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Mycological Progress

, Volume 11, Issue 3, pp 655–688 | Cite as

Cylindrocarpon root rot: multi-gene analysis reveals novel species within the Ilyonectria radicicola species complex

  • Ana Cabral
  • Johannes Z. Groenewald
  • Cecília Rego
  • Helena Oliveira
  • Pedro W. Crous
Open Access
Original Article

Abstract

Ilyonectria radicicola and its Cylindrocarpon-like anamorph represent a species complex that is commonly associated with root rot disease symptoms on a range of hosts. During the course of this study, several species could be distinguished from I. radicicola sensu stricto based on morphological and culture characteristics. DNA sequence analysis of the partial β-tubulin, histone H3, translation elongation factor 1-α and nuclear ribosomal RNA-Internal Transcribed Spacer (nrRNA-ITS) genes were employed to provide further support for the morphological species resolved among 68 isolates associated with root rot disease symptoms. Of the various loci screened, nrRNA-ITS sequences were the least informative, while histone H3 sequences were the most informative, resolving the same number of species as the combined dataset across the four genes. Within the Ilyonectria radicicola species complex, 12 new taxa are delineated occurring on a diverse range of hosts, the most common being Cyclamen, Lilium, Panax, Pseudotsuga, Quercus and Vitis.

Keywords

Cylindrocarpon root rot Nectria-like fungi Phylogeny Systematics 

Introduction

The genus Cylindrocarpon was introduced in 1913 by Wollenweber, with C. cylindroides as type. Cylindrocarpon and Cylindrocarpon-like species have since been commonly associated with root and decay of woody and herbaceous plants (Domsch et al. 2007). Cylindrocarpon root rot causes losses up to 30% on ginseng (Panax quinquefolium) (Seifert et al. 2003), and plays an important role in black foot rot of grapevines (Halleen et al. 2004, 2006), apple replant disease (Tewoldemedhin et al. 2010), and beech cankers (Castlebury et al. 2006), to name but a few hosts of economic importance.

In his taxonomic revision of Cylindrocarpon, Booth (1966) divided this genus into four groups based on the presence or absence of microconidia or chlamydospores. Booth’s group 4 represents Neonectria s. str., as it accommodates the type species N. ramulariae (anamorph: C. obtusiusculum). Most of the teleomorphs of Cylindrocarpon species have since this date been classified in Neonectria (Brayford et al. 2004; Halleen et al. 2004, 2006; Mantiri et al. 2001; Rossman et al. 1999). Several phylogenetic studies have, however, revealed that Neonectria/Cylindrocarpon is paraphyletic (Castlebury et al. 2006; Halleen et al. 2004, 2006; Hirooka et al. 2005; Mantiri et al. 2001). The first step in resolving this issue was taken by Halleen et al. (2004), who proposed Campylocarpon for species resembling Cylindrocarpon with 3–5-septate, curved macroconidia, and lacking microconidia. A further phylogenetic study (Chaverri et al. 2011) divided the Neonectria complex into four genera based on a combination of characters linked to perithecial anatomy and conidial septation: Ilyonectria, Neonectria/Cylindrocarpon s. str., Rugonectria and Thelonectria. In this study, a single generic name was proposed for each clade in an attempt to move towards a single nomenclature for pleomorphic fungi, meaning that the Cylindrocarpon-like anamorphs of Ilyonectria, Rugonectria and Thelonectria were placed in teleomorph genera, as recently done with other groups of pleomorphic fungi (Crous et al. 2006, 2007, 2009a; Gräfenhan et al. 2011; Lombard et al. 2010; Schroers et al. 2011).

Cylindrocarpon root rot is commonly associated with “Cylindrocarpondestructans in the literature (Halleen et al. 2004; Samuels and Brayford 1990). This fungus was originally described as Ramularia destructans from roots of ginseng (Panax quinquefolium) collected in the USA (Zinssmeister 1918). Furthermore, it has been linked to the teleomorph Ilyonectria radicicola (Booth 1966; Chaverri et al. 2011; Samuels and Brayford 1990), which Gerlach and Nilsson (1963) described from rotting bulbs of Cyclamen persicum collected in Sweden. Samuels and Brayford (1990) commented on the morphological variation in collections of I. radicicola and its anamorph “C.” destructans. Seifert et al. (2003) showed that there was more than one “C.” destructans-like species occurring on Panax, and that none of the resolved clades correlated to the ex-type strain of I. radicicola, leading Halleen et al. (2006) to question the purported anamorph/teleomorph link between I. radicicola (from Cyclamen, Sweden) and “C.” destructans (from Panax, USA). Based on a phylogenetic analysis of ITS nrRNA gene sequences, Schroers et al. (2008) concluded that the I. radicicola complex includes “C.” destructans, “C.” destructans var. crassum, I. coprosmae, I. liriodendri, N. austroradicicola and N. macroconidialis.

The aim of the present study was to elucidate the morphological variation present within the I. radicicola complex, and to link fresh collections to older names introduced for species in this complex. This was addressed by combining morphological and culture characteristics with DNA sequence data derived from the Internal Transcribed Spacers (ITS) of the nrRNA gene operon, and partial β-tubulin (TUB), histone H3 (HIS), and translation elongation factor 1-α (TEF) genes.

Materials and methods

Isolates

This study (Table 1) included 42 “C.destructans s. lat. isolates [including the ex-type strains of I. radicicola (CBS 264.65) and “C.destructans f.sp. panacis (CBS 124662), “C.destructans var. destructans and “C.destructans var. crassum], six “C.didymum isolates, six I. liriodendri isolates, one N. macroconidialis isolate and one I. coprosmae isolate, all deposited at the CBS-KNAW Fungal Biodiversity Centre, Utrecht, the Netherlands (CBS). Also included are two isolates that were previously identified as Ramularia mors-panacis (CBS 306.35) and R. panacicola (CBS 307.35) by Hildebrand (1935).
Table 1

Details pertaining to isolates investigated during this study

Species

Strain numbera

Collected/isolated by, year

Isolated from

Location

GenBank accession numbers

ITS

TUB

H3

EF1

Campylocarpon fasciculare, Holotype

CBS 112613; STE-U 3970; C 76

F. Halleen, 2000

Vitis vinifera, trunk of young grapevine showing decline symptoms; scion Cabernet Sauvignon; rootstock Richter 99

South Africa, Western Cape, Riebeeck Kasteel

AY677301

AY677221

JF735502

JF735691

Campylocarpon pseudofasciculare, Holotype

CBS 112679; STE-U 5472; HJS-1227

F. Halleen, 2000

Vitis vinifera, roots, asymptomatic nursery grapevine plant; scion Sultana; rootstock Ramsey

South Africa, Western Cape, Wellington

AY677306

AY677214

JF735503

JF735692

Neonectria macroconidialis

CBS 119596; ICMP 9349; IMI 332705; GJS 85-59

G.J. Samuels, 1985

Astelia sp.

New Zealand, Gisborne, Urewera National Park

JF735259

JF735372

JF735504

JF735693

Ilyonectria coprosmae

CBS 119606; GJS 85-39

G.J. Samuels, 1985

Metrosideros sp.

Canada, Ontario

JF735260

JF735373

JF735505

JF735694

Ilyonectria radicicola, type strain

CBS 264.65

L. Nilsson, 1961

Cyclamen persicum

Sweden, Skåne, Bjärred

AY677273

AY677256

JF735506

JF735695

Ilyonectria liriodendri, type strain of “C.liriodendri

CBS 110.81; IMI 303645

J.D. MacDonald & E.E. Butler, 1978

Liriodendron tulipifera, root

USA, California, Yolo Co., Davis

DQ178163

DQ178170

JF735507

JF735696

Ilyonectria liriodendri

CBS 117526; Cy68

C. Rego, 1999

Vitis vinifera, asymptomatic rootstocks; rootstock 99 R, clone 179 F

Portugal, Ribatejo e Oeste

DQ178164

DQ178171

JF735508

JF735697

Ilyonectria liriodendri

CBS 117527; Cy76

C. Rego, 1999

Vitis vinifera, asymptomatic rootstocks; rootstock 110 R, clone 164E

Portugal, Ribatejo e Oeste

DQ178165

DQ178172

JF735509

JF735698

Ilyonectria liriodendri

CBS 117640; IMI 357400; Cy1

C. Rego, 1992

Vitis vinifera, 4-year-old plant showing decline symptoms; scion Seara Nova; rootstock 99R

Portugal, Torres Vedras, Dois Portos

DQ178166

DQ178173

JF735510

JF735699

Ilyonectria liriodendri

CBS 112596; STE-U 3994; C 14

F. Halleen, 1999

Vitis vinifera, roots

South Africa, Western Cape, De Wet

AY677264

AY677239

JF735511

JF735700

Ilyonectria liriodendri

CBS 112607; STE-U 3986; C 81

F. Halleen, 2000

Vitis vinifera, basal end of trunk

South Africa, Western Cape, Robertson

AY677269

AY677241

JF735512

JF735701

Ilyonectria liriodendri

Cy164

C. Rego, 1997

Malus domestica; cultivar Lysgolden; rootstock MM106

Portugal, Porto de Mós, Valbom

AM419079

AM419112

JF735513

JF735702

Ilyonectria liriodendri

Cy122

W.D. Gubler

Vitis vinifera

USA, California

JF735261

JF735374

JF735514

JF735703

Ilyonectria liriodendri

Cy190

N. Cruz, 2005

Vitis vinifera, basal end of 6-year-old plant; scion Alvarinho; rootstock 196-17

Portugal, Monção, Cortes

JF735262

JF735375

JF735515

JF735704

Ilyonectria liriodendri

Cy232

L. Inácio & J. Henriques, 2007

Quercus suber, stem

Portugal, Macedo de Cavaleiros

JF735263

JF735376

JF735516

JF735705

Ilyonectria robusta

CBS 321.34

-

Loroglossum hircinum, root

Tunisia, Tunis

AY677275

AY677253

JF735517

JF735706

Ilyonectria robusta, type strain of Ramularia robusta

CBS 308.35

A.A. Hildebrand

Panax quinquefolium

Canada, Ontario

JF735264

JF735377

JF735518

JF735707

Ilyonectria robusta

CBS 773.83

J. Hemelraad

water, in aquarium with Anodonta

Netherlands, Utrecht

AY677276

AY677254

JF735519

JF735708

Ilyonectria robusta

CBS 605.92

R. Schröer, 1992

Tilia petiolaris, root

Germany, Hamburg

EF607078

EF607065

JF735520

JF735709

Ilyonectria robusta

CBS 117813; IFFF 84

E. Halmschlager, 1993

Quercus robur, root

Austria, Niederweiden

-

JF735378

-

-

Ilyonectria robusta

CBS 117814; IFFF 85

E. Halmschlager, 1993

Quercus sp., root

Austria, Patzmannsdorf

JF735265

JF735379

JF735521

JF735710

Ilyonectria robusta

CBS 117815; IFFF 86

E. Halmschlager, 1993

Quercus sp., root

Austria, Patzmannsdorf

JF735266

JF735380

JF735522

JF735711

Ilyonectria robusta

CBS 117817; IFFF 88

E. Halmschlager, 1993

Quercus sp., root

Austria, Patzmannsdorf

-

JF735381

-

-

Ilyonectria robusta

CBS 117818; IFFF 89

E. Halmschlager, 1993

Quercus sp., root

Austria, Patzmannsdorf

JF735267

JF735382

JF735523

JF735712

Ilyonectria robusta

CBS 117819; IFFF 90

E. Halmschlager, 1993

Quercus robur, root

Austria, Niederweiden

-

JF735383

-

-

Ilyonectria robusta

CBS 117820; IFFF 91

E. Halmschlager, 1993

Quercus robur, root

Austria, Niederweiden

JF735268

JF735384

JF735524

JF735713

Ilyonectria robusta

CBS 117821; IFFF 93

E. Halmschlager, 1993

Quercus robur, root

Austria, Niederweiden

JF735269

JF735385

JF735525

JF735714

Ilyonectria robusta

CBS 117822; IFFF 94

E. Halmschlager, 1993

Quercus robur, root

Austria, Niederweiden

JF735270

JF735386

JF735526

JF735715

Ilyonectria robusta

CBS 117823; IFFF 95

E. Halmschlager, 1993

Quercus robur, root

Austria, Niederweiden

JF735271

JF735387

JF735527

JF735716

Ilyonectria robusta

CD1666

R. D. Reeleder, 1998

Panax quinquefolium

Canada, Nova Scotia

AY295331

JF735388

JF735528

JF735717

Ilyonectria robusta

CPC 13532; DAOM 139398; K 18-3A

-

Prunus cerasus cultivar Montmorency

Canada, Ontario

AY295330

JF735389

JF735529

JF735718

Ilyonectria robusta

Cy23

C. Rego, 1997

Vitis sp. rootstock 99R clone 179 F in nursery

Portugal, Ribatejo e Oeste

AJ875333

AM419093

JF735530

JF735719

Ilyonectria robusta

Cy158

C. Rego & T. Nascimento, 2004

Vitis vinifera, 1-year-old, died before sprouting; scion Alicante Bouschet; rootstock 1103P

Portugal, Lamego, Cambres

JF735272

JF735390

JF735531

JF735720

Ilyonectria robusta

CBS 129084; Cy192

N. Cruz, 2005

Vitis vinifera, basal end of 25-year-old plant; scion Alicante; rootstock 196-17

Portugal, Monção

JF735273

JF735391

JF735532

JF735721

Ilyonectria robusta

Cy231

F. Caetano, 2005

Thymus sp.

Portugal, Lisbon

JF735274

JF735392

JF735533

JF735722

Ilyonectria crassa

CBS 139.30

W.F. van Hell, 1930

Lilium sp., bulb

Netherlands

JF735275

JF735393

JF735534

JF735723

Ilyonectria crassa

CBS 158.31; IMI 061536; NRRL 6149

1930

Narcissus sp., root

Netherlands

JF735276

JF735394

JF735535

JF735724

Ilyonectria crassa

CBS 129083; NSAC-SH-1

S. Hong, 1998

Panax quinquefolium

Canada, Nova Scotia

AY295311

JF735395

JF735536

JF735725

Ilyonectria crassa

NSAC-SH-2

S. Hong, 1998

Panax quinquefolium

Canada, Nova Scotia

AY295313

JF735396

JF735537

JF735726

Ilyonectria crassa

NSAC-SH-2.5

S. Hong, 1998

Panax quinquefolium

Canada, Nova Scotia

AY295314

JF735397

JF735538

JF735727

Cylindrocarpon” sp.

CBS 120370; CR 20

P. Axelrood, 1998

Pseudotsuga menziesii

Canada, British Columbia

AY295317

JF735398

JF735539

JF735728

Ilyonectria rufa Authentic strain of Coleomyces rufus

CBS 153.37

F. Moreau, 1937

Dune sand

France

AY677271

AY677251

JF735540

JF735729

Ilyonectria rufa

CBS 156.47; IAM 14673; JCM 23100

-

Azalea indica

Belgium, Amandsberg

AY677272

AY677252

JF735541

JF735730

Ilyonectria rufa

CBS 640.77

F. Gourbière, 1977

Abies alba

France, Villeurbanne

JF735277

JF735399

JF735542

JF735731

Ilyonectria rufa

CBS 120371; CR 26

P. Axelrood, 1998

Pseudotsuga menziesii

Canada, British Columbia

AY295318

JF735400

JF735543

JF735732

Ilyonectria rufa

CBS 120372; CR 29

P. Axelrood, 1998

Pseudotsuga menziesii

Canada, British Columbia

JF735278

JF735401

JF735544

JF735733

Ilyonectria rufa

CPC 13536; DAOM 226721; CR36

P. Axelrood, 1998

Pseudotsuga menziesii

Canada, British Columbia

JF735279

JF735402

JF735545

JF735734

Ilyonectria rufa

94-1628

R.C. Hamelin, 1994

Picea glauca

Canada, Quebec

AY295315

JF735403

JF735546

JF735735

Ilyonectria mors-panacis

CBS 120359; CD1561

R. D. Reeleder, 1996

Panax quinquefolium

Canada, Ontario

AY295309

JF735404

JF735547

JF735736

Ilyonectria mors-panacis

CBS 120360; CD1567

R. D. Reeleder, 1996

Panax quinquefolium

Canada, Ontario

-

AY297200

-

-

Ilyonectria mors-panacis

CBS 120361; CD1596

R. D. Reeleder, 1996

Panax quinquefolium

Canada, Ontario

JF735280

JF735405

JF735548

JF735737

Ilyonectria mors-panacis

CBS 120362; CD1598

R. D. Reeleder, 1996

Panax quinquefolium

Canada, Ontario

-

AY297202

-

-

Ilyonectria mors-panacis

CBS 120363; CD1635

R. D. Reeleder, 1997

Panax quinquefolium

Canada, Ontario

-

AY297204

-

-

Ilyonectria mors-panacis

CBS 120364; CD1636

R. D. Reeleder, 1997

Panax quinquefolium

Canada, Ontario

JF735281

JF735406

JF735549

JF735738

Ilyonectria mors-panacis

CBS 120365; CD1637

R. D. Reeleder, 1997

Panax quinquefolium

Canada, Ontario

JF735282

JF735407

JF735550

JF735739

Ilyonectria mors-panacis

CBS 120366; CD 1639

R. D. Reeleder, 1997

Panax quinquefolium

Canada, Ontario

JF735283

JF735408

JF735551

JF735740

Ilyonectria mors-panacis

CBS 120367; CD1640

R. D. Reeleder, 1997

Panax quinquefolium

Canada, Ontario

AY295321

JF735409

JF735552

JF735741

Ilyonectria mors-panacis

CBS 120368; CD1641

R. D. Reeleder, 1997

Panax quinquefolium

Canada, Ontario

JF735284

JF735410

JF735553

JF735742

Ilyonectria mors-panacis

CBS 120369; CD1642

R. D. Reeleder, 1997

Panax quinquefolium

Canada, Ontario

JF735285

JF735411

JF735554

JF735743

Ilyonectria mors-panacis

CPC 13535; DAOM 221059; CD 0265

R. D. Reeleder, 1989

Panax quinquefolium

Canada, Ontario

JF735286

JF735412

JF735555

JF735744

Ilyonectria mors-panacis

CPC 13537; DAOM 226727; CD 1570

R. D. Reeleder, 1996

Panax quinquefolium

Canada, Ontario

JF735287

JF735413

JF735556

JF735745

Ilyonectria mors-panacis, type of Ramularia mors-panacis

CBS 306.35

A.A. Hildebrand

Panax quinquefolium

Canada, Ontario

JF735288

JF735414

JF735557

JF735746

Ilyonectria mors-panacis

CBS 307.35

A.A. Hildebrand

Panax quinquefolium

Canada, Ontario

JF735289

JF735415

JF735558

JF735747

Ilyonectria mors-panacis, type of “C.destructans f.sp. panacis

CBS 124662; NBRC 31881; SUF 811

Y. Miyazawa

Panax ginseng

Japan, Nagano, Kitasaku-gun

JF735290

JF735416

JF735559

JF735748

Ilyonectria pseudodestructans

CPC 13534; DAOM 150670; Berkenkamp 1

B. Berkenkamp, 1974

Poa pratensis

Canada, Alberta, Lacombe

AY295319

JF735417

JF735560

JF735749

Ilyonectria pseudodestructans

CBS 117812; IFFF 83

E. Halmschlager, 1993

Quercus sp., root

Austria, Patzmannsdorf

JF735291

JF735418

JF735561

JF735750

Ilyonectria pseudodestructans

CBS 117824; IFFF 98

E. Halmschlager, 1993

Quercus sp., root

Austria, Patzmannsdorf

JF735292

JF735419

JF735562

JF735751

Ilyonectria pseudodestructans

CBS 129081; Cy20

C. Rego, 1996

Vitis vinifera, 4-year-old, showing decline symtoms, scion Malvasia Fina; rootstock 1103P

Portugal, Gouveia, São Paio

AJ875330

AM419091

JF735563

JF735752

Ilyonectria pseudodestructans

Cy22

C. Rego, 1996

Vitis vinifera, 5-year-old, showing decline symtoms, scion Aragonez; rootstock 99R

Portugal, Viseu, Silgueiros

AJ875331

AM419092

JF735564

JF735753

Ilyonectria europaea

Cy131

P. Lecomte & S. Chamont, 2000

Actinidia chinensis 'Hayward', internal lesion of stem

France, St. Chicq-du-Gaue

AM419067

AM419103

JF735565

JF735754

Ilyonectria europaea

Cy155

C. Rego & H. Oliveira, 2004

Vitis vinifera, 2-year-old, showing decline symtoms, scion Alfrocheiro; rootstock SO4

Portugal, Alter do Chão

JF735293

JF735420

JF735566

JF735755

Ilyonectria europaea

CBS 129078; Cy241

C. Rego, 2008

Vitis vinifera, basal end of a 2-year-old plant; scion Petit Verdot; rootstock 110R

Portugal, Vidigueira

JF735294

JF735421

JF735567

JF735756

Ilyonectria europaea

CBS 537.92

V. Demoulin, 1992

Aesculus hippocastanum, wood

Belgium, Liège

EF607079

EF607064

JF735568

JF735757

Ilyonectria europaea

CBS 102892; No.5/97-12

W. Leibinger, 1997

Phragmites australis, stem

Germany, Lake Constance

JF735295

JF735422

JF735569

JF735758

Ilyonectria lusitanica

CBS 129080; Cy197

N. Cruz, 2005

Vitis vinifera, below grafting zone, 6-year-old plant; scion Alvarinho; rootstock 196-17

Portugal, Melgaço, Alvaredo

JF735296

JF735423

JF735570

JF735759

Ilyonectria venezuelensis

CBS 102032; ATCC 208837; AR2553

A. Rossman, 1985

Bark

Venezuela, Amazonas, Cerro de la Neblina

AM419059

AY677255

JF735571

JF735760

Ilyonectria panacis

CBS 129079; CDC-N-9a

K. F. Chang, 1998

Panax quinquefolium

Canada, Alberta

AY295316

JF735424

JF735572

JF735761

Ilyonectria liliigena

CBS 189.49; IMI 113882

M.A.A. Schipper

Lilium regale, bulb

Netherlands, Hoorn

JF735297

JF735425

JF735573

JF735762

Ilyonectria liliigena

CBS 732.74

G.J. Bollen, 1973

Lilium sp.

Netherlands, Heemskerk

JF735298

JF735426

JF735574

JF735763

Ilyonectria liliigena

CBS 304.85

G.J. Bollen, 1985

Lilium sp., bulb

Netherlands

JF735299

JF735427

JF735575

JF735764

Ilyonectria liliigena

CBS 305.85

G.J. Bollen, 1985

Lilium sp., bulb

Netherlands

JF735300

JF735428

JF735576

JF735765

Ilyonectria gamsii

CBS 940.97

J.T. Poll, 1997

Soil

Netherlands, Lelystad

AM419065

AM419089

JF735577

JF735766

Cylindrocarpon” sp.

Cy228

F. Caetano, 2003

Ficus sp.

Portugal, Lisbon

JF735301

JF735429

JF735578

JF735767

Ilyonectria anthuriicola

CBS 564.95; PD 95/1577

R. Pieters, 1995

Anthurium sp., root

Netherlands, Bleiswijk

JF735302

JF735430

JF735579

JF735768

Ilyonectria vitis

CBS 129082; Cy233

C. Rego, 2008

Vitis vinifera, basal end of a 2-year-old plant; scion Touriga Nacional; rootstock 110R

Portugal, Vidigueira

JF735303

JF735431

JF735580

JF735769

Ilyonectria cyclaminicola

CBS 302.93

M. Hooftman, 1993

Cyclamen sp., bulb

Netherlands, Roelofarendsveen

JF735304

JF735432

JF735581

JF735770

Cylindrocarpon pauciseptatum

CBS 100819; LYN 16202/2

H.M. Dance, 1998

Erica melanthera, root

New Zealand, Tauranga

EF607090

EF607067

JF735582

JF735771

Cylindrocarpon pauciseptatum

CBS 113550

2003

Vitis sp, blackening areas in wood and base of trunk

New Zealand, Keesbury Estate

EF607080

EF607069

JF735583

JF735772

Cylindrocarpon pauciseptatum

CBS 120497; KIS 10763

H.-J. Schroers, 2006

Vitis sp.brownish spots of healthy looking root of ca. 12-year-old, possibly dead, in vineyard

Slovenia, Mrzlak

EF607085

EF607071

JF735584

JF735773

Cylindrocarpon pauciseptatum

CBS 120498; KIS 10775

M. Žerjav, 2006

Vitis sp., decayed secondary roots with black areas of 3-year-old, dead

Slovenia, Ljutomer

EF607087

EF607072

JF735585

JF735774

Cylindrocarpon pauciseptatum

CBS 120499; KIS 10780

M. Žerjav, 2006

Vitis sp., decayed secondary roots with black areas of 3-year-old, dead

Slovenia, Ljutomer

EF607084

EF607074

JF735586

JF735775

Cylindrocarpon pauciseptatum, type

CBS 120171; KIS 10467

M. Žerjav, 2005

Vitis sp., partly decayed roots of 4-year-old plant, still living but badly shooting; in vineyard

Slovenia, Krŝko

EF607089

EF607066

JF735587

JF735776

Cylindrocarpon pauciseptatum

CBS 120172; KIS 10729

M. Žerjav, 2006

Vitis sp., strongly decayed, blackish brown root of ca. 9-year-old plant, possibly dead; in vineyard

Slovenia, Žužemberk

EF607086

EF607070

JF735588

JF735777

Cylindrocarpon pauciseptatum

CBS 120173; KIS 10468

M. Žerjav, 2005

Vitis sp., partly decayed roots of 4-year-old plant, still living but badly shooting; in vineyard

Slovenia, Krŝko

EF607088

EF607068

JF735589

JF735778

Cylindrocarpon pauciseptatum

Cy196

N. Cruz, 2005

Vitis vinifera, basal end of 4-year-old plant; scion Alvarinho; rootstock 196-17

Portugal, Melgaço/Monção

JF735305

JF735433

JF735590

JF735779

Cylindrocarpon pauciseptatum

Cy217

A. Cabral, 2007

Vitis vinifera, asymptomatic; scion Gouveio

Portugal, Torres Vedras

JF735306

JF735434

JF735591

JF735780

Cylindrocarpon pauciseptatum

Cy238

C. Rego, 2008

Vitis vinifera, basal end of a 2-year-old plant; scion Petit Verdot; rootstock 110R

Portugal, Vidigueira

JF735307

JF735435

JF735592

JF735781

Cylindrocarpon” sp 1

CBS 162.89

M. Barth, 1988

Hordeum vulgare, root

Netherlands, Noordoostpolder, Marknesse, Lovinkhoeve

AM419060

AM419084

JF735610

JF735799

Cylindrocarpon” sp 2

Cy108

C. Rego, 1999

Vitis vinifera, basal end of a 4-year-old plant showing decline symptoms; scion Aragonez; rootstock SO4

Portugal, Nelas

JF735316

AM419100

JF735611

JF735800

Cylindrocarpon” sp 2

Cy200

N. Cruz, 2005

Vitis vinifera, basal end of a 16-year-old plant; scion Alvarinho; rootstock 196-17

Portugal, Melgaço

JF735317

JF735445

JF735612

JF735801

Cylindrocarpon” sp 2

CBS 159.34; IMI 113891; MUCL 4084; VKM F-2656

H.W. Wollenweber, 1934

 

Germany

JF735318

JF735446

JF735613

JF735802

Cylindrocarpon” sp 2

CBS 173.37; IMI 090176

T.R. Peace, 1937

Pinus laricio, associated with dieback

UK, England, Devon, Haldon

JF735319

JF735447

JF735614

JF735803

Cylindrocarpon” sp. 3

Cy135

C. Rego & T. Nascimento, 2003

Vitis vinifera, basal end of a 1.5-year-old plant showing decline symptoms; scion Aragonez; rootstock 3309 C

Portugal, Estremoz

AM419069

AM419105

JF735615

JF735804

Cylindrocarpon” sp. 3

Cy144

C. Rego & T. Nascimento, 2003

Vitis vinifera, grafting zone of a 1.5-year- old plant showing decline symptoms; scion Aragonez; rootstock 3309 C

Portugal, Estremoz

AM419074

AM419107

JF735616

JF735805

Cylindrocarpon” sp. 3

CBS 129085; Cy145

C. Rego & T. Nascimento, 2003

Vitis vinifera, basal end of a 1.5-year-old plant showing decline symptoms; scion Aragonez; rootstock 3309 C

Portugal, Estremoz

JF735320

JF735448

JF735617

JF735806

Cylindrocarpon” sp. 3

Cy146

C. Rego & T. Nascimento, 2003

Vitis vinifera, grafting zone of a 1.5-year-old plant showing decline symptoms; scion Aragonez; rootstock 3309 C

Portugal, Estremoz

JF735321

JF735449

JF735618

JF735807

Cylindrocarpon” sp. 3

Cy147

C. Rego & T. Nascimento, 2003

Vitis vinifera, grafting zone of a 1.5-year-old plant showing decline symptoms; scion Aragonez; rootstock 3309 C

Portugal, Estremoz

JF735322

JF735450

JF735619

JF735808

Cylindrocarpon” sp. 3

Cy148

C. Rego & T. Nascimento, 2003

Vitis vinifera, basal end of a 1.5-year-old plant showing decline symptoms; scion Aragonez; rootstock 3309 C

Portugal, Estremoz

JF735323

JF735451

JF735620

JF735809

Cylindrocarpon” sp. 3

Cy149

C. Rego & T. Nascimento, 2003

Vitis vinifera, basal end of a 1.5-year-old plant showing decline symptoms; scion Aragonez; rootstock 3309 C

Portugal, Estremoz

JF735324

JF735452

JF735621

JF735810

Cylindrocarpon” sp. 3

Cy150

C. Rego & T. Nascimento, 2003

Vitis vinifera, basal end of a 1.5-year-old plant showing decline symptoms; scion Aragonez; rootstock 3309 C

Portugal, Estremoz

JF735325

JF735453

JF735622

JF735811

Cylindrocarpon” sp. 3

Cy151

C. Rego & T. Nascimento, 2003

Vitis vinifera, asymptomatic 1.5-year-old plant; scion Aragonez; rootstock 3309 C

Portugal, Estremoz

JF735326

JF735454

JF735623

JF735812

Cylindrocarpon” sp. 3

Cy152

C. Rego & T. Nascimento, 2003

Vitis vinifera, asymptomatic 1.5-year-old plant; scion Aragonez; rootstock 3309 C

Portugal, Estremoz

JF735327

JF735455

JF735624

JF735813

Cylindrocarpon” sp. 3

Cy153

C. Rego & T. Nascimento, 2003

Vitis vinifera, asymptomatic 1.5-year-old plant; scion Aragonez; rootstock 3309 C

Portugal, Estremoz

JF735328

JF735456

JF735625

JF735814

Cylindrocarpon” sp. 3

Cy243

C. Rego, 2008

Vitis vinifera, basal end of a 2-year-old-plant; scion Touriga Nacioal; rootstock 110R

Portugal, Vidigueira

JF735329

JF735457

JF735626

JF735815

Cylindrocarpon” sp. 3

CPC 13539; 94–1685; CCFC226730

R. C. Hamelin, 1994

Picea glauca

Canada, Quebec

JF735330

JF735458

JF735627

JF735816

Cylindrocarpon” sp. 5

Cy133; IAFM Cy9-1

J. Armengol

Vitis vinifera

Spain, Valencia, L'Alcudia

JF735331

JF735459

JF735628

JF735817

Cylindrocarpon” sp. 5

Cy134; IAFM Cy20-1

J. Armengol

Vitis vinifera

Spain, Ciudad Real, Villarubia de los Ojos

JF735332

AM419104

JF735629

JF735818

Cylindrocarpon” sp. 5

CBS 129087; Cy159

A. Cabral & H. Oliveira, 2004

Vitis vinifera, basal end of a 3-year-old plant with root discolouration and decline symptoms; scion Sangiovese; rootstock 1103P

Portugal, Alcácer do Sal, Torrão

JF735333

AM419111

JF735630

JF735819

Cylindrocarpon” sp. 6

CBS 112593; STE-U 3990; C 107

F. Halleen, 2000

Vitis vinifera, roots of an asymptomatic nursery plant; scion Pinotage; rootstock 101–14 Mgt

South Africa, Western Cape, Wellington, Voorgroenberg

AY677281

AY677236

JF735631

JF735820

Cylindrocarpon” sp. 6

CBS 112608; STE-U 3987; C 62

F. Halleen, 2000

Vitis vinifera, roots, scion Chardonnay; rootstock 101–14 Mgt

South Africa, Western Cape, Citrusdal

AY677288

AY677235

JF735632

JF735821

Cylindrocarpon” sp. 6

CBS 113552; STE-U 5713; HJS-1306; NZ C 41

R. Bonfiglioli, 2003

Vitis sp. decline of nursery plants dead rootstocks

New Zealand, Candy P New Ground

JF735334

AY677237

JF735633

JF735822

Cylindrocarpon” sp. 6

Cy115

W.D. Gubler

Vitis vinifera

USA, California

JF735335

JF735460

JF735634

JF735823

Cylindrocarpon” sp. 6

Cy116

W.D. Gubler

Vitis vinifera

USA, California

AJ875322

JF735461

JF735635

JF735824

Cylindrocarpon” sp. 6

Cy117

W.D. Gubler

Vitis vinifera

USA, California

AJ875321

JF735462

JF735636

JF735825

Cylindrocarpon” sp. 6

Cy119

W.D. Gubler

Vitis vinifera

USA, California

JF735336

JF735463

JF735637

JF735826

Cylindrocarpon” sp. 6

Cy124

W.D. Gubler

Vitis vinifera

USA, California

JF735337

JF735464

JF735638

JF735827

Cylindrocarpon” sp. 6

Cy125

W.D. Gubler

Vitis vinifera

USA, California

AM419066

JF735465

JF735639

JF735828

Cylindrocarpon” sp. 6

Cy129

W.D. Gubler

Vitis vinifera

USA, California

JF735338

JF735466

JF735640

JF735829

Cylindrocarpon” sp. 6

Cy130

W.D. Gubler

Vitis vinifera

USA, California

JF735339

JF735467

JF735641

JF735830

Cylindrocarpon” sp. 6

Cy230

F. Caetano, 2005

Festuca duriuscula

Portugal, Lisbon

JF735340

JF735468

JF735642

JF735831

Ilyonectria macrodidyma

CBS 112594; STE-U 3991; C 111

F. Halleen, 2000

Vitis vinifera, roots of an asymptomatic nursery plant; scion Pinotage; rootstock Richter 99

South Africa, Western Cape, Malmesbury, Jakkalsfontein

AY677282

AY677231

JF735643

JF735832

Ilyonectria macrodidyma

CBS 112601; STE-U 3983; C 82

F. Halleen, 1999

Vitis vinifera, roots with black foot symtoms; scion Pinotage; rootstock US 8-7

South Africa, Western Cape, Tulbagh

AY677284

AY677229

JF735644

JF735833

Ilyonectria macrodidyma

CBS 112603; STE-U 4007; C 8

F. Halleen, 1999

Vitis vinifera, trunk of a plant showing decline symptoms, scion Sauvignon blanc; rootstock Richter 110

South Africa, Western Cape, Darling

AY677285

JF735469

JF735645

JF735834

Ilyonectria macrodidyma

CBS 112605; STE-U 3984; C 106

F. Halleen, 2000

Vitis vinifera, basal end of an asymptomatic nursery plant; scion Sultana; rootstock 143-B Mgt

South Africa, Western Cape, Malmesbury, Jakkalsfontein

AY677287

AY677230

JF735646

JF735835

Ilyonectria macrodidyma, holotype of C. macrodidymum

CBS 112615; STE-U 3976; C 98

F. Halleen, 2000

Vitis vinifera, roots, asymptomatic nursery grapevine plant scion Sultana; rootstock 143-B Mgt

South Africa, Western Cape, Malmesbury, Jakkalsfontein

AY677290

AY677233

JF735647

JF735836

Ilyonectria macrodidyma

Cy123

W.D. Gubler

Vitis vinifera

USA, California

JF735341

JF735470

JF735648

JF735837

Ilyonectria macrodidyma

Cy128

W.D. Gubler

Vitis vinifera

USA, California

JF735342

JF735471

JF735649

JF735838

Ilyonectria macrodidyma

Cy139

C. Rego & T. Nascimento, 2003

Vitis vinifera, basal end of a 1.5-year-old plant showing decline symptoms; scion Aragonez; rootstock 3309 C

Portugal, Estremoz

AM419071

AM419106

JF735650

JF735839

Ilyonectria macrodidyma

Cy140

C. Rego & T. Nascimento, 2003

Vitis vinifera, grafting zone of a 1.5-year-old plant showing decline symptoms; scion Aragonez; rootstock 3309 C

Portugal, Estremoz

JF735343

JF735472

JF735651

JF735840

Ilyonectria macrodidyma

Cy175

C. Rego, 2004

Vitis vinifera, basal discolouration in rootstocks; scion Touriga Nacional; rootstock 1103P

Portugal, Torre de Moncorvo

JF735344

JF735473

JF735652

JF735841

Ilyonectria macrodidyma

Cy181

C. Rego, 2005

Vitis vinifera, scion 140-Ru; rootstock Aragonês

Portugal, Alcácer do Sal

JF735345

JF735474

JF735653

JF735842

Ilyonectria macrodidyma

Cy216

A. Cabral, 2007

Vitis vinifera, asymptomatic; scion Marssanne

Portugal, Torres Vedras

JF735346

JF735475

JF735654

JF735843

Ilyonectria macrodidyma

Cy244

C. Rego, 2008

Vitis vinifera, basal end of a 2-year-old plant; scion Petit Verdot; rootstock 110R

Portugal, Vidigueira

JF735347

JF735476

JF735655

JF735844

Ilyonectria macrodidyma

Cy258

C. Rego, 2008

Vitis vinifera, basal end of a 2-year-old plant; scion Cabernet Sauvignon; rootstock 110R

Portugal, Vidigueira

JF735348

JF735477

JF735656

JF735845

Cylindrocarpon” sp.4

CBS 119.41

H.C. Koning

Fragaria sp., root

Netherlands, Baarn

JF735349

JF735478

JF735657

JF735846

Cylindrocarpon” sp.4

CBS 188.49

J.A. von Arx

Abies nordmanniana, root

Netherlands, Egmond

AM419063

AM419087

JF735658

JF735847

Cylindrocarpon” sp.4

CBS 112604; STE-U 4004; C 10

F. Halleen, 1999

Vitis vinifera, roots; scion Cabernet Sauvignon ; rootstock 101–14 Mgt

South Africa, Western Cape, Paarl

AY677286

AY677227

JF735659

JF735848

Cylindrocarpon” sp.4

CBS 112609; STE-U 3969; HJS-1217

M. Sweetingham, 1979

Vitis sp., dark brown discoloration in trunk; scion Cabernet Sauvignon

Australia, Tasmania, Bream Creek

AY677289

AY677226

JF735660

JF735849

Cylindrocarpon” sp.4

CBS 113555; STE-U 5715; HJS-1309; NZ C 60

R. Bonfiglioli, 2003

Vitis sp., blackening areas in wood and roots; scion Pinot Noir; rootstock 101-14

New Zealand, Fiddlers Green

JF735350

AY677234

JF735661

JF735850

Cylindrocarpon” sp.4

CBS 112598; STE-U 3997; C 115

F. Halleen, 2000

Vitis vinifera, roots of an asymptomatic plant; scion Sultana; rootstock Ramsey

South Africa, Western Cape, Wellington, Lelienfontein

JF735351

JF735479

JF735662

JF735851

Cylindrocarpon” sp.4

CPC 13533; CCFC 144524; Dias 2B

H.F. Dias, 1972

Vitis vinifera, Concord Bradt grapes, roots and stems

Canada, Ontario

AY295332

JF735480

JF735663

JF735852

Cylindrocarpon” sp.4

Cy69

C. Rego, 1999

Vitis vinifera, asymptomatic rootstocks; rootstock SO4, clone 102 F

Portugal, Ribatejo e Oeste

AJ875332

AM419095

JF735664

JF735853

Cylindrocarpon” sp.4

Cy71

C. Rego, 1999

Vitis vinifera, asymptomatic rootstocks; rootstock 99R, clone 96 F

Portugal, Ribatejo e Oeste

AJ875335

AM419096

JF735665

JF735854

Cylindrocarpon” sp.4

Cy72

C. Rego, 1999

Vitis vinifera, asymptomatic rootstocks; rootstock clone 113 F

Portugal, Ribatejo e Oeste

AJ875336

AM419097

JF735666

JF735855

Cylindrocarpon” sp.4

Cy75

C. Rego, 1999

Vitis vinifera, asymptomatic rootstocks; rootstock 99R

Portugal, Ribatejo e Oeste

AJ875334

AM419098

JF735667

JF735856

Cylindrocarpon” sp.4

Cy96

E. Halmschlager

Quercus sp., root

Austria, Patzmannsdorf

JF735352

JF735481

JF735668

JF735857

Cylindrocarpon” sp.4

Cy97

E. Halmschlager

Quercus sp., root

Austria, Patzmannsdorf

JF735353

JF735482

JF735669

JF735858

Cylindrocarpon” sp.4

Cy118

W.D. Gubler

Vitis vinifera

USA, California

JF735354

JF735483

JF735670

JF735859

Cylindrocarpon” sp.4

Cy120

W.D. Gubler

Vitis vinifera

USA, California

AJ875320

AM419101

JF735671

JF735860

Cylindrocarpon” sp.4

Cy132; IAFM Cy1-1

J. Armengol

Vitis vinifera

Spain, Alicante

JF735355

JF735484

JF735672

JF735861

Cylindrocarpon” sp.4

Cy136

C. Rego & T. Nascimento, 2003

Vitis vinifera, basal end of a 1.5-year-old plant showing decline symptoms; scion Aragonez; rootstock 3309 C

Portugal, Estremoz

JF735356

JF735485

JF735673

JF735862

Cylindrocarpon” sp.4

Cy137

C. Rego & T. Nascimento, 2003

Vitis vinifera, basal end of a 1.5-year-old plant showing decline symptoms; scion Aragonez; rootstock 3309 C

Portugal, Estremoz

AM419070

JF735486

JF735674

JF735863

Cylindrocarpon” sp.4

Cy138

C. Rego & T. Nascimento, 2003

Vitis vinifera, basal end of a 1.5-year-old plant showing decline symptoms; scion Aragonez; rootstock 3309 C

Portugal, Estremoz

JF735357

JF735487

JF735675

JF735864

Cylindrocarpon” sp.4

Cy141

C. Rego & T. Nascimento, 2003

Vitis vinifera, basal end of a 1.5-year-old plant showing decline symptoms; scion Aragonez; rootstock 3309 C

Portugal, Estremoz

JF735358

JF735488

JF735676

JF735865

Cylindrocarpon” sp.4

Cy142

C. Rego & T. Nascimento, 2003

Vitis vinifera, grafting zone of a 1.5-year-old plant showing decline symptoms; scion Aragonez; rootstock 3309 C

Portugal, Estremoz

JF735359

JF735489

JF735677

JF735866

Cylindrocarpon” sp.4

Cy143

C. Rego & T. Nascimento, 2003

Vitis vinifera, basal end of a 1.5-year-old plant showing decline symptoms; scion Aragonez; rootstock 3309 C

Portugal, Estremoz

JF735360

JF735490

JF735678

JF735867

Cylindrocarpon” sp.4

Cy157

H. Oliveira, 2004

Vitis vinifera, scion Touriga Nacional; rootstock 99R

Portugal, Alenquer

AM419077

AM419110

JF735679

JF735868

Cylindrocarpon” sp.4

Cy214

A. Cabral, 2007

Vitis vinifera, asymptomatic; scion Grenache

Portugal, Torres Vedras

JF735361

JF735491

JF735680

JF735869

Cylindrocarpon” sp.4

CBS 129086; Cy218

A. Cabral, 2007

Vitis vinifera, asymptomatic; scion Chenin

Portugal, Torres Vedras

JF735362

JF735492

JF735681

JF735870

Cylindrocarpon” sp.4

Cy221

L. Leandro

Fragaria x ananassa

USA, North Caroline, Asheville

JF735363

JF735493

JF735682

JF735871

Cylindrocarpon” sp.4

Cy222

L. Leandro

Fragaria x ananassa

USA, North Caroline, Asheville

JF735364

JF735494

JF735683

JF735872

Cylindrocarpon” sp.4

Cy223

L. Leandro

Fragaria x ananassa

USA, North Caroline, Asheville

JF735365

JF735495

JF735684

JF735873

Cylindrocarpon” sp.4

Cy235

C. Rego, 2008

Vitis vinifera, basal end of a 2-year-old plant; scion Cabernet Sauvignon; rootstock 110R

Portugal, Vidigueira

JF735366

JF735496

JF735685

JF735874

Cylindrocarpon” sp.4

Cy237

C. Rego, 2008

Vitis vinifera, basal end of a 2-year-old plant; scion Chardonnay; rootstock 110R

Portugal, Vidigueira

JF735367

JF735497

JF735686

JF735875

Cylindrocarpon” sp.4

Cy240

C. Rego, 2008

Vitis vinifera, basal end of a 2-year-old plant; scion Touriga Nacional; rootstock 140RU

Portugal, Vidigueira

JF735368

JF735498

JF735687

JF735876

Cylindrocarpon” sp.4

Cy246

C. Rego, 2008

Vitis vinifera, basal end of a 2-year-old plant; scion Antão Vaz; rootstock 110R

Portugal, Vidigueira

JF735369

JF735499

JF735688

JF735877

Cylindrocarpon” sp.4

Cy260

C. Rego, 2008

Vitis vinifera, basal end of a 2-year-old plant; scion Cabernet Sauvignon; rootstock 110R

Portugal, Vidigueira

JF735370

JF735500

JF735689

JF735878

Cylindrocarpon” sp.4

Cy262

C. Rego, 2008

Vitis vinifera, basal end of a 2-year-old plant; scion Cabernet Sauvignon; rootstock 110R

Portugal, Vidigueira

JF735371

JF735501

JF735690

JF735879

Neonectria major, type strain

CBS 240.29; IMI 113909

H.W. Wollenweber

Alnus incana, canker

Norway

JF735308

DQ789872

JF735593

JF735782

Neonectria ditissima, authentic strain of C. willkommii

CBS 226.31; IMI 113922

H.W. Wollenweber

Fagus sylvatica

Germany, Tharandt

JF735309

DQ789869

JF735594

JF735783

Neonectria ditissima, representative strain of N. galligena

CBS 835.97

W. Gams, 1997

Salix cinerea, dead branch of still living tree

Belgium, Marais de Sampant

JF735310

DQ789880

JF735595

JF735784

Neonectria ditissima

Cy169

H. Oliveira, 1997

Malus domestica

Portugal, Alcobaça

AM419080

AM419113

JF735596

JF735785

Neonectria ditissima

Cy172

T. Nascimento, 2004

Malus domestica; scion Oregon; rootstock MM107

Portugal, Caldas da Rainha

AM419081

AM419114

JF735597

JF735786

Neonectria neomacrospora, representative strain

CBS 118984; GJS 03-28

L. Reitman, 2005

Arceuthobium tsugense, parasiting Abies balsams

Canada, British Columbia, Vancouver Island, Spider Lake

JF735311

DQ789882

JF735598

JF735787

Cylindrocarpon cylindroides, representative strain

CBS 324.61; DSM 62489; IMB 9628

J.A. von Arx

Abies concolor

Netherlands, Zwolle

JF735312

DQ789875

JF735599

JF735788

Cylindrocarpon cylindroides

CBS 503.67

F. Roll-Hansen

Abies alba, wood

Norway, Hordaland, Fana

AY677261

JF735436

JF735600

JF735789

Cylindrocarpon sp.

CPC 13545; DAOM 185212; # 5

J.A. Traquair & B. Harrison, 1982

Pyrus sp.

Canada, Ontario, Harrow

AY295303

JF735437

JF735601

JF735790

Neonectria ramulariae, authentic strain of C. obtusiusculum (=C. magnusianum)

CBS 151.29; IMI 113894; MUCL 28083; MUCL 28094

H.W. Wollenweber

Malus sylvestris, fruit

UK, England, Cambridge

JF735313

JF735438

JF735602

JF735791

Neonectria ramulariae

CBS 182.36; IMI 113893; UPSC 1903

H.W. Wollenweber

Malus sylvestris, fruit

-

JF735314

JF735439

JF735603

JF735792

Cylindrocarpon sp.

CR21

P. Axelrood

Pseudotsuga menziesii

Canada, British Columbia

JF735315

JF735440

JF735604

JF735793

Cylindrocarpon sp.

CPC 13530; DAOM 185722; JAT 1591

J.A. Traquair, 1983

Pyrus sp., lesions on seedlings

Canada, Ontario, Harrow

AY295302

JF735441

JF735605

JF735794

Cylindrocarpon sp.

CPC 13531; CCFC 226722; DAOM 226722; CR6

P. Axelrood

Pseudotsuga menziesii, root

Canada, British Columbia

AY295301

JF735442

JF735606

JF735795

Cylindrocarpon obtusisporum

CBS 183.36; IMI 113895

H.W. Wollenweber, 1936

Solanum tuberosum, tuber

Germany

AM419061

AM419085

JF735607

JF735796

Cylindrocarpon obtusisporum

CPC 13544; DAOM 182772; JAT 1366

J.A. Traquair, 1982

Prunus armenica, twigs

Canada, Ontario, Ruthven

AY295306

JF735443

JF735608

JF735797

Cylindrocarpon obtusisporum

94-1356

R. C. Hamelin, 1994

Picea mariana

Canada, Quebec

AY295304

JF735444

JF735609

JF735798

a ATCC American Type Culture Collection, USA; CBS CBS-KNAW Fungal Biodiversity Centre (Centraalbureau voor Schimmelcultures), Utrecht, The Netherlands; CCFC Canadian Collection of Fungal Cultures, Agriculture and Agri-Food Canada, Ottawa, Canada; CPC Culture collection of Pedro Crous, housed at CBS; Cy Cylindrocarpon collection housed at Laboratório de Patologia Vegetal “Veríssimo de Almeida” - ISA, Lisbon, Portugal; DAOM Agriculture and Agri-Food Canada National Mycological Herbarium, Canada; DSM Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH, Braunschweig, Germany; GJS Gary J. Samuels collection; HJS Hans-Josef Schroers collection; IAFM Instituto Agroforestal Mediterráneo, Universidad Politécnica de Valencia, Spain; IAM Institute of Molecular and Cellular Biosciences, The University of Tokyo, Japan; ICMP International Collection of Microorganisms from Plants, Auckland, New Zealand; IFFF Institute of Forest Entomology, Forest Pathology and Forest Protection, Austria; IMI International Mycological Institute, CABI-Bioscience, Egham, Bakeham Lane, U.K.; JAT J. A. Traquair collection; JCM Japan Collection of Microorganisms, Japan; KIS Agricultural Institute of Slovenia, Ljubljana, Slovenia; LYN Lynchburg College, Biology Department, USA; MUCL Mycothèque de l’Université Catholique de Louvain, Belgium; NBRC NITE Biological Resource Center, Japan; NRRL Agricultural Research Service Culture Collection, USA; STE-U Stellenbosch University, South Africa; UPSC Fungal Culture Collection at the Botanical Museum, Uppsala University, Uppsala, Sweden; VKM All-Russian Collection of Microorganisms, Russia.

Besides those, 10 “Cylindrocarpon” spp. isolates were obtained in Portugal from grapevine plants showing decline symptoms, either 1- to 6-year-old plants in vineyards (Cy22, Cy155, Cy158, Cy190, CBS 129078, CBS 129080, CBS 129081, CBS 129082) or from rootstock nurseries (Cy23), and from a 25-year-old grapevine plant with esca symptoms (CBS 129084). Furthermore, isolates were obtained from a young Malus domestica (Cy164) and from the stem of a young Quercus suber (Cy232) plant, both showing decline symptoms, and from Thymus sp. (Cy231) and Ficus sp. (Cy228). One isolate (Cy131) was made available by P. Lecomte (Institut National de la Recherche Agronomique, Bordeaux-Aquitaine, France) and was obtained from an internal lesion of a stem of Actinidia chinensis ‘Hayward’. Another isolate (Cy122) was made available by W.D. Gubler (University of California, Davis, USA) and was obtained from Vitis sp. All of these isolates are stored in a culture collection at the Laboratório de Patologia Vegetal “Veríssimo de Almeida” (LPVVA-ISA, Lisbon, Portugal).

An additional 25 “C.destructans isolates used during this study were made available by K.A. Seifert (Agriculture and Agri-Food, Canada), and were isolated from commercial Panax quinquefolium gardens (CBS 120359–120369, CBS 129079, CBS 129083, CD1666, CPC 13535, CPC 13537, NSAC-SH2, NSAC-SH2.5), Picea glauca (94–1628, CPC 13539), Poa pratensis (CPC 13534), Pseudotsuga menziesii (CBS 120370–120372, CPC 13536) and Prunus cerasus (CPC 13532) (Seifert et al. 2003).

Another 109 isolates were also included in the analysis to add phylogenetic support to this study and represent strains of the following taxa: C. cylindroides, C. obtusisporum, C. pauciseptatum, species 1 to 6 (Mostert et al., in preparation; Cabral et al., in preparation), I. macrodidyma, N. ditissima, N. major, N. neomacrospora and N. ramulariae.

DNA isolation, sequencing and phylogenetic analysis

For each isolate, genomic DNA was isolated from mycelium following the protocol of Möller et al. (1992), adapted by Crous et al. (2009b). Sequencing of the ITS and part of the β-tubulin (TUB), histone H3 (HIS) and translation elongation factor 1-α (TEF) genes was performed after PCR amplification using 1× PCR buffer (Bioline, London, UK), 1.5 mM MgCl2, 32 μM of each dNTPs, 0.24 μM of each primer, 0.5 units Taq DNA Polymerase (Bioline), and 1 μl of diluted gDNA in a final volume of 12.5 μl. The cycle conditions in a iCycler thermocycler (BioRad, Hercules, USA) were 94°C for 5 min, followed by 40 cycles at 94°C for 30 s, 52°C for 30 s and 72°C for 80 s, and a final elongation at 72°C for 10 min. Primers were V9G (de Hoog and Gerrits van den Ende 1998) and ITS4 (White et al. 1990) for ITS, T1 (O’Donnell and Cigelnik 1997) and Bt-2b (Glass and Donaldson 1995) for TUB, CYLH3F and CYLH3R (Crous et al. 2004b) for HIS, and EF1 and EF2 (O’Donnell et al. 1998) or CylEF-1 (5’- ATG GGT AAG GAV GAV AAG AC-3’; J.Z. Groenewald, unpublished) and CylEF-R2 (Crous et al. 2004b) for TEF. For TEF, the following modifications were made to the amplification protocol: 2.0 mM of MgCl2, 40 μM of each dNTPs and addition of 5% of Dimethyl sulfoxide (DMSO; Sigma-Aldrich, Zwijndrecht, Netherlands).

After confirmation by agarose gel electrophoresis, amplicons were sequenced in both directions with the corresponding PCR primers and a DYEnamic ET Terminator Cycle Sequencing Kit (Amersham Biosciences, Diegem, Belgium) according to the manufacturer’s recommendations. The products were analysed on an ABI Prism 3700 DNA Sequencer (Perkin-Elmer, Waltham, USA).

Sequences were assembled and edited to resolve ambiguities, using the EditSeq and SeqMan modules of the Lasergene software package (DNAStar, Madison, USA). Consensus sequences for all isolates were compiled into a single file (Fasta format) and aligned using CLUSTAL X v. 2.0.11 (Larkin et al. 2007). Following manual adjustment of the alignment by eye where necessary, the alignment was subjected to phylogenetic analyses as described by Crous et al. (2004b). Novel sequences were lodged in GenBank (Table 1), taxonomic novelties in MycoBank (Crous et al. 2004a), and the alignments and phylogenetic trees in TreeBASE (http://www.treebase.org).

Morphology

Isolates were grown for up to 5 weeks at 20°C on synthetic nutrient poor agar (SNA; Nirenberg 1976) with and without two 1-cm2 filter paper pieces, carnation leaf agar (CLA; Crous et al. 2009b), potato-dextrose agar (PDA; Difco, Detroit, USA) and oatmeal agar (OA; Crous et al. 2009b) under continuous n-UV light (NUV, 400–315 nm; Blacklight-Blue; Sylvania, Capelle a/d Ijssel, Netherlands).

Measurements were done on a 1-cm2 agar plug removed from the colony margin, placed on a microscope slide, to which a drop of water and coverslip were added. For each isolate, 30 measurements were obtained for each structure. Measurements were done at ×1,000 magnification using a Nikon Eclipse 80i microscope, or a Leica DM2500. Images were captured using a Nikon DS-Fi1 digital camera with NIS-Elements Software, or a Leica DFC295 digital camera with the Leica Application Suite. Measurements for length and width of conidia and ascospores are given as (Minimum) Lower Limit of a 95% Confidence Interval – Upper Limit of a 95% Confidence Interval (Maximum). For other measurements, only the extreme values are given.

Culture characteristics (texture, density, colour, growth front, transparency and zonation) were described on PDA after incubation at 20°C in the dark for 14 days. Colour (surface and reverse) was described using the colour chart of Rayner (1970). Cardinal temperatures for growth were assessed by inoculating 90-mm-diam PDA dishes with a 3-mm-diam plug cut from the edge of an actively growing colony. Growth was determined after 7 days in two orthogonal directions. Trials were conducted at various temperatures (4, 10, 15, 18, 20, 22, 25, 30 and 35°C) with three replicate plates per strain at each temperature.

To induce the formation of perithecia, isolates were crossed in 60-mm-diam Petri dishes containing a minimal salts medium supplemented with two sterile birch toothpicks (Guerber and Correll 2001). The plates were incubated at 20°C under n-UV light for 8–20 weeks. Two strains were considered sexually compatible if perithecia were formed that exuded masses of viable ascospores. The colour reaction of the perithecia was checked in 3% KOH and in lactic acid. For sectioning, perithecia were mounted in Jung Tissue Freezing Medium (Leica) or in Arabian Gum, and cut in 10- to 15-μm-thick sections using a Leica cryostat CM3050 S or CM1850 at −20°C.

Results

Phylogeny

Amplification products of approximately 700 bases (ITS), 650 bases (TUB), 500 bases (HIS) and 600–800 bases (TEF) were obtained for the isolates listed in Table 1. The manually adjusted combined alignment contains 189 sequences (including the two outgroup sequences) and the statistical parameters for the combined and individual analyses are presented in Table 2. For the combined analysis, only a maximum of 1,000 equally most parsimonious trees were saved, the first of which is presented as Fig. 1. Phylogenetic trees derived from the individual loci are available in TreeBASE. The combined analysis of the four genes enabled the identification of 37 species. However, the analysis of HIS data alone was enough to resolve these taxa. Sequences of TEF could not distinguish species 6, I. robusta, I. europaea, I. lusitanica, I. rufa and N. ditissima; whereas sequences of TUB could not separate I. robusta, species 4, and 6, while “I.” macrodidyma, species 5, I. liliigena and I. pseudodesctructans were supported by low bootstrap values, and CBS 120370 clustered apart from the remaining isolates of I. crassa. Of all loci screened, ITS proved to be the least informative, being unable to resolve 22 of the species in this study. Neighbour-Joining (NJ) analyses using the three substitution models, as well as the parsimony analysis, yielded trees with similar topology and bootstrap support values for the individual and combined gene analyses. The trees obtained supported the same clades, sometimes with rearrangements in the order of these clades between the different analyses (data not shown). The results of the phylogenetic analyses are highlighted below under the taxonomic notes or in the Discussion, where applicable.
Table 2

Statistical information on the individual datasets and number of equally most parsimonious trees for each locus [Internal Transcribed Spacers (ITS) of the nuclear ribosomal RNA gene operon, and partial β-tubulin (TUB), histone H3 (HIS) and translation elongation factor 1-α (TEF) genes]

 

ITS

TUB

HIS

TEF

Combined

Aligned characters (including gaps)

475

502

440

696

2,113

Parsimony-informative characters

122

212

215

364

913

Variable and parsimony-uninformative characters

31

38

11

43

123

Constant characters

322

252

214

289

1,077

Equally most parsimonious trees obtained

136

384

1

60

1,000

Tree length

294

603

1,095

1,149

3,259

Consistency index (Cl)

0.718

0.660

0.468

0.611

0.559

Retention index (RI)

0.978

0.972

0.946

0.966

0.959

Rescaled Consistency index (RC)

0.702

0.642

0.442

0.590

0.537

Fig. 1

The first of 1,000 equally most parsimonious trees obtained from the combined ITS, TUB, HIS and TEF sequence alignment of Cylindrocarpon isolates and relatives with a heuristic search using PAUP v. 4.0b10. The tree was rooted using Campylocarpon isolates as outgroup sequences and bootstrap support values are indicated near the nodes, where”ns” designates not supported. Ex-type strains are indicated in bold. Newly described species are indicated by blue boxes. Scale bar shows 10 changes

Taxonomy

The present study treats isolates that have been freshly collected, or previously identified and maintained in culture collections as “Cylindrocarpon destructans”, meaning cylindrical, rarely curved, 3-septate macroconidia with obtuse apices, abundant microconidia and chlamydospores (Samuels and Brayford 1990). The latter species has in the past been acknowledged as anamorph of I. radicicola (Booth 1966; Chaverri et al. 2011; Samuels and Brayford 1990). However, an examination of the neotype of “C.” destructans in this study [CUP-011985, conidia (18.0)23.0–30.0(35.0) × (6.0)6.5(7.0) μm], found conidia to be considerably smaller than those of I. radicicola (24.0)33.1(47.0) × (4.9)6.4(7.8) μm (Gerlach and Nilsson 1963) (also confirmed in the present study by examination of CBS 264.65, ex-type), revealing them to represent two distinct species. Furthermore, based on the phylogenetic and morphological data obtained in the present study, several novel species could be distinguished that are phylogenetically distinct from I. radicicola, and morphologically distinct based on a range of characters linked to culture characteristics, conidiophores, macro- and microconidium morphology. Some of these could be linked to older names, or taxa long regarded as potential syonyms of “destructans”, which could now be resurrected. These taxa are treated below:

Ilyonectria anthuriicola A. Cabral & Crous, sp. nov. (Fig. 2)

MycoBank 560108.
Fig. 2

Ilyonectria anthuriicola (CBS 564.95). ac Simple conidiophores on aerial mycelium. dg Conidiophores giving rise to microconidia, formed on mycelium at agar surface, penicillately mono- or bi-verticillate. hl Micro- and macroconidia. m Chlamydospores in mycelium. Bars 10 μm

Etymology: Named after its host, Anthurium.

Cylindrocarpi destructantis morphologice simile, sed longitudine media conidiorum longiore, 29.5–32.2 μm, distinguitur.

Conidiophores simple or complex to sporodochial. Simple conidiophores arising laterally or terminally from aerial mycelium, solitary to loosely aggregated, unbranched or sparsely branched bearing up to three phialides, 1–3-septate, 40–95 μm long; phialides monophialidic, more or less cylindrical but slightly tapering towards the tip, 10.5–20.5 μm long, 2.5–3.5 μm wide at the base, 3.0–4.5 μm at widest point, 1.5–2.5 μm near the aperture. Conidiophores giving rise to microconidia, formed on mycelium at agar surface, penicillately mono- or bi-verticillate; phialides monophialidic, narrowly flask-shaped, typically with widest point near the middle, 8–15 μm long, 2.0–3.0 μm wide at the base, 2.5–4.5 μm at widest point, 1.0–2.0 μm near the apex. Sporodochial conidiophores irregularly branched; phialides cylindrical, mostly widest near the middle. Macroconidia formed in flat domes of slimy masses, (1–)3-septate, straight or minutely curved, cylindrical with both ends more or less obtusely rounded, mostly without a visible hilum; 1-septate, (20.0)23.5–26.7(29.0) × (5.5)5.9–6.8(7.0) μm (average = 25.1 × 6.4 μm), with a length:width ratio of 3.6–4.8; 2-septate, (25.0)26.6–29.3(32.0) × (6.5)6.8–7.8(8.5) μm (av. = 27.9 × 7.3 μm), with a length:width ratio of 3.2–4.8; 3-septate, (25.0)29.5–32.2(38.0) × (6.0)7.5–8.1(9.0) μm (av. = 30.8 × 7.8 μm) with a length:width ratio of 3.1–5.2. Microconidia 0(−1)-septate, subglobose to ovoid, rarely ellipsoid, mostly with a visible centrally located or slightly laterally displaced hilum; aseptate microconidia, (4.9)5.0–8.1(12.0) × (4.0)4.3–5.5(6.5) μm (av. = 6.5 × 4.9 μm), with a length:width ratio of 1.0–1.8; 1-septate, (11.0)11.6–16.7(18.0) × (5.0)5.4–6.1(6.0) μm (av. = 14.1 × 5.8 μm), with a length:width ratio 1.8–3.0. Chlamydospores globose to subglobose to ellipsoid, 8–14 × 7–12 μm, smooth, but often appearing rough due to deposits, thick-walled, formed intercalary in chains or in clumps and also in the cells of macroconidia, hyaline, becoming golden-brown.

Holotype: Netherlands, Bleiswijk, root rot of Anthurium sp., 1995, coll./isol. R. Pieters, holotype CBS H-20555, culture ex-type CBS 564.95.

Culture characteristics: Mycelium felty with average density. Surface on OA chestnut, with aerial mycelium sparse, saffron; margin pure yellow to orange. Surface on PDA, chestnut with saffron aerial mycelium, growth at margin luteous; zonation absent, transparency homogeneous, margin even; reverse similar to surface, but chestnut to cinnamon on OA, and chestnut on PDA. Colonies on PDA do not grow at 4°C after 7 days. Optimum temperature 20°C when colonies reach 25–27 mm, after 7 days. Colony diam was 20–22 mm at 25°C, after 7 days. Hardly grows at 30°C (2 mm colony diam after 7 days).

Isolate studied: CBS 564.95 (Table 1).

Host and distribution: Roots of Anthurium sp. (Netherlands).

Ilyonectria crassa (Wollenw.) A. Cabral & Crous, comb. et stat. nov. (Fig. 3)

MycoBank 560109.
Fig. 3

Ilyonectria crassa (CBS 129083). ac Simple conidiophores on aerial mycelium. dg Micro- and macroconidia. hi Chlamydospores and macroconidia. Bars 10 μm

Basionym: Cylindrocarpon radicicola var. crassum Wollenw., Z. Parasitenkunde 3: 495. 1931.

Cylindrocarpon destructans var. crassum (Wollenw.) C. Booth, Mycol. Pap. 104: 37. 1966.

Conidiophores simple or complex, to sporodochial. Simple conidiophores arising laterally or terminally from aerial mycelium, solitary to loosely aggregated, unbranched or sparsely branched bearing up to two phialides, rarely consisting only of phialides, 1–4-septate, 40–180 μm long; phialides monophialidic, cylindrical to subulate, 20–55 μm long, 2.5–4.0 μm wide at the base, 1.5–2.0 μm near the apex. Complex conidiophores aggregated in small sporodochia (on carnation leaf), repeatedly and irregularly branched; phialides more or less cylindrical, but tapering slightly in the upper part towards the apex, or narrowly flask-shaped, mostly with widest point near the middle, 17–24 μm long, 2.0–3.0 μm wide at the base, 2.5–3.5 μm at the widest point, and 1.5–2.5 μm wide near the apex. Macroconidia predominating, formed on both type of conidiophores, on SNA formed in flat domes of slimy masses, 1–3-septate, straight, cylindrical, but may narrow towards the tip, more or less broadly rounded, and the base appearing somewhat acute due to the presence of the hilum, mostly centrally located; 1-septate, (21.0)25.7–27.3(34.0) × (4.5)5.0–5.3(6.5) μm (av. = 26.5 × 5.1 μm), with a length:width ratio of 3.8–6.7; 2-septate, (23.0)28.5–30.3(37.0) × (4.5)5.3–5.6(6.5) μm (av. = 29.4 × 5.4 μm)with a length:width ratio of 4.2–6.7; 3-septate, (29.0)34.1–36.0(49.0) × (5.0)5.6–5.8(7.0) μm (av. = 35.1 × 5.7 μm), with a length:width ratio of 4.8–8.9. Microconidia 0–1-septate, ellipsoid to subcylindrical, more or less straight, with a visible, truncate hilum; aseptate microconidia, (7.0)9.7–10.9(15.0) × (3.0)3.3–3.6(4.5) μm (av. = 10.3 × 3.5 μm), with a length:width ratio of 1.8–4.3; 1-septate, (12.0)14.2–15.2(19.0) × (3.0)3.8–4.2(5.0) μm (av. = 14.7 × 4.0 μm), with a length:width ratio 2.7–5.0. Conidia formed in heads on simple conidiophores or as white (OA) or unpigmented (SNA) masses. Chlamydospores globose to subglobose to cylindrical, 7–15 × 6–10 μm, smooth, but often appearing rough due to deposits, thick-walled, terminal on short lateral branches, rarely intercalary, single, in chains or in clumps, and also in the cells of the macroconidia, hyaline, becoming pale brown.

Lectotype: The Netherlands, on Lilium bulbs, Dec. 1930, coll./isol. W.F. van Hell, lectotype designated here CBS H-20556, culture ex-lectotype CBS 139.30.

Culture characteristics: Mycelium cottony to felty with average to strong density. Surface on OA cinnamon, with aerial mycelium sparse, buff. Surface on PDA saffron with aerial mycelium sparse buff to saffron to pale luteous. No zonation was observed, transparency was homogeneous and growth at margin even. Reverse similar to surface, except in colour, saffron to cinnamon on OA, and chestnut to sienna on PDA. Colonies on PDA grow 5–8 mm diam at 4°C after 7 days. Optimum temperature at 20°C, when colonies reach 31–46 mm diam, after 7 days. Colony diam was 19–34 mm at 25°C, after 7 days. No growth was observed at 30°C.

Isolates studied: CBS 139.30; CBS 158.31; CBS 129083; NSAC-SH-2; NSAC-SH-2.5 (Table 1).

Hosts and distribution: Lilium sp. (bulbs), Narcissus sp. (roots) (Netherlands), Panax quinquefolium (roots) (Canada).

Notes: In the original description, Wollenweber (1931) cites Cylindrocarpon radicicola var. crassum as occurring on roots of Ulmus, Taxus and Lilium in Europe (Germany and the Netherlands). He did not designate any type specimen. However, he specifically refers to a culture sent to him by Prof. J. Westerdijk on Lilium from the CBS in the Netherlands in 1930, which was regarded as authentic for the species. This culture is represented by CBS 139.30 (accessioned in 1930, from Lilium, the Netherlands), and thus we designate a dried, sporulating culture as lectotype for the species.

Ilyonectria cyclaminicola A. Cabral & Crous, sp. nov. (Fig. 4)

MycoBank 560110.
Fig. 4

Ilyonectria cyclaminicola (CBS302.93). a, b Simple conidiophores on aerial mycelium. c Pennicilate conidiophores with aseptate microconidia. d Sporodochial conidiophore on carnation leaf agar. e Isolated chlamydospores formed in lateral branches. fj Micro- and macroconidia. Bars 10 μm

Etymology: Named after the host from which it was isolated, Cyclamen sp.

Cylindrocarpi destructantis morphologice simile, sed longitudine media conidiorum longiore, 26.9–31.9 μm, distinguitur.

Conidiophores simple or complex to sporodochial. Simple conidiophores arising laterally or terminally from aerial mycelium, solitary to loosely aggregated, unbranched or sparsely branched, bearing up to two phialides, 1–3-septate, 60–120 μm long; phialides monophialidic, more or less cylindrical but slightly tapering towards the tip, 20–60 μm long, 2.0–4.0 μm wide at the base, 3.0–4.5 μm at widest point, 1.5–2.5 μm near the aperture. Conidiophores giving rise to microconidia formed by mycelium at agar surface, penicillate to mono-verticillate; phialides monophialidic, more or less cylindrical, but with slight taper towards the tip, 19–34 μm long, 1.5–2.5 μm wide at the base, 2.0–3.0 μm at widest point, 1.0–2.0 μm near the apex. Sporodochial conidiophores irregularly branched; phialides more or less cylindrical, but slightly tapering towards the tip, or narrowly flask-shaped, with widest point near the base, 14–26 μm long, 2.5–3.5 μm wide at the base 3.0–4.0 μm at widest point, 1.0–2.0 μm near the apex. Macroconidia formed in flat domes of slimy masses, 1(−3)-septate, straight or minutely curved, cylindrical with both ends more or less broadly rounded, sometimes with a constriction at the septa, mostly without a visible hilum; 1-septate, (19.2)21.3–23.6(29.8) × (4.4)5.4–6.0(7.3) μm (av. = 22.5 × 5.7 μm), with a length:width ratio of 3.4–5.5; 2-septate, (23.8)24.0–28.4(29.8) × (5.0)5.5–7.3(8.0) μm (av. = 26.2 × 6.4 μm), with a length:width ratio of 3.1–5.1; 3-septate, (25.3)26.9–31.9(33.6) × (5.8)5.9–6.5(6.9) μm (av. = 29.4 × 6.2 μm), with a length:width ratio of 3.7–5.6. Microconidia formed in heads or on the agar surface, 0–1-septate, subglobose to ovoid to subcylindrical, mostly with a visible, centrally located or slightly laterally displaced hilum; aseptate microconidia, (3.9)7.6–8.9(12.9) × (2.2)3.6–3.9(5.4) μm (av. = 8.2 × 3.7 μm), with a length:width ratio of 1.2– 3.4; 1-septate, (11.5)13.8–15.2(17.5) × (3.7)4.6–4.9(5.5) μm (av. = 14.5 × 4.7 μm), with a length:width ratio of 2.3–3.9. Chlamydospores globose to subglobose, 10–18 × 10–16 μm, smooth, but often appearing rough due to deposits, thick-walled, formed in lateral branches, rarely intercalary, mostly isolated, hyaline, becoming medium brown.

Holotype: Netherlands, Roelofarendsveen, NAKS laboratory, Cyclamen bulb, May 1993, coll./isol. M. Hooftman, iden. E.J. Hermanides-Nijhof, holotype CBS H-20557, culture ex-type CBS 302.93.

Culture characteristics: Mycelium felty with average density. Surface on OA sepia to chestnut. Surface on PDA sepia to chestnut, with sparse, rust, aerial mycelium; no zonation was observed, and transparency was homogeneous; margins predominantly even. Reverse similar to surface, except in colour, sepia to dark brick on OA and chestnut on PDA. Colonies on PDA do not grow at 4°C after 7 days. Optimum temperature at 22°C, when colonies reach 68–70 mm diam, after 7 days. Colony diam was 63–64 mm at 25°C, after 7 days. No growth was observed at 30°C.

Isolate studied: CBS 302.93 (Table 1).

Host and distribution: Bulb of Cyclamen sp. (Netherlands).

Ilyonectria europaea A. Cabral, Rego & Crous, sp. nov. (Fig. 5)

MycoBank 560103.
Fig. 5

Ilyonectria europaea (CBS 129078). ac Simple conidiophores on aerial mycelium. d Sporodochial conidiophore on carnation leaf agar. e Chlamydospores in aerial mycelium. fi Micro- and macroconidia. Bars 10 μm

Etymology: Named after the European continent, where this fungus appears to be widely distributed.

Ilyonectriae robustae morphologice similis, sed longitudine media macroconidiorum breviore, 29.7–31.5 μm, distinguitur.

Conidiophores simple or complex to sporodochial. Simple conidiophores arising laterally or terminally from aerial mycelium, solitary to loosely aggregated, unbranched or sparsely branched, bearing up to three phialides, 1–3-septate, 50–120 μm long; phialides monophialidic, cylindrical to subulate, 26–60 μm long, 2.5–3.5 μm wide at the base, 1.5–2.5 μm near the apex. Complex conidiophores aggregated in small sporodochia (on carnation leaf), repeatedly and irregularly branched. Macroconidia predominating, formed on both type of conidiophores, on SNA formed in flat domes of slimy masses, 1(−3)-septate, straight or minutely curved, cylindrical with both ends more or less broadly rounded, but may narrow towards the tip, mostly without a visible hilum; 1-septate, (16.4)21.9–23.4(34.0) × (4.0)5.2–5.6(7.8) μm (av. = 22.7 × 5.4 μm), with a length:width ratio of 3.2–5.4; 2-septate, (22.0)26.4–28.1(34.0) × (4.4)5.9–6.4(8.0) μm (av. = 27.2 × 6.1 μm), with a length:width ratio of 3.4–6.4; 3-septate, (22.0)29.7–31.5(40.0) × (5.0)6.5–6.9(8.6) μm (av. = 30.6 × 6.7 μm), with a length:width ratio of 3.5–6.0. Microconidia 0–1-septate, ellipsoid to ovoid, more or less straight, without a visible hilum; aseptate microconidia sometimes curved towards one end, (3.0)8.5–9.8(17.0) × (1.7)3.3–3.5(5.0) μm (av. = 9.1 × 3.4 μm), with a length:width ratio of 1.5–3.4; 1-septate, (9.2)13.4–14.6(18.9) × (3.0)4.0–4.4(5.9) μm (av. = 14.0 × 4.2 μm), with a length:width ratio 2.6–4.0. Conidia formed in heads or on simple conidiophores as white (OA) or unpigmented (SNA) masses. Chlamydospores globose to subglobose, 9–14 × 7–14 μm, smooth, but often appearing rough due to deposits, thick-walled, terminal on short or long lateral branches or intercalary, single, in chains or in clumps, golden-brown.

Holotype: Portugal, Vidigueira, at basal end of a 2-year-old Vitis vinifera plant; scion Petit Verdot, rootstock 110R, 2008, coll./isol. C. Rego, holotype CBS H-20558, culture ex-type CBS 129078 = Cy241 = CPC 19165.

Culture characteristics: Mycelium felty with average density. Surface on OA chestnut, with saffron aerial mycelium. Sienna to saffron on PDA, with luteous aerial mycelium. Concentric zonation, with homogeneous transparency, margins predominantly even. Reverse similar to surface, except in the colour; sepia on OA, and chestnut to umber on PDA. Colonies on PDA grow poorly, 1–5 mm diam at 4°C after 7 days. Optimum temperature for growth is 22°C, when colonies reach 43–57 mm diam, after 7 days. Colony diam was 37–47 mm at 25°C, after 7 days. No growth was observed at 30°C.

Isolates studied: Cy131; Cy155; CBS 537.92; CBS 102892; CBS 129078 (Table 1).

Hosts and distribution: Actinidia chinensis ‘Hayward’ (internal lesion of stem) (France), Aesculus hippocastanum (wood) (Belgium), Phragmites australis (stem) (Germany), Vitis vinifera (Portugal).

Ilyonectria gamsii A. Cabral & Crous, sp. nov. (Fig. 6)

MycoBank 560112.
Fig. 6

Ilyonectria gamsii (CBS 940.97). ac Simple conidiophores on aerial mycelium. dh Micro- and macroconidia. i Chlamydospores on mycelium. Bars 10 μm

Etymology: Named after Prof. dr. Walter Gams, who has made a major contribution to our knowledge of Hypocrealean soil fungi.

Ilyonectriae panacis morphologice similis, sed longitudine media macroconidiorum breviore, 34.3–38.5 μm, distinguitur.

Conidiophores simple or complex to sporodochial. Simple conidiophores arising laterally or terminally from aerial mycelium, solitary to loosely aggregated, unbranched or sparsely branched, bearing up to two phialides, 1–3-septate, 50–150 μm long; phialides monophialidic, cylindrical to subulate, 30–60 μm long, 2.5–3.5 μm wide at the base, 1.5–2.0 μm near the aperture. Sporodochial conidiophores irregularly branched; phialides cylindrical, mostly widest near the base. Macroconidia predominating, formed on simple conidiophores, on SNA formed in flat domes of slimy masses, 1–3-septate, straight, cylindrical with both ends broadly rounded, with mostly visible, centrally located hilum; 1-septate, (22.0)25.7–27.9(33.0) × (4.0)5.1–5.5(6.0) μm (av. = 26.8 × 5.3 μm), with a length:width ratio of 4.3–6.2; 2-septate, (25.0)28.2–31.7(39.0) × (5.0)5.5–5.9(6.5) μm (av. = 29.9 × 5.7 μm), with a length:width ratio of 4.2–7.1; 3-septate, (24.0)34.3–38.5(44.0) × (5.0)5.9–6.3(7.0) μm (av. = 36.4 × 6.1 μm), with a length:width ratio of 4.3–7.3. Microconidia 0–1-septate, ellipsoid to subcylindrical, more or less straight, mostly with a visible hilum; aseptate microconidia (4.0)6.9–8.0(10.0) × (3.0)4.0–4.5(5.0) μm (av. = 7.4 × 4.3 μm), with a length:width ratio of 1.3–2.9; 1-septate, (8.0)12.9–15.7(18.0) × (4.0)4.2–4.7(5.5) μm (av. = 14.3 × 4.4 μm), with a length:width ratio 1.8–4.0. Chlamydospores globose to subglobose to ellipsoidal, 8–14 × 7–12 μm, smooth, but often appearing rough due to deposits, thick-walled, mostly intercalary, rarely terminal on short lateral branches, single, in chains or in clumps, hyaline, becoming medium brown.

Holotype: Netherlands, Lelystad, soil, June 1997, coll./isol. J.T. Poll, iden. W. Gams, holotype CBS H-20559, culture ex-type CBS 940.97.

Culture characteristics: Mycelium cottony, dense. Surface on OA cinnamon, with sparse, buff aerial mycelium, on PDA umber to chestnut, with buff to saffron aerial mycelium; zonation absent, transparency homogeneous, margin even; reverse similar to surface, but chestnut on PDA. Colonies on PDA grow 6–7 mm diam at 4°C after 7 days. Optimum temperature at 22°C when colonies reach 44–45 mm diam, after 7 days. Colony diam is 22–24 mm at 25°C, after 7 days. No growth observed at 30°C.

Isolate studied: CBS 940.97 (Table 1).

Habitat and distribution: Soil (Netherlands).

Ilyonectria liliigena A. Cabral & Crous, sp. nov. (Fig. 7)

MycoBank 560114.
Fig. 7

Ilyonectria liliigena (CBS 189.49). ad Simple conidiophores on aerial mycelium. e Chlamydospores on mycelium. fi Micro- and macroconidia. Bars 10 μm

Etymology: Named after its host, Lilium regale.

Ilyonectriae panacis morphologice similis, sed longitudine media macroconidiorum 3-septatorum breviore, 27.9–29.8 μm, distinguitur.

Conidiophores simple or complex or sporodochial. Simple conidiophores arising laterally or terminally from aerial mycelium, solitary to loosely aggregated, unbranched or sparsely branched, bearing up to two phialides, 1–4-septate, 50–170 μm long; phialides monophialidic, cylindrical to subulate, 30–65 μm long, 2.0–3.5 μm wide at the base, 1.5–2.0 μm near the apex. Sporodochial conidiophores irregularly branched; phialides cylindrical, mostly widest near the base. Macroconidia predominating, formed on simple conidiophores, on SNA formed in flat domes of slimy masses, 1(−3)-septate, straight or frequently minutely curved, cylindrical or sometimes typically minutely widening towards the tip, therefore appearing somewhat clavate, mostly without a visible hilum; 1-septate, (19.0)22.9–24.6(30.0) × (3.3)4.2–4.5(5.2) μm (av. = 23.8 × 4.3 μm), with a length:width ratio of 4.0–7.0; 2-septate, (21.0)26.1–27.7(32.1) × (4.0)4.7–5(5.7) μm (av. = 26.9 × 4.9 μm)with a length:width ratio of 3.8–7.0; 3-septate, (23.9)27.9–29.8(35.0) × (3.9)4.7–5.1(6.0) μm (av. = 28.9 × 4.9 μm), with a length:width ratio of 4.0–8.3. Microconidia 0–1-septate, ellipsoidal to subcylindrical, more or less straight, mostly with a visible hilum; aseptate, microconidia (5.9)8.9–10.3(17.0) × (2.5)3.0–3.2(4.4) μm (av. = 9.6 × 3.1 μm), with a length:width ratio of 2.0–4.6; 1-septate, (10.0)12.9–14.3(18.0) × (2.5)3.3–3.6(4.5) μm (av. = 13.6 × 3.4 μm), with a length:width ratio 2.8–5.6. Conidia formed in heads on simple conidiophores or as white (OA) or unpigmented (SNA) masses. Chlamydospores globose to subglobose, 6–14 × 5–12 μm, smooth but often appearing rough due to deposits, thick-walled, mostly in terminal on short lateral branches or rarely intercalary, single, in chains or in clumps, hyaline, becoming slightly brown at margins.

Holotype: Netherlands, Hoorn, bulb rot of Lilium regale, 1949, coll./isol. M.A.A. Schipper, holotype CBS H-20560, culture ex-type CBS 189.49.

Culture characteristics: Mycelium felty, with an average to strong density. Surface on OA sienna, with sparse, saffron, aerial mycelium. Surface on PDA sepia to cinnamon, with saffron to buff aerial mycelium. Zonation absent or concentric, with homogeneous transparency. Margins were even, or sometimes slightly uneven. Reverse similar to surface, except in colour; on OA pale vinaceous to cinnamon; on PDA buff to saffron to chestnut. Colonies on PDA grew poorly (1–4 mm diam) at 4°C after 7 days. Optimum temperature at 22°C, when colonies reach 34–45 mm diam, after 7 days. Colony diam was 16–29 mm at 25°C, after 7 days. No growth was observed at 30°C.

Isolates studied: CBS 189.49; CBS 732.74; CBS 304.85; CBS 305.85 (Table 1).

Host and distribution: Lilium regale bulbs (Netherlands).

Ilyonectria lusitanica A. Cabral, Rego & Crous, sp. nov. (Fig. 8)

MycoBank 560105.
Fig. 8

Ilyonectria lusitanica (CBS 129080). ac Simple conidiophores of the aerial mycelium. d Chlamydospores on mycelium. eg Micro- and macroconidia. Bars (a)  20 μm, (bg) 10 μm

Etymology: Named after the Latin name for the country from where it was collected, Portugal.

Ilyonectriae europaeae morphologice similis, sed longitudine media macroconidiorum breviore, 25–28.4 μm, distinguitur.

Conidiophores simple or complex, sporodochial. Simple conidiophores arising laterally or terminally from aerial mycelium, solitary to loosely aggregated, unbranched or sparsely branched, bearing up to two phialides, 1–4-septate, 60–220 μm long; phialides monophialidic, cylindrical to subulate, 20–70 μm long, 2.5–3.5 μm wide at the base, 1.5–2.5 μm near the aperture. Complex conidiophores aggregated in small sporodochia, repeatedly and irregularly branched. Macroconidia predominating, formed by both type of conidiophores, on SNA formed in flat domes of slimy masses, 1(−3)-septate, straight or minutely curved, cylindrical with both ends more or less broadly rounded, but may narrow towards the tip, without a visible hilum, and may have a constriction on the septa in older cultures; 1-septate, (14.0)17.3–18.8(21.0) × (4.0)4.6–5(5.5) μm (av. = 18.1 × 4.8 μm), with a length:width ratio of 2.8–4.8; 2-septate, (18.0)20.5–22.1(27.0) × (4.0)4.9–5.2(6.0) μm (av. = 21.3 × 5.1 μm), with a length:width ratio of 3.5–5.4; 3-septate, (18.0)25.0–28.4(38.0) × (4.5)5.2–5.5(6.0) μm (av. = 26.7 × 5.4 μm), with a length:width ratio of 3.6–6.8. Microconidia 0–1-septate, ellipsoid to ovoid, more or less straight, without a visible hilum, and may have a constriction at the septum; aseptate, (5.0)6.9–8.2(10.0) × (2.5)3.0–3.3(4.0) μm (av. = 7.6 × 3.2 μm), with a length:width ratio of 1.7–3.3; 1-septate, (8.0)10.0–11.0(14.0) × (3.0)3.4–3.7(4.0) μm (av. = 10.5 × 3.6 μm), with a length:width ratio 2.0–3.7. Conidia formed in heads on simple conidiophores or as white (OA) or unpigmented (SNA) masses. Chlamydospores rarely observed, globose to subglobose to cylindrical, 9–13 × 7–11 μm, smooth, but often appearing rough due to deposits, thick-walled, intercalary, hyaline, becoming slightly brown at the margin.

Holotype: Portugal, Melgaço, Alvaredo, on Vitis vinifera, below grafting zone, 6–year-old plant; scion Alvarinho on rootstock 196–17, 2005, coll./isol. N. Cruz, holotype CBS H-20563, culture ex-type CBS 129080 = Cy197 = CPC 19166.

Culture characteristics: Mycelium felty with average density. Surface on OA cinnamon, with aerial mycelium sparse, buff. Surface on PDA, cinnamon, with sparse, ochreous to buff aerial mycelium. Zonation absent, transparency homogeneous, margin even. Reverse similar to surface but buff to cinnamon on OA, and chestnut to cinnamon on PDA. Colonies on PDA grow 5–6 mm at 4°C after 7 days. Optimum temperature between 20 and 22°C, with colonies reaching 42–46 mm and 43–46 mm, respectively, after 7 days. Colony diam was 31–32 mm at 25°C, after 7 days. No growth observed at 30°C.

Isolate studied: CBS 129080 (Table 1).

Host and distribution: Vitis vinifera (Portugal).

Ilyonectria mors-panacis (A.A. Hildebr.) A. Cabral & Crous, comb. nov. (Fig. 9)

MycoBank 560115.
Fig. 9

Ilyonectria mors-panacis (CBS120363). a, b Simple conidiophores on aerial mycelium. cg Micro- and macroconidia. hj Chlamydospores on mycelium. Bars 10 μm

Basionym: Ramularia mors-panacis A.A. Hildebr., Can. J. Res. 12: 101. 1935.

= Cylindrocarpon panacis Matuo & Miyaz., Trans. Mycol. Soc. Japan 9: 111. 1969.

Cylindrocarpon destructans f.sp. panacis Matuo & Miyaz., Ann. Phytopath. Soc. Japan 50: 390. 1984.

Conidiophores simple or complex, sporodochial. Simple conidiophores arising laterally or terminally from aerial mycelium, solitary to loosely aggregated, unbranched or sparsely branched, rarely consisting only of phialides, 1–3-septate, 45–170 μm long; phialides monophialidic, cylindrical to subulate, 23–55 μm long, 2.0–3.0 μm wide at the base, 1.5–3.0 μm near the apex. Complex conidiophores aggregated in small sporodochia, repeatedly and irregularly branched. Macroconidia predominating, formed on simple conidiophores, on SNA formed in flat domes of slimy masses, 1(−3)-septate, straight, cylindrical with both ends more or less broadly rounded, mostly without a hilum; 1-septate, (21.0)28.2–31.6(40.0) × (5.0)5.8–6.3(7.5) μm (av. = 29.9 × 6.1 μm), with a length:width ratio of 3.3–7.0; 2-septate, (28.0)30.5–38.4(42.0) × (5.0)5.9–6.4–7.0(7.1) μm (av. = 34.4 × 6.4 μm), with a length:width ratio of 4.0–6.0; 3-septate, (37.8)39.0–44.2(45.0) × (6.9)7.0–7.5(7.5) μm (av. = 41.0 × 7.2 μm), with a length:width ratio of 5.3–6.0. Microconidia 0–1-septate, ellipsoid to subcylindrical, more or less straight, without a visible hilum; aseptate, (5.0)8.9–10.4(17.0) × (2.5)3.6–3.9(5.0) μm (av. = 9.6 × 3.8 μm), with a length:width ratio of 1.3–3.4; 1-septate, (9.0)12.5–14.1(19.0) × (3.5)4.4–4.8(5.5) μm (av. = 13.3 × 4.6 μm), with a length:width ratio 2.0–4.0. Conidia formed in heads on simple conidiophores or as white, creamy (OA) or hyaline (SNA) masses. Chlamydospores globose to subglobose, 8–16 × 7–15 μm, smooth, but often appearing rough due to deposits, thick-walled, terminal on short lateral branches or intercalary, single, in chains or in clumps, hyaline, becoming medium brown.

Lectotype: Canada, Ontario, on living roots of Panax quinquefolium, June 1935, A.A. Hildebrand, lectotype designated here CBS H-20561, culture ex-lectotype CBS 306.35.

Culture characteristics: Mycelium felty with an average density. Surface on OA and PDA chestnut, with sparse, buff to rosy-buff to cinnamon or saffron aerial mycelium. Concentric zonation, with homogeneous transparency, and even margins. Reverse similar to surface, ochreous to fulvous, or sepia to dark vinaceous on OA, and chestnut to sienna on PDA. Colonies on PDA grow 3–9 mm diam at 4°C after 7 days. Optimum temperature for growth is 18°C, when colonies reach 22–40 mm diam, after 7 days. Colony diam was 31–40 mm at 25°C after 7 days. No growth was observed at 30°C.

Isolates studied: CBS 306.35; CBS 307.35; CBS 120359; CBS 120360; CBS 120361; CBS 120362; CBS 120363; CBS 120364; CBS 120365; CBS 120366; CBS 120367; CBS 120368; CBS 120369; CBS 124662; CPC 13535; CPC 13537 (Table 1).

Hosts and distribution: Panax ginseng (Japan), P. quinquefolium (Canada).

Notes: Ilyonectria mors-panacis is distinct from “C.” destructans (anamorph: “C”. destructans, neotype CUP-011985, conidia (18.0)23.0–30.0(35.0) × (6.0)6.5(7.0) μm) in having larger conidia, and indistinct hila (being prominent, flat, 2 μm diam in I. radicicola; see also Samuels and Brayford 1990, Fig. 1). “Ramularia” panacicola is distinct by also having shorter conidia than I. mors-panacis, 5.5–34.2 × 2.5–7.2 μm (Zinssmeister 1918), and appears to be another potential synonym of “C.destructans. However, no authentic material could be located of “R.panacicola, and the only isolate deposited under this name was a Canadian strain collected by Hildebrand (1935), which in fact represented I. mors-panacis (Fig. 1). The oldest name for the species on Panax treated here, therefore, is R. mors-panacis (CBS 306.35), with the Japanese collections (“C.panacis ≡ “C.destructans f.sp. panacis, CBS 124662 = NBRC 31881) being later synonyms (see Fig. 1).

Ilyonectria panacis A. Cabral & Crous, sp. nov. (Fig. 10)

MycoBank 560104.
Fig. 10

Ilyonectria panacis (CBS 129079). ac Simple, unbranched or sparsely branched conidiophores on aerial mycelium. d, e Chlamydospores on mycelium. fi Micro- and macroconidia. Bars 10 μm

Etymology: Named after its host, Panax quinquefolium.

Ilyonectriae liliigenae morphologice similis, sed longitudine media macroconidiorum longiore, 31–35 μm, distinguitur.

Conidiophores simple or complex, sporodochial. Simple conidiophores arising laterally or terminally from aerial mycelium, solitary to loosely aggregated, unbranched or sparsely branched bearing up to three phialides, 1–5-septate, 60–220 μm long; phialides monophialidic, cylindrical to subulate, 20–65 μm long, 2.5–3.0 μm wide at the base, 1.5–2.0 μm near the aperture. Complex conidiophores aggregated in small sporodochia, repeatedly and irregularly branched. Macroconidia predominating, formed on both type of conidiophores, on SNA formed in flat domes of slimy masses, 1(−3)-septate, straight, cylindrical with both ends more or less broadly rounded, mostly with a visible centrally located hilum; 1-septate, (20.0)23.7–25.9(32.0) × (4.0)4.7–5.0(5.5) μm (av. = 24.8 × 4.8 μm), with a length:width ratio of 4.0–6.0; 2-septate, (23.0)27.0–30.3(37.0) × (4.8)5.0–5.4(6.0) μm (av. = 28.7 × 5.2 μm), with a length:width ratio of 4.6–6.7; 3-septate, (27.0)31.2–35.0(49.0) × (5.0)5.4–5.8(6.0) μm (av. = 33.1 × 5.6 μm), with a length:width ratio of 4.9–8.2. Microconidia 0–1-septate, ellipsoid to ovoid to subcylindrical, more or less straight, mostly with a visible hilum; aseptate, (6.0)8.0–9.8(13.0) × (3.5)3.7–3.9(4.0) μm (av. = 8.9 × 3.8 μm), with a length:width ratio of 1.7–3.3; 1-septate, (8.0)11.3–13.7(16.0) × (3.5)3.8–4.2(4.5) μm (av. = 12.5 × 4.0 μm), with a length:width ratio 1.8–4.3. Conidia formed in heads on simple conidiophores or as white (OA) or unpigmented (SNA) masses. Chlamydospores globose to subglobose to ellipsoidal, 8–14 × 6–10 μm, smooth, but often appearing rough due to deposits, thick-walled, terminal on short lateral branches or intercalary, single, in chains or in clumps, hyaline, becoming medium brown.

Holotype: Canada, Alberta, Panax quinquefolium, 1998, coll./isol. K. F. Chang, holotype CBS H-20562, culture ex-type CBS 129079 = CDC-N-9A = CPC 19167.

Culture characteristics: Mycelium felty with strong density. Surface on OA chestnut to sienna, with aerial mycelium sparse, vinaceous-buff. Surface on PDA chestnut to cinnamon, with aerial mycelium sparse, buff to saffron. No zonation was observed, and transparency was homogeneous; margins predominantly even. Reverse similar to surface, except in the colour, fawn to cinnamon on OA, and chestnut on PDA. Colonies on PDA grow 5 mm diam at 4°C after 7 days. Optimum temperature at 20°C, with colonies reaching 40–42 mm diam, after 7 days. Colony diam was 15 mm at 25°C after 7 days. No growth observed at 30°C.

Isolate studied: CBS 129079 (Table 1).

Host and distribution: Panax quinquefolium (Canada).

Notes: Several species have in the past been described on Panax in the genera Ramularia and Cylindrocarpon. The only unresolved species is “C.” destructans (and its potential synonym, “Ramularia” panacicola, see above). “Cylindrocarpon” destructans is clearly different from I. panacis, which has larger conidia, (27.0)31.2–33.1–35.0(49.0) × (5.0)5.4–5.6–5.8(6.0) μm.

Ilyonectria pseudodestructans A. Cabral, Rego & Crous, sp. nov. (Fig. 11)

MycoBank 560106.
Fig. 11

Ilyonectria pseudodestructans (all from CBS 129081, except g and e from CBS117824). ad Simple, unbranched or sparsely branched conidiophores on aerial mycelium. eg Chlamydospores on mycelium and macroconidia. hl Micro- and macroconidia. Bars 10 μm

Etymology: Named after its morphological similarity to “Cylindrocarpon” destructans.

Ilyonectriae crassae morphologice similis, sed macroconidiis clavatis distinguitur.

Conidiophores simple or complex, sporodochial. Simple conidiophores arising laterally or terminally from aerial mycelium, solitary to loosely aggregated, unbranched or sparsely branched, bearing up to two phialides, 1–3-septate, 50–180 μm long; phialides monophialidic, cylindrical to subulate, 30–58 μm long, 2.5–3.5 μm wide at the base, 1.5–2.0 μm near the aperture. Complex conidiophores aggregated in small sporodochia, repeatedly and irregularly branched. Macroconidia predominating, formed by simple conidiophores, on SNA formed in flat domes of slimy masses, 1–3(−4)-septate, straight, typically clavate, mostly centrally located hilum; 1-septate, (19.0)25.8–27.5(35.0) × (4.0)5.0–5.3(6.5) μm (av. = 26.7 × 5.2 μm), with a length:width ratio of 3.8–6.6; 2-septate, (23.0)30.0–31.7(38.0) × (5.0)5.3–5.5(6.0) μm (av. = 30.9 × 5.4 μm), with a length:width ratio of 4.3–7.4; 3-septate, (28.0)34.2–36.2(48.0) × (5.0)5.9–6.2(7.0) μm (av. = 35.2 × 6.0 μm), with a length:width ratio of 4.6–7.4. Microconidia 0–1-septate, ellipsoid to ovoid to subcylindrical, more or less straight, with a visible, centrally located hilum; aseptate (6.0)10.5–11.8(15.0) × (3.0)3.6–3.8(4.5) μm (av. = 11.2 × 3.7 μm), with a length:width ratio of 1.5–4.3; 1-septate, (10.0)14.6–15.6(18.0) × (3.0)4.1–4.4(5.0) μm (av. = 15.1 × 4.2 μm), with a length:width ratio of 2.4–5.0. Conidia formed in heads on simple conidiophores or as white (OA) or unpigmented (SNA) masses. Chlamydospores globose to subglobose to ellipsoid, 9–18 × 8–14 μm, smooth but often appearing rough due to deposits, thick-walled, terminal on short lateral branches or intercalary, in chains or in clumps, and also in the cells of macroconidia, hyaline, becoming medium brown.

Holotype: Portugal, São Paio, Gouveia, Vitis vinifera, 4-year-old, showing decline symptoms, scion Malvasia fina; rootstock 1103P, 1996, coll./isol. C. Rego, holotype CBS H-20564, culture ex-type CBS 129081 = Cy20 = CPC 19164.

Culture characteristics: Mycelium felty, with average to strong density. Surface on OA cinnamon, with sparse, buff to saffron or chestnut to sienna aerial mycelium. Surface on PDA cinnamon to vinaceous, with sparse, saffron to buff or chestnut to sienna aerial mycelium. Zonation absent, with homogeneous transparency; margins even. Reverse similar to surface, except in colour, sepia to cinnamon on OA and chestnut to cinnamon on PDA. Colonies on PDA grow poorly (4–6 mm diam), at 4°C after 7 days. Optimum temperature between 20–22°C, when colonies reach 32–44 mm and 37–41 mm diam, respectively, after 7 days. Colony diam was 22–29 mm at 25°C after 7 days. No growth was observed at 30°C.

Isolates studied: CPC 13534; CBS 117812; CBS 117824; CBS 129081; Cy22 (Table 1).

Hosts and distribution: Poa pratensis (Canada), Quercus sp. (Austria), Vitis vinifera (Portugal).

Notes: Ilyonectria pseudodestructans is reminiscent of “Cylindrocarpon” destructans, in having a similar conidial morphology (3-septate, with central, truncate hilum). However, conidia of I. pseudodestructans are somewhat longer than those of I. radicicola.

Ilyonectria robusta (A.A. Hildebr.) A. Cabral & Crous, comb. nov. (Figs. 12 and 13)

MycoBank 560113.
Fig. 12

Ilyonectria robusta (a, b from CPC 13532 × CBS 308.35; ck from CPC 13532 × CBS 117813). a, b Development of perithecia on the surface of a birch toothpick or agar. ce Perithecium mounted in lactic acid. d Ostiolar area. e Surface view of perithecium wall region. fh Longitudinal sections of perithecia showing details of ostiole and wall. ik Asci and ascospores. Bars (ac) 50 μm; (d, f) 20 μm; (e, gk) 10 μm

Fig. 13

Ilyonectria robusta (All from CBS 129084, except f from CBS 605.92). ac Simple conidiophores on aerial mycelium. d Sporodochial conidiophore on carnation leaf agar. e Chlamydospores on mycelium fi Micro- and macroconidia. Bars 10 μm

Basionym: Ramularia robusta A.A. Hildebr. Can. J. Res. 12: 102. 1935.

Perithecia formed heterothallically in vitro, disposed solitarily or in groups, developing directly on the agar surface or on sterile pieces of birch wood, ovoid to obpyriform, with a flattened apex, up to 70 μm wide, orange to red, becoming purple-red in 3 % KOH (positive colour reaction), smooth to warted, up to 250 μm diam and high; perithecial wall consisting of two regions; outer region 11–36 μm thick, composed of 1–3 layers of angular to subglobose cells, 10–30 × 6–24 μm; cell walls up to 1 μm thick; inner region 8–14 μm thick, composed of cells that are flat in transverse optical section and angular to oval in subsurface optical face view, 5–11 × 2.5–5 μm; Asci narrowly clavate to cylindrical, 40–50 × 4.5–6 μm, 8-spored; apex subtruncate, with a minutely visible ring. Ascospores medianly 1-septate, ellipsoid to oblong-ellipsoid, somewhat tapering towards both ends, smooth to finely warted, frequently guttulate, hyaline, (8.2)9.4–9.7–10.0(11.5) × (2.5)2.9–3.0–3.1(3.7) μm. Conidiophores simple or complex or sporodochial. Simple conidiophores arising laterally or terminally from aerial mycelium, solitary to loosely aggregated, unbranched or sparsely branched, bearing up to three phialides, 1–4-septate, 55–160 μm long; phialides monophialidic, cylindrical to subulate, 20–60 μm long, 2.0–3.0 μm wide at the base, 1.5–2.0 μm near the apex. Complex conidiophores aggregated in small sporodochia (on carnation leaf agar; Crous et al. 2009b), repeatedly and irregularly branched; phialides more or less cylindrical, but tapering slightly in the upper part towards the apex, or narrowly flask-shaped, mostly with widest point near the middle, 15–20 μm long, 2.5–3.5 μm wide at the base, 3.0–4.0 μm at the widest point, and 1.0–2.0 μm wide near the apex. Macroconidia predominating, formed on simple conidiophores, on SNA formed in flat domes of slimy masses, 1–3-septate, straight, minutely curved or sometimes distorted, cylindrical with both ends more or less broadly rounded, but may narrow towards the tip, mostly without a visible hilum; 1-septate, (15.0)22.8–24.6(35.0) × (4.5)6.3–6.7(8.0) μm (av. = 23.7 × 6.5 μm), with a length:width ratio of 2.7–5.2; 2-septate, (20.0)26.2–28.1(38.0) × (5.0)6.9–7.2(8.0) μm (av. = 27.2 × 7.0 μm), with a length:width ratio of 2.9–5.2; 3-septate, (24)32.3–34.7(58) × (6.0)7.2–7.5(9.0) μm (av. = 33.5 × 7.4 μm), with a length:width ratio of 3.1–7.3. Microconidia 0–1-septate, ellipsoid to ovoid to subcylindrical, more or less straight, without a visible hilum; aseptate, (4.0)8.0–9.3(14.0) × (2.5)3.6–4.0(5.5) μm (av. = 8.7 × 3.8 μm), with a length:width ratio of 1.3–4.0; 1-septate, (9.0)13.5–14.7(18.0) × (3.5)4.7–5.1(6.0) μm (av. = 14.1 × 4.9 μm), with a length:width ratio 1.5–4.5. Conidia formed in heads on simple conidiophores or as white (OA) or unpigmented (SNA) masses. Chlamydospores globose to subglobose, 7–14 × 6–13 μm, smooth, but often appearing rough due to deposits, thick-walled, mostly occurring intercalary in chains, hyaline, becoming golden-brown.

Lecto- and teleotype: Canada, Ontario, on living roots of Panax quinquefolium, 1935, A.A. Hildebrand, lectotype designated here CBS H-20565, as dried culture of CBS 308.35; teleotype designated here CBS H-20566, including fertile perithecia of the teleomorph (CPC 13532 × CBS 308.35), culture ex-lectotype CBS 308.35.

Fertile matings: Perithecia observed after 4 wk in crossings of strains: CPC 13532 × CBS 308.35, CPC 13532 × CBS 773.83, CPC 13532 × CBS 605.92, CPC 13532 × CBS 117813, CBS 129084 × CBS 308.35, CBS 129084 × CBS 605.92, CBS 129084 × CBS 117813.

Culture characteristics: Mycelium felty with an average density. Surface on OA sienna to sepia with aerial mycelium sparse, buff. Surface on PDA cinnamon, with aerial mycelium buff to cinnamon, or rosy buff on PDA. Zonation absent to concentric, with homogeneous transparency; margins predominantly even, but sometimes uneven. Reverse similar to surface, except in the colour, sienna on OA and chestnut at the centre, and sienna to orange towards the margin on PDA. Colonies on PDA grow 4–7 mm at 4°C after 7 days. Optimum temperature at 22°C when colonies reach 40–52 mm diam, after 7 days. Colony diam was 35–48 mm at 25°C after 7 days. No growth to slight growth (0–2 mm) was observed at 30°C.

Isolates studied: CBS 321.34; CBS 308.35; CBS 773.83; CBS 605.92; CBS 117813; CBS 117814; CBS 117815; CBS 117817; CBS 117818; CBS 117819; CBS 117820; CBS 117821; CBS 117822; CBS 117823; CBS 129084; CD1666; CPC 13532; Cy23; Cy158; Cy231 (Table 1).

Hosts and distribution: Loroglossum hircinum (root) (Tunisia), Panax quinquefolium (root) (Canada), Prunus cerasus, Thymus sp., Vitis vinifera (basal end of rootstock) (Portugal), Quercus robur (root), Quercus sp. (root) (Austria), Tilia petiolaris (rootstock) (Germany), water (in aquarium with Anodonta sp.) (Netherlands).

Notes: When Hildebrand (1935) described Ramularia robusta from living roots of Panax quinquefolium in Ontario, Canada, he did not indicate a type specimen. However, he deposited an original culture in the CBS. A sporulating, dried-down culture is thus herewith designated as lectotype, and a new name proposed in Ilyonectria, with a teleotype represented by a fertile mating between CPC 13532 × CBS 308.35.

Ilyonectria rufa A. Cabral & Crous, sp. nov. (Fig. 14)

MycoBank 560116.
Fig. 14

Ilyonectria rufa (All from CBS 156.47, except c from CBS 120372). ac Simple, sparsely branched conidiophores on aerial mycelium. df Chlamydospores in mycelium and in macroconidia. gk Micro- and macroconidia. Bars 10 μm

Etymology: The epithet “rufa” referring to “Coleomyces rufus”, a provisional name proposed for this species by Moreau and Moreau (1937).

Ilyonectriae crassae morphologice similis, sed macroconidiis brevioribus, 28–31.2 μm longis, distinguitur.

Conidiophores simple or complex, sporodochial. Simple conidiophores arising laterally or terminally from aerial mycelium, solitary to loosely aggregated, unbranched or sparsely branched, bearing up to two phialides, 1–5-septate, 55–210 μm long; phialides monophialidic, cylindrical to subulate, 20–57 μm long, 2.5–3.5 μm wide at the base, 1.5–2.0 μm near the aperture. Complex conidiophores aggregated in small sporodochia, repeatedly and irregularly branched. Macroconidia predominating, formed on both types of conidiophores, on SNA formed in flat domes of slimy masses, 1(−3)-septate, straight, cylindrical with both ends broadly round, mostly centrally located hilum; 1-septate, (17.0)22.3–23.8(29.0) × (4.0)5.1–5.4(6.0) μm (av. = 23.1 × 5.3 μm), with a length:width ratio of 3.1–5.6; 2-septate, (19.0)24.5–26.6(32.0) × (4.0)5.2–5.5(6.5) μm (av. = 25.5 × 5.4 μm), with a length:width ratio of 3.4–6.0; 3-septate, (23.0)28.6–31.2(37.0) × (5.0)5.5––5.9(7.0) μm (av. = 29.9 × 5.7 μm), with a length:width ratio of 3.4–7.2. Microconidia 0–1-septate, ellipsoid to subcylindrical, more or less straight, with a visible, centrally located hilum; aseptate, (4.0)8.4–9.8(15.0) × (3.0)3.5–3.8(5.0) μm (av. = 9.1 × 3.6 μm), with a length:width ratio of 1.3–4.0; 1-septate, (9.0)12.1–13.3(17.0) × (3.0)4.2–4.6(5.5) μm (av. = 12.7 × 4.4 μm), with a length:width ratio 2.2–3.8. Conidia formed in heads on simple conidiophores or as white (OA) or unpigmented (SNA) masses. Chlamydospores globose to subglobose to cylindrical, 7–12 × 6–9 μm, smooth, but often appearing rough due to deposits, thick-walled, terminal on short, lateral branches, or intercalary, single, in chains or in clumps, and also in the cells of the macroconidia, hyaline, becoming slightly brown in the outer wall.

Holotype: France, dune sand, Feb. 1937, coll./isol. F. Moreau, holotype CBS H-20567, culture ex-type CBS 153.37.

Culture characteristics: For CBS 153.37, CBS 156.47, CPC 13536 and 94–1628: Mycelium felty with average to strong density. Surface on OA buff to saffron, aerial mycelium sparse, buff. On PDA rosy-buff to cinnamon, with aerial mycelium sparse, buff to rosy-buff or pale luteus in the centre. For CBS 640.77, CBS 120371 and CBS 120372: Mycelium felty, with low to average density. Surface on OA cinnamon to sienna, aerial mycelium sparse, saffron to cinnamon. On PDA saffron to cinnamon, with aerial mycelium cinnamon to rust. Zonation absent or concentric, with homogeneous transparency; margins even or sometimes uneven. Reverse similar, except in colour, saffron on OA, and cinnamon to rosy-buff on PDA, or sienna with pigments, pale vinaceous in OA and umber to chestnut on PDA. Colonies on PDA grow poorly, (2–7 mm diam) at 4°C, after 7 days. Optimum temperature between 20–22°C, when colonies reach 28–42 mm, 31–46 mm diam, respectively, after 7 days. Colony diam was 19–24 mm at 25°C after 7 days. No growth observed at 30°C.

Isolates studied: CBS 153.37; CBS 156.47; CBS 640.77; CBS 120371; CBS 120372; CPC 13536; 94–1628 (Table 1).

Hosts and distribution: Azalea indica (Belgium), dune sand (France), Picea glauca, Pseudotsuga menziesii (Canada).

Notes: The genus Coleomyces represents a later synonym of Cylindrocarpon (Booth 1966). However, Coleomyces, which is based on C. rufus (Moreau and Moreau 1937), was published as “ad interim”, suggesting that Moreau and Moreau were planning to validate the name later, which was not the case. Based on the International Code of Botanical Nomenclature (Art. 34.1, Ex. 6), Chaverri et al. (2011) correctly chose to ignore the name. However, an original strain of C. rufus was deposited in the CBS (CBS 153.37), and the species epithet is herewith validated for the species.

Ilyonectria venezuelensis A. Cabral & Crous, sp. nov. (Fig. 15)

MycoBank 560117.
Fig. 15

Ilyonectria venezuelensis (CBS 102032). a, b Simple conidiophores on aerial mycelium. ce Conidiophores bearing terminal, penicillate phialides. fj Micro- and macroconidia. Bars 10 μm

Etymology: Named after the country from where it was collected, Venezuela.

Ilyonectriae robustae morphologice similis, sed conidiophoris cum verticillo terminali phialidum distinguitur.

Conidiophores simple or complex, sporodochial. Simple conidiophores arising laterally or terminally from aerial mycelium or from agar surface, solitary to loosely aggregated, unbranched, or bearing terminal, penicillate phialides, 1–4-septate, 35–200 μm long; phialides monophialidic, cylindrical to subulate, 40–60 μm long, 2.5–3.5 μm wide at the base, 1.5–2.5 μm near the apex, or narrowly flask-shaped, 16–40 μm long, 2.0–3.0 μm wide at the base, 3.0–4.0 μm at the widest point, 1.5–2.5 μm near the apex. Complex conidiophores aggregated in small sporodochia, repeatedly and irregularly branched. Macroconidia predominating, formed by both types of conidiophores, on SNA formed in flat domes of slimy masses, 1–3-septate, straight or minutely curved, cylindrical with both ends more or less broadly rounded, but may narrow towards the tip, mostly without a visible hilum; 1-septate, (22.0)24.6–27.3(35.0) × (5.0)5.3–5.7(6.5) μm (av. = 26.0 × 5.5 μm), with a length:width ratio of 3.8–7.0; 2-septate, (25.0)26.3–37.4(44.0) × (5.9)6.0–6.6(7.0) μm (av. = 31.9 × 6.3 μm), with a length:width ratio of 4.2–6.8; 3-septate, (28.0)36.5–41.7(50.0) × (6.0)6.9–7.5(8.0) μm (av. = 39.1 × 7.2 μm), with a length:width ratio of 4.0–6.7. Microconidia 0–1-septate, ellipsoid to ovoid, more or less straight, without a visible hilum; aseptate, (5.0)8.4–10.5(13.0) × (3.0)3.3–3.7(4.0) μm (av. = 9.5 × 3.5 μm), with a length:width ratio of 1.7–3.4; 1-septate, (11.0)14.5–16.3(19.0) × (3.5)3.9–4.3(5.0) μm (av. = 15.4 × 4.1 μm), with a length:width ratio 2.8–4.8. Conidia formed in heads on simple conidiophores or as white (OA) or unpigmented (SNA) masses. Chlamydospores ovoid to ellipsoidal, 6–13 × 5–7 μm, smooth, but often appearing rough due to deposits, thick-walled, terminal on short lateral branches or intercalary, single, in chains or in clumps, hyaline, becoming slightly brown at the margin.

Holotype: Venezuela, Amazonas, Cerro de la Neblina, tree bark, 1985, coll./isol. A. Rossman, holotype CBS H-20568, culture ex-type CBS 102032.

Culture characteristics: Mycelium cottony with average to strong density. Surface on OA saffron, with aerial mycelium sparse, buff, on PDA buff to saffron, with aerial mycelium saffron to pale luteous; zonation absent, transparency homogeneous, margin even; reverse similar to surface, but saffron to cinnamon on PDA. Colonies on PDA grow poorly (2–3 mm) at 4°C, after 7 days. Optimum temperature at 20°C, with colonies reaching 49 mm diam, after 7 days. Colony diam was 35–36 mm at 25°C after 7 days. No growth was observed at 30°C.

Isolate studied: CBS 102032 (Table 1).

Host and distribution: Tree bark (Venezuela).

Ilyonectria vitis A. Cabral, Rego & Crous, sp. nov. (Fig. 16)

MycoBank 560107.
Fig. 16

Ilyonectria vitis (CBS 129082). ac Simple conidiophores on aerial mycelium. dg Conidiophores forming microconidia arising from mycelium at agar surface, reduced to a stipe with a terminal arrangement of phialides, ranging from 2 to a dense cluster; sparsely branched or penicillate. il Micro- and macroconidia. m Chlamydospores on mycelium. Bars 10 μm

Etymology: Named after the host from which it was collected, Vitis vinifera.

Ilyonectriae anthuriicolae morphologice similis, sed longitudine media macroconidiorum longiore, 41.6–43.5 μm, distinguitur.

Conidiophores simple or complex or sporodochial. Simple conidiophores arising laterally or terminally from aerial mycelium, solitary to loosely aggregated, unbranched or sparsely branched, bearing up to three phialides, 1–3 septate, 30–70 μm long; monophialides more or less cylindrical, but tapering slightly towards the tip, 11–21 μm long, 2.0–3.0 μm wide at the base, 3.0–4.5 μm at widest point, 1.5–2.5 μm near the apex. Conidiophores forming microconidia arising from mycelium at agar surface, reduced to monophialides, or a stipe with a terminal arrangement of phialides, ranging from 2 to a dense cluster; sparsely branched or penicillate; monophialides narrowly flask-shaped, typically with widest point near the middle, 10–17 μm long, 1.5–3.0 μm wide at the base, 2.5–4.0 μm at widest point, 1.0–2.0 μm near the apex. Sporodochial conidiophores irregularly branched; phialides more or less cylindrical but slightly tapering towards the tip, or narrowly flask-shaped, with widest point near the middle, 14–20 μm long, 2.5–3.5 μm wide at the base, 3.0–4.5 μm at widest point, 1.5–2.5 μm near the apex. Macroconidia formed in flat domes of slimy masses, predominantly 3-septate, rarely 1–2- or 4-septate, straight or minutely curved, cylindrical with both ends more or less broadly rounded, mostly without a visible hilum; 3-septate conidia (34.9)41.6–43.5(51.6) × (6.2)7.9–8.2(9.5) μm (av. =42.5 × 8.0 μm), with a length:width ratio of 3.9–6.7. Microconidia on SNA formed in heads, aseptate, subglobose to ovoid, rarely ellipsoid, mostly with a visible, centrally located or slightly laterally displaced hilum, (3.7)4.9–5.4(6.7) × (3.2)3.7–4.0(4.6) μm (av. = 5.1 × 3.9 μm), with a length:width ratio of 1.1–1.7. Chlamydospores globose to subglobose to ellipsoid, 9–18 × 6–13 μm, smooth, but often appearing rough due to deposits, thick-walled, formed intercalary in chains or in clumps, and also in the cells of macroconidia, hyaline, becoming golden-brown.

Holotype: Portugal, Vidigueira, Vitis vinifera, basal end of a 2-year-old plant; scion Touriga Nacional; rootstock 110R, 2008, coll./isol. C. Rego, holotype CBS H-20569, culture ex-type CBS 129082 = Cy233 = CPC 19168.

Culture characteristics: Mycelium felty with density low to average. Surface on OA sienna, with sparse, saffron aerial mycelium, and luteous growth at margin. Surface on PDA chestnut, with sienna aerial mycelium, with luteous margin. Zonation was absent (OA) or concentric (PDA), transparency was homogeneous (PDA) or not (OA). Growth at margin even to uneven. Reverse similar to surface, except in colour, sienna to saffron on OA, and chestnut to umber on PDA. Colonies on PDA do not grow at 4°C after 7 days. Optimum temperature at 20°C, when colonies reach 29–30 mm diam, after 7 days. Colony diam was 39–40 mm at 25°C and 8–9 mm at 30°C after 7 days. No growth was observed at 35°C.

Isolate studied: CBS 129082 (Table 1).

Host and distribution: Vitis vinifera (Portugal).

Key to species treated

  1. 1

    Growth at margin on OA after 14 days at 20°C, lacking yellow pigmentation

     
  2. 2

    Colony diameter on PDA after 7 days at 25°C < 30 mm

     
  3. 3

    Macroconidia forming chlamydospores

     
  4. 4

    Macroconidia 1–3-septate, 3-septate macroconidia mean range 34.1–36.2 μm long

     
  5. 5

    Macroconidia cylindrical, with the base appearing somewhat acute I. crassa

     
  6. 5*

    Macroconidia clavate I. pseudodestructans

     
  7. 4*

    Macroconidia predominantly 1-septate; 3-septate macroconidia smaller, mean range 28–31.2 μm long I. rufa

     
  8. 3*

    Macroconidia lacking chlamydospores

     
  9. 6

    Macroconidia predominantly curved I. liliigena

     
  10. 6*

    Macroconidia straight

     
  11. 7

    Macroconidia lacking visible hilum I. mors-panacis

     
  12. 7*

    Macroconidia with a visible, centrally located hilum

     
  13. 8

    Three-septate macroconidia mean range 31.0–35.0 μm long I. panacis

     
  14. 8*

    Three-septate macroconidia mean range 34.3–38.5 μm long I. gamsii

     
  15. 2*

    Colony diameter after 7 days at 25°C >30 mm

     
  16. 9

    Colony diameter after 7 days at 25°C, >50 mm I. cyclaminicola

     
  17. 9*

    Colony diameter after 7 days at 25°C, 30–50 mm

     
  18. 10

    Conidiophores bearing a terminal whorl of phialides I. venezuelensis

     
  19. 10*

    Conidiophores unbranched, or different from above

     
  20. 11

    Teleomorph known, and can be induced in culture

     
  21. 12

    Three-septate macroconidia mean range 32.3–34.7 μm long; ascospores mean range 9.4–10.0 μm long I. robusta

     
  22. 12*

    Three-septate macroconidia mean range 30.0–36.0 μm long; ascospores mean range longer, 10–13 μm long I. radicicola a

     
  23. 11*

    Teleomorph unknown

     
  24. 13

    Mean range of 3-septate macroconidia, 29.7–31.5 × 6.5–6.9 μm I. europaea

     
  25. 13*

    Mean range of 3-septate macroconidia smaller, 25.0–28.4 × 5.2–5.5 μm I. lusitanica

     
  26. 1*

    Yellow pigmentation present at margin

     
  27. 14

    Macroconidia 3-septate, mean range 29.5–32.2 μm long I. anthuriicola

     
  28. 14*

    Macroconidia 3-septate, mean range 41.6–43.5 μm long I. vitis

     

aNo authentic cultures of “C.” destructans, conidia (18.0–)23.0–30.0(−35.0) × (6.0–)6.5(−7.0) μm, are presently available.

Discussion

“Cylindrocarpon” destructans is a cosmopolitan soil-borne pathogen causing disease on a wide number of herbaceous and woody plant species (Samuels and Brayford 1990). The wide morphological and pathogenic amplitude of “C.” destructans makes it a commonly identified species, with many diseases from the Cylindrocarpon-complex being attributed to it, and ranking at the top of all “Cylindrocarpon” spp. deposited in the NCBI nucleotide database.

In this study, “C.” destructans isolates from the CBS culture collection (deposited under the wider concept of the species) were analysed using a multigene approach in order to clarify taxonomic aspects of this species complex. Molecular analyses show that these isolates cluster in various clades supported by high bootstrap support values. A previous study (Seifert et al. 2003) included a subset of the strains used here, and already highlighted the existence of unexpected divergence in “C.destructans, as opposed to a large homogeneity in e.g. Neonectria ditissima. Several species have in recent years been separated from the “C.destructans complex, including “C.macroconidialis, “C.coprosmae and “C.austroradicicola based on morphological (Samuels and Brayford 1990) and molecular characters (Schroers et al. 2008; Seifert et al. 2003). Furthermore, several isolates causing black foot disease of grapevine, previously considered as “C.destructans, were recently identified as I. liriodendri (Chaverri et al. 2011; Halleen et al. 2006), along with the ex-type strain from Liriodendron tulipifera (CBS 110.81) and a strain from Cyclamen (CBS 301.93). In this study, two further strains isolated from young Malus domestica and Quercus suber trees showing decline symptoms were also identified as I. liriodendri.

Altogether, we analysed 68 strains putatively belonging to “C.destructans, but none of them clustered together with the ex-type culture of I. radicicola (CBS 264.65), suggesting that this species may not be as common as previously accepted. Halleen et al. (2006) identified a single strain (IMI 313237, isolated from arecoid palm) clustering with CBS 264.65. This also raises questions relating to the correlation between the anamorph, “C.destructans, and its purported teleomorph, I. radicicola.

“Nectria” radicicola was described by Gerlach and Nilson (1963) from decayed leaves, flowers stalks and corms of Cyclamen persicum collected in Sweden, with a “Cylindrocarpon” anamorph they identified as “C.” radicicola.

In 1924, Wollenweber introduced “C.radicicola (McAlpine) Wollenw. as a new combination, based on Septocylindrium radicicola McAlpine (1899), described from Citrus trees in Australia. Later, Wollenweber (1928) noted that Septocylindrium radicicola, with catenulate conidia, was different from “C.radicicola, and the name was therefore based on Wollenweber’s (1928) description. Because of this confusion in names, Booth (1966) suggested that “C.radicicola should be dropped, and that the name to be used as anamorph for I. radicicola should be “C.destructans [originally described by Zinssmeister (1918) on Panax quinquefolia from Wisconsin, USA]. Furthermore, Booth (1966) designated a neotype for “C.destructans, obtained from P. quinquefolia in USA, KY, Washington Co., Springfield; collected by W.B. Edwards in 1922, available as herbarium material in Cornell Plant Pathology Herbarium, CUP-011985. This specimen was re-examined in the present study [conidia (18.0)23.0–30.0(35.0) × (6.0)6.5(7.0) μm] [original description by Zinssmeister (1918), 9.0–32.4 × 3.2–8.1 μm], thus revealing conidia to be smaller than those formed by I. radicicola, which are (24.0)33.1(47.0) × (4.9)6.4(7.8) μm. From these observations, we conclude that “C.destructans, which occurs on P. quinquefolia in the USA, represents yet another species distinct from I. radicicola, which is not yet represented in our phylogenetic tree (Fig. 1).

A strain deposited in CBS culture collection by Hildebrand in 1935, as “Ramularia” mors-panacis, was found to represent original material (ex-type CBS 306.35), collected from living roots of Panax quinquefolium in Ontario, Canada. The epithet “mors-panacis” is therefore resurrected for this clade, while the Japanese collection identified as “Cylindrocarpon destructans” f. sp. panacis (ex-type CBS 124662 = NBRC 31881), isolated from Panax ginseng in Japan, is treated as synonym.

The ex-type strain of “Ramularia” robusta (CBS 308.35), isolated from living roots of Panax quinquefolium in Ontario, Canada, can be resurrected for a large clade representing isolates from a range of hosts and continents. Similarly, an authentic strain of “C.destructans var. crassum (Booth 1966; Wollenweber 1931) is available for a species occurring on Panax and Pseudotsuga in Canada, Lilium and Narcissus in the Netherlands, and can thus be resurrected as I. crassa. Although strain CBS 120370 clustered together with other strains of I. crassa for most genes (no nucleotide differences in ITS and TEF, two nucleotide differences in HIS), this strain was not included in that species because of an 8-bp difference in TUB, a slower growth rate (e.g., 21 mm colony diam at 20°C for 7 days, as opposed to 31–46 mm for other isolates), a lower conidial length:width ratio [e.g. for 3-septate conidia 4.0–6.5, as opposed to 4.8–8.9 and smaller conidia (e.g. 3-septate conidia ranging from (26.0)31.2–34.0(40.0) × (6.0)6.6–7.1(8.0) μm (av. 32.6 × 6.9 μm) , as opposed to (29.0)34.1–36(49.0) × (5.0)5.6–5.8(7.0) μm (av. 35.1 × 5.7 μm)]. Further studies should thus be conducted in order to clarify the taxonomy of this strain.

Ilyonectria anthuriicola and I. vitis are very similar in morphology to “C.pauciseptatum. These species all have predominantly 3-septate macroconidia after 10 days in culture. Ilyonectria anthuriicola is easier to distinguish than “C.pauciseptatum as the 3-septate conidia are smaller and narrower, (25.0)29.5–32.2(38.0) × (6.0)7.5–8.1(9.0) μm, while in “C.pauciseptatum they are (37.0)42.0–47.0(54.0) × (7.0)8.5–9.5(10.0) (Schroers et al. 2008). For I. vitis 3-septate macroconidia are of similar size to those of “C.pauciseptatum, (34.9)41.6–43.5(51.6) × (6.2)7.9–8.2(9.5), making it difficult to distinguish them based on this character. Growth rate at 20°C is slower in I. vitis than in “C.pauciseptatum, and they also differ regarding colony characteristics and colours. Morphologically, however, they remain difficult to distinguish.

In this study, the genetic structure of the I. radicicola complex was analysed using a multi-locus approach along with morphological and culture characters. Three major groups were identified based on this approach, each group containing several species. Although we have been able to clarify several aspects related to the host range and distribution of taxa in the I. radicicola species complex, further collections, especially from Panax in the USA, will be essential to elucidate the status of “C.” destructans.

Notes

Acknowledgements

The first author would like to thank Dr M.C. Silva (Coffee Rust Research Centre / Tropical Research Institute, Portugal) for making equipment available, and for assisting with the use of the cryostat for perithecial sectioning. Drs F. Caetano (Instituto Superior de Agronomia, Portugal), J. Armengol (Univ. Politécnica de Valencia, Spain), K.A. Seifert (Agriculture and Agri-Food, Canada), M.L. Inácio (Instituto Nacional de Recursos Biológicos/INIA, Portugal) P. Lecomte (Institut National de la Recherche Agronomique, Bordeaux-Aquitaine, France) and W.D. Gubler (Univ. California, Davis, USA) are thanked for making several strains available, without which this study would not have been possible. The curator of CUP is acknowledged for making the neotype “C.” destructans available for examination. This article is part of a PhD dissertation (Instituto Superior de Agronomia, Technical University of Lisbon, Portugal). This work was partially supported by Fundação para a Ciência e a Tecnologia, Portugal (grant number SFRH/BD/24790/2005; project PTDC/AGR-AAM/099324/2008).

Open Access

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Copyright information

© The Author(s) 2011

Authors and Affiliations

  • Ana Cabral
    • 1
  • Johannes Z. Groenewald
    • 2
  • Cecília Rego
    • 1
  • Helena Oliveira
    • 1
  • Pedro W. Crous
    • 2
    • 3
    • 4
  1. 1.CEER-Biosystems Engineering, Instituto Superior de AgronomiaTechnical University of LisbonLisboaPortugal
  2. 2.CBS-KNAW Fungal Biodiversity CentreUtrechtThe Netherlands
  3. 3.Microbiology, Department of BiologyUtrecht UniversityUtrechtThe Netherlands
  4. 4.Laboratory of PhytopathologyWageningen University and Research Centre (WUR)WageningenThe Netherlands

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