KUMCC). Facesoffungi and Index Fungorum numbers are registered (Jayasiri et al. 2015, Index Fungorum 2016). DNA extraction, PCR amplification and sequencing DNA extraction was performed from fresh fungal mycelia growing on MEA at room temperature (18−25°C) for 3 weeks. The genomic DNA was obtained using a E.Z.N.A.TM Fungal DNA MiniKit (Omega Biotech, CA, USA) following the manufacturer’s instructions. Polymerase chain reactions (PCR) were carried out using primer pairs of ITS5 and ITS4 to amplify the internal transcribed spacer region (ITS1-5.8S-ITS2) (White et al. 1990), and the large subunit rDNA (28S, LSU) was amplified with primers LROR and LR5 (Vilgalys & Hester 1990). The amplification reaction was performed in 50 μl reaction volume containing 2 µl of DNA template, 2 µl of each forward and reverse primers, 25 µl of 2 × Bench TopTMTaq Master Mix (mixture of Taq DNA Polymerase (recombinant): 0.05 units/µL, MgCl2: 4 mM, and dNTPs (dATP, dCTP, dGTP, dTTP): 0.4 mM) and 19 µl of double-distilled water (ddH2O) (sterilized water). The PCR thermal cycle program for ITS gene amplification were set as: initially 95 °C for 3 min, followed by 30 cycles of denaturation at 95 °C for 1 min, annealing at 51 °C for 1 min, elongation at 72 °C for 45 s, and final extension at 72 °C for 10 min. The PCR thermal cycle program for LSU gene amplification were provided as: initially 95 °C for 3 min, followed by 34 cycles of denaturation at 95 °C for 30 s, annealing at 51 °C for 50 s, elongation at 72 °C for 1 min, and final extension at 72 °C for 10 min. The quality of PCR products were checked by using 1% agarose gel electrophoresis stained with ethidium bromide. Purification and sequencing of PCR product were carried out at Life Biotechnology Co., Shanghai, China. Phylogenetic analysis Blast searches were made to identify the closest matches in GenBank and recently published sequences in of Castlebury et al. (2002), Voglmayr et al. (2012), Voglmayr & Jaklitsch (2014) and Maharachchikumbura et al. (2015, 2016). 90 • Phytotaxa 270 (2) © 2016 Magnolia Press NORPHANPHOUN ET AL. Combined analyses of ITS and LSU sequence data of 62 taxa (Table 1) from Diaporthales were downloaded from GenBank and Magnaporthe salvinii (CBS 243.76) and M. grisea (GAD1) were used as outgroup taxa. Additionally, the datasets were optimized manually as detailed in Castlebury et al. (2002), Voglmayr et al. (2012), Voglmayr & Jaklitsch (2014) and Maharachchikumbura et al. (2015, 2016). The combined sequence alignments were obtained from MEGA7 version 7.0.14 (Kumar et al. 2015) and ambiguously aligned regions were excluded, gaps were treated as missing data which performed in BioEdit v. 7.2 (Hall 1999). Phylogenetic trees were inferred with maximum likelihood (ML), maximum parsimony (MP) and Bayesian inference (BI). TABLE 1. GenBank accession numbers of the sequences used in phylogenetic analyses. Species Strain ITS LSU Anisogramma anomala 529478 EU683064 EU683066 Anisogramma virgultorum 529479 EU683062 EU683065 Cainiella johansonii Kruys 731 (UPS) - JF701920 Chapeckia nigrospora AR 3809 - EU683068 Cryphonectria parasitica CMW 7048 JN942325 JN940858 Cryptodiaporthe aesculi AFTOL-ID 1238 - DQ836905 Cryptodiaporthe salicella AR3455 - AF408345 Cryptometrion aestuescens CMW 18790 GQ369458 HQ730869 Cryptometrion parasitica ATCC 38755 AY141856 EU199123 Cryptosporella hypodermia AR3552 EU199181 AF408346 Cryptosporella hypodermia AFTOL-ID 2124 - DQ862028 Cytospora elaeagni CFCC 89633 KF765677 KF765693 Cytospora parasitica MFLUCC 14-1055 KT459408 KT459409 Cytospora tanaitica MFLUCC 14-1057 - - Diaporthales sp. YMJ 1364 JX570889 JX570891 Diaporthales sp. BCC00200 - EF622231 Diaporthe eres AFTOL-ID 935 DQ491514 - Diaporthe eres AR3538 - AF408350 Diaporthe detrusa AR3424 - AF408349 Ditopella ditopa AR3423 - AF408360 Gnomonia gnomon CBS 199.53 AY818956 - Hapalocystis occidentalis WU 24705 - AY616231 Harknessia australiensis CPC 15029 JQ706085 JQ706211 Harknessia ellipsoidea CPC 17111 JQ706087 JQ706213 Harknessia eucalypti CBS 342.97 AY720745 AF408363 Harknessia pseudohawaiiensis CPC 17379 JQ706111 JQ706234 Hercospora tiliae AR3526 - AF408365 Lamproconium desmazieri MFLUCC 14-1047 KX430132 KX430133 Lamproconium desmazieri MFLUCC 15-0870 KX430134 KX430135 Lamproconium desmazieri MFLUCC 15-0871 KX430136 KX430137 Lamproconium desmazieri MFLUCC 15-0872 KX430138 KX430139 ...Continued on next page LAMPRoCoNIUM DESMAzIERI Phytotaxa 270 (2) © 2016 Magnolia Press • 91 TABLE 1. (Continued) Species Strain ITS LSU Lamproconium desmazieri MFLUCC 15-0873 KX430140 KX430141 Leucostoma niveum AR 3413 JX438624 NG_027590 Luteocirrhus shearii CBS 130776 KC197021 KC197019 Magnaporthe grisea GAD1 - JQ920470 Magnaporthe salvinii CBS 243.76 KM484861 DQ341498 Melanconiella ellisii BPI 843491 JQ926268 JQ926268 Melanconiella spodiaea MSH JQ926298 JQ926298 Melanconiella spodiaea SPOD JQ926300 - Melanconis alni AR3500 - AF408371 Melanconis alni AR3748 EU199195 EU199130 Melanconis desmazieri AR3525 - AF408372 Melanconis desmazieri AR3827 JX522735 - Melanconis desmazieri CBS 109780 JX522736 - Melanconis marginalis AR3442 - AF408373 Melanconis stilbostoma AR3501 - AF408374 ophiovalsa betulae AR3524 - AF408375 Ophiovalsa suffusa AR3496 - AF408376 Phragmoporthe conformis AR3632 - AF408377 Pilidiella castaneicola CBS 143.97 - AF408378 Pilidiella diplodiella STE-U 3708 AY339323 AY339284 Pilidiella wangiensis CPC 19397 JX069873 JX069857 Plagiostoma euphorbiae CBS 340.78 EU199198 AF408382 Pseudoplagiostoma eucalypti CBS 124807 GU973512 GU973606 Pseudoplagiostoma oldii CBS 124808 GU973534 GU973609 Pseudoplagiostoma variabile CBS 113067 GU973536 GU973611 Pseudovalsa longipes AR3541 - EU683072 Pseudovalsa modonia AR 3558 - EU683073 Rossmania ukurunduensis AR 3484 - EU683075 Schizoparme straminea CBS 149.22 - AF362569 Schizoparme straminea STE-U 3932 AY339348 AY339296 Stegonsporium protopyriforme CBS 117041 NR_126119 - Stilbospora macrosperma CBS 121883 JX517290 JX517299 Stilbospora macrosperma CBS 121695 JX517288 JX517297 Sydowiella fenestrans AR 3777 - EU683078 Thailandiomyces bisetulosus BCC00018 - EF622230 Tirisporella beccariana BCC36737 - JQ655450 Note. The ex-type strains are in bold. 92 • Phytotaxa 270 (2) © 2016 Magnolia Press NORPHANPHOUN ET AL. FIGURE 1. Maximum likelihood (ML) majority rule consensus tree of combined ITS and LSU sequence data based on MP, ML and Bayesian analyses. Values above the branches indicate maximum parsimony and maximum likelihood bootstrap ≥70%, (MPBS/MLBS). Values at the third positions, respectively, above or below the branches represent posterior probabilities (BI PP ≥ 0.95) from Bayesian inference analysis. The tree is rooted to Magnaporthe salvinii and M. grisea. Strain numbers are given following the taxon names. The new sequences resulting from this study are in blue. Synonyms are in red. Ex-type strains are in black bold. LAMPRoCoNIUM DESMAzIERI Phytotaxa 270 (2) © 2016 Magnolia Press • 93 Maximum-likelihood (ML) analysis was performed in RAxML (Stamatakis 2006) implemented in raxmlGUI v.1.3 (Silvestro & Michalak 2012). The 1000 rapid bootstrap replicates were run with generalized time reversible GTRGAMMA model of nucleotide substitution and searches for model selected for ML were applied. Trees were visualized using FigTree v1.4.0 (http://tree.bio.ed.ac.uk/software/figtree/, Rambaut 2012). Maximum parsimony (MP) analysis was performed using PAUP (Phylogenetic Analysis Using Parsimony) v. 4.0b10 (Swofford 2002). The trees were inferred using the heuristic search option with tree bisection-reconnection (TBR) as the branch swapping algorithm and 1000 random sequence additions. Maxtrees were setup to 5000, branches of zero length were collapsed and all multiple parsimonious trees were saved. Descriptive tree statistics for parsimony tree length [TL], consistency index [CI], retention index [RI], rescaled consistency index [RC] and homoplasy index [HI] were calculated for the Maximum Parsimonious Tree (MPT). The robustness of the most parsimonious trees were evaluated by 1000 bootstrap replications, each with ten replicates of random stepwise addition of taxa (Felsenstein 1985). The Kishino-Hasegawa tests (KHT) (Kishino & Hasegawa 1989) were performed to determine whether the trees were significantly different. Trees were viewed in TreeView v.1.6.6 (Page 1996). Bayesian inference (BI) analysis was performed using the Markov Chain Monte Carlo (MCMC) method with MrBayes 3.2.2 (Ronquist et al. 2012). The best-fit nucleotide substitution models for each dataset were separately determined using MrModeltest version 2.2 (Nylander 2004). GTR+I+G were selected as best-fitting models for the ITS and LSU datasets. The Markov Chain Monte Carlo sampling (MCMC) analyses, with four chains, were run, started from random tree topology and lasted 5,000,000 generations and sampled every 100 generations (Nylander et al. 2008). The Tracer v. 1.5.0 program was used to check the effective sampling sizes (ESS) that should be above 200, the stable likelihood plateaus and burn–in value (Rambaut & Drummond 2009). The first 5000 generations were excluded as burn-in and tree were visualized using FigTree v1.4.0 (http://tree.bio.ed.ac.uk/software/figtree/, Rambaut 2012). The phylograms are visualized in FigTree v1.4.0 (http://tree.bio.ed.ac.uk/software/figtree/, Rambaut 2012) and made in Adobe Illustrator CS6 and Adobe Photoshop CS6 Extended version 13.0 × 64. Sequences data from this study are deposited in GenBank. Results Phylogenetic analyses The phylogenetic tree based on combined analysis of ITS and LSU sequence data was used to resolve the relationships in Lamproconium in Diaporthales. The phylogenetic analyses were obtained from maximum likelihood (ML), maximum parsimony (MP) and Bayesian analyses. The alignment comprised 67 taxa and 1534 total characters including gaps. Parsimony analyses indicate that 920 characters were constant, 141 variable characters were parsimony uninformative and 473 characters were parsimony informative. The parsimony analysis of the data matrix resulted in two equally parsimonious trees and the first tree (TL = 518, CI = 0.530, RI = 0.801, RC = 0.425, HI = 0.470) is shown in Fig. 1. The Bayesian analysis resulted in the same topology as the MP trees. The phylogenetic results from Fig. 1 are discussed in the notes. Five isolates (MFLUCC 14-1047, MFLUCC 15-0870, MFLUCC 15-0871, MFLUCC 15-0872 and MFLUCC 15-0873) grouped together with three strains of Melanconis desmazieri (AR3525, AR3827 and CBS 109780) in the combined phylogeny with 100% ML, 70% MP bootstrap support and 1.0 PP support (Fig. 1). Our isolate grouped close to Hercospora tiliae, but as a distinct lineage with 97% ML, 76% MP support and 1.0 PP in the combined phylogeny (Fig. 1). There are two genera with nine strains constituting the family Lamproconiaceae, based on the multi-gene phylogeny and with support from morphological observations. Taxonomy Lamproconiaceae C. Norphanphoun, T.C. Wen & K.D. Hyde, fam. nov. Index Fungorum number: IF552187, Facesoffungi number: FoF 02248 Pathogen and saprobe on dying twigs and branches. Sexual morph: Undetermined. Asexual morph: Conidiomata pycnidial, solitary, partly immersed in host tissue, uniloculate, multiloculate or convoluted, dark blue (Lamproconium), 94 • Phytotaxa 270 (2) © 2016 Magnolia Press NORPHANPHOUN ET AL. dark blackish brown (Hercospora), erumpent in the centre. Pycnidium thick-walled, thin at inner layer, hyaline (Lamproconium), dark brown (Hercospora), comprising wall cells of textura angularis (Lamproconium) or textura intricata (Hercospora). ostiole absent, dehiscence irregular. Paraphyses interspersed with conidiophores. Conidiophores filiform or cylindrical, pale bluish or hyaline, septate, branched, smooth-walled, formed at the base of conidiomatal wall. Conidiogenous cells holoblastic, cylindrical to subcylindrical, each forming a single conidium at the conidiophore apex, or annellidic, colourless to olivaceous, smooth-walled. Conidia fusiform, ellipsoid, thickwalled, contents granular, aseptate, bluish to glistening dark blue (Lamproconium), hyaline (Hercospora), smoothwalled, produced in mucilage but without a distinct mucilaginous envelope or appendage. Type genus:—Lamproconium (Grove) Grove. Notes:—The order Diaporthales comprises 12 families, viz. Cryphonectriaceae Gryzenh. & M.J. Wingf., Diaporthaceae Höhn. ex Wehm., Gnomoniaceae G. Winter, Harknessiaceae Crous, Melanconidaceae G. Winter, Pseudoplagiostomataceae Cheew., M.J. Wingf. & Crous, Pseudovalsaceae M.E. Barr, Schizoparmaceae Rossman, D.F. Farr & Castl., Stilbosporaceae Link, Sydowiellaceae Lar.N. Vassiljeva, Tirisporellaceae Suetrong et al. and Valsaceae Tul. & C. Tul. (Maharachchikumbura 2015, 2016). The family Lamproconiaceae is established to accommodate Lamproconium and Hercospora and is introduced based on morphology and phylogenetic analyses. Lamproconiaceae forms a robust clade basal to Sydowiellaceae and Stilbosporaceae in the combined ITS and LSU phylogeny (Fig. 1). It is morphologically different in conidial form from the asexual morphs of Sydowiellaceae and Stilbosporaceae. Species of Sydowiellaceae have been reported as Melanconis-like and is allied with Hercospora, as shown in the earlier studies (Castlebury et al. 2002, Rossman et al. 2007). Nevertheless, Hercospora is a distinct genus in that the ostioles from individual fruiting bodies converging within the stroma and emerge as one ostiole. Hercospora tiliae, with its unusual asexual morph groups with Melanconis desmazieri, also from Tilia (Castlebury et al. 2002, Rossman et al. 2007, Petrak 1938, Castlebury et al. 2002, Voglmayr et al. 2012, Voglmayr & Jaklitsch 2014, Maharachchikumbura et al. 2015, 2016). Thus Hercospora falls outside Sydowiellaceae and belongs in Lamproconiaceae. The asexual species of Stilbosporaceae and Lamproconiaceae are coelomycetes. The conidia of Lamproconiaceae species are aseptate, fusiform or ellipsoid, with granular contents, and hyaline or bluish to glistening dark blue, while in Stilbosporaceae conidia are cylindrical, clavate to pyriform, eu- or distoseptate, with or without oblique or longitudinal septa and brown (Maharachchikumbura et al. 2016). Lamproconium (Grove) Grove, British Stem- and Leaf-Fungi (Coelomycetes) (Cambridge) 2: 321 (1937) Melanconium sect. Lamproconium Grove, Bull. Misc. Inf., Kew: 161 (1918) Type species:—Lamproconium desmazieri (Berk. & Broome) Grove. Lamproconium desmazieri (Berk. & Broome) Grove [as ‘desmazieri’], British Stem- and Leaf-Fungi (Coelomycetes) (Cambridge) 2: 321 (1937) Figs. 2–4 Discella desmazieri Berk. & Broome, Ann. Mag. nat. Hist., Ser. 2 5: 377 (1850) Melanconium desmazieri (Berk. & Broome) Sacc., Michelia 2(no. 7): 355 (1881) Melanconis desmazieri Petr., Annls. mycol. 36(1): 55 (1938) Facesoffungi number: FoF02249 Pathogen causing canker on branches or twigs of lime trees (Tilia spp.). Lime cankers associated with L. desmazieri, produced splitting and longitudinal breakage of the outer branches, the symptom will appear as localized, sunken, slightly discolored, dark blue to black lesions on branches discoloration and necrosis of the branches. Branch/top dieback associated with L. desmazieri in having black terminal dead shoots, apex downwards initially discoloration; becoming wilted, with brown to dark brown discoloration at the base, midrib, and finally becoming dry and dead. Sexual morph: Undetermined. Asexual morph: Conidiomata 800–1000 × 400–550 µm diam., pycnidial, solitary, partly immersed in host tissue, uniloculate, dark blue, with a raised centre. Pycnidium 50–70 μm, with multi-layered wall, thin at inner layer, hyaline, comprising wall cells of textura angularis. Paraphyses interspersed within conidiophores. Conidiophores 30–120 µm, arising from the outermost wall layer at the basal of pycnidium, filiform or cylindrical, pale bluish to hyaline, septate, branched, smooth-walled. Conidiogenous cells cylindrical to subcylindrical, annellidic, with flared periclinal thickenings in the collarette zone, colourless to olivaceous, smooth-walled. Conidia 22–28.5 × LAMPRoCoNIUM DESMAzIERI Phytotaxa 270 (2) © 2016 Magnolia Press • 95 8–10 µm (x̅ = 25.25 × 9 µm, n = 30), fusiform, ellipsoid, infrequently slightly curved, aseptate, initially hyaline, bluish to glistening dark blue at maturity, narrowly rounded at ends, smooth-walled. FIGURE 2. Lamproconium desmazieri (= Melaconium desmazieri) (redrawn from Grove 1918). A. Conidia. B. Conidiophores and developing conidia. FIGURE 3. Dieback disease caused by Lamproconium desmazieresi (MFLU 14-0780, reference specimen). A, B. Tilia cordata with conidiomata on twigs and branches. C. Immersed conidiomata on branch. Material examined:—RUSSIA. Rostov region: Krasnosulinsky district, Donskoye forestry, artificial forest, on dead branches of Tilia cordata Mill. (Tiliaceae), 21 May 2014, T. Bulgakov (MFLU 14-0780, reference specimen designated here, PDD); living culture, MFLUCC 14-1047, KUMCC. RUSSIA. Rostov region: Shakhty city, Central urban microdistrict, Central Park, parkland, on dying brunches (necrotrophic) of T. tomentosa Moench, 9 July 2015, T. Bulgakov (MFLU 15-1940, PDD); living culture, MFLUCC 15-0870, KUMCC. RUSSIA. Rostov region: Krasnosulinsky district, Donskoye forestry, ravine forest, on dead branches of T. cordata, 18 June 2015, T. Bulgakov (MFLU 15-2037, PDD); living culture, MFLUCC 15-0871, KUMCC. RUSSIA. Rostov region: Rostovon-Don city, territory of Southern Federal University, parkland, on dead and dying branches of T. cordata, 23 April 96 • Phytotaxa 270 (2) © 2016 Magnolia Press NORPHANPHOUN ET AL. FIGURE 4. Lamproconium desmazieresi (MFLU 14-0780, reference specimen). A–C. Conidiomata on host. D. Cross section of a conidioma. E. Peridium and raised host tissue. F. Apex of conidioma. G–I. Conidiogenous cells with attached conidia (note: annellations at the tip of the conidiogenous cell). J. Immature conidium. K–M. Mature conidia. N. Germinating conidium. Scale bars: A, B = 2 mm, C = 1 mm, D = 500 µm, E = 100 µm, F = 200 µm, G, H = 30 µm, I = 20 µm, J–M = 10 µm, N = 30 µm. LAMPRoCoNIUM DESMAzIERI Phytotaxa 270 (2) © 2016 Magnolia Press • 97 2015, T. Bulgakov (MFLU 15-2111, PDD); living culture, MFLUCC 15-0872, KUMCC. RUSSIA. Rostov region: Krasnosulinsky district, Donskoye forestry, ravine forest, on dying branches of T. cordata, 18 June 2015, T. Bulgakov (MFLU 15-2192, PDD); living culture, MFLUCC 15-0873, KUMCC. Notes:—Lamproconium was introduced as a section of Melanconium by Grove (1918) and as a subgenus (Index Fungorum 2016), with Melanconium desmazieri as the type species. The taxon with bright coloured spores was collected on living twigs of Tilia sp. in the UK. Grove (1937) had raised the subgenus to generic rank. In this study, we have determined our collections as having fusiform, ellipsoid, infrequently slightly curved, aseptate and glistening, dark blue conidia, with narrowly rounded ends (22–28.5 × 8–10 µm). The morphology of our collections is similar to Lamproconium desmazieri (Table 2). Therefore, we introduce our collections as belonging to the genus Lamproconium. Petrak (1938) reported Melanconis desmazieri as the sexual morph of Melanconium desmazieri, also from Tilia sp. In the phylogenetic study of Castlebury et al. (2002), based on LSU sequence data, Melanconis desmazieri fell outside Melanconidaceae sensu stricto and grouped with Hercospora tiliae. These taxa were therefore placed in Diaporthales genera incertae sedis (Castlebury et al. 2002, Voglmayr et al. 2012, Voglmayr & Jaklitsch 2014). Phylogenetic analyses in this study generated from maximum likelihood, maximum parsimony and Bayesian analyses using combined ITS and LSU sequence data from 67 taxa (including our new strains), indicate that L. desmazieri belongs with Hercospora tiliae as a distinct lineage of Diaporthales (Fig. 1). Hence, we synonymize M. desmazieri under L. desmazieri and designate one of our collections as a reference specimen for L. desmazieri. Hercospora Fr., Syst. orb. veg. (Lundae) 1: 119 (1825) Possible synonyms (See Index Fungorum 2016) Facesoffungi number: FoF02250 Saprobic on branches and twigs of temperate trees. Sexual morph: Stromatic tissues prosenchymatous around perithecia, delimited externally by blackened dense pseudoparenchymatous zone, interior whitish, composed of interwoven hyphae mixed with substrate cells. Ascomata perithecial, few, small, circinate, beaks converging, becoming united and erumpent through stroma surface as single large opening. Asci 8-spored, unitunicate, broadly cylindrical. Ascospores hyaline, broadly ellipsoid, one septate, wall smooth, without gelcoating, with narrow terminal and median appendages in some species. Asexual morph: Rabenhorstia sp., Stromata prosenchymatous. Conidiomata pycnidial, uniloculate, ostiolate, ostiole surrounded by a superficial cap of sterile tissues. Conidiophores elongate. Conidia hyaline ovoid to ellipsoid, one-celled. Type species:—Hercospora tiliae (Pers.) Tul. & C. Tul. Notes:—Fries (1825) listed Sphaeria tiliae Pers., and Sphaeria atrovirens Alb. & Schwein., as two of the species in Hercospora. However morphologically S. tiliae has hyaline ascospores while S. atrovirens comprising opaque ascospores. Tulasne and Tulasne (1863) accepted H. tiliae as the type species of Hercospora. Petrak (1938) and Ruhland (1900) implicated Rabenhorstia tiliae (Pers.) Fr., as the asexual morph of Hercospora tiliae. Fourteen species listed under Hercospora (Index Fungorum, 2016). Hercospora tiliae (Pers.) Tul. & C. Tul., Select. fung. carpol. (Paris) 2: 154 (1863) Figs. 5–6 Possible synonyms (See Index Fungorum 2016) Facesoffungi number: FoF02452 Saprobic on branches and twigs of Tilia sp. Sexual morph: Stromata 700–800 μm wide, prosenchymatous around perithecia, delimited externally by greenish-blackened dense pseudoparenchymatous zone, interior whitish, composed of interwoven hyphae mixed with substrate cells, 3–5 ascomata in a stromata. Ascomata 1–1.05 mm high, 0.24–0.34 mm diam., (x̅ = 1.02 × 334 μm, n = 10), perithecial, small, aggregated, scattered, globose to subglobose, light brown to dark brown, coriaceous, ostiolate, papillate. Papilla 625–645 μm high, 190–290 μm diam., (x̅ = 640 × 250 μm, n = 10), converging and erumpent through stroma surface as single, large opening, wide at the top, narrowing towards the base, dark brown region around base of papilla. Peridium 10–20 μm wide (x̅ = 16 μm, n = 10), comprises light brown, compressed, cells of textura angularis. Asci 140–175 μm × 17–24 μm diam., (x̅ = 160 × 21 μm, n = 10), 8spored, unitunicate, cylindrical, short-stalked, J- apical apparatus. Ascospores 20–25 μm × 9–11 μm diam., (x̅ = 23 × 98 • Phytotaxa 270 (2) © 2016 Magnolia Press NORPHANPHOUN ET AL. 10 μm, n = 10), uniseriate, broadly ellipsoid, 1-septate, not or lightly constricted at the septa, hyaline, smooth. Asexual morph: Stromata prosenchymatous. Conidiomata pycnidial, uniloculate, ostiolate, ostiole surrounded by a superficial cap of sterile tissues. Conidiophores elongate. Conidia 14–16.5 × 4.5–6.5 μm (x̅ = 15 × 5 μm, n = 10), hyaline ovoid to ellipsoid, one-celled. FIGURE 5. Hercospora tiliae (F148711, reference specimen). A. Packet of herbarium specimen. B. Herbarium specimens. C. Cross section of ascomata. D. Peridium. E. Papilla. F–H. Asci in water. I–K. Ascospores. Scale bars: C = 200 µm, D, F–H = 40 µm, E = 100 µm. LAMPRoCoNIUM DESMAzIERI Phytotaxa 270 (2) © 2016 Magnolia Press • 99 Material examined:—SWEDEN. Uppland: Upl. Stockholm: Roslagstull Stockholm, on bark of Tilia sp., L. Romell, 1 April 1887, F148711 (S). FIGURE 6. Hercospora tiliae (redrawn from Sutton 1980) A. Conidia. B. Conidiophores and developing conidia. C. Conidioma. Scale bars: A = 10 µm, B = 20 µm. Notes:—Hercospora Fr. comprises 15 species (Index Fungorum 2016) with Hercospora tiliae as the type. The characters of the genus include eustromatic, immersed, subepidermal, dark blackish brown, separate, uniloculate, multiloculate or convoluted and thick-walled conidiomata; conidiophores branched extensively at the base, less so above, hyaline, septate, smooth, often developing in mucilage, formed at the base and sides of the conidiomatal wall; and ellipsoid, thick-walled, hyaline, aseptate conidia (18–20 × 6–7.5 µm) (Petrak 1938, Sutton 1980). Phylogenetic studies (Castlebury et al. 2002, Rossman et al. 2007, Voglmayr et al. 2012, Voglmayr & Jaklitsch 2014) based on LSU sequence data, placed H. tiliae in Diaporthales, genera incertae sedis, where it grouped with Melanconis desmazierii. In the present study based on maximum likelihood, maximum parsimony and Bayesian analyses of combined ITS and LSU sequence data, Lamproconium desmazierii (= Melanconis desmazieri) and Hercospora tiliae, clustered in Lamproconiaceae fam. nov. 100 • Phytotaxa 270 (2) © 2016 Magnolia Press NORPHANPHOUN ET AL. Acknowledgements We would like to thank the Mushroom Research Foundation, Chiang Rai, Thailand, and the Science Research Foundation of Guizhou University (No. 201309) for supporting this research. Kevin D. Hyde thanks the Chinese Academy of Sciences, project number 2013T2S0030, for award of Visiting Professorship for Senior International Scientists at Kunming Institute of Botany. Chada Norphanphoun thanks Rungtiwa Pookamsak, Saowaluck Tibpromma, Anupama Daranagama, Chathumini Jayasiri and Yuan-Pin Xiao for helpful comments and advice on the manuscript. References Berkeley, M.J. & Broome, C.E. (1850) Notices of British fungi (380-437). 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