Diaporthe (=Phomopsis) is a cosmopolitan genus of fungi comprised of endophytes, plant pathogens, and saprobes occurring on a wide range of annual and perennial hosts, including economically important crops (Uecker 1988; Farr and Rossman 2014; Udayanga et al. 2011). The genus belongs to class Sordariomycetes, order Diaporthales, and the family Diaporthaceae, typified by the species Diaporthe eres Nitschke (Wehmeyer 1933). With the change to one scientific name for fungi (McNeill et al. 2012), Diaporthe has priority, being the older generic name compared to Phomopsis. Many species are able to colonize diverse hosts as opportunists; some species are host specific and multiple species can even co-occur on the same host (Mostert et al. 2001; Farr et al. 2002a; Crous and Groenewald 2005). Species of Diaporthe cause cankers, diebacks, root rots, fruit rots, leaf spots, blights and wilts on a wide range of plant hosts including some economically important hosts and have been the subject of considerable phytopathological research. Examples of diseases on major crops include Diaporthe/Phomopsis complex causing soybean seed decay, pod and stem blight and cankers, sunflower stem canker (D. helianthi), dead arm of grapevines (D. ampelina) and melanosis in Citrus (D. citri) (Van Niekerk et al. 2005; Santos et al. 2011; Thompson et al. 2011; Udayanga et al. 2014a, b). In addition, several species of Diaporthe are known from clinical reports of immunocompromised patients, although these pathogens are only provisionally identified to species level (Garcia-Reyne et al. 2011; Mattei et al. 2013). Diaporthe comprises a major component of endophytes in tropical and temperate trees, and several species have been used in secondary metabolite research (Isaka et al. 2001; Li et al. 2010a, b; Kaul et al. 2012).


Species identification and numbers


The Genealogical Concordance Phylogenetic Species Recognition (GCPSR) has been applied in the genus Diaporthe to define the species boundaries in recent studies (Udayanga et al. 2012b; Gomes et al. 2013; Tan et al. 2012). Therefore species delimitation is currently based on DNA sequence data and comparison of morphological characters (Santos and Phillips 2009; Santos et al. 2010; Diogo et al. 2010; Udayanga et al. 2014a, b). Although the genus Diaporthe has received much attention, few phylogenetic studies have thus far been conducted; hence the taxonomy of some of the species in this genus is still uncertain including many of the common plant pathogens. Index Fungorum lists 892 Diaporthe names and 983 Phomopsis names whereas MycoBank (2014) lists 919 Diaporthe names and 1,040 Phomopsis names. However, the names available in the literature are mostly applied based on host association and morphology except fewer species described in the last two decades based on DNA sequence data. Ex-type cultures are available for less than 100 species known despite a large number of species listed in databases and literature. The delimitation of species within the genus Diaporthe improved once DNA sequence data were incorporated (Castlebury and Mengistu 2006; Van Rensburg et al. 2006; Santos et al. 2010; Udayanga et al. 2012b, 2014a, b) since this facilitates obtaining detailed insight into complex evolutionary relationships.


Molecular phylogeny

Since the first molecular phylogenetic study in Diaporthe (Rehner and Uecker 1994), rDNA ITS, partial sequences  of translation elongation factor 1-α (TEF) and  mating  type genes (MAT 1-1-1/1-2-1) have commonly been used in molecular phylogenetic studies in this genus (Van Niekerk et al. 2005;  Van  Rensburg  et  al.  2006;  Santos et al. 2010; Udayanga et al. 2011; Sun et al. 2012). Udayanga et al. (2012a) used ITS, TEF, β- tubulin and CAL genes with a selected set of ex-type cultures and additional isolates to infer the phylogeny of the genus. In a parallel study, a multi-marker phylogeny was effectively used to describe novel species in Diaporthe based on fresh collections from Thailand (Udayanga et al. 2012b). Gomes et al. (2013) used a Brazilian collection of isolates and existing ex-type cultures for a combined phylogenetic analysis of five genetic markers which included ITS, TEF, β- tubulin, CAL and HIS. They introduced several novel taxa from Brazilian collections from medicinal plants with one epitype for Diaporthe anarcardi from Kenya. Udayanga et al. (2014a, b) revisited the Diaporthe species associated with Citrus worldwide with a comprehensive assessment of the genes including ITS, TEF, β- tubulin, CAL and ACT. The study revisited several important phytopathogens including  D.  citri,  D.  cytosporella,  D.  forniculina  and D. rudis, with the epitypes designated with modern The clarification of  D. foeniculaina and D. rudis revealed the potential extensive host association of some species.

Udayanga et al. (2014a) further emphasized that ITS alone can cause much confusion in defining closely related taxa, which has also been noted by several previous researchers regarding closely related species in Diaporthe (Farr et al. 2002a, b; Murali et al. 2006; Santos et al. 2010). The variation of ITS sequences can result in superfluous, multiple terminal branches in combined analyses, even when other gene regions do not support these distinctions (Udayanga et al. 2014a, b). The TEF gene is informative when it comes to clarifying species limits in Diaporthe (Table, Fig.).


Table  Diaporthe. Details of the isolates used in the phylogenetic tree

Species Isolate Host GeneBank accession numbers  
      ITS β-tubulin TEF 1-α CAL
Diaporthe acaciigena CBS 129521* Acacia retinodes KC343005 KC343973 KC343731 KC343247
D. alleghaniensis CBS 495.72* Betula alleghaniensis KC343007 KC343975 KC343733 KC343249
D. alnea CBS 146.46* Alnus sp. KC343008 KC343976 KC343734 KC343250
D. ambigua CBS 114015* Pyrus communis KC343010 KC343978 KC343736 KC343252
D. ampelina CBS 114016* Vitis vinifera AF230751 JX275452 AY745056 AY230751
D. amygdali CBS 126679* Prunus dulcis KC343022 KC343990 AY343748 KC343264
D. anacardii CBS 720.97* Anacardium ocidentale KC343024 KC343992 KC343750 KC343266
D. angelicae CBS 111592* Heracleum sphondylium KC343027 KC343995 KC343753 KC343269
D. aquatica IFRDCC 3051* JQ797437
D. arecae CBS 161.64* Areca catechu KC343032 KC344000 KC343758 KC343274
D. arengae CBS 114979* Arenga engleri KC343034 KC344002 KC343760 KC343276
D. aspalathi CBS 117169* Aspalathus linearis KC343036 KC344004 KC343762 KC343278
D. australafricana CBS 111886* Vitis vinifera KC343038 KC344006 KC343764 KC343280
D. beilharziae BRIP 54792* Indigofera australis JX862529 KF170921 JX862535
D. bicincta CBS 121004* Juglans sp. KC343134 KC344102 KC343860 KC343376
D. brasiliensis CBS 133183* Aspidosperma tomentosum KC343042 KC344010 KC343768 KC343284
D. caulivora CBS 127268* Glycine max KC343045 KC344013 KC343771 KC343287
D. celastrina CBS 139.27* Celastrus sp KC343047 KC344015 KC343773 KC343289
D. citri CBS 135422* Citrus sp. KC843311 KC843187 KC843071 KC843157
D. citriasiana ZJUD 30* Citrus sp. JQ954645 KC357459 JQ954663 KC357491
D. citrichinensis ZJUD 34* Citrus sp. JQ954648   JQ954666 KC357494
D. crotalariae CBS 162.33* Crotalaria spectabilis KC343056 KC344024 KC343782 KC343298
D. cuppatea CBS 117499* Aspalathus linearis KC343057 KC344025 KC343783 KC343299
D. cynaroidis CBS 122676* Protea cynaroides KC343058 KC344026 KC343784 KC343300
D. cytosporella FAU461* Citrus limon KC843307 KC843221 KC843116 KC843141
D. endophytica CBS 133811* Schinus terebinthifolius KC343065 KC343065 KC343791 KC343307
D. eres AR5193* Ulmus Sp. KJ210529 KJ420799 KJ210550 KJ434999
P. cotoneastri CBS 439.82* Cotoneaster sp. KC343090 KC344058 KC343816 KC343332
D. fraxini-angustifoliae BRIP 54781* Fraxinus angustifolia JX862528 KF170920 JX862534
D. foeniculina CBS 123208* Foeniculum vulgare KC343104 KC344072 KC343830 KC343346
D. foeniculina CBS 123209* Foeniculum vulgare KC343105 KC344073 KC343831 KC343347
D. foeniculina CBS 187.27 * Camellia sinensis KC343107 KC344075 KC343833 KC343349
D. ganjae CBS 180.91* Cannabis sativa KC343112 KC344080 KC343838 KC343354
D. gulyae BRIP 54025* Helianthus annuus JF431299 JN645803
D. helianthi CBS 592.81* Helianthus annuus KC343115 KC344083 KC343841 KC343357
D. helicis AR5211* Hedera helix KJ210538 KJ420828 KJ210559 KJ435043
D. hickoriae CBS 145.26* Carya glabra KC343118 KC344086 KC343844 KC343360
D. hongkongensis CBS 115448* Dichroa febrífuga KC343119 KC344087 KC343845 KC343361
D. inconspicua CBS 133813* Maytenus ilicifolia KC343123 KC344091 KC343849 KC343365
D. infecunda CBS 133812* Schinus terebinthifolius KC343126 KC344094 KC343852 KC343852
D. kochmanii BRIP 54033* Helianthus annuus JF431295 JN645809
D. kongii BRIP 54031* Helianthus annuus JF431301 JN645797
D. longispora CBS 194.36* Ribes sp. KC343135 KC344103 KC343861 KC343377
D. lusitanicae CBS 123212* Foeniculum vulgare KC343136 KC344104 KC343862 KC343378
D. mayteni CBS 133185* Maytenus ilicifolia KC343139 KC344107 KC343865 KC343381
D. melonis CBS 507.78 * Glycine soja KC343141 KC344109 KC343867 KC343383
D. musigena CBS 129519* Musa sp. KC343143 KC344111 KC343869 KC343385
D. neoarctii CBS 109490* Ambrosia trifida KC343145 KC344113 KC343871 KC343387
D. nothofagi BRIP 54801* Nothofagus cunninghamii JX862530 KF170922 JX862536
D. novem CBS 127270* Glycine max KC343155 KC344123 KC343881 KC343397
D. oxe CBS 133186* Maytenus ilicifolia KC343164 KC344132 KC343890 KC343406
D. paranensis CBS 133184* Maytenus ilicifolia KC343171 KC344139 KC343897 KC343413
D. pascoei BRIP 54847* Persea americana JX862532 KF170924 JX862538
D. perjuncta CBS 109745* Ulmus glabra KC343172 KC344140 KC343898 KC343414
D. pseudomangiferae CBS 101339* Mangifera indica KC343181 KC344149 KC343907 KC343423
D. pseudophoenicicola CBS 462.69* Mangifera indica KC343183 KC344151 KC343909 KC343425
D. psoraleae CBS 136412* Psoralea pinnata KF777158 KF777251 KF777245
D. psoraleae-pinnatae CBS 136413 Psoralea pinnata KF777159 KF777252
D. pterocarpi MFLUCC 10-0571* Pterocarpus indicus JQ619899 JX275460 JX275416 JX197451
D. pterocarpicola MFLUCC 10-0580* Pterocarpus indicus JQ619887 JX275441 JX275403 JX197433
D. pulla CBS 338.89* Hedera helix KC343152 KC344120 KC343878 KC343394
D. raonikayaporum CBS 133182* Spondias mombin KC343188 KC344156 KC343914 KC343430
D. rudis CBS 109291* Laburnum anagyroides KC843331 KC843177 KC843090 KC843146
D. rudis CBS 113201* Vitis vinifera KC343234 KC344202 KC343960 KC343476
D. saccarata CBS 116311* Protea repens KC343190 KC344158 KC343916 KC343432
D. salicicola BRIP 54825* Salix purpurea JX862531 JX862531 JX862537
D. schini CBS 133181* Schinus terebinthifolius KC343191 KC344159 KC343917 KC343433
D. sclerotioides CBS 296.67* Cucumis sativus KC343193 KC344161 KC343919 KC343435
D. siamensis MFLUCC 10-0573a* Dasymaschalon sp. JQ619879 JX275429 JX275393
D. terebinthifolii CBS 133180* Schinus terebinthifolius KC343216 KC344184 KC343942 KC343458
D. thunbergii MFLUCC 10-0576* Thunbergia grandifolia JQ619893 JX275449 JX275409 JX197440
D. toxica CBS 534.93* Lupinus angustifolius KC343220 KC344188 KC343946 KC343462
Diaporthella corylina CBS 121124* Corylus sp. KC343004 KC343972 KC343730 KC343246
P. lithocarpus CGMCC 3.15175* Lithocarpus glabra KC153104 KC153095
P. mahothocarpus CGMCC 3.15181* Lithocarpus glabra KC153096 KC153087
P. ternstroemia CGMCC 3.15183* Ternstroemia gymnanthera KC153098 KC153089


Fig.  Phylogram generated from parsimony analysis based on„ combined ITS, EF1-α, β- tubulin, and CAL sequenced data of Diaporthe. Parsimony bootstrap support values and Bayesian posterior probabilities greater than 50 % are indicated above the nodes. The ex-type (ex-epitype) and voucher strains are in bold. The tree is rooted with Diaporthella corylina CBS 121124.



ITS and TEF are recommended for preliminary identification of the species (Castlebury et al. 2001; Castlebury 2005; Santos and Phillips 2009; Santos et al. 2010). ITS, TEF, β- tubulin, CAL, HIS and ACT should be used in the combined analysis (selection of 4–5 genes), with recommended primers in relevant publications (Udayanga et al. 2012b, 2014a, b; Gomes et al. 2013).

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