Taxonomy of Arthrinium minutisporum sp. nov., Pezicula neosporulosa, and Acrocalymma pterocarpi: New Records from Soil in Korea

Figures & data

Table 1. List of species used in this study and their GenBank accession numbers for phylogenetic analysis.

		GenBank accession numbers
Species	Strain numbers	ITS	LSU	TUB2	TEF1-α	RPB2
Arthrinium arundinis	CBS 106.12	KF144883	–	KF144973	KF145015	–
A. aureum	CBS 244.83	AB220251	–	KF144981	KF145023	–
A. balearicum	CBS 145129^T	MK014869		MK017975	MK017946	–
A. bambusae	LC7106	KY494718	–	KY705186	KY806204	–
A. camelliae-sinensis	LC5007	KY494704	–	KY705173	KY705103	–
A. descalsii	CBS 145130^T	MK014870		MK017975	MK017947	–
A. dichotomanthi	LC4950	KY494697	–	KY705167	KY705096	–
A. gaoyouense	CFCC52301	MH197124	–	MH236789	MH236793	–
A. guizhouense	LC5318	KY494708	–	KY705177	KY705107	–
A. hydei	CBS 114990^T	KF144890	–	KF144982	KF145024	–
A. hydei	LC7105	KY494717	–	KY705185	KY705116	–
A. jiangxiense	LC4577	KY494693	–	KY705163	KY705092	–
A. kogelbergense	CBS 113332	KF144891	–	KF144983	KF145025	–
A. locutum-pollinis	LC11683	MF939595	–	MF939622	MF939616	–
A. malaysianum	CBS 102053^T	KF144896	–	KF144988	KF145030	–
A. marii	CBS 497.90	AB220252	–	KF144993	KF145035	–
A. minutisporum	17E-042^T	LC517882	–	LC518888	LC518889	–
A. obovatum	LC4940	KY494696	–	KY705166	KY705095	–
A. phaeospermum	CBS 114314	KF144904	–	KF144996	KF145038	–
A. phragmites	CBS 135458	KF144909	–	KF145001	KF145043	–
A. pseudoparenchymaticum	LC7234	KY494743	–	KY705211	KY705139	–
A. pseudospegazzinii	CBS 102052^T	KF144911	–	KF145002	KF145045	–
A. pterospermum	CPC 20193^T	KF144913	–	KF145004	KF145046	–
A. qinlingense	CFCC 52303	MH197120	–	MH236791	MH236795	–
A. rasikravindrae	LC7115	KY494721		KY705189	KY705118	–
A. sacchari	CBS 212.30	KF144916	–	KF145005	KF145047	–
A. saccharicola	CBS 191.73	KF144920	–	KF145009	KF145051	–
A. subroseum	LC7291	KY494751	–	KY705219	KY705147	–
A. thailandicum	LC5630	KY494714	–	KY806200	KY705113	–
A. xenocordella	CBS 595.66	KF144926	–	KF145013	KF145055	–
Seiridium phylicae	CPC 19965	LT853093	–	LT853240	LT853190	–
Pezicula acericola	CBS 245.97	KR859098	KR858889	–	–	KF376213
P. aurantiaca	CBS 201.46	KR859102	KR858893	–	–	KF376210
P. carpinea	CBS 921.96	KR859107	KR858898	–	–	KF376159
P. chiangraiensis	MFLUCC 15-0170^T	KU310621	KU310622			KU310623
P. cinnamomea	CBS 626.96	KR859152	KR858944	–	–	KF376162
P. cornina	CBS 285.39	KR859163	KR858955	–	–	KR859333
P. corylina	CBS 249.97	KR859168	KR858960	–	–	KF376161
P. eucrita	CBS 259.97	KR859179	KR858971	–	–	KF376205
P. fagacearum	CBS 112400^T	NR155654	KR858993	–	–	KR859335
P. microspore	CBS 124641^T	NR155655	KR859004	–	–	KR859337
P. neocinnamomea	CBS 100248^T	NR155656	KR859005	–	–	KF376209
P. neoheterochroma	CBS 127388^T	NR155657	NG058178	–	–	KR859338
P. neosporulosa	CBS 101.96^T	NR138003	MH874192	–	–	KF376193
P. neosporulosa	17E-039	LC517883	LC517884	–	–	LC518890
P. ocellata	CBS 949.97	KR859233	KR859025	–	–	KF376215
P. pseudocinnamomea	CBS 101000^T	NR155658	KR859027	–	–	KR859340
P. rubi	CBS 253.97	KR859250	KR859042	–	–	KF376204
P. sporulosa	CBS 224.96^T	NR137161	MH874199	–	–	KF376201
Rhizodermea veluwensis	CBS 110605^T	KR859282	KR859076	–	–	KR859353
Acrocalymma aquatic	MFLUCC 11-0208^T	JX276951	NG042698	–	–	–
A. cycadis	CPC 17345	KJ869124	KJ869181	–	–	–
A. ficus	CBS 317.76^T	KP170619	KP170712	–	–	–
A. medicaginis	CPC 24340^ET	KP170620	KP170713	–	–	–
A. pterocarpi	MFLUCC 17-0926^T	MK347732	MK347949	–	–	–
A. pterocarpi	NC13-171	LC517880	LC517881	–	–	–
A. vagum	REF096	JN859316	JN859474	–	–	–
A. walker	CBS 257.93	AF383965	FJ795454	–	–	–
Boeremia exigua	CBS 431.74	FJ427001	JX681074	–	–	–

ITS: internal transcribed spacer regions; LSU: 28S rDNA gene; TUB2: β-tubulin; TEF1: translation elongation factor 1-alpha; RPB2: RNA polymerase II. The newly generated sequences were indicated in bold.

Figure 1. Cultural and morphological characteristics of 17E-042. (A) Colonies were grown on potato dextrose agar; (B) Malt extract agar; (C) Oatmeal agar at 25 °C for 7 days, reverse and obverse side view, respectively. (D) Colonies on oatmeal agar; (E–G) Conidiogenous cells give rise to conidia; (H–J) Sterile cells; (K–L) Conidia. Scale bars: E–L = 10 μm.

Table 2. Morphological comparison, habitat, and origin of Arthrinium minutisporum sp. nov. with the closest species of Arthrinium.

Sl. No.	Strain Name	Conidiogenous cells (μm)	Conidia (μm)	Habitat	Origin	References
1	A.minutisporum (17E-042)	5.0–7.5 × 3.6–6.0	5.7–8.2 × 4.6–7.0	Soil	Korea: Sancheong-gun	This study
2	A. phragmites (CBS135458^ET)	12.0– 15.0 × 3.0–5.0	9.0–10.0(–12.0) × (5.0–)6.0(–7.0)	Culms of Phragmites australis	Italy: Bomarzo	[7]
3	A. aureum (CBS 244.83^ET)	N/A	10.0 − 30.0 × 10.0 − 15.0	Air	Spain: Barcelona	[46]
4	A. hydei (CBS 114990^T)	5.0–8.0 × 4.0–5.0	(15.0–)17.0–19.0(–22.0) × (10.0–)11.0–12.0(–14.0)	Culms of Bambusa tuldoides	Hong Kong: Tai Po Kau	[7]
5	A. pterospermum (CPC 20194)	5.0–10.0 × 4.0–5.0	15.0–25.0 × 8.0–10.0	Leaf of Lepidosperma gladiatum	Australia: Adelaide	[7]
6	A. xenocordella (CBS 478.86^ET)	5.0–7.0 × 4.0–5.0	(7.0–)9.0–10.0(–11.0) × 6.0–7.0	Soil from roadway	Zimbabwe: Matopos	[7]
7	A. kogelbergense (CBS 113333^ET)	5.0–12.0 × 4.0–5.0	(8.0–)9.0–10.0 × 7.0–8.0 (–9.0) × 4.0–5.0	Dead culms of Restionaceae	South Africa	[7]
8	A. phaeospermum (CBS 142.55)	5.0–10.0 × 3.0–5.0	(9.0–)10.0(–12.0) × 6.0–7.0	Soil	Japan: Tiba prefecture	[7]
9	A. saccharicola (CBS 831.71)	5.0–10.0 × 3.0–5.0	(7.0–)8.0–9.0(–10.0) × (4.0–)5.0(–6.0)	Unknown	The Netherlands	[7]
10	A. arundinis (CBS 133509)	6.0–12.0 × 3.0–4.0	(5.0–)6.0–7.0 × 3.0–4.0	Aspergillus flavus sclerotium buried in a sandy field	USA: Kilbourne	[7]
11	A. rasikravindrae (NFCCI 2144^T)	N/A	10.0–15.0 × 6.0–10.5	Soil	Norway	[5]
12	A. balearicum (CBS 145129^ET)	not studied	not studied	Undetermined Poaceae	Spain	[3]
13	A. descalsii (CBS 145130^ET)	5.0 × 4.0	(5.0–)7.0–8.0 × 6.0–7.0	Dead stems of Ampelodesmos mauritanicus	Spain	[3]

N/A: not available in previous references.

Figure 2. Neighbor-joining phylogenetic tree of 17E-042 based on combined sequences data (ITS+TUB2+TEF1) showing the relationship between Arthrinium minutisporum sp. nov., and the closest Arthrinium spp. The tree was rooted using Seiridium phylicae CPC 19965 as an outgroup. The numbers above the branches represent the bootstrap values (ML/MP/NJ) obtained for 1000 replicates (values smaller than 50% were not shown). The isolated strain of this study is indicated in bold. Bar, 0.02 substitutions per nucleotide position.

Figure 3. Cultural and morphological characteristics of 17E-039. (A) Colonies were grown on potato dextrose agar; (B) Malt extract agar; (C) Oatmeal agar at 25 °C for 14 days, reverse and obverse side view, respectively. (D–F) Conidiomata forming on oatmeal agar; (G) Macroconidiogenous cells; (H) Microconidiogenous cells; (I–K) Macroconidia; (L, M) Microconidia. Arrows indicate microconidiogenous cells. Scale bars: D–F = 500 μm; G–M = 10 μm.

Table 3. Morphological characteristics of the strain 17E-039 with reference to Pezicula neosporulosa.

Characteristics		Strain 17E-039^a	Pezicula neosporulosa^b
Conidiomata	Shape and size	Globose or sub-globose, initially closed and covered by gray to white hyphae, later opening to form a disc-like structure of 0.5–1.2 mm diameter, which produces first clear water droplets, then pale yellow to dark yellow conidial slime.	Globose or sub-globose, initially closed and covered by gray to white hyphae, later opening to form a disc-like structure of 1–3 mm diameter, which produces first clear water droplets, then milky white to pale yellow conidial slime.
Conidiogenous cells (Diam.) (μm)	Macro	7.2–23.1 × 4.5–9.1	7.5–14.0 × 4.0–5.5
	Micro	7.5–14.7 × 1.5–2.5	7.5–15.0 × 1.0–2.0
Conidia (μm)	Macro	20.4–34.3 × 8.0–12.0	24.0–36.5 × 9.0–12.0
	Micro	7.6–15.0 × 1.2–1.7	8.0–14 × 1.5–2.5

^aFungal strain studied in this paper.

^bSource of the description [10].

Figure 4. Neighbor-joining phylogenetic tree of 17E-039 based on combined sequences data (ITS + LSU+RPB2) showing the relationships between P. neosporulosa and the closest Pezicula spp. The tree was rooted using Rhizodermea veluwensis CBS 110605^T as an outgroup. The numbers above the branches represent the bootstrap values (ML/MP/NJ) obtained for 1000 replicates (values smaller than 60% were not shown). The isolated strain of this study is indicated in bold. Bar, 0.01 substitutions per nucleotide position.

Figure 5. Cultural and morphological characteristics of NC13-171. (A) Colony on potato dextrose agar; (B) Malt extract agar; (C) Oatmeal agar at 25 °C in 14 days, reverse and obverse side view, respectively. (D) Structures formed in culture; (E–G) Chlamydospores. Scale bars: D = 500 μm; E–G = 10 μm.

Figure 6. Neighbor-joining phylogenetic tree based on the concatenated ITS region and partial of 28S rDNA sequences showing the phylogenetic position of Acrocalymma pterocarpi NC13-171 among members of the genus Acrocalymma. The tree was rooted using Boeremia exigua CBS 431.74 as an outgroup. The numbers above the branches represent the bootstrap values (ML/MP/NJ) obtained for 1000 replicates (values smaller than 50% were not shown). The isolated strain of this study is indicated in bold. Bar, 0.01 substitutions per nucleotide position.

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