DNA isolation, amplification and phylogenetic analysis

Colonies were cultivated on 2 % potato-dextrose agar (PDA), and genomic DNA extraction was undertaken using the UltraClean™ Microbial DNA Kit (MO Bio, Carlsbad, CA, USA) according to manufacturer’s instructions. Using 20 isolates, we screened nine loci, of which the five more informative loci were selected for multi-gene analyses.

The primers ITS5 and ITS4 (White et al. 1990) were used to amplify the internal transcribed spacer region (ITS) of the nuclear ribosomal RNA gene operon, including the 3’ end of the 18S nrRNA, the first internal transcribed spacer region, the 5.8S nrRNA gene; the second internal transcribed spacer region and the 5’ end of the 28S nrRNA gene. The primers EF1-728F and EF1-986R (Carbone & Kohn 1999) were used to amplify part of the translation elongation factor 1-α gene (TEF1) and the primers ACT-512F and ACT-783R (Carbone & Kohn 1999) were used to amplify part of the actin gene (ACT). The primers Gpd1-LM and Gpd2-LM (Myllys et al. 2002) were used to amplify part of the glyceraldehyde-3-phosphate dehydrogenase (GPDH) gene, and part of the calmodulin (CAL) gene was sequenced using the primers CAL-228F and CAL-737R (Carbone & Kohn 1999). The primers CYLH3F (Crous et al. 2004b) and H3-1b (Glass & Donaldson 1995) were used to amplify part of the histone H3 (HIS) gene, and the primers T1 (O’Donnell & Cigelnik 1997) and Bt-2b (Glass & Donaldson 1995) to amplify part of the β-tubulin gene (TUB). The primers NMS1 and NMS2 (Li et al. 1994) were used to amplify an internal region of the mitochondrial SSU (mtSSU). The partial large subunit nrDNA (LSU) was sequenced using the primers LSU1Fd (Crous et al. 2009a) and LR5 (Vilgalys & Hester 1990).

Amplification reactions had a total reaction volume of 12.5 μL which was composed of 1× PCR buffer (Bioline GmbH, Luckenwalde, Germany), 5.6 % DMSO (v/v), 20 μM dNTPs, 0.2 μM of each forward and reverse primers, 0.25 U of BioTaq Taq DNA polymerase (Bioline GmbH, Luckenwalde, Germany), and 10 ng of genomic DNA. PCR conditions were the same for all loci, except for the MgCl₂ concentration: 2 mM MgCl₂ for the genes LSU and TEF1, 1.5 mM MgCl₂ for the genes ACT, GPDH, mtSSU, ITS and TUB, and 1 mM MgCl₂ for CAL and HIS genes. The PCR conditions were: start step of 2 min at 94 °C, followed by 40 cycles of 30 s at 94 °C, 1 min at adequate annealing temperature, and 1 min at 72 °C, followed by a finishing step of 3 min at 72 °C and a cool down step to 4 °C. The annealing temperature varied for each gene: 61 °C (ACT, GPDH, mtSSU); 58 °C (CAL, ITS, HIS); 55 °C (TEF1, TUB) and 48 °C (LSU).

However, some of these primer pairs failed to amplify with some isolates included in this study, and therefore additional combinations were used. The amplification reaction and cycle conditions were the same except the annealing temperature and MgCl₂ concentration. For the amplification of TEF1 with primers EF1-728F and EF2 (O’Donnell et al. 1998), 52 °C and 2 mM MgCl₂; TUB with primers T1 (O’Donnell & Cigelnik 1997) and CYLTUB1R (Crous et al. 2004b), 50 °C and 1 mM MgCl₂; CAL with primers CAL-228F and CAL2Rd (Quaedvlieg et al. 2011, Groenewald et al. 2013), 58 °C and 1 mM MgCl₂.

Amplicons were sequenced using both PCR primers with a BigDye Terminator Cycle Sequencing Kit v. 3.1 (Applied Biosystems, Foster City, CA, USA) according to the manufacturer’s instructions, and sequences were analysed on an ABI Prism 3700 DNA Sequencer (Perkin-Elmer, Norwalk, Foster City, CA, USA). The consensus sequences were visually inspected using MEGA v. 5 software (Tamura et al. 2011). The alignment of obtained sequences was performed using the online MAFFT interface (Katoh & Toh 2008; http://mafft.cbrc.jp/alignment/server).

For the phylogenetic analyses based on Maximum Likelihood and Bayesian inference, we chose the best evolutionary models for each data partition using the software MrModelTest v. 2.3 (Nylander 2004). MrBayes v. 3.1.1 (Ronquist & Huelsenbeck 2003) was used to generate the phylogenetic trees under optimal criteria per data partition. The heating parameter was set at 0.3 and the Markov Chain Monte Carlo (MCMC) analysis of four chains was started in parallel from a random tree topology and lasted until the average standard deviation of split frequencies came below 0.01. Trees were saved each 10 000 generations and the resulting phylogenetic tree (Fig. 1) was printed with Geneious v. 5.5.4 (Drummond et al. 2011) and the layout of the tree was done in Adobe Illustrator v. CS5.1. Diaporthella corylina (CBS 121124) was used as outgroup in the phylogenetic analyses based on its position as sister family in Diaporthales (Vasilyeva et al. 2007). New sequences generated in this study were deposited in NCBIs GenBank nucleotide database (www.ncbi.nlm.nih.gov; Table 1) and the alignment and phylogenetic tree in TreeBASE (study S13943; www.treebase.org).

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Fig. 1

Consensus phylogram of 22 104 trees resulting from a Bayesian analysis of the combined 5-gene sequence alignment. Clades are numbered on the right of the boxes and Diaporthe species names in purple reflect new combinations and in red new species. Strain accession numbers are followed by the original species name (black, when applicable), the isolation source (green) and country of origin (blue). Accession numbers and names in bold represent strains known to be ex-type strains or are considered to be authentic for the species. Red dots indicate strains from medicinal plants and yellow dots from humans. Bayesian posterior probabilities are shown at the nodes and the scale bar represents the expected changes per site. The tree was rooted to Diaporthella corylina (strain CBS 121124).

Locus resolution and SNP detection

Neighbour-joining analyses using the general time-reversible substitution model were applied to each data partition individually to check the stability and robustness of each species clade under each dataset using PAUP v. 4.0b10 (Swofford 2003) (TreeBASE study S13943). Alignment gaps were treated as missing data and all characters were unordered and of equal weight. Any ties were broken randomly when encountered. The robustness of the trees obtained was evaluated by 1 000 bootstrap replications (Hillis & Bull 1993). In the present study, both the analysis of the combined alignment (Fig. 1) and of the individual loci were used to determine the species boundaries. For each clade in the combined analysis, the position of the members of that clade was determined in the phylogenetic tree obtained from each of the individual loci to check whether these members still represent a single clade in the individual gene tree. In this way the robustness of a given clade could be evaluated together with the posterior probability value of that clade. A species was only counted if it was distinct from its closest relatives and the species clade contained all the associated strains.

Unique fixed nucleotide positions are used to characterise and describe several sterile species (see applicable species notes). For each sterile species that was described, the closest phylogenetic neighbour(s) were selected from Fig. 1 and this focused dataset was subjected to SNP analyses. These single nucleotide polymorphisms (SNPs) were determined for each aligned data partition using DnaSP v. 5.00.07 (Librado & Rozas 2009).

Locus resolution and SNP detection

The mtSSU and LSU regions had very few informative sites for the tested strains and were therefore not selected as good markers at species level. The ACT and GPDH regions were also discarded as suitable candidates for the multi-gene analyses because of their long branch lengths which made unambiguous alignments impossible. These four loci were therefore not used for further amplification and sequencing on the complete dataset. The remaining five loci had varied success for species identification and some phylogenetic lineages were more prone to less variability than others. Fifty-eight of the 95 species could be identified by all five loci. The loci are treated individually below:

• CAL – The locus could distinguish 74 of the 95 species (78 % success). It had difficulty separating: D. endophytica and D. phaseolorum (clades 4, 5); D. angelicae, D. arctii and D. subordinaria (clades 17–19); D. alleghaniensis, D. alnea, D. celastrina, D. eres, D. juglandina, D. neilliae and D. nobilis (clades 60–62, 64–67); and D. eugeniae, D. musigena, D. perseae, D. pseudomangiferae, D. pseudophoenicicola, Diaporthe sp. 6 and Diaporthe sp. 7 (clades 82–86, 88, 89). A single strain each of D. angelicae (clade 17), D. novem (clade 22) and D. terebinthifolii (clade 9) clustered separate from the other strains of the species.

• HIS – The locus could distinguish 84 of the 95 species (88 % success). It had difficulty separating: D. australafricana and D. viticola (clades 49, 50); D. celastrina, D. eres and D. nobilis (clades 66, 67, 62); D. arecae and D. perseae (clades 86, 87); and D. pseudophoenicicola and Diaporthe sp. 8 (clades 89, 90). A single strain each of D. endophytica (clade 5) and D. terebinthifolii (clade 9) clustered separate from the other strains of the species. This is the only locus that can distinguish D. angelicae (clade 17).

• ITS – The locus could distinguish 75 of the 95 species (79 % success). It had difficulty separating: D. angelicae, D. arctii and D. subordinaria (clades 17–19); D. cynaroides and D. viticola (clades 48, 50); D. alnea, D. neilliae and D. nobilis (clades 60–62); D. arengae, D. eugeniae and D. pseudomangiferae (clades 81–83); D. arecae and D. perseae (clades 86, 87); and D. aspalathi and D. woodii (clades 93, 94). A single strain each of D. arecae (clade 87), D. inconspicua (clade 75), D. novem (clade 22) and D. terebinthifolii (clade 9), and two strains each of D. impulsa (clade 51) and D. infecunda (clade 23), clustered separate from the other strains of the species. This is the only locus that can distinguish D. celastrina (clade 17) and D. eres (clade 67).

• TEF1 – The locus could distinguish 72 of the 95 species (76 % success). It had difficulty separating: D. tecomae, D. terebinthifolii (clades 9, 10); D. angelicae, D. arctii and D. subordinaria (clades 17–19); D. australafricana and D. viticola (clades 49, 50); D. celastrina and D. juglandina (clades 65, 66); D. eres and D. nobilis (clades 67, 62); D. chamaeropis, D. cinerascens and D. foeniculaceae (clades 76–78); and D. arengae, D. arecae, D. eugeniae, D. musigena, D. perseae, D. pseudomangiferae, D. pseudophoenicicola, Diaporthe sp. 6 and Diaporthe sp. 8 (clades 81–87, 89, 90).

• TUB – The locus could distinguish 84 of the 95 species (88 % success). It had difficulty separating: D. endophytica and D. phaseolorum (clades 4, 5); D. alleghaniensis, D. celastrina, D. eres, D. juglandina, D. nobilis and D. vaccinii (clades 62–67); and D. aspalathi and D. woodii (clades 93, 94). A single strain of D. angelicae (clade 17) clustered separate from the other strains of the species. This is the only locus that can distinguish D. perseae (clade 86).

Descriptions based on DNA characters are provided for three species in the Taxonomy section, namely D. endophytica (clade 5), D. inconspicua (clade 75) and D. infecunda (clade 23). Diaporthe endophytica (clade 5) was compared to D. phaseolorum (clade 4); D. inconspicua (clade 75) to D. anacardii (clade 69), D. chamaeropis (clade 77), D. cinerascens (clade 76), D. elaeagni (clade 73), D. foeniculacea (clade 78), D. hickoriae (clade 72), D. oncostoma (clade 70), D. saccarata (clade 71) and D. stictica (clade 74); and D. infecunda (clade 23) to D. angelicae (clade 17), D. arctii (clade 19), D. cuppatea (clade 20), D. lusitanicae (clade 21), D. neoarctii (clade 16), D. novem (clade 22) and D. subordinaria (clade 18).

Taxonomy

The multigene analyses resulted in 95 well-supported clades correlating to 243 isolates of Diaporthe (Table 1, Fig. 1). Fifteen new species are described, nine of which were isolated from medicinal plants (Aspidosperma tomentosum, Maytenus ilicifolia, Schinus terebinthifolius, Spondias mombin) in Brazil (clades 5, 9, 11, 23, 30, 31, 34, 35 and 36). Twenty-eight clades contain ex-type strains of presently known species, or strains accepted as authentic for the species name or which could be designated as epitypes in the present study, and were therefore well-resolved (7, 8, 12, 14, 17, 20–22, 24, 26–28, 40, 42, 43, 45, 48–50, 63, 64, 69, 71, 72, 84 and 91–93). The sexual-asexual relationship was resolved for several taxa, and is reported below. New combinations in Diaporthe are introduced below for several Phomopsis names that represented well-resolved taxa. Several potential epitypes were identified during this study, which are discussed below.

Diaporthe acaciigena Crous, Pascoe & Jacq. Edwards, Persoonia 26: 123. 2011

Specimen examined. Australia, Victoria, Otway Ranges, Anglesea, S38°23′21.7″ E144°11′12.7″, on leaves of Acacia retinodes, 16 Oct. 2009, P.W. Crous, I.G. Pascoe & J. Edwards (holotype CBS H-20581, ex-type culture CPC 17622 = CBS 129521).

Notes — Clade 43 contains the ex-type culture of D. acaciigena isolated from Acacia retinodes in Australia. This species is morphologically similar to D. amygdali (clade 42) (Crous et al. 2011), and closely related to D. pustulata (clade 44).

Diaporthe acerina (Peck) Sacc., Syll. Fung. (Abellini) 1: 611. 1882

Basionym. Valsa acerina Peck, Ann. Rep. N.Y. State Mus. Nat. Hist. 28: 73. 1876. 1874.

Specimen examined. Unknown, from Acer saccharum, Sept. 1927, L.E. Wehmeyer (CBS 137.27).

Notes — Clade 39 is represented by D. acerina, isolated from Acer saccharum. This species is genetically similar to D. perjuncta (clade 40). It is known to occur in Europe and North America on dead limbs and trunks of Acer pseudoplatanus, A. saccharinum, A. saccharum, A. spicatum, and Acer sp. (Aceraceae) (Spielman 1985, Farr et al. 1989).

Diaporthe alleghaniensis R.H. Arnold, Canad. J. Bot. 45: 787. 1967

Specimen examined. Canada, Ontario, on branches of Betula alleghaniensis, June 1972, R.H. Arnold (ex-type culture CBS 495.72 = ATCC 24097 = DAOM 45776).

Notes — Clade 64 contains the ex-type strain of D. alleghaniensis, isolated from Betula alleghaniensis in Canada. Diaporthe alleghaniensis causes canker and dieback of B. alleghaniensis, B. lenta, B. papyrifera and B. pendula in Canada (Arnold 1975), but has also been reported from Japan (Farr & Rossman 2012).

Diaporthe alnea Fuckel, Jahrb. Nassauischen Vereins Naturk. 23–24: 207. 1870 (1869–1870)

• = Phomopsis alnea Höhn., Sber. Akad. Wiss. Wien, Math.-naturw. Kl., Abt. 1 115: 681 (33 of repr.). 1906.

Specimens examined. Unknown, on Alnus sp., June 1946, S. Truter (CBS 146.46); on Alnus sp., Aug. 1947, S. Truter (CBS 159.47).

Notes — Clade 61 consists of two isolates from Alnus (presumably collected in the Netherlands). Diaporthe alnea causes dieback of Alnus glutinosa (alder) and A. incana (grey alder). It has been reported from Europe, Russia and the USA (Munk 1957, Oak & Dorset 1983, Moricca 2002, Mel’nik et al. 2008, Farr & Rossman 2012).

Diaporthe ambigua Nitschke, Pyrenomycetes Germanici 2: 311. 1870

Specimens examined. Italy, Sicily, Catania, on Platanus acerifolia, G. Granata (CBS 127746 = IMI 395956); Perugia, on Helianthus annuus, Mar. 1987, A. Zazzerini (CBS 187.87). – Portugal, Vale Andeiro, on Foeniculum vulgare, J.M. Santos (CBS 123210 = Di-C003/10, CBS 123211 = Di-C002/9). – South Africa, Western Cape Province, from Pyrus communis, deposited 2002, S. Denman (ex-epitype culture CBS 114015 = CPC 2657); Western Cape Province, on crown of Aspalathus linearis, 15 May 1997, J.C. Janse van Rensburg (CBS 117167 = CPC 5414).

Notes — Clade 26 represents D. ambigua, which contains two isolates previously misidentified as D. scabra (CBS 127746) and D. helianthi (CBS 187.87), and four isolates of D. ambigua, including the ex-epitype culture. Diaporthe ambigua is an important pathogen of Malus domestica, Prunus salicina and Pyrus communis in South African fruit orchards. Infection by D. ambigua is associated with sunken lesions with longitudinal cracks on affected fruit trees. The fungus rapidly kills nursery rootstocks, but also kills mature rootstocks over a longer period of time (Smit et al. 1996). This species is also found as saprobe on wild fennel (Santos & Phillips 2009). It has been reported on Aspalathus linearis (van Rensburg et al. 2006), Foeniculum vulgare, Malus domestica (Smit et al. 1996, Santos & Phillips 2009), Malus sylvestris (Crous et al. 2000), Prunus spp. (Smit et al. 1996, Mostert et al. 2001a), Pyrus communis (Nitschke 1867), Pyrus ussuriensis (Tai 1979) and Vitis vinifera (van Niekerk et al. 2005). It is widely distributed, and is known from China, Cuba (Tai 1979), Germany (Nitschke 1867), South Africa (Smit et al. 1996), UK (Dennis 1986) and the USA (Washington) (Shaw 1973).

Diaporthe ampelina (Berk. & M.A. Curtis) R.R. Gomes, C. Glienke & Crous, comb. nov. — MycoBank MB802922

Basionym. Phoma ampelina Berk. & M.A. Curtis, Grevillea 2, 18: 81. 1873.

• ≡ Phomopsis ampelina (Berk. & M.A. Curt.) Grove, Bull. Misc. Inform. Kew 4: 184. 1919.

• = Phoma viticola Sacc., Michelia 2: 92. 1880.

• ≡ Phomopsis viticola (Sacc.) Sacc., Ann. Mycol. 13: 118. 1915.

• = Fusicoccum viticolum Reddick, Cornell Univ. Agric. Exp. Sta. Bull. 263: 331. 1909.

• ≡ Phomopsis viticola (Reddick) Goid., Atti Reale Accad. Naz. Lincei 26: 107. 1937.

• = Phomopsis viticola Sacc. var. ampelopsidis Grove, Bull. Misc. Inform. Kew 4: 183. 1919.

• = Diaporthe neoviticola Udayanga, Crous & K.D. Hyde, Fung. Diversity 56: 166. 2012 (a nom. nov. based on Phoma viticola Sacc.).

Conidiomata pycnidial, eustromatic, subepidermal, brown to black, scattered or aggregated, globose, flask-like to conical, outer surface smooth, convoluted to unilocular, singly ostiolate, up to 430 μm wide and 190–300 μm tall, including short necks which rarely occur. Pycnidial wall consisting out of two regions of textura angularis; the outer region brown, 2–3 cells thick, 5–7 μm wide, inner region brown, 3–4 cells thick, 7–15 μm wide, with the outside cells compressed. Conidial mass globose or in cirrhi, white, pale-yellow to yellow, but predominantly pale-yellow. Alpha conidiophores cylindrical, some filiform, rarely septate and branched, 5–35 × 1–3 μm (av. = 25 × 2 μm). Alpha conidiogenous cells subcylindrical, tapering towards the apex, collarettes and periclinal thickening present, 3–19 × 1–2 μm (av. = 10 × 1.5 μm). Alpha conidia commonly found, fusoid-ellipsoidal, apex acutely rounded, base obtuse to subtruncate, multi-guttulate with guttules grouped at the polar ends, rarely biguttulate, (7–)9.5–10.5(–13) × (1.5–)2–3(–3.5) μm (av. = 10 × 2.5 μm). Beta conidiophores ampulliform to subcylindrical, rarely branched, 10–34 × 1–2 μm (av. = 26 × 1.5 μm). Beta conidiogenous cells subcylindrical, tapering towards the apex, collarette and periclinal thickening present, 7–14 × 1–2 μm (av. = 11–1.5 μm). Beta conidia less common than alpha conidia, straight, curved or hamate, 20–25 × 0.5–1 μm (av. = 23–1 μm). Gamma conidia rarely observed, fusoid to subcylindrical, apex acutely rounded, base subtruncate, multi-guttulate, 12–18 × 1.5–2 μm (av. = 15 × 2 μm). Description adapted from Mostert et al. (2001a).

Specimens examined. France, Bordeaux, Naujan-et-Postiac, on Vitis vinifera (Cabernet Sauvignon grapevine), May 1998, P. Larignon (PREM 56460 neotype, ex-neotype culture CBS 114016). – Italy, Perugia, on Vitis vinifera, May 1980, A. Zazzerini (CBS 267.80 = CPC 2671). – Turkey, from Vitis vinifera, 1 Dec. 2001, M. Erkan (CBS 114867 = CPC 4708). – USA, California, on Vitis vinifera, J.D. Cucuzza (CBS 111888 = ATCC 48153 = CPC 2673).

Notes — Grove (1919) distinguished P. ampelina (K 58408) from P. viticola by its external appearance on the host. However, Mostert et al. (2001a) re-examined the type specimen, and found alpha conidia to be ellipsoid-fusoid, 8–12 × 2.5–3.5 μm, within the range of P. viticola (Mostert et al. 2001a: f. 29), and thus considered them to be synonymous. Udayanga et al. (2012) proposed D. neoviticola as a nom. nov. for P. viticola, but this name is superfluous, as the older epithet ‘ampelina’ has precedence and should be adopted.

Diaporthe ampelina (clade 53) is a well-resolved species. It causes cane and leaf spot and infections of pruning wounds of Vitis and Ampelopsidis spp. (Vitaceae). Several species of Diaporthe can infect the host and cause variable symptoms in different parts of the vine (canes, leaves and fruits) causing considerable confusion in the taxonomy of these species on grapevine (Phillips 1999, Scheper et al. 2000, Mostert et al. 2001a). Merrin et al. (1995) studied the variation of Diaporthe in Australia using morphology. They identified two taxa (Phomopsis taxon 1 and taxon 2), which cause cane and leaf blight of Vitis spp.; and taxon 2 was identified as showing more resemblance to P. viticola. Mostert et al. (2001a) studied the species occurring on grapevines in South Africa using morphological, cultural, molecular and pathological characterisation and clarified the taxonomy of this complex. Diaporthe ampelina (= Phomopsis viticola, D. neoviticola, Phomopsis taxon 2 from Australia) was found to be the cause of cane and leaf spot disease, and was neotypified. Although the sexual morph has never been reported, Santos et al. (2010) found both MAT loci to be present in this species, and showed that it is heterothallic. However, the sexual morph could not be induced in culture by crossing opposing mating types.

Diaporthe amygdali (Delacr.) Udayanga, Crous & K.D. Hyde, Fung. Diversity 56: 166. 2012

Basionym. Fusicoccum amygdali Delacr., Bull. Soc. Mycol. France 21: 280. 1905.

• ≡ Phomopsis amygdali (Delacr.) J.J. Tuset & M.T. Portilla, Canad. J. Bot. 67, 5: 1280. 1989.

Specimens examined. Portugal, Mirandela, from Prunus dulcis, 2010, E. Diogo (ex-epitype culture CBS 126679); Tavira, on Prunus dulcis, 2010, E. Diogo (CBS 126680). – South Africa, Western Cape Province, on Vitis vinifera, 1 Mar. 1997, L. Mostert (CBS 111811 = CPC 2632); Western Cape Province, in wood on Prunus salicina, 2008, U. Damm (CBS 120840 = CPC 5833). – USA, Georgia, cankers on Prunus persica, Mar. 1994, W. Uddin (CBS 115620 = FAU 1005).

Notes — Diaporthe amygdali (clade 42) is the causal agent of twig canker and blight of almonds (Prunus dulcis) and peach (P. persica) wherever these hosts are grown (Diogo et al. 2010). It was first described as Fusicoccum amygdali causing cankers on almonds in France (Delacroix 1905). Tuset & Portilla (1989) re-examined the type specimen of F. amygdali and, based on morphology and symptomatology, they considered that it would be best accommodated in the genus Phomopsis. Clade 42 contains the ex-epitype strain (CBS 126679), five Phomopsis amygdali isolates from Prunus dulcis in Portugal, from P. persica in USA, P. salicina in South Africa, and from Vitis vinifera in South Africa.

Diaporthe anacardii (Early & Punith.) R.R. Gomes, C. Glienke & Crous, comb. nov. — MycoBank MB802923; Fig. 2

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Fig. 2

Diaporthe anacardii (CBS 720.97). a. Conidiomata sporulating on PNA; b. conidiomata sporulating on PDA; c, d. conidiogenous cells; e. beta conidia; f. alpha conidia. — Scale bars = 10 μm.

Basionym. Phomopsis anacardii Early & Punith., Trans. Brit. Mycol. Soc. 59, 2: 345. 1972.

Conidiomata pycnidial, sporulating profusely on OA, globose, up to 600 μm diam, multilocular, black, erumpent; cream conidial droplets exuding from central ostioles; walls consisting of 3–6 layers of medium brown textura angularis. Conidiophores hyaline, smooth, 1–3-septate, branched, densely aggregated, cylindrical, straight to sinuous, 10–25 × 2–3 μm. Conidiogenous cells 9–16 × 1.5–2 μm, phialidic, cylindrical to cymbiform, terminal and lateral, with slight taper towards apex, 1–1.5 μm diam, with visible periclinal thickening; collarette slightly flared, up to 2 μm long when present. Paraphyses rarely present, hyaline, smooth, 1–3-septate, cylindrical with obtuse ends, extending above conidiophores. Alpha conidia aseptate, hyaline, smooth, guttulate, fusoid to ellipsoid, tapering towards both ends, straight, apex subobtuse, base bluntly rounded with flattened hilum, (6.5–)7–8(–9) × (2–)3(–3.5) μm. Gamma conidia not observed. Beta conidia spindle-shaped, aseptate, smooth, hyaline, apex subacutely rounded, base truncate, tapering from lower third towards apex, curved, (15–)20–25 × 1.5(–2) μm.

Culture characteristics — Colonies covering dish after 2 wk in the dark at 25 °C. On OA dirty white with moderate aerial mycelium and patches of iron-grey. On PDA having patches of dirty white and umber, reverse bay with patches of umber. On MEA having patches of dirty white and olivaceous-grey, reverse umber with patches of olivaceous-grey.

Specimens examined. East Africa, on Anacardium occidentale, Apr. 1997, M. Puccioni (epitype designated here CBS H-21101, culture ex-epitype CBS 720.97). – Kenya, on Anacardium occidentale, 4 Dec. 1969, M.P. Early (holotype IMI 144866).

Notes — Phomopsis anacardii (clade 69 as D. anacardii) was described from Anacardi occidentalis in Kenya, and also recorded from Nigeria, Guinea and Cuba (Early & Punithalingam 1972).

Diaporthe angelicae (Berk.) D.F. Farr & Castl., Mycoscience 44: 204. 2003. — Fig. 3

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Fig. 3

Diaporthe angelicae (CBS 111591). a, b. Transverse section through conidiomata, showing conidiomatal wall; c, d. conidiogenous cells; e. alpha and beta conidia; f. conidiogenous cells giving rise to beta conidia; g. beta conidia. — Scale bars: a = 140 μm, all others = 10 μm.

Basionym. Sphaeria angelicae Berk., Mag. Zool. Bot.: 28. 1837.

• ≡ Diaporthopsis angelicae (Berk.) Wehm., The genus Diaporthe Nitschke: 228. 1933.

• ≡ Mazzantia angelicae (Berk.) Lar. N. Vassiljeva, Pyrenomycetes of the Russia Far East. I. Gnomoniaceae: 49. 1993.

• = Leptosphaeria nigrella Auersw., Mycol. Eur. Pyr. 5/6, pl. 12, f. 163. 1869.

• ≡ Diaporthe nigrella (Auersw.) Niessl, Beitr.: 51. 1872.

• ≡ Diaporthopsis nigrella (Auersw.) Fabre, Ann. Sci. Nat., Bot. 6 15: 35. 1883.

Conidiomata pycnidial, globose to ellipsoidal, aggregated or scattered, dark brown to black, immersed, ostiolate, 100–281 μm wide, 70–200 μm tall, lacking necks, with outer surface covered in hyphae; pycnidal wall consisting of brown, thick-walled cells of textura angularis; conidial mass globose or exuding in cirrhi, white to pale luteous or pale yellow. Conidiophores hyaline, subcylindrical, rarely branched, tapering towards the apex, aseptate, (12–)13–16(–18) × 3(–4) μm. Conidiogenous cells hyaline, subcylindrical, straight to curved, tapering towards the apex, collarette not flared, periclinal thickening inconspicuous, 8–10(–11) × 3(–3.5) μm. Alpha conidia hyaline, oblong to ellipsoid, apex bluntly rounded, base obtuse to subtruncate, bi- to multi-guttulate, (7–)8–10(–11) × 3(–4) μm. Beta conidia hyaline, smooth, spindle shaped, slightly curved, (19–)22–26(–28) × (1–)2 μm. Gamma conidia not observed (CBS 111592).

Culture characteristics — See Castlebury et al. (2003).

Specimens examined. Austria, Karnten, St. Margareten, decaying stems of Heracleum sphondylium, Aug. 2001, A.Y. Rossman (CBS 111591 = AR 3724); Niederosterreich, Ottenstein, decaying stems of Heracleum sphondylium, Aug. 2001, A.Y. Rossman (ex-epitype culture CBS 111592 = AR3776). – France, Bretagne, La Ville Borée, near Quessoy, on seeds of Heracleum sphondylium, 27 July 1990, H.A. van der Aa (CBS 501.90); sea dunes near Seignose le Penon, on Eryngium maritimum, leaf spots, 10 June 1986, H.A. van der Aa (CBS 344.86). – Italy, San Casciano, Prov., Florence, twig blight of Foeniculum vulgare, July 1996, L. Mugnai (CBS 100871). – Portugal, Malveira da Serra, Sintra, on Foeniculum vulgare, A.J.L. Phillips (CBS 123215 = Ph-C133/1).

Notes — Diaporthe angelicae (clade 17) is known to cause stem decay in several hosts including Heracleum sphondylium (Apiaceae) and Foeniculum vulgare (Apiaceae) in Europe and North America (Santos & Phillips 2009). Wehmeyer (1933) not only linked the conidial form of Phomopsis asteriscus to the sexual state Diaporthopsis angelicae, but also stated that Diaporthe berkeleyi was a synonym of Diaporthopsis angelicae. However, Castlebury et al. (2003) showed that Diaporthopsis is a synonym of Diaporthe, and also designated an epitype for D. angelicae.

Diaporthe arctii (Lasch) Nitschke, Pyrenomycetes Germanici 2: 268. 1870

Basionym. Sphaeria arctii Lasch, in Rabenh., Klotzsch. Herb. Vivum Mycol.: no. 1046. 1846.

• ≡ Phomopsis arctii (Lasch) Traverso, Fl. Ital. Crypt., Pars 1: Fungi. Pyrenomycetae. Xylariaceae, Valsaceae, Ceratostomataceae: 226. 1906.

Specimen examined. Unknown, from Arctium sp., Sept. 1925, A.W. Archer (CBS 136.25).

Notes — There are several clades that contain isolates previously identified as D. arctii (clades 16, 19, part 2 and 67, part 4). We suspect that clade 19 may represent the real D. arctii, as it is basal to D. subordinaria, and Wehmeyer (1933) regarded the latter (from Plantago lanceolata) as synonym of D. arctii (from Arctium).

Diaporthe arecae (H.C. Srivast., Zakia & Govindar.) R.R. Gomes, C. Glienke & Crous, comb. nov. — MycoBank MB802924

Basionym. Subramanella arecae H.C. Srivast., Zakia & Govindar., Mycologia 54, 1: 7. 1962.

Specimens examined. India, on fruit of Areca catechu, Feb. 1964, H.C. Srivastava (isotype CBS H-7808, ex-isotype culture CBS 161.64). – Suriname, on fruits of Citrus sp., Oct. 1975, I. Block (CBS 535.75).

Notes — The Diaporthe isolate from citrus (CBS 535.75) could well be distinct, but more strains are required to resolve this clade (clade 87).

Diaporthe arengae R.R. Gomes, C. Glienke & Crous, sp. nov. — MycoBank MB802925; Fig. 4

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Fig. 4

Diaporthe arengae (CBS 114979). a. Conidiomata sporulating on PNA; b, c. conidiogenous cells; d. beta conidia; e, f. alpha conidia. — Scale bars = 10 μm.

Etymology. Named after the host genus from which it was collected, Arenga.

Pycnidia in culture on PNA sporulating poorly, subglobose, up to 250 μm diam, black, erumpent; cream conidial droplets exuding from central ostiole; walls consisting of 3–6 layers of medium brown textura angularis. Conidiophores hyaline in upper region, pale brown at base, smooth, 0–6-septate, branched, densely aggregated, cylindrical, straight to sinuous, 10–60 × 2.5–4 μm. Conidiogenous cells 8–15 × 1.5–2.5 μm, phialidic, cylindrical, terminal and lateral, with slight taper towards apex, 1–1.5 μm diam, with visible periclinal thickening; collarette not flared, up to 2 μm long when present. Paraphyses not observed. Alpha conidia aseptate, hyaline, guttulate, fusoid-ellipsoid, tapering towards both ends, apex subobtuse, base with flattened hilum, (5–)6–7(–9) × (2–)2.5(–3) μm. Gamma conidia not observed. Beta conidia rarely observed, subcylindrical, aseptate, smooth, hyaline, apex bluntly rounded, base truncate, tapering absent to very slight, curved, 20–25 × 1.5 μm.

Culture characteristics — Colonies covering dish after 2 wk in the dark at 25 °C. On MEA surface with fluffy aerial mycelium, pale luteous, in reverse orange with patches of sienna. On OA umber with patches of sienna and saffron, in reverse umber with patches of saffron. On PDA surface with fluffy white aerial mycelium, umber with patches of saffron, in reverse umber with patches of pale luteous to luteous.

Specimen examined. Hong Kong, Victoria Peak, from Arenga engleri, 7 Oct. 1999, K.D. Hyde (holotype CBS H-21104, culture ex-type CBS 114979 = HKUCC 5527).

Notes — The Diaporthe species occurring on palms are summarised by Fröhlich et al. (1997). Diaporthe arengae (clade 81) is distinguished from known species based on a combination of its conidial morphology and host.

Diaporthe aspalathi E. Jansen, Castl. & Crous, Stud. Mycol. 55: 71. 2006

Basionym. Diaporthe phaseolorum var. meridionalis F.A. Fernández, Mycologia 88: 438. 1996 (non D. meridionalis Sacc., Syll. Fung. 1: 638. 1878).

Specimens examined. South Africa, Western Cape Province, Clanwilliam, Langebergpunt, in branch on Aspalathus linearis, J.C. Janse van Rensburg (ex-type culture CBS 117169 = CPC 5428); in crown on Aspalathus linearis, 17 Oct. 1997, J.C. Janse van Rensburg (CBS 117168 = CPC 5420); on Aspalathus linearis, 2 Dec. 1996, S. Lamprecht (CBS 117500 = CPC 5408).

Notes — Diaporthe aspalathi (clade 93) causes soybean stem canker in the South-eastern USA (Fernández & Hanlin 1996), and is not closely related to D. phaseolorum as might be expected. Although morphologically similar, this species clustered apart from the reference strain of D. phaseolorum (clade 4). Diaporthe aspalathi is also the main causal organism of canker and dieback of rooibos (Aspalathus linearis), and not D. phaseolorum as reported earlier (Smit & Knox-Davies 1989a, b, van Rensburg et al. 2006).

Diaporthe australafricana Crous & Van Niekerk, Australas. Pl. Pathol. 34: 33. 2005

Specimens examined. Australia, on Vitis vinifera, 1 July 1995, R.W.A. Schepers (ex-type culture CBS 111886 = CPC 2676). – South Africa, on V. vinifera, 1 Nov. 1997, L. Mostert (CBS 113487 = CPC 2655).

Notes — Clade 49 contains two isolates of D. australafricana, one of them being the ex-type strain (CBS 111886), which is a sibling species of D. viticola in clade 50 (van Niekerk et al. 2005). Both species were described from Vitis vinifera, but D. australafricana is thus far only known from grapevines in Australia and South Africa.

Diaporthe batatas Harter & E.C. Field, Phytopathology 2: 121. 1912

Specimen examined. USA, on Ipomoea batatas, Feb. 1921, L.L. Harter (CBS 122.21).

Notes — Clade 8 consists of a single strain of D. batatas isolated from Ipomoea batatas in the USA. This species and D. phaseolorum have in the past been considered as varieties, namely D. phaseolorum var. batatatis and D. phaseolorum var. batatae. However, the genetic data revealed no homology between the two species. Although it is not certain if CBS 122.21 (culture sterile) is an ex-type strain of D. batatas, it is regarded as authentic for the name.

Diaporthe beckhausii Nitschke, Pyrenomycetes Germanici 2: 295. 1870

Specimen examined. Unknown, from Viburnum sp., Sept. 1927, L.E. Wehmeyer (CBS 138.27).

Notes — Clade 47 is represented by D. beckhausii, which was isolated from Viburnum sp. (origin unknown, presumably North America, whereas the species was originally described from Viburnum collected in Germany). Diaporthe beckhausii is known from woody stems of Betula sp., Cydonia japonica, Elaeagnus angustifolia, Halesia sp., Menispermum canadense, Menispermum sp., V. opulus, Viburnum sp. and V. tinus in temperate North America and Europe (Farr & Rossman 2012).

Diaporthe brasiliensis R.R. Gomes, C. Glienke & Crous, sp. nov. — MycoBank MB802926; Fig. 5

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Fig. 5

Diaporthe brasiliensis (CBS 133183). a. Conidiomata sporulating on PDA; b, c. transverse section through conidiomata, showing conidiomatal wall; d, e. conidiogenous cells; f, g. alpha conidia. — Scale bars: b = 80 μm, all others = 10 μm.

Etymology. Named after the country where it was collected, Brazil.

Conidiomata pycnidial, globose to conical, immersed, scattered or aggregated, brown to black, ostiolate, 70–160 μm wide, 60–140 μm tall, necks 60–130 μm tall, outer surface smooth; pycnidal wall consisting of brown, thick-walled cells of textura angularis; conidial mass globose, white to pale-luteous. Conidiophores hyaline, cylindrical, filiform, straight to curved, 1–3-septate, (17–)20–27(–30) × 2(–4) μm. Alpha conidiogenous cells hyaline, cylindrical, filiform, straight to curved, collarette flared, with slight periclinal thickening, (7–)8–12(–14) × 2(–3) μm. Alpha conidia hyaline, ellipsoid to irregular, apex bluntly rounded, base obtuse to subtruncate, bi- to multi-guttulate, 6–7(–8) × 2–3 μm. Beta and gamma conidia not observed.

Culture characteristics — Colonies on PDA flat, with an entire edge, surface mycelium dense and felty, buff, grey-olivaceous or olivaceous-grey; colonies covering dish after 2 wk at 25 °C in the dark; reverse olivaceous, dull green, olivaceous-buff. On OA raised, entire edge, surface mycelium dense felty, smoke-grey to grey-olivaceous; reverse purplish grey to pale purplish grey, grey olivaceous or olivaceous buff. On MEA raised, with an entire edge, buff, smoke-grey, with patches of olivaceous-grey and vinaceous-buff; reverse dark mouse-grey, buff.

Specimens examined. Brazil, Rio de Janeiro, endophytic species isolated from leaf of Aspidosperma tomentosum (popular name Peroba-do-campo), July 2007, K. Rodriguez (holotype CBS H-21100, ex-type culture CBS 133183 = LGMF 924 = CPC 20300); same collection details (LGMF 926 = CPC 20302).

Notes — Endophytic isolates (clade 36) from a medicinal plant in Brazil.

Diaporthe carpini (Pers.) Fuckel, Jahrb. Nassauischen Vereins Naturk. 23–24: 205. 1870 (1869–1870)

Basionym. Sphaeria carpini Pers., Syn. Meth. Fung. (Göttingen) 1: 39. 1801.

Specimen examined. Sweden, Skåne, S. Mellby par., Stenshuvud, on Carpinus betulus, 14 Apr. 1989, K. & L. Holm (CBS 114437 = UPSC 2980).

Notes — Diaporthe carpini (clade 55) is known from several European countries, where it occurs on Carpinus spp.

Diaporthe caulivora (Athow & Caldwell) J.M. Santos, Vrandečić & A.J.L. Phillips, Persoonia 27: 13. 2011

Basionym. Diaporthe phaseolorum var. caulivora Athow & Caldwell, Phytopathology 44: 323. 1954.

Specimens examined. Canada, Ontario, in mature stem on Glycine soja, Mar. 1955, A.A. Hildebrand (CBS 178.55 = ATCC 12048 = CECT 2023). – Croatia, in stem on Glycine max, K. Vrandečić (ex-neotype culture CBS 127268).

Notes — Clade 91 is represented by two isolates of D. caulivora on Glycine soja and G. max, respectively obtained from Canada (CBS 178.55) and Croatia (ex-neotype: CBS 127268). The soybean canker species complex was recently treated by Santos et al. (2011).

Diaporthe celastrina Ellis & Barthol., J. Mycol. 8, 4: 173. 1902

Specimen examined. Unknown, on Celastrus scandens, Sept. 1927, L.E. Wehmeyer (CBS 139.27).

Notes — Strains from the USA are required to confirm the identity of this culture (clade 66).

Diaporthe chamaeropis (Cooke) R.R. Gomes, C. Glienke & Crous, comb. nov. — MycoBank MB802927; Fig. 6

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Fig. 6

Diaporthe chamaeropis (CBS 454.81). a. Conidiomata sporulating on PDA; b. conidiomata sporulating on PNA; c–e. conidiogenous cells; f. alpha conidia; g. beta conidia. — Scale bars = 10 μm.

Basionym. Phoma chamaeropis Cooke, Grevillea 13 (no. 68): 95. 1885.

• ≡ Phomopsis chamaeropsis (Cooke) Petr., as ‘Phomopsis chamaeropis’, Ann. Mycol. 17, 2/6: 83. 1920 (1919).

Conidiomata pycnidial in culture on PNA, globose, up to 400 μm diam (up to 600 μm diam on OA), black, erumpent; cream conidial droplets exuding from central ostioles; walls consisting of 3–6 layers of medium brown textura angularis. Conidiophores hyaline, smooth, 1–5-septate, branched, densely aggregated, cylindrical, straight to sinuous, 10–50 × 2–2.5 μm. Conidiogenous cells 10–20 × 1.5–2 μm, phialidic, cylindrical, terminal and lateral, with slight taper towards apex, 1–1.5 μm diam, with visible periclinal thickening; collarette not observed. Paraphyses not observed. Alpha conidia aseptate, hyaline, smooth, guttulate, fusoid to ellipsoid, tapering towards both ends, straight, apex subobtuse, base subtruncate, (5–)6–8(–9) × 2(–2.5) μm. Gamma conidia not observed. Beta conidia spindle-shaped, aseptate, smooth, hyaline, apex acutely rounded, base truncate, tapering from lower third towards apex, curved, (20–)22–27(–30) × 1.5(–2) μm.

Culture characteristics — Colonies covering dish after 2 wk in the dark at 25 °C. On OA with moderate aerial mycelium, surface dirty white with patches of pale olivaceous-grey, reverse with patches of dirty white and sienna. On MEA surface dirty white with patches of olivaceous-grey, reverse sienna, with patches of luteous. On PDA surface olivaceous-grey with patches of dirty white, reverse iron-grey.

Specimens examined. Croatia, Rab, slope behind Hotel ‘Imperial’, on dead branch of Spartium junceum, July 1970, J.A. von Arx (CBS 753.70). – Greece, Thessaloniki, dead part of leaf of Chamaerops humilis, Aug. 1981, H.A. van der Aa (CBS 454.81).

Notes — Conidial dimensions closely fit those provided in the original description (on Chamaerops humulis from Czechoslovakia; Uecker 1988), suggesting that these cultures (clade 77) could be authentic for the name.

Diaporthe cinerascens Sacc., Syll. Fung. (Abellini) 1: 679. 1882. — Fig. 7

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Fig. 7

Diaporthe cinerascens (CBS 719.96). a. Conidiomata sporulating on PDA; b, c. conidiogenous cells; d. alpha conidia. — Scale bars = 10 μm.

• = Phoma cinerescens Sacc., Michelia 1 (no. 5): 521. 1879.

• ≡ Phomopsis cinerascens (Sacc.) Traverso, Fl. Ital. Crypt. Pyrenomycetae 2, 1: 278. 1906.

Conidiomata pycnidial, sporulating poorly on MEA, globose, up to 300 μm diam, black, erumpent; creamy-luteous conidial droplets exuding from central ostioles; walls consisting of 3–6 layers of medium brown textura angularis. Conidiophores hyaline, smooth, 1–3-septate, branched, densely aggregated, cylindrical, straight to sinuous, 17–30 × 2–3 μm. Conidiogenous cells 8–18 × 2–3 μm, phialidic, cylindrical, terminal and lateral, with slight taper towards apex, 1.5–2 μm diam, with visible periclinal thickening; collarette mostly absent, slightly flared when present, up to 2 μm long. Paraphyses not observed. Alpha conidia aseptate, hyaline, smooth, guttulate, fusoid to ellipsoid, tapering towards both ends, straight, apex subobtuse, base subtruncate, 7–8(–9) × (2.5–)3 μm. Gamma conidia aseptate, hyaline, smooth, ellipsoid-fusoid, apex acutely rounded, base subtruncate, 8–12 × 3 μm. Beta conidia not observed.

Culture characteristics — Colonies covering dish after 2 wk in the dark at 25 °C. On MEA with profuse aerial mycelium, surface dirty white, reverse ochreous with patches of umber. On PDA with sparse aerial mycelium, surface olivaceous-grey, reverse iron-grey. On OA surface with moderate aerial mycelium, olivaceous-grey to pale olivaceous-grey.

Specimen examined. Bulgaria, Kostinbrod, Plant Protection Institute, on branch of Ficus carica, 1995, E. Ilieva (CBS 719.96).

Notes — Diaporthe cinerascens (clade 76) represents a European species occurring on Ficus, so the present culture could be authentic for the name, as the conidial dimenions match those provided in the original description. This species was orginally associated with canker and dieback of Ficus spp. in Italy (Saccardo 1879), and the causal organism identified as Phomopsis cinerascens (sexual morph: Diaporthe cinerascens) by Grove (1935). Diaporthe cinerascens affects all commercial figs in California (Ogawa & English 1991), and is found in several geographical locations of the world (Hampson 1981, Anderson & Hartman 1983, Benschop et al. 1984, Banihashemi & Javadi 2009). Ficus spp. are important exotic garden ornamentals across the USA and Canada as well as in the tropics.

Diaporthe citri F.A. Wolf, J. Agric. Res. 33, 7: 625. 1926

• = Phomopsis citri H.S. Fawc., Phytopathology 2, 3: 109. 1912.

Specimens examined. Brazil, on seed of Glycine max, A. Almeida EMBRAPA/PR (LGMF 946 = CPC 20322). – Italy, unknown host, June 1939, G. Goidánich (CBS 199.39). – Suriname, Paramaribo, on decaying fruit of Citrus sinensis, Apr. 1932, N.J. van Suchtelen (CBS 230.52).

Notes — Clade 6 is represented by three isolates. One isolate (CBS 199.39) was previously identified as D. conorum from Italy, while another originates from soybean seed collected in Brazil (LGMF 946), and the third isolate is from Citrus sinensis in Suriname (CBS 230.52). Because D. conorum is regarded as synonym of D. eres (clade 67), we tentatively refer to this clade as D. citri, awaiting more isolates from Citrus. Diaporthe citri is a serious pathogen that is widely distributed, and associated with melanosis and stem-end rot of citrus fruits (Punithalingam & Holliday 1973, McKenzie 1992, Mondal et al. 2007, Farr & Rossman 2012).

Diaporthe convolvuli (Ormeno-Nuñez, Reeleder & A.K. Watson) R.R. Gomes, C. Glienke & Crous, comb. nov. — MycoBank MB802928

Basionym. Phomopsis convolvuli Ormeno-Nuñez, Reeleder & A.K. Watson, Canad. J. Bot. 66, 11: 2232. 1988.

Specimen examined. Turkey, isolated from leaves with anthracnose on Convolvulus arvensis, D. Berner (CBS 124654 = DP 0727).

Notes — Phomopsis convolvuli (clade 3) was originally described from diseased leaves of Convolvulus arvensis in Québec (Ormeno-Nuñez et al. 1988). The isolate of Phomopsis convolvuli studied here (CBS 124654), was found causing anthracnose on field bindweed (Convolvulus arvensis), a troublesome perennial weed to many important agricultural crops in the world, and was considered potentially useful as biological control agent (Kuleci et al. 2009).

Diaporthe crataegi (Curr.) Fuckel, Jahrb. Nassauischen Vereins Naturk. 23–24: 204. 1870

Basionym. Valsa crataegi Curr., Trans. Linn. Soc. London 22: 278. 1858.

Specimen examined. Sweden, Skåne, Trolle-Ljungby par., Tosteberga, on Crataegus oxyacantha, 15 Apr. 1989, K. & L. Holm (CBS 114435 = UPSC 2938).

Notes — Clade 41 is represented by D. crataegi isolated from Crataegus oxyacantha in Sweden. The species is common on C. chrysocarpa, C. laevigata and C. oxyacantha in Canada and Europe (Farr & Rossman 2012).

Diaporthe crotalariae G.F. Weber, Phytopathology 23: 602. 1933

• = Phomopsis crotalariae G.F. Weber, Phytopathology 23: 602. 1933.

Specimen examined. USA, on Crotalaria spectabilis, Oct. 1933, G.F. Weber (ex-type culture CBS 162.33).

Notes — Clade 92 contains the ex-type strain (CBS 162.33) of D. crotalariae isolated from Crotalaria spectabilis in the USA.

Diaporthe cuppatea (E. Jansen, Lampr. & Crous) Udayanga, Crous & K.D. Hyde, Fung. Diversity 56: 166. 2012

Basionym. Phomopsis cuppatea E. Jansen, Lampr. & Crous, Stud. Mycol. 55: 72. 2006.

Specimen examined. South Africa, Western Cape Province, on Aspalathus linearis, 2006, J. Janse van Rensburg (holotype CBS H-19687, ex-type culture CBS 117499 = STE-U 5431 = CPC 5431).

Notes — Diaporthe cuppatea (clade 20) is known only from the original collection made from dying branches of Aspalathus linearis in South Africa (van Rensburg et al. 2006).

Diaporthe cynaroidis Marinc., M.J. Wingf. & Crous, CBS Biodiversity Ser. (Utrecht) 7: 39. 2008

Specimen examined. South Africa, Western Cape Province, on leaf litter of Protea cynaroides, 26 June 2000, S. Marincowitz (ex-type culture CBS 122676 = CMW 22190 = CPC 13180).

Notes — Clade 48 contains the ex-type culture of D. cynaroidis (CBS 122676), which was isolated from Protea cynaroides in South Africa (Marincowitz et al. 2008). This species is closely related to D. australafricana and D. viticola (clades 49 and 50, respectively).

Diaporthe decedens (Pers.) Fuckel, Jahrb. Nassauischen Vereins Naturk. 23–24: 30. 1871

Basionym. Sphaeria tessella var. decedens Pers., Syn. Meth. Fung. (Göttingen) 1: 48. 1801.

Specimens examined. Austria, on Corylus avellana, Oct. 2001, W. Jaklitsch (CBS 109772 = AR 3459). – Sweden, Öland, Kastlösa par., on Corylus avellana, 7 June 1989, K. & L. Holm (CBS 114281 = UPSC 2957).

Notes — Diaporthe decedens represents a European species on Corylus. Clade 68 consists of two isolates obtained on Corylus avellana from Austria and Sweden.

Diaporthe detrusa (Fr.) Fuckel, Jahrb. Nassauischen Vereins Naturk. 23–24: 205. 1870 (1869–1870)

Basionym. Sphaeria detrusa Fr., in Kunze & Schmidt, Mykologische Hefte (Leipzig) 2: 43. 1823.

• = Phoma detrusa Sacc., Michelia 2: 96. 1880.

• ≡ Phomopsis detrusa (Sacc.) Traverso, Fl. Ital. Crypt. Pars 1: Fungi. Pyrenomycetae. Xylariaceae, Valsaceae, Ceratostomataceae 1, 1: 195. 1906.

Specimens examined. Austria, on Berberis vulgaris, Oct. 2001, A.Y. Rossman (CBS 109770 = AR 3424). – Sweden, Uppland, Hållnäs par., on Berberis vulgaris, 14 May 1991, K. & L. Holm (CBS 114652 = UPSC 3371). – Unknown, on Berberis vulgaris, Sept. 1927, L.E. Wehmeyer (CBS 140.27).

Notes — Clade 54 contains three isolates of D. detrusa obtained from Berberis vulgaris in Austria, Sweden and one of them with an unknown origin (presumably North America). This European species is known to also occur in the USA (Farr & Rossman 2012).

Diaporthe elaeagni Rehm, Syll. Fung. 14: 546. 1899. — Fig. 8

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Fig. 8

Diaporthe elaeagni (CBS 504.72). a. Conidiomata sporulating on PNA; b. conidiomata sporulating on PDA; c, d. conidiogenous cells; e. beta conidia. — Scale bars = 10 μm.

• ?= Phoma elaeagni Sacc., Michelia 1, 3: 354. 1878.

• ≡ Phomopsis elaeagni (Sacc.) Petr., Ann. Mycol. 19, 1–2: 48. 1921.

Specimen examined. Netherlands, Maassluis, on twig of Elaeagnus sp., May 1972, J. Gremmen (CBS 504.72).

Notes — In culture CBS 504.72 (clade 73) primarily produces beta conidia (spindle shaped, 16–22 × 2 μm, thus wider than seen on average in most other species); alpha conidia rarely observed, fusoid-ellipsoidal, 7–10 × 2–3 μm, thus correlating with dimensions of Phomopsis elaeagni (Sacc.) Petr., which is a homonym of P. elaeagni Sacc. Furthermore, conidial dimensions of the asexual state of D. elaeagni are not known. Additional collections and type studies are thus required to resolve the complex occurring on Elaeagnus.

Diaporthe endophytica R.R. Gomes, C. Glienke & Crous, sp. nov. — MycoBank MB802929

Etymology. Named after its endophytic growth habit.

Cultures sterile. Diaporthe endophytica (clade 5) differs from its closest phylogenetic neighbour, D. phaseolorum (clade 4), by unique fixed alleles in five loci based on alignments of the separate loci deposited in TreeBase as study S13943: ITS positions 357 (C), 359 (G), 360 (T), 368 (A), 369 (A), 371 (A), 372 (G) and 373 (G); TUB positions 135 (C) and 592 (T); CAL position 145 (G); TEF1 positions 18 (G), 26 (T), 40 (T), 42 (A), 63 (A), 124 (A), 175 (A) and 343 (A); HIS position 369 (C).

Culture characteristics — Colonies with sparse aerial mycelium, covering the dish after 2 wk at 25 °C. On PDA buff, honey to isabelline; reverse smoke-grey. On OA smoke-grey to olivaceous-grey. On MEA buff with umber patches; reverse dark mouse-grey, with patches of isabelline.

Specimens examined. Brazil, endophytic in leaf on Schinus terebinthifolius, July 2007, J. Lima (LGMF 911 = CPC 20287, LGMF 919 = CPC 20295), (holotype CBS H-21107, culture ex-type LGMF 916 = CPC 20292 = CBS 133811); endophytic in petiole on Maytenus ilicifolia, July 2007, R.R. Gomes (LGMF 928 = CPC 20304, LGMF 934 = CPC 20310, LGMF 935 = CPC 20311, LGMF 937 = CPC 20313); in seed on Glycine max, A. Almeida EMBRAPA/PR (LGMF 948 = CPC 20324).

Notes — Clade 5 represents a distinct lineage, containing eight sterile isolates originating from Brazil. Four of them were isolated from Maytenus ilicifolia, three from S. terebinthifolius and one from soybean seeds. Isolates could not be induced to sporulate on any of the media defined in this study, nor on sterilised plant host tissue placed on WA.

Diaporthe eres Nitschke, Pyrenomycetes Germanici 2: 245. 1870

• = Phomopsis cotoneastri Punith., Trans. Brit. Mycol. Soc. 60, 1: 157. 1973.

• = Phoma oblonga Desm., Ann. Nat. Sci. Bot. 20: 218. 1853.

• ≡ Phomopsis oblonga (Desm.) Traverso, Fl. Ital. Crypt. Pars 1: Fungi. Pyrenomycetae. Xylariaceae, Valsaceae, Ceratostomataceae: 248. 1906.

Specimens examined. Austria, on Acer campestre, Oct. 2001, W. Jaklitsch (CBS 109767 = AR 3538 = WJ 1643). – Germany, Monheim, on leaf spot of Hordeum sp., 5 Aug. 1984, M. Hossfeld (CBS 841.84). – Italy, Milano, on twig of Juglans regia, Dec. 1980, M. Bisiach (CBS 102.81). – Latvia, on Rhododendron sp., I. Apine (CBS 129168). – Netherlands, Oostvoorne, on dead stems of Arctium sp., 13 Dec. 1984, M. de Nooij (CBS 110.85); Soest, Dalweg, on fallen fruit of Fraxinus sp., 21 Feb. 1999, G. Verkley (CBS 101742); Veldhoven, on dead branch of Sorbus aucuparia, Nov. 1973, W.M. Loerakker (CBS 287.74); Baarn, garden Chopinlaan, on dead branch of Wisteria sinensis, 6 June 1983, H.A. van der Aa (CBS 528.83); Soest, inside house, on Abutilon sp., 26 Mar. 1997, A. Aptroot (CBS 688.97); Baarn, potted plant, on cladodes of Opuntia sp., 23 Sept. 1996, H.A. van der Aa (CBS 365.97); on Alliaria officinalis, Feb. 1962, G.H. Boerema (CBS 445.62); Baarn, on dead leaf of Ilex aquifolium, 11 June 1967, H.A. van der Aa (CBS 370.67 = MUCL 9931); Prov. Zuid-Holland, Huize Oud-Poelgeest, Oegstgeest, dieback of Ilex aquifolium, 21 Nov. 1994, G.J.M. Verkley (CBS 694.94); Baarn, garden Eemnesserweg 90, on dead stem of Rumex hydrolapathum, 19 Mar. 1996, H.A. van der Aa (CBS 485.96); Baarn, Cantonspark, on withering leaf of Magnolia × soulangeana, 23 Oct. 1968, H.A. van der Aa (CBS 791.68); Zuid-Holland, Ridderkerk, Huys ten Donck, on leaf tip of Osmanthus aquifolium, 7 May 1977, H.A. van der Aa (CBS 297.77); on Phaseolus vulgaris, Sept. 1950, Goossens (CBS 422.50); Baarn, on dead stem of Allium giganteum, May 1985, H.A. van der Aa (CBS 283.85); from Laburnum × watereri ‘Vossii’, Apr. 1935, I. de Boer (CBS 267.55); Boskoop, nursery, dying twigs of Skimmia japonica, Nov. 1981, H.A. v. Kesteren (CBS 122.82). – UK, Scotland, on living and dead twig of Fraxinus excelsior, Feb. 1938, J.A. MacDonald (CBS 250.38); on Cotoneaster sp., 1971, H. Butin (ex-type culture of P. crotoneaster CBS 439.82 = BBA P-407 = IMI 162181a); Oxford, on Picea abies seedling, Nov. 1937, T.R. Peace (CBS 186.37). – Unknown, on rotten fruit of Malus sylvestris, May 1961, Geigy (CBS 375.61); May 1932, W.G. Hutchinson (CBS 267.32).

Notes — Diaporthe eres (clade 67) is the type species of the genus Diaporthe, and is present in several hosts, though it is known to be morphologically highly variable (Castlebury et al. 2002). Wehmeyer (1933) described this species on more than 60 hosts, and listed several synonymies based on morphological data. A detailed morphological study is required to designate a suitable epitype strain for D. eres, and to resolve the status of all its purported synonyms.

Diaporthe eugeniae (Punith.) R.R. Gomes, C. Glienke & Crous, comb. nov. — MycoBank MB802930

Basionym. Phomopsis eugeniae Punith., Trans. Brit. Mycol. Soc. 63, 2: 232. 1974.

Specimens examined. West Sumatra, on Eugenia aromatica, May 1973, J. Waller (holotype IMI 177560); Lampung, on leaf of Eugenia aromatica, July 1982, R. Kasim (CBS 444.82).

Notes — Diaporthe eugeniae (clade 83) was originally described on Eugenia aromatica from West Sumatra. Although the present isolate could be authentic for the name, it unfortunately proved to be sterile.

Diaporthe fibrosa (Pers.) Fuckel, Jahrb. Nassauischen Vereins Naturk. 23–24: 204. 1870 (1869–1870)

Basionym. Sphaeria fibrosa Pers., Syn. Meth. Fung. (Göttingen) 1: 40. 1801.

Specimens examined. Austria, Vienna, on Rhamnus cathartica, Oct. 2001, A.Y. Rossman (CBS 109751 = AR 3425). – Sweden, Uppland, Dalby par., Hässleborg, on Rhamnus cathartica, 10 Mar. 1987, K. & L. Holm (CBS 113830 = UPSC 2117).

Notes — Clade 52 consists of two isolates from Rhamnus cathartica collected in Sweden and Austria. Diaporthe fibrosa was originally described from Europe on Rhamnus, so these cultures may well prove to be authentic for the name.

Diaporthe foeniculacea Niessl, in von Thümen, Contr. Ad. Fl. Myc. Lusit. 2: 30. 1880. — Fig. 9

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Fig. 9

Diaporthe foeniculacea (CBS 111554). a. Conidiomata sporulating on PDA; b, c. transverse section through conidiomata, showing conidiomatal wall; d–f. conidiogenous cells; g. beta conidia; h. alpha conidia. — Scale bars: b = 250 μm, all others = 10 μm.

• = Phoma foeniculina Sacc., Syll. Fung. 3: 125. 1884.

• ≡ Phomopsis foeniculina (Sacc.) Câmara, Agron. Lusit. 9: 104. 1947.

• = Phomopsis theicola Curzi, Atti Ist. Bot. Univ. Pavia, 3 sér., 3: 65. 1927.

• = Diaporthe neotheicola A.J.L. Phillips & J.M. Santos, Fung. Diversity 34: 120. 2009.

Conidiomata pycnidial, eustromatic, multilocular, immersed, ostiolate, dark brown, scattered or aggregated, 350–890 μm wide, 160–320 μm tall, necks absent, outer surface covered with hyphae; pycnidal wall consisting of brown, thick-walled cells of textura angularis; conidial mass globose to conical and exuding in cirrhi, yellow to reddish brown. Conidiophores hyaline, subcylindrical and cylindrical, filiform, branched above the septa, tapering towards the apex, 1–3-septate, (19–)20–28(–32) × 2(–3) μm. Conidiogenous cells hyaline, subcylindrical and filiform, straight, slightly tapering towards the apex, collarette not flared, prominent periclinal thickening, (10–)11–15(–17) × 2(–3) μm. Alpha conidia hyaline, oblong to ellipsoidal, apex bluntly rouded, base obtuse to subtruncate, bi- to multi-guttulate (6–)7–9 × 2(–3) μm. Beta conidia hyaline, smooth, slightly curved, (26–)28–32(–34) × 1(–2) μm. Gamma conidia not observed (based on isolate CBS 111554).

Specimens examined. India, Calcutta, unknown host, Feb. 1948, S.R. Bose (CBS 400.48). – Italy, on leaves and branches of Camellia sinensis, Oct. 1927, M. Curzi (ex-type culture of P. theicola CBS 187.27); Perugia, on Diospyros kaki, June 1956, M. Ribaldi (CBS 287.56); Apulia, near Bari, on Prunus amygdalus, winter 1974/75, A. Ciccarone (CBS 171.78). – Netherlands, Baarn, ‘Madoera’, back frond, on Wisteria sinensis, 24 Apr. 1969, H.A. van der Aa (CBS 357.69). – New Zealand, Waikato region, on Pyrus pyrifolia, 2001, W. Kandula (CBS 116957). – Portugal, near Lisbon, São Marcos, base of senescent stem of Foeniculum vulgare, Apr. 2002, A.J.L. Phillips (CBS 111554); Évora, Foeniculum vulgare, 1 Nov. 2007, A.J.L. Phillips (ex-type cultures of Diaporthe neotheicola CBS 123209, CBS 123208); Pedras del Rei, near Tavira, on Bougainvillea spectabilis, 15 June 1988, H.A. van der Aa (CBS 603.88); Madeira, Serra da Agua, base of senescent stem of Foeniculum vulgare, Aug. 2001, A.J.L. Phillips (CBS 111553).

Notes — Diaporthe foeniculacea (clade 78) was originally described from Foeniculum vulgare in Portugal, and represents an older name for D. theicola and D. neotheicola. There are many described species that occur on Foeniculum vulgare (wild fennel). Among them, P. theicola and its teleomorph D. neotheicola (Santos & Phillips 2009), and D. foeniculacea, the causal agent of stem necrosis of fennel. Phillips (2003) redescribed D. foeniculacea, and established the sexual-asexual connection between D. foeniculacea and Phomopsis foeniculina. The synonymy of D. neotheicola under D. foeniculacea is based on the fact that the cultures matching the original descriptions are in fact genetically identical. However, as there are no ex-type strains of D. foeniculacea, this synonymy strongly relies on the earlier opinion of Phillips (2003). Either way, this matter can only be resolved once an epitype has been designated for D. foeniculacea, fixing the application of the name. We recommend that additional collections linked to stem necrosis of fennel in Portugal are obtained, before this decision is made.

Diaporthe ganjae (McPartl.) R.R. Gomes, C. Glienke & Crous, comb. nov. — MycoBank MB802932

Basionym. Phomopsis ganjae McPartl., Mycotaxon 18, 2: 527. 1983.

Specimen examined. USA, Illinois, Hannah City, dead leaf of Cannabis sativa, deposited Mar. 1991, J.M. McPartland (holotype HA 10987, ex-type culture ILLS 43621 = CBS 180.91).

Notes — Diaporthe ganjae (clade 24) is known only from the original collection taken from wilted, dead leaves of Cannabis sativa in Illinois, USA (McPartland 1983). Phylogenetically D. ganjae is closely related to an isolate identified as D. manihotia (CBS 505.76), isolated from Manihot utilissima in Rwanda (clade 25).

Diaporthe gardeniae (Buddin & Wakef.) R.R. Gomes, C. Glienke & Crous, comb. nov. — MycoBank MB802933

Basionym. Phomopsis gardeniae Buddin & Wakef., Gard. Chron., ser. 3 103: 45. 1938.

• = Phomopsis gardeniae H.N. Hansen & Barrett, Mycologia 30, 1: 18. 1938 (homonym).

Specimen examined. Italy, on stem of Gardenia florida, June 1956, M. Ribaldi (CBS 288.56).

Notes — Diaporthe gardeniae (clade 59) causes gardenia canker in Gardenia jasminoides, G. lucida and Gardenia sp. (Farr & Rossman 2012). This disease is considered as serious (Tilford 1934, Huber 1936, Miller 1961). It was originally observed in 1894 in England (Cooke 1894), and has since been reported from the USA (Preston 1945) and India (Mathur 1979). All parts of the plant are susceptible to infection, including roots, stems and leaves (McKenzie et al. 1940), although cankered stems are the most diagnostic symptoms for this disease.

Diaporthe helianthi Munt.-Cvetk., Mihaljč. & M. Petrov, Nova Hedwigia 34: 433. 1981

• = Phomopsis helianthi Munt.-Cvetk., Mihaljč. & M. Petrov, Nova Hedwigia 34: 433. 1981.

Specimens examined. Serbia, Vojvodina, overwintering stem on Helianthus annuus, 1980, M. Muntañola-Cvetkovic (ex-type culture CBS 592.81 = CBS H-1540). – Unknown, on seed of H. annuus, June 1994, Vanderhave Res., Rilland, Netherlands (CBS 344.94).

Notes — Diaporthe helianthi (clade 14) is associated worldwide with stem canker and grey spot disease of sunflower (Helianthus annuus) (Muntañola-Cvetkovic′ et al. 1981). Yield reductions of up to 40 % have been recorded in Europe (Masirevic & Gulya 1992) including the former Yugoslavia as well as France where it was considered a major pathogen of sunflower (Battilani et al. 2003, Debaeke et al. 2003). Diaporthe helianthi is also widespread in the sunflower growing regions of the USA (Gulya et al. 1997). The wide geographic distribution, and high genetic variability of the pathogen lead to the evolution of new strains that could be more aggressive, causing large yield losses and a decline in disease control (Pecchia et al. 2004, Rekab et al. 2004).

Diaporthe cf. heveae 1

Specimen examined. Brazil, São Paulo, from Hevea brasiliensis, Apr. 1997, D.S. Attili (CBS 852.97) (originally identified as Phomopsis heveae).

Notes — Diaporthe heveae and Phomopsis heveae were both described from Hevea in Sri Lanka, and could represent the same species. Two isolates deposited in CBS under this name, CBS 852.97 (from Hevea brasiliensis in Brazil) and CBS 681.84 (from Hevea brasiliensis in India) were shown to represent two distinct species (clades 46 and 80, respectively). However, as both were found to be sterile, their taxonomy could not be resolved.

Diaporthe cf. heveae 2

Specimen examined. India, Kerala, Kottayam, in leaf on Hevea brasiliensis, Sept. 1984, K. Jayarathnam (CBS 681.84).

Notes — Isolate CBS 681.84 (clade 80, P. heveae from Hevea brasiliensis in India) is sterile, and thus its taxonomy could not be resolved. Diaporthe heveae has been reported from Brazil, China, India, Indonesia, Malaysia, Sri Lanka and Thailand (Holliday 1980, Zhuang 2001, Udayanga et al. 2011).

Diaporthe hickoriae Wehm., Monogr. Gen. Diaporthe Nitschke & Segreg., Univ. Michigan Stud., Sci. Ser. 9: 149. 1933

Specimen examined. USA, Michigan, on Carya glabra, June 1926, L.E. Wehmeyer (ex-type culture CBS 145.26).

Notes — Diaporthe hickoriae (clade 72) occurs on the bark of Carya glabra in the USA (Wehmeyer 1933).

Diaporthe hongkongensis R.R. Gomes, C. Glienke & Crous, sp. nov. — MycoBank MB802934; Fig. 10

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Fig. 10

Diaporthe hongkongensis (CBS 115448). a, b. Conidiomata sporulating on PDA; c, d. conidiogenous cells; e. beta conidia; f. alpha conidia. — Scale bars = 10 μm.

Etymology. Named after the location where it was collected, Hong Kong.

Conidiomata pycnidial, superficial to embedded on PDA, solitary to aggregated, globose with central ostiole, exuding a creamy conidial cirrhus; pycnidial up to 200 μm diam; wall of 3–6 layers of brown textura angularis. Conidiophores lining the inner cavity, reduced to conidiogenous cells. Conidiogenous cells phialidic, hyaline, smooth, ampulliform to subcylindrical with prominent apical taper, 5–12 × 2–4 μm; apex with periclinal thickening and minute collarette, 1 μm long. Paraphyses intermingled among conidiophores, hyaline, smooth, frequently branched below, up to 4-septate, with clavate terminal cell, up to 80 μm long, apex 2–8 μm diam. Alpha conidia hyaline, smooth, granular to guttulate, aseptate, fusiform, tapering towards both ends, mostly straight, apex acutely rounded, base truncate, (5–)6–7(–8) × (2–)2.5(–3) μm. Gamma conidia aseptate, hyaline, smooth, ellipsoid-fusoid, apex subobtuse, base truncate, 10–13 × 2 μm. Beta conidia aseptate, hyaline, smooth, spindle-shaped, apex acutely rounded, base truncate, widest in mid region, mostly curved in upper part, 18–22 × 1.5–2 μm.

Culture characteristics — Colonies covering dish after 2 wk in the dark at 25 °C, with moderate aerial mycelium. On OA surface dirty white with patches of pale olivaceous-grey, reverse dirty white with patches of olivaceous-grey and iron-grey. On PDA surface iron-grey, with patches of dirty white, reverse iron-grey. On MEA surface dirty white with patches of olivaceous-grey, reverse iron-grey with patches of dirty white.

Specimen examined. Hong Kong, Tai Po Kau, on fruit of Dichroa febrifuga, 20 Feb. 2002, K.D. Hyde (holotype CBS H-21103, culture ex-type CBS 115448 = HKUCC 9104).

Notes — Isolate CBS 115448 (clade 79; reported as Phomopsis pittospori on Dichroa febrifuga from Hong Kong) is morphologically distinct from P. pittospori (from Pittosporum twigs in California; alpha conidia 6–8 × 1.5 μm, beta conidia 18–20 × 1 μm), with wider alpha and beta conidia.

Diaporthe hordei (Punith.) R.R. Gomes, C. Glienke & Crous, comb. nov. — MycoBank MB802935

Basionym. Phomopsis hordei Punith., Trans. Brit. Mycol. Soc. 64, 3: 428. 1975.

Specimen examined. Norway, Fellesbygget, As, on root of Hordeum vulgare, Oct. 1992, L. Sundheim (CBS 481.92).

Notes — Diaporthe hordei (clade 13) was described from Hordeum vulgare in the UK. Although the present culture could be authentic (from Hordeum collected in Norway), it proved to be sterile, so its morphology could not be confirmed.

Diaporthe impulsa (Cooke & Peck) Sacc., Syll. Fung. (Abellini) 1: 618. 1882

Basionym. Valsa impulsa Cooke & Peck, Ann. Rep. N.Y. State Mus. Nat. Hist. 27: 109. 1875 (1874).

Specimens examined. Sweden, Uppland, Dalby par., Jerusalem, on Sorbus aucuparia, 24 Oct. 1989, K. & L. Holm (CBS 114434 = UPSC 3052). – Unknown, on Sorbus americana, Sept. 1927, L.E. Wehmeyer (CBS 141.27).

Notes — Clade 51 is represented by two isolates of D. impulsa occurring on Sorbus spp. Diaporthe impulsa is a known pathogen of Sorbus spp., and has a wide geographic distribution (Farr & Rossman 2012). It was originally described from Sorbus in the USA, thus CBS 141.27 may well prove to be a good reference strain for the species.

Diaporthe inconspicua R.R. Gomes, C. Glienke & Crous, sp. nov. — MycoBank MB802936

Etymology. Referring to its inconspicuous nature, growing as endophyte in host tissue.

Cultures sterile. Diaporthe inconspicua (clade 75) differs from its closest phylogenetic neighbours, clade 68–74 and 76–78, by unique fixed alleles in four loci based on alignments of the separate loci deposited in TreeBase as study S13943: TUB positions 33 (A), 102–104 and 106–111 (indels), 127 (G), 149 (C), 151 (A), 195 (C), 204 (T), 357 (G), 446 (G), 449 (C), 465 (T), 484 (T), 559 (A), 592 (A), 629 (T), 653 (T), 708 (C), 732 (C), 754 (A), 763 (C), 784 (A) and 787 (G); CAL positions 28 (C), 102 (G), 114 (T), 148 (T), 152 (T), 153 (A), 157 (C), 170 (G), 199 (C) and 281 (C); TEF1 positions 9 (T), 16 (A), 22 (A), 29 (G), 30 (G), 81 (C), 86 (C), 87 (A), 88 (A), 89 (T), 131 (A), 275 (A), 298 (C) and 315 (T); HIS positions 139 (T), 211 (T), 244 (T) and 408 (T).

Culture characteristics — Colonies covering the dish after 2 wk in the dark at 25 °C. On OA spreading, flat with sparse aerial mycelium, surface cream in centre, umber in outer region. On PDA surface and reverse cream to dirty white with sparse aerial mycelium. On MEA with sparse aerial mycelium, surface becoming folded, dirty white in centre, sienna in outer region, and luteous in reverse.

Specimens examined. Brazil, on petiole of Maytenus ilicifolia, July 2007, R.R. Gomes (holotype CBS H-21102, ex-type culture LGMF 930 = CPC 20306 = CBS 133813); same collection details (LGMF 931 = CPC 20307); on Spondias mombin, 2007, K. Rodriguez (LGMF 922 = CPC 20298).

Notes — Sterile endophytic isolates (clade 75) from medicinal plants in Brazil.

Diaporthe infecunda R.R. Gomes, C. Glienke & Crous, sp. nov. — MycoBank MB802937

Etymology. Named after its sterile growth in culture.

Cultures sterile. Diaporthe infecunda (clade 23) differs from its closest phylogenetic neighbours, clade 16–22, by unique fixed alleles in five loci based on alignments of the separate loci deposited in TreeBase as study S13943: ITS positions 108 (T), 279 (C), 292 (G), 359 (C) and 360 (G); TUB positions 11 (indel), 106 (G), 138 (T), 140 (A), 153 (G), 155 (T), 184 (A), 197 (G), 202 (C), 302 (A), 354 (A), 369–374 (indels), 398 (G), 407 (indel), 414 (C), 422 (T), 424 (G), 425 (C), 432 (G), 452 (C), 454 (C), 458 (C), 461 (G), 479 (T), 482 (T), 486 (C), 540 (T), 572 (C), 622 (A), 694 (T), 696 (T), 697 (G), 716 (C), 728 (C), 776 (G), 778 (G) and 796 (C); CAL positions 64 (T), 83 (T), 104 (G), 146 (C), 151 (C), 155 (G), 159 (C), 172 (C), 176 (T), 179 (A), 184 (G), 197 (T), 206 (T), 212 (C) and 221 (T); TEF1 positions 6 (A), 9 (G), 13 (G), 16 (C), 21 (A), 30 (G), 32 (indel), 39 (A), 40 (A), 41 (G), 42 (T), 43 (A), 79 (G), 83 (T), 90 (T), 92 (T), 96 (A), 97 (C), 106 (C), 116 (A), 120 (C), 123 (A), 127 (A), 132 (A), 135 (G), 173 (G), 255 (T), 284 (A), 294 (C), 299 (C) and 309 (A); HIS positions 173 (T), 196 (T), 197 (G), 199 (C/T), 221 (C), 222 (C), 230 (G), 263 (C), 264 (T), 268 (C), 273 (T) and 279 (C).

Culture characteristics — Colonies covering the dish after 2 wk in the dark at 25 °C. On PDA surface umber with patches of white, reverse chestnut. On MEA surface dirty white, reverse umber. On OA surface with patches of dirty white and umber.

Specimens examined. Brazil, on leaf of Schinus terebinthifolius, July 2007, J. Lima (holotype CBS H-21095, ex-type culture LGMF 906 = CPC 20282 = CBS 133812); additional isolates with same collection details (LGMF 908 = CPC 20284, LGMF 912 = CPC 20288, LGMF 917 = CPC 20293, LGMF 918 = CPC 20294, LGMF 920 = CPC 20296); in petiole of Maytenus ilicifolia, July 2007, R.R. Gomes (LGMF 933 = CPC 20309, LGMF 940 = CPC 20316).

Notes — Clade 23 represents endophytic isolates from leaves of medicinal plants growing in Brazil. It consists of eight isolates, two from Maytenus ilicifolia, and six from Schinus terebinthifolius.

Diaporthe juglandina (Fuckel) Nitschke, Pyrenomycetes Germanici 2: 281. 1870

Basionym. Aglaospora juglandina Fuckel, Fungi Rhenani Exsicc., suppl. 7 (no. 2101–2200): no. 2159. 1868.

Specimen examined. USA, Tennessee, Great Smoky Mts National Park, dead wood of Juglans sp., L. Vasilyeva (CBS 121004).

Notes — Diaporthe juglandina (clade 65) represents a European taxon described from Juglans. European collections are required to confirm whether this name can be applied to the clade.

Diaporthe longispora (Wehm.) R.R. Gomes, C. Glienke & Crous, comb. nov. — MycoBank MB802938

Basionym. Diaporthe strumella var. longispora Wehm., Mycologia 28, 1: 46. 1936.

Specimen examined. Canada, Ontario, Toronto, on Ribes sp., May 1936, L.E. Wehmeyer (ex-type culture CBS 194.36).

Notes — Clade 27 comprises the ex-type culture of D. strumella var. longispora isolated from Ribes sp., and forms a sister clade with D. sclerotioides (clade 28). Diaporthe strumella is found on woody limbs, especially of Ribes spp. in temperate North America and Europe (Farr & Rossman 2012). As D. strumella var. longispora is morphologically clearly a distinct species, we elevate this variety to species status.

Diaporthe lusitanicae A.J.L. Phillips & J.M. Santos, Fung. Diversity 34: 118. 2009

Specimen examined. Portugal, Lisbon, Oeiras, Estação Agronómica Nacional, stem of Foeniculum vulgare, 14 Aug. 2007, J.M. Santos (ex-type cultures CBS 123212 = Di-C001/5, CBS 123213 = Di-C001/3).

Notes — This species (clade 21) was described in 2009 on senescent stems of Foeniculum vulgare (wild fennel) in Portugal by Santos & Phillips (2009).

Diaporthe manihotia Punith., Kavaka 3: 29. 1976 (1975)

• = Phomopsis manihotis Swarup, L.S. Chauhan & Tripathi, Mycopathol. Mycol. Appl. 28, 4: 345. 1966.

Specimen examined. Rwanda, on leaves of Manihot utilissima, 9 July 1976, J. Semal (CBS 505.76).

Notes — Phomopsis manihotis (clade 25 as D. manihotia) causes leaf spot of cassava (Manihot esculenta), though the disease is also referred to as Phomopsis blight of tapioca. Severe infection leads to defoliation and stem lesions. Affected areas become shrivelled with numerous pycnidia embedded in the tissue. On severely infected stems the bark starts to gradually peel off, leading to partial or total girdling. The disease is known from Africa (Ethiopia, Nigeria), Asia (India), Central America and West Indies (S.E. Dominica), and South America (Colombia) (Sarbhoy et al. 1971, Mathur 1979, Farr & Rossman 2012).

Diaporthe mayteni R.R. Gomes, C. Glienke & Crous, sp. nov. — MycoBank MB802939; Fig. 11

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Fig. 11

Diaporthe mayteni (CBS 133185). a. Conidiomata sporulating on PNA; b, c. transverse section through conidiomata, showing conidiomatal wall; d, e. conidiogenous cells; f. beta conidia; g. alpha conidia. — Scale bars: b = 85 μm, all others = 10 μm.

Etymology. Named after the host genus from which it was collected, Maytenus.

Conidiomata pycnidial, globose, immersed, scattered and aggregated, brown to black, ostiolate, 70–230 μm wide, 40–150 μm tall, with short necks, 40–140 μm; outer surface smooth or covered in hyphae; pycnidal wall consisting of brown, thick-walled cells of textura angularis; conidial mass globose or exuding in cirrhi; predominantly yellow, pale luteous to cream. Conidiophores hyaline, subcylindrical to cylindrical, rarely branched above the septa, tapering towards the apex, 1–3-septate, (10–)13–27(–36) × (2–)3(–4) μm. Conidiogenous cells hyaline, subcylindrical, rarely tapering towards the apex, collarette present and not flared, with prominent periclinal thickening, (5–)6–10(–13) × 2(–3) μm. Alpha conidia hyaline, oblong to ellipsoid, apex bluntly rounded, base obtuse; biguttulate, (5–)6(–7) × (2–)3 μm. Beta and gamma conidia absent.

Culture characteristics — Colonies on PDA flat, with entire edge, cottony, olivaceous buff, with primrose aerial mycelium in concentric rings, with olivaceous patches; colonies reaching 66 mm diam after 2 wk at 25 °C; reverse olivaceous buff and greenish olivaceous. On OA flat, with entire edge, cottony appressed, buff, white, the center of the colony pale olivaceous-grey, patches isabelline and luteous; colonies reaching 56 mm diam; reverse buff and pale olivaceous grey. On MEA flat, with entire edge, aerial mycelium cottony, white to pale olivaceous grey or olivaceous buff; colonies reaching 37 mm diam; reverse hazel, ochreous, with patches greenish black and olivaceous black.

Specimen examined. Brazil, Paraná, Colombo, endophytic species isolated from petiole of Maytenus ilicifolia (popular name Espinheira Santa), July 2007, R.R. Gomes (holotype CBS H-21096, ex-type culture CBS 133185 = LGMF 938 = CPC 20314).

Notes — Diaporthe mayteni (clade 30) grows endophytically in Maytenus ilicifolia in Brazil.

Diaporthe megalospora Ellis & Everh., Proc. Acad. Nat. Sci. Philadelphia 42: 235. 1890

Specimen examined. Unknown, from Sambucus canadensis, Sept. 1927, L.E. Wehmeyer (CBS 143.27).

Notes — Diaporthe megalospora (clade 15) is known on Sambucus canadensis from North America (Wehmeyer 1933, Hanlin 1963, Farr & Rossman 2012). Fresh collections are required to designate an epitype, and fix the genetic application of the name.

Diaporthe melonis Beraha & M.J. O’Brien, Phytopathol. Z. 94, 3: 205. 1979

• = Phomopsis cucurbitae McKeen, Canad. J. Bot. 35: 46. 1957.

Specimens examined. Indonesia, Java, Muneng, Exp. Station, on Glycine soja, Sept. 1987, H. Vermeulen (CBS 435.87). – USA, Texas, Rio Grande Valley, on Cucumis melo, 1978, L. Beraha & M.J. O’Brien (ex-isotype culture CBS 507.78, specimen derived from culture CBS H-891).

Notes — Clade 7 represents D. melonis (Beraha & O’Brien 1979), and contains the ex-isotype culture, and one isolate previously identified as D. phaseolorum var. sojae (though the two isolates are not identical). Diaporthe melonis is frequently reported on soybean (Santos et al. 2011). Phomopsis cucurbitae (treated here as synonym) is reported to have a cosmopolitan distribution, and to cause black rot disease of greenhouse cucumbers (McKeen 1957, Punithalingam & Holliday 1975, Ohsawa & Kobayashi 1989).

Diaporthe musigena Crous & R.G. Shivas, Persoonia 26: 119. 2011

Specimen examined. Australia, Queensland, Brisbane Botanical Garden, on leaves of Musa sp., 14 July 2009, P.W. Crous & R.G. Shivas (ex-type culture CBS 129519 = CPC 17026).

Notes — Clade 84 represents D. musigena, isolated from Musa sp. in Australia (Crous et al. 2011).

Diaporthe neilliae Peck, Ann. Rep. N.Y. State Mus. Nat. Hist. 39: 52. 1887 (1886)

Specimen examined. Unknown, on Spiraea sp., Sep. 1927, L.E. Wehmeyer (CBS 144.27).

Notes — Diaporthe neilliae (clade 60) was originally described from Spiraea sp. from North America. The origin of the present isolate, however, remains unclear (presumably North America).

Diaporthe neoarctii R.R. Gomes, C. Glienke & Crous, sp. nov. — MycoBank MB802940, Fig. 12

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Fig. 12

Diaporthe neoarctii (CBS 109490). a. Conidiomata sporulating on PDA; b, c. transverse section through conidiomata, showing conidiomatal wall; d–f. conidiogenous cells; g. alpha conidia. — Scale bars: b = 225 μm, all others = 10 μm.

Etymology. Named after its superficial resemblance to Diaporthe arctii.

Conidiomata pycnidial, ampulliform to finger-like, aggregated, dark brown to black, immersed, ostiolate, 300–450 μm wide, 200–670 μm tall, with prominent necks 240–560 μm long, outer surface covered with hyphae; pycnidal wall consisting of brown, thick-walled cells of textura angularis; conidial mass globose, pale yellow. Conidiophores hyaline, ampulliform to subcylindrical, filiform, branched above the septa, tapering towards the apex, rarely septate, (12–)13–17(–18) × (2–)3 μm. Conidiogenous cells hyaline, subcylindrical, filiform, straight, tapering towards the apex, collarette flared, periclinal thickening prominent, (10–)11–13(–14) × (1.5–)2(–3) μm. Alpha conidia hyaline, fusoid, apex acute, base obtusely rounded to subtruncate, bi- to multi-guttulate, (9–)11–13(–14) × 3(–4) μm. Beta and gamma conidia not observed.

Culture characteristics — Colonies with sparse aerial mycelium, covering the dish after 2 wk in the dark at 25 °C. On MEA umber with patches of greyish sepia, umber in reverse. On PDA fuscous-black on surface and in reverse.

Specimen examined. USA, New Jersey, isolated from Ambrosia trifida, May 2001, G. Bills (holotype CBS H-21094, ex-type culture CBS 109490 = GB 6421 = AR 3450).

Notes — Isolates originally identified as D. arctii cluster in clades 19 and 67 (Fig. 1). Diaporthe neoarctii (clade 16) was isolated from Ambrosia trifida in New Jersey, USA, and differs morphologically from the ex-type culture of D. arctii (alpha conidia 7 × 3–3.5 μm) (clade 19). Based on these differences D. neoarctii is described as a novel species.

Phylogenetic species recognition by genealogical concordance

Taylor et al. (2000) developed the Genealogical Concordance Phylogenetic Species Recognition (GCPSR) concept to define the limits of sexual species, using the phylogenetic concordance of multiple unlinked genes. This concept has proved greatly useful in fungi, because it is more finely discriminating than other species concepts, as several species are unable to be crossed, or cannot be recognised due to the lack of distinguishing morphological characters or sterility (Reynolds 1993, Taylor et al. 2000, Cai et al. 2011). The adoption of genealogical concordance for species recognition in Diaporthe enabled us to distinguish species that were otherwise not possible to identify due to either sterility, or the loss of specific character states. For instance, D. viticola and D. australafricana are two closely related species (clades 50 and 49, respectively) associated with grapevines. They are morphologically similar, but occur on different continents (van Niekerk et al. 2005). These species have probably accumulated genetic differences due to their geographical isolation. Several cryptic species were recently described in other genera using the GCPSR criterion, some of which are consistent with allopatric divergence, because these species occupy non-overlapping areas separated by geographic barriers, e.g. in Cladosporium (Bensch et al. 2012), Colletotrichum (Damm et al. 2012a, b), Harknessia (Crous et al. 2012), Ilyonectria (Cabral et al. 2012a, b) and Phyllosticta (Glienke et al. 2011), to name but a few. Using the GCPSR concept it is possible to define the genetic variation observed in some species, but still insufficient to establish them as distinct species, since genetic flow still occurs between them. For example, isolates of clades 79–90 clustered differently based on analyses of the different genes, probably because of recent gene flow among them.

We have compared the location and monophyly of the strains in each clade in the phylogenetic tree of the combined alignment (Fig. 1) to those phylogenetic trees obtained from the individual loci to determine the species boundaries and species resolution. The five loci selected for the Bayesian phylogeny have a similar resolution for species discrimination, ranging from TEF1 resolving 72 out of the 95 species, to HIS and TUB resolving 84 of the 95 species.

The ITS region, which is often considered to be less than optimal for closely related species, was not much better or worse (resolving 75 of the 95 species) than the other included loci. However, given the recent acceptance of the ITS region as official fungal barcode (Schoch et al. 2012) and its intermediate resolving power in the present study, this locus should not be discarded from future studies. Also, TEF1, which has in the past been used as additional locus for phylogenetic studies of Diaporthe, performed the worst in this study (resolving 72 of the 95 species), although this was not much worse than ITS and CAL (resolving 75 and 74 of the 95 species, respectively). The HIS and TUB regions appear to have the best resolution for species discrimination in the present study and therefore are good candidates as secondary markers to the commonly used ITS region. Similar results were also reported for ITS, CAL, TEF1 and TUB by Udayanga et al. (2012), who suggested that TUB be considered as secondary phylogenetic marker for Diaporthe.

We thank the technical staff, Arien van Iperen (cultures), Marjan Vermaas (photographic plates) and Mieke Starink-Willemse (DNA isolation, amplification and sequencing) for their invaluable assistance. We are grateful to the Brazilian agency CAPES for financial support to Renata R. Gomes.