The endophyte Allantophomopsis cytisporea is associated with snow blight on Calluna vulgaris in the Alps—An effect of climate change?

Figures & data

Table 1. Sampling sites, collection dates and plant species collected.

Geographic origin	Coordinates^a	Altitude (m.a.s.l.)	Hosts examined^b	Collection date	Collector
Arosa, Sandböden	46°48ʹ06.6ʺ N 9°40ʹ23.8ʺ E	2,050	Juniperus communis subsp. nana, Picea abies, Pinus mugo subsp. mugo	28 June 2006	M. Schneider
Arosa, Maraner Hauptji	46°47ʹ48.9ʺ N 9°40ʹ15.8ʺ E	2,150	Calluna vulgaris	5 June 2015	O. Holdenrieder
Davos, Dischma	46°45ʹ12.4ʺ N 9°54ʹ5.9ʺ E	1,840	Calluna vulgaris, Erica carnea	3 June 2015	N. Luschka
Davos, Sertig	46°43ʹ28.5ʺ N 9°51ʹ3.4ʺ E	1,870	Calluna vulgaris	14 May 2015	N. Luschka
Davos, Kühalptal	46°42ʹ44.3ʺ N 9°51ʹ37.8ʺ E	1,980	Juniperus communis subsp. nana, Erica carnea	14 May 2015	N. Luschka
Davos, Rinerhorn	46°45ʹ5.0ʺ N 9°49ʹ7.5ʺ E	2,080	Erica carnea	6 June 2015	N. Luschka
Davos, Geisswaldji	46°49ʹ59.9ʺ N 9°52ʹ42.3ʺ E	1,800	Calluna vulgaris	24 May 2015	N. Luschka
Göscheneralp	46°40ʹ9.8ʺ N 8°32ʹ38.4ʺ E	1,820	Juniperus communis subsp. nana, Picea abies	3 August 2012	A. Gross
Lötschental	46°27ʹ14.3ʺ N 7°54ʹ34.7ʺ E	2,328	Juniperus communis subsp. nana, Picea abies, Calluna vulgaris	26 July 2012	O. Holdenrieder
San Bernardino	46°28ʹ49.2ʺ N 9°10ʹ31.5ʺ E	1,930	Calluna vulgaris	3 April 2002	T. N. Sieber
San Bernardino	46°28ʹ49.2ʺ N 9°10ʹ31.5ʺ E	1,930	Calluna vulgaris, Juniperus communis subsp. nana, Pinus mugo subsp. mugo	23 June 2012	T. N. Sieber
San Bernardino	46°28ʹ49.2ʺ N 9°10ʹ31.5ʺ E	1,930	Calluna vulgaris, Pinus mugo subsp. mugo	8 June 2014	T. N. Sieber

^aCollections were made within a radius of 200 m.

^bSamples only collected from plants with shoots covered by superficial mycelia. Calluna vulgaris occurred at all sites but was not colonized by snow molds at all sites and was therefore not collected at all sites.

Figure 1. (a) The San Bernardino site (1930 m a.s.l.) during snowmelt on 8 June 2014; arrowheads indicate places where felty brown to black mycelial mats of Allantophomospsis cytisporea as shown in (b), (c), and (d) can be observed. (b) Mycelial mat right at the edge of melting snow. (c) Mycelial mat covering horsts of dead leaves of the grass Nardus stricta. (d) Mycelial mat covering Calluna vulgaris. (e) Shoot blight on C. vulgaris observed at the San Bernardino site on 24 August 2014. (f) Close-up of (e).

Table 2. Allantophomopsis strains isolated from mycelial mats covering shoots of various plant species at various sites in the Swiss Alps during snow melt.

Strain number^a	Geographic origin	Altitude (m.a.s.l.)	Host	Growth rate on MEA at 20°C (mmd^−)^b	GenBank accession number
A_AR_1_06	Arosa, Sandböden	2,050	Juniperus communis subsp. nana	6.3	FJ904499
A_AR_2_06	Arosa, Sandböden	2,050	Juniperus communis subsp. nana	6.6	MK790117
A_AR_3_06	Arosa, Sandböden	2,050	Juniperus communis subsp. nana	6.9	MK790118
A_SB_4_02	San Bernardino	1,930	Calluna vulgaris	7.5	MK790119
A_SB_5_02	San Bernardino	1,930	Calluna vulgaris	7.2	MK790120
A_SB_6_12	San Bernardino	1,930	Pinus mugo subsp. mugo	7.2	MK790121
A_SB_7_12	San Bernardino	1,930	Calluna vulgaris	7.0	MK790122
A_SB_8_12	San Bernardino	1,930	Juniperus communis subsp. nana	6.5	MK790123
A_SB_9_12	San Bernardino	1,930	Juniperus communis subsp. nana	6.8	MK790124
A_LT_11_12	Lötschental	2,328	Calluna vulgaris	n.a.	MK790116
A_SB_12_14	San Bernardino	1,930	Calluna vulgaris	n.a.	MK790129
A_SB_14_14	San Bernardino	1,930	Calluna vulgaris	5.5	MK790125
A_SB_15_14	San Bernardino	1,930	Calluna vulgaris	n.a.	MK790130
A_SB_18_14	San Bernardino	1,930	Pinus mugo subsp. mugo	6.8	MK790126
A_SB_19_14	San Bernardino	1,930	Pinus mugo subsp. mugo	6.5	MK790127
A_SB_20_14	San Bernardino	1,930	Pinus mugo subsp. mugo	6.9	MK790128
A_DA_21_15	Davos, Geisswaldji	1,800	Calluna vulgaris	6.8	MK790134
A_DA_22_15	Davos, Geisswaldji	1,800	Calluna vulgaris	5.5	MK790131
A_DA_23_15	Davos, Geisswaldji	1,800	Calluna vulgaris	n.a.	MK790133
A_DA_24_15	Davos, Geisswaldji	1,800	Calluna vulgaris	5.6	MK790132

^aStrains deposited at UAMH Center for Global Microfungal Biodiversity, Toronto, Canada.  ^bn.a. = not available.

Table 3. Collection details and GenBank accession numbers of isolates included in the phylogenetic analysis.

Fungal name^a	ITS sequence GenBank accession number	Strain number	Host	Substrate	Geographic origin	Lifestyle/disease	Reference
Allantophomopsiella pseudotsugae	KJ663826	CBS 321.53	Picea abies	Dead bark	Norway	Dieback	Crous et al. (2014)
Allantophomopsiella pseudotsugae	KJ663829	CBS 841.91	Pinus sp.	n.a.	Germany	n.a.	Crous et al. (2014)
Allantophomopsiella pseudotsugae	KJ663827	CBS 437.71	Pinus sylvestris	n.a.	The Netherlands	n.a.	Crous et al. (2014)
Allantophomopsiella pseudotsugae	JN033384	CBS 322.53	Picea abies	Dead bark	Sunnfjord, Norway	Saprophyte	Han et al. (2014)
Allantophomopsis cytisporea	KJ663830	CBS 262.85	Conifer	Root	Germany	n.a.	Crous et al. (2014)
Allantophomopsis cytisporea	KJ663822	CBS 109.22	Vaccinium macrocarpon	Leaf	United States	Black rot	Crous et al. (2014)
Allantophomopsis cytisporea	KR873228	CBS 140061 ex-type	Vaccinium macrocarpon	Berry	Aluksne, Latvia	Black rot	Crous et al. (2015)
Allantophomopsis lunata	NR132922	CBS 137781 ex-type	Vaccinium macrocarpon	Berry	New Jersey	Black rot	Crous et al. (2015)
Allantophomopsis lycopodina	AB041243	CBS 361.68	Rhododendron sp.	Litter	The Netherlands	Saprophyte	Okane et al. (2001)
Allantophomopsis sp.	KJ663839	CBS 322.36	Pinus radiata	n.a.	New Zealand	n.a.	Crous et al. (2015)
Fungal sp.	FM172911	agrAP150	Calluna vulgaris	Root hair	Bavaria, Germany	Endophyte	n.a.
Fungal sp.	FM200723	AP652	Calluna vulgaris	Leaf	Bavaria, Germany	Endophyte	n.a.
Fungal sp.	HM123110	AZ0370	Pinus arizonica var. arizonica	Dead needle	Arizona	Saprophyte	U’ren et al. (2010)
Fungal sp.	HM123479	AZ0780	Physcia caesia	Lichen thallus	Arizona	Endolichenophyte	U’ren et al. (2010)
Phacidiopycnis sp.	HM595538	M5, ZLY-2010b	Abies beshanzuensis	Needle	Baishanzu National Nature Reserve, China	Endophyte	Yuan et al. (2011)
Phacidiopycnis washingtonensis	AY608644	CLX2152	Malus domestica	Fruit	Chelan, Washington	Postharvest disease	Xiao et al. (2005)
Phacidiopycnis washingtonensis	JF732919	OVB10-001	Malus domestica	Fruit	Germany	Postharvest disease	Weber (2011)
Phacidiopycnis washingtonensis	JQ013491	JKM54	Pinus radiata	n.a.	New Zealand	n.a.	n.a.
Gremmenia infestans	U92305		Pinus cembra	Shoot	Obersulzbachtal, Germany	Snow blight	Gernandt et al. (1997)
Phacidium lacerum	FR717225	ZK205/08	Picea abies	Litter	Czech Republic	Saprophyte	Zifcakova et al. (2011)
Phacidium lacerum	FR837911	ZK20/08	Picea abies	Needle	Czech Republic	Endophyte	Koukol et al. (2012)
Phacidium lacerum	KJ663842	CBS 338.70	Ilex aquifolium	Litter	The Netherlands	Saprophyte	Crous et al. (2014)
Phacidium lacerum	KJ663846	CBS 761.73	Pinus sylvestris	Dead needle	France	Saprophyte	Crous et al. (2014)
Phacidium lacerum	KJ663843	CBS 400.81	Juniperus communis	Living needle	France	Endophyte	Crous et al. (2014)
Phacidium lauri	KJ663850	CBS 308.68	Prunus laurocerasus	Litter	The Netherlands	Saprophyte	Crous et al. (2014)
Phacidium pseudophacidioides	KJ663853	CBS 590.69	Ilex aquifolium	Litter	The Netherlands	Saprophyte	Crous et al. (2014)
Potebniamyces pyri	AY608641	CBS 339.78	Pyrus communis	Fruit	British Columbia, Canada	Postharvest disease	Xiao et al. (2005)
Potebniamyces pyri	KJ663823	CBS 282.55	Pyrus communis	Peduncle	The Netherlands	Saprophyte	Crous et al. (2014)
Potebniamyces pyri	KJ663859	CBS 322.63	Pyrus communis	Bark	The Netherlands	Saprophyte	Crous et al. (2014)
Pseudophacidium ledi	KJ663860	CBS 377.59	Picea abies	n.a.	Switzerland	n.a.	Crous et al. (2014)

n.a. = not available.

^aCurrent name according to Index fungorum (http://www.indexfungorum.org/Names/Names.asp), accessed 12 March 2019.

Figure 2. (a) Mycelium of Allantophomopsis cytisporea colonizing dead tissue of a Calluna vulgaris leaf (scale bar: 20 µm); the bright meandering lines are the cell walls of the puzzle piece–like epidermal cells of C. vulgaris. (b) Interaction between Herpotrichia pinetorum (strain 95-Fi; Schneider et al. 2009) to the left and A. cytisporea (strain A_DA_21_15) to the right after 21 days at 4°C on MEA. Conidia of A. cytisporea strains (c) A_SB_8_12 and (d) A_SB_6_12 in H₂O stained with fountain pen ink (Pelikan, blue) and observed using phase contrast optics (scale bar: 10 µm); arrowheads indicate the slimy appendages of the conidia.

Figure 3. The most parsimonious tree showing the phylogenetic relationships among snow mold strains collected for this study (bold) and related species as inferred from ITS1-5.8s-ITS2 (489 character states including gaps) sequences. The scale bar shows the number of changes, and bootstrap support values of greater than 50 percent from 100 replicates are shown at the nodes. Taxon names are preceeded by GenBank accession numbers. The species boundaries are delimited with colored blocks. The tree was rooted to Gremmenia (Phacidium) infestans (GenBank accession U92305).

Table 4. Frequency (percentage) of incubated mycelial samples giving rise to Allantophomopsis cytisporea, Herpotrichia pinetorum, and other fungi.

Host	Number of samples (n)	Allantophomopsis cytisporea	Herpotrichia pinetorum	Other fungi^a	No growth
Calluna vulgaris^b	68	16.2	10.3	51.5	22.1
Erica carnea	9	0	0	88.9	11.1
Juniperus communis subsp. nana	99	5.1	84.0^c	7.1^d	3.0
Picea abies	6	0	100.0	0	0
Pinus mugo subsp. mugo	18	22.2	16.7	11.1	50.0

^aIncluding yeasts, Cladosporium spp., Mucor hiemalis, Phaeosphaeria lycopodina (GenBank accession number: MK790186), Truncatella angustata (MK790184), Botrytis cinerea, and Mycocentrospora sp. (MK790185).

^bA few samples consisted of a surface-sterilized (1 minute, 96 percent ethanol) leaf.

^cTwo Herpotrichia isolates were Neopeckia coulteri (FJ904496, FJ904497).

^dOne Herpotrichia sp. (FJ904498) isolate, not conspecific with either N. coulteri or H. pinetorum (see Schneider et al. [2009] for GenBank accessions).

Figure 4. Growth rates of two Allantophomopsis cytisporea strains and Herpotrichia pinetorum at various temperatures.

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