Phyllosticta

Pers., Traité sur les Champignons Comestibles (Paris): 55. 147. 1818. Fig. 46.

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Fig. 46. Phyllosticta spp. A−E. Disease symptoms. A. Aloe with dead leaf tips that harbour Phyllosticta aloeicola. B. Symptoms on Citrus maxima caused by Phyllosticta citrimaxima. C. Water-soaked lesions on banana fruit associated with freckle disease on banana caused by Phyllosticta sp. D. Symptomatic leaf of Cussonia sp. caused by Phyllosticta cussoniae. E. Symptoms on lemon leaf caused by Phyllosticta sp. F−H. Sexual morphs. F, G. Asci and ascospores of Phyllosticta abieticola (ex-type CBS 112067). H. Ascospores of Phyllosticta capitalensis (ex-epitype CBS 128856). I−U. Asexual morphs. I. Conidiomata sporulating on OA of Phyllosticta cussoniae (ex-epitype CPC 14873). J. Vertical section through conidioma of Phyllosticta rhaphiolepidis (ex-type MUCC 432). K. Conidiomatal wall of textura angularis of Phyllosticta rhaphiolepidis (ex-type MUCC 432). L. Conidioma with ostiole (arrowed) of Phyllosticta cordylinophila (ex-neotype CPC 20261). M, N. Conidiogenous cells giving rise to conidia. M. Phyllosticta foliorum (ex-neotype CBS 447.68). N. Phyllosticta capitalensis (ex-epitype CBS 128856). O−Q. Conidia. O. Phyllosticta aloeicola (CPC 20677). P. Phyllosticta podocarpicola (ex-type CBS 728.79). Q. Phyllosticta capitalensis (ex-epitype CBS 128856). R, S. Appressoria of Phyllosticta mangifera-indica (ex-type CPC 20274). T, U. Spermatia. T. Phyllosticta cussoniae (ex-epitype CPC 14873). U. Phyllosticta leucothoicola (ex-type MUCC 553). Scale bars: I = 25 μm; others = 10 μm. Pictures A, B, D, F–M, O, P, R–U taken from Wikee et al. (2013b); C from Wong et al. (2012); N, Q from Glienke et al. (2011).

Synonym: Guignardia Viala & Ravaz, Bull. Soc. mycol. Fr. 8: 63. 1892.

Classification: Dothideomycetes, Dothideomycetidae, Botryosphaeriales, Phyllostictaceae.

Type species: Phyllosticta convallariae Pers., nom. inval. (= Phyllosticta cruenta (Fr.) J. Kickx f.). Reference strain: CBS 858.71.

DNA barcode (genus): LSU.

DNA barcodes (species): ITS, act, gapdh, tef1. Table 15. Fig. 47.

Table 15. DNA barcodes of accepted Phyllosticta spp.

Species	Isolates1	GenBank accession number2				References
		ITS	act	gapdh	tef1
Phyllosticta abieticola	CBS 112067T	KF170306	KF289238	–	–	Wikee et al. (2013b)
Phy. alliacea	MUCC 0014T	AB454263	AB704207	–	–	Motohashi et al., 2009, Ando et al., 2013
Phy. aloeicola	CBS 136058T	KF154280	KF289311	KF289124	KF289193	Wikee et al. (2013b)
Phy. ampelicida	ATCC 200578NT	KC193586	KC193581	KC193584	–	Zhang et al. (2013b)
Phy. ardisiicola	MUCC 0031T	AB454274	AB704216	–	–	Motohashi et al., 2009, Ando et al., 2013
Phy. aristolochiicola	BRIP 53316aT	JX486129	–	–	–	Crous et al. (2012a)
Phy. aspidistricola	MUCC 0010T	AB454260	AB704204	–	–	Motohashi et al., 2009, Ando et al., 2013
Phy. beaumarisii	CBS 535.87T	AY042927	KF306232	KF289074	KF289170	Baayen et al., 2002, Wikee et al., 2013b
Phy. bifrenariae	CBS 128855T	JF343565	JF343649	JF343744	JF343586	Glienke et al. (2011)
Phy. brazillianiae	CBS 129060T	JF343572	JF343656	JF343758	JF343593	Glienke et al. (2011)
Phy. capitalensis	CBS 128856ET	JF261465	JF343647	JF343776	JF261507	Glienke et al. (2011)
Phy. carissicola	CPC 25665T	KT950849	KT950872	KT950876	KT950879	Crous et al. (2015e)
Phy. carochlae	CGMCC 3.17317T	KJ847422	KJ847430	KJ847438	KJ847444	Zhou et al. (2015)
Phy. catimbauensis	URM 7672T	MF466160	MF466157	–	MF466155	Crous et al. (2017b)
Phy. cavendishii	BRIP 55420IsoT	JQ743562	–	–	–	Wong et al. (2012)
Phy. citriasiana	CBS 120486T	FJ538360	FJ538476	JF343686	FJ538418	Wulandari et al., 2009, Glienke et al., 2011
Phy. citribraziliensis	CBS 100098T	FJ538352	FJ538468	JF343691	FJ538410	Wulandari et al., 2009, Glienke et al., 2011
Phy. citricarpa	CBS 127454ET	JF343583	JF343667	JF343771	JF343604	Glienke et al. (2011)
Phy. citrichinaensis	CBS 130529T	JN791597	JN791526	–	JN791452	Wang et al. (2011)
Phy. citrimaxima	CBS 136059T	KF170304	KF289300	KF289157	KF289222	Wikee et al. (2013b)
Phy. concentrica	CBS 937.70ET	FJ538350	KF289257	JF411745	FJ538408	Wulandari et al., 2009, Glienke et al., 2011, Wikee et al., 2013b
Phy. cordylinophila	CBS 136244NT	KF170287	KF289295	KF289076	KF289172	Wikee et al. (2013b)
Phy. cornicola	CBS 111639	KF170307	KF289234	–	–	Wikee et al. (2013b)
Phy. cruenta	CBS 858.71	MG934458	MG934465	MG934474	MG934501	Present study
Phy. cussonia	CBS 136060ET	JF343578	JF343662	JF343764	JF343599	Glienke et al. (2011)
Phy. elongata	CBS 126.22T	FJ538353	FJ538469	KF289164	FJ538411	Wulandari et al., 2009, Wikee et al., 2013b
Phy. ericarum	CBS 132534T	KF206170	KF289291	KF289162	KF289227	Wikee et al. (2013b)
Phy. eugeniae	CBS 445.82	AY042926	KF289246	KF289139	KF289208	Baayen et al., 2002, Wikee et al., 2013b
Phy. fallopiae	MUCC 0113T	AB454307	AB704228	–	–	Motohashi et al., 2009, Ando et al., 2013
Phy. foliorum	CBS 447.68NT	KF170309	KF289247	KF289132	KF289201	Wikee et al. (2013b)
Phy. gaultheriae	CBS 447.70T	JN692543	KF289248	JN692508	JN692531	Su & Cai (2012)
Phy. hamamelidis	MUCC 149	KF170289	KF289309	–	–	Wikee et al. (2013b)
Phy. hostae	CGMCC 3.14355T	JN692535	JN692511	JN692503	JN692523	Su & Cai (2012)
Phy. hubeiensis	CGMCC 3.14986T	JX025037	JX025032	JX025027	JX025042	Zhang et al. (2013a)
Phy. hymenocallidicola	CBS 131309T	JQ044423	KF289242	KF289142	KF289211	Crous et al., 2011b, Wikee et al., 2013b
Phy. hypoglossi	CBS 434.92NT	FJ538367	FJ538483	JF343695	FJ538425	Wulandari et al., 2009, Glienke et al., 2011, Wikee et al., 2013b
Phy. ilicis-aquifolii	CGMCC 3.14358T	JN692538	JN692514	–	JN692526	Su & Cai (2012)
Phy. iridigena	CBS 143410T	MG934459	MG934466	–	MG934502	Present study
Phy. kerriae	MUCC 0017T	AB454266	AB704209	–	KC342576	Motohashi et al., 2009, Ando et al., 2013, Wikee et al., 2013a
Phy. leucothoicola	CBS 136073T	AB454370	KF289310	–	–	Motohashi et al., 2009, Wikee et al., 2013b
Phy. ligustricola	MUCC 0024T	AB454269	AB704212	–	–	Motohashi et al., 2009, Ando et al., 2013
Phy. maculata	CBS 132581ET	JQ743570	–	–	–	Wong et al. (2012)
Phy. mangiferae-indicae	CBS 136061T	KF170305	KF289296	KF289121	KF289190	Wikee et al. (2013b)
Phy. mimusopisicola	CBS 138899T	KP004447	–	–	–	Crous et al. (2014d)
Phy. minima	CBS 585.84NT	KF206176	KF289249	KF289135	KF289204	Wikee et al. (2013b)
Phy. musarum	BRIP 55434IsoET	JQ743584	–	–	–	Wong et al. (2012)
Phy. neopyrolae	CBS 134750T	AB454318	AB704233	–	–	Motohashi et al., 2009, Ando et al., 2013
Phy. owaniana	CBS 776.97ET	FJ538368	KF289254	JF343767	FJ538426	Wulandari et al., 2009, Glienke et al., 2011, Wikee et al., 2013b
Phy. pachysandricola	MUCC 124T	AB454317	AB704232	–	–	Motohashi et al., 2009, Ando et al., 2013
Phy. paracapitalensis	CBS 141353T	KY855622	KY855677	KY855735	KY855951	Guarnaccia et al. (2017)
Phy. paracitricarpa	CBS 141357T	KY855635	KY855690	KY855748	KY855964	Guarnaccia et al. (2017)
Phy. parthenocisii	CBS 111645	JN692542	JN692518	–	JN692530	Su & Cai (2012)
Phy. partricuspidatae	NBRC 9466T	KJ847424	KJ847432	KJ847440	KJ847446	Zhou et al. (2015)
Phy. paxistimae	CBS 112527T	KF206172	KF289239	KF289140	KF289209	Wikee et al. (2013b)
Phy. persooniae	CBS 143409T	MG934460	MG934467	MG934475	MG934503	Present study
Phy. philoprina	CBS 587.69	KF154278	KF289250	KF289137	KF289206	Wikee et al. (2013b)
Phy. podocarpi	CBS 111646	AF312013	KC357670	KF289169	KC357671	Wikee et al. (2013b), Carroll (unpubl. data), Wikee (unpubl. data)
Phy. podocarpicola	CBS 728.79T	KF206173	KF289252	KF289134	KF289203	Wikee et al. (2013b)
Phy. pseudotsugae	CBS 111649	KF154277	KF289236	KF289167	KF289231	Wikee et al. (2013b)
Phy. rhaphiolepidis	MUCC 432T	DQ632660	AB704242	–	DQ632724	Andjic et al., 2007, Ando et al., 2013
Phy. rubella	CBS 111635T	KF206171	KF289233	KF289129	KF289198	Wikee et al. (2013b)
Phy. schimae	CGMCC 3.14354T	JN692534	JN692510	JN692506	JN692522	Su & Cai (2012)
Phy. schimicola	CGMCC 3.17319T	KJ847426	KJ847434	KJ854895	KJ847448	Zhou et al. (2015)
Phy. sphaeropsoidea	CBS 756.70	AY042934	KF289253	KF289133	KF289202	Baayen et al., 2002, Wikee et al., 2013b
Phy. spinarum	CBS 292.90	JF343585	JF343669	JF343773	JF343606	Glienke et al. (2011)
Phy. styracicola	CGMCC 3.14985T	JX052040	JX025035	JX025030	JX025045	Zhang et al. (2013a)
Phy. telopeae	CBS 777.97T	KF206205	KF289255	KF289141	KF289210	Wikee et al. (2013b)
Phy. vaccinii	ATCC 46255ET	KC193585	KC193580	KC193583	KC193582	Zhang et al. (2013b)
Phy. vacciniicola	CBS 136062T	KF170312	KF289287	KF289165	KF289229	Wikee et al. (2013b)
Phy. vitis-rotundifoliae	CGMCC 3.17322T	KJ847428	KJ847436	KJ847442	KJ847450	Zhou et al. (2015)
Phy. yuccae	CBS 117136	JN692541	JN692517	JN692507	JN692529	Su & Cai (2012)

1

ATCC: American Type Culture Collection, Virginia, USA; BRIP: Queensland Plant Pathology Herbarium, Brisbane, Australia; CBS: Westerdijk Fungal Biodiversity Institute, Utrecht, The Netherlands; CGMCC: Chinese General Microbiological Culture Collection Center, Beijing, China; CPC: Culture collection of Pedro Crous, housed at Westerdijk Fungal Biodiversity Institute; MUCC: Murdoch University, Perth, Western Australia; NBRC: Biological Resource Center, NITE, Chiva, Japan; URM: Culture Collection Mycobank, Prof. Maria Auxiliadora Cavalcanti, Federal University of Pernambuco, Recife, Brazil. ^{T, ET, IsoT, IsoET} and ^NT indicate ex-type, ex-epitype, ex-isotype, ex-isoepitype and ex-neotype strains, respectively.

2

ITS: internal transcribed spacers and intervening 5.8S nrDNA; act: partial actin gene; gapdh: partial glyceraldehyde-3-phosphate dehydrogenase gene; tef1: partial translation elongation factor 1-alpha gene.

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Fig. 47. RAxML phylogram obtained from the combined ITS (492 bp), act (291 bp), gapdh (629 bp) and tef1 (341 bp) sequence alignment of all the accepted species of Phyllosticta. The tree was rooted to Peyronellaea obtusa CMW8232. The novelties proposed in this study are indicated in bold. RAxML bootstrap support (BS) values above 70 % and Bayesian posterior probability scores above 0.95 are shown at the nodes. GenBank accession numbers are indicated in Table 15. ^{T, ET, IsoT, IsoET} and ^NT indicate ex-type, ex-epitype, ex-isotype, ex-isoepitype and ex-neotype strains, respectively. TreeBASE: S21899.

Ascomata pseudothecial, separate to gregarious, globose to subglobose, brown to black, unilocular with a central ostiole. Pseudoparaphyses mostly absent at maturity, filamentous, branched, septate when present. Asci bitunicate, fissitunicate, clavate to subcylindrical, 8-spored, fasciculate, stipitate, with an ocular chamber. Ascospores bi- to triseriate, hyaline, guttulate to granular, aseptate, ellipsoid, ellipsoid-fusoid to limoniform, smooth-walled, usually with mucilaginous caps at ends, or surrounded by a mucilaginous sheath. Conidiomata and spermatogonia pycnidial, immersed, subepidermal to erumpent, unilocular, rarely multilocular, glabrous, ostiolate, dark brown to black; ostiole circular to oval; conidiomatal wall thick-walled, dark brown, textura angularis, with inner layers of hyaline to pale brown, thin-walled, textura prismatica to angularis. Conidiophores lining cavity of conidioma, reduced to conidiogenous cells, invested in mucus. Conidiogenous cells discrete, producing macroconidia and spermatia, also produced in separate spermatogonia, ampulliform, lageniform, doliiform to subcylindrical, hyaline, smooth, proliferating percurrently near apex, invested in a mucoid layer. Conidia ellipsoid-fusoid to obovoid or ovoid, rarely subcylindrical, aseptate, broadly rounded at apex, often tapering strongly toward base, unicellular, hyaline, smooth-walled, guttulate to granular, often enclosed in a persistent mucilaginous sheath, and bearing an unbranched, tapering, straight to curved, mucoid apical appendage. Spermatogenous cells ampulliform to lageniform or subcylindrical, hyaline, smooth, phialidic. Spermatia hyaline, smooth, granular, subcylindrical or dumbbell-shaped, with rounded or blunt ends (adapted from Wikee et al. 2013b).

Culture characteristics: Colonies on MEA, OA and PDA after 2 wk in dark at 27 °C erumpent or flat, spreading with sparse or moderate aerial mycelium; on MEA, OA and PDA surface frequently iron-grey or olivaceous grey, less frequently greenish to dark green; reverse iron-grey, olivaceous grey or black.

Optimal media and cultivation conditions: PNA, OA, PDA and SNA under near-ultraviolet light at 27 °C to induce sporulation.

Distribution: Worldwide.

Hosts: Wide range of hosts from trees to ornamentals.

Disease symptoms: Leaf spots and various fruit diseases.

Notes: Phyllosticta was introduced by Persoon (1818), with Phy. convallariae designated as type species (Donk 1968). However, this species was invalid because it lacked a description. Therefore, Phy. cruenta, which is a synonym of Phy. convallariae, was designated as type of the genus (van der Aa & Vanev 2002). There is no available type material for this species, which was described from Polygonatum multiflorum collected in Germany. A strain deposited in CBS previously identified as Guignardia reticulata, which is the sexual morph of Phy. cruenta, was isolated from Polygonatum odoratum in the Czech Republic, being a potential neotype for Phy. cruenta. However, this strain is sterile and we have chosen to consider it as a reference strain since we could not confirm its identification based on morphology.

 Phyllosticta includes plant pathogenic species that cause diseases of significant economic importance. For example, Phy. citricarpa is the responsible for citrus black spot, which is considered a quarantine pest in Europe and the USA (Baayen et al., 2002, Glienke et al., 2011, Guarnaccia et al., 2017). Other examples include the Phy. ampelicida species complex that causes black rot disease on grapevines (Wicht et al., 2012, Carstens et al., 2017), and the Phy. musarum species complex that is responsible for banana freckle disease (Pu et al., 2008, Wong et al., 2012).

 Phoma and Phyllosticta have been difficult to distinguish since both genera were recognised as pycnidial fungi producing unicellular, hyaline conidia. Subsequent molecular data enabled the discrimination of both genera, as well as the fact that Phyllosticta was linked to its sexual morph, Guignardia (Glienke et al., 2011, Wikee et al., 2011, Wikee et al., 2013b, Wong et al., 2012, Zhou et al., 2015, Guarnaccia et al., 2017).

 Phyllosticta was formerly placed in the Botryosphaeriaceae, together with Botryosphaeria (Schoch et al. 2006). However, Wikee et al. (2013b) showed that it represents a different phylogenetic lineage, for which the family name Phyllostictaceae (Fries 1849) was resurrected.

References: van der Aa 1973 (morphology and pathogenicity); van der Aa & Vanev 2002 (synonyms, collection information and notes); Glienke et al., 2011, Wong et al., 2012, Wikee et al., 2013b, Zhou et al., 2015, Guarnaccia et al., 2017 (ecology, morphology and phylogeny); Wikee et al. 2011 (pathogenicity and phylogeny).

Phyllosticta iridigena

Y. Marín & Crous, sp. nov. MycoBank MB823971. Fig. 48.

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Fig. 48. Phyllosticta iridigena (ex-type CBS 143410). A. Conidiomata sporulating on SNA. B–E. Conidiogenous cells giving rise to conidia. F. Conidia. Scale bars: A = 200 μm, B–F = 10 μm.

Etymology: Name reflects the host it was isolated from, Iris.

Conidiomata 90–200 μm diam, pycnidial, solitary, globose, dark brown, with central ostiole; conidiomatal wall of 3–8 layers of brown textura angularis. Conidiophores lining cavity, reduced to conidiogenous cells. Conidiogenous cells 4–7 × 4–6 μm, doliiform, hyaline, smooth, proliferating percurrently at apex. Conidia (10–)12–13(–15) × (7–)8(–9) μm, solitary, ellipsoid to obovoid, aseptate, smooth, hyaline, guttulate, granular; conidia encased in a mucoid sheath 2–3 μm diam, and a single apical mucoid appendage, 7–15 × 2 μm, tapering to acutely rounded apex.

Culture characteristics: Colonies flat, spreading, with moderate aerial mycelium and smooth, feathery margins, reaching 45 mm diam after 2 wk at 25 °C. On MEA surface pale olivaceous grey; reverse iron-grey. On PDA surface and reverse olivaceous grey. On OA surface pale olivaceous grey with diffuse yellow pigment in agar.

Material examined: South Africa, on Iris sp. (Iridaceae), 16 Jan. 2010, P.W. Crous (holotype CBS H-23385, culture ex-type CBS 143410 = CPC 32669).

Notes: This species clusters in a well-supported clade (95 % BS / 1 PP) with Phy. hypoglossi and Phy. cussoniae. Phyllosticta hypoglossi produces longer conidiogenous cells (10–15 μm) and wider conidia [(9–)10(–11) μm] than Phy. iridigena. Moreover, these three species are isolated from different hosts, i.e. Phy. hypoglossi from Ruscus (Ruscaceae), Phy. cussoniae from Cussonia (Araliaceae) and Phy. iridigena from Iris (Iridaceae). Phyllosticta cussonia and Phy. iridigena have been found in the same country, South Africa, while Phy. hypoglossi is an European species.

Phyllosticta persooniae

Y. Marín & Crous, sp. nov. MycoBank MB823972. Fig. 49.

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Fig. 49. Phyllosticta persooniae (ex-type CBS 143409). A. Conidiomata sporulating on OA. B, C. Conidiogenous cells giving rise to conidia. D. Conidia. Scale bars: A = 200 μm, B–D = 10 μm.

Etymology: Name reflects the host genus Persoonia from which it was isolated.

Conidiomata 200–300 μm diam, pycnidial, solitary, globose, dark brown, with central ostiole; conidiomatal wall of 3–8 layers of brown textura angularis. Conidiophores 10–18 × 6–7 μm, lining cavity, 0–1-septate, subcylindrical, hyaline, smooth, rarely branched. Conidiogenous cells 9–17 × 4–5 μm, terminal, subcylindrical, hyaline, smooth, proliferating percurrently at apex. Conidia (9–)10–11(–12) × (7–)8(–9) μm, solitary, ellipsoid to obovoid, aseptate, smooth, hyaline, guttulate, granular; conidia encased in a mucoid sheath that is inconspicuous and dissolves at maturity, but with a single apical mucoid appendage, 7–15 × 2–3 μm, tapering to acutely rounded apex.

Culture characteristics: Colonies flat to erumpent, spreading, with sparse to moderate aerial mycelium and feathery, lobate margins, reaching 30 mm diam after 2 wk at 25 °C. On MEA surface smoke grey; reverse olivaceous grey. On PDA surface and reverse olivaceous grey. On OA surface pale mouse grey.

Material examined: Australia, New South Wales, South East Forests National Park, Nunnock Swamp, on Persoonia sp. (Proteaceae), 28 Nov. 2016, P.W. Crous (holotype CBS H-23386, culture ex-type CBS 143409 = CPC 32603).

Notes: Phyllosticta persooniae is phylogenetically distant from all other species of Phyllosticta, the most closely related species being Phy. foliorum. Morphologically, these can be distinguished by the size of their conidia. Phyllostica foliorum is characterised by its larger conidia i.e. (12–)13–14(–15) × (9–)10(–11) ìm vs. (9–)10–11(–12) × (7–)8(–9) μm in Phy. persooniae. Moreover, Phy. foliorum has never been found on Persoonia (Proteaceae) or in Australia, which is the host and distribution of Phy. persooniae (Farr & Rossman 2017). Most species of Phyllosticta are host-specific.

Authors: Y. Marin-Felix & P.W. Crous