Lizel Mostert
Department of Plant Pathology
University of Stellenbosch
Stellenbosch

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Petri disease and esca are serious diseases of young and old vines in most countries where grapevines (Vitis spp.) are cultivated. 

The disease can manifest as a sudden collapse of the foliage, but more often takes the form of a slow decline accompanied by weak growth, various leaf symptoms (interveinal chlorosis, leaf necrosis, wilting) and gradual death of grapevines.  Internal symptoms of Petri disease can normally be seen when transverse or longitudinal cuts are made in the trunk and shoots.  These include black spots (Fig. 1a) and dark brown to black streaking of the xylem tissues.  Petri disease is caused by a combination of Phaeomoniella chlamydospora and several species of Phaeoacremonium (Scheck et al., 1998; Mugnai et al., 1999; Groenewald et al., 2001).  Esca can be typically identified by internal wood decay (Fig. 1b), as well as symptoms on leaves (Fig. 1c) and berries.  Various types of wood deterioration are observed when a transverse cut is made.  Symptoms on the leaves consist of interveinal regions of chlorotic and yellowish tissue that turns yellow-brown or red-brown, and have also been described as “tiger stripes.”  Fungi that have been associated with esca symptoms include Pa. chlamydospora, several species of Phaeoacremonium as well as the wood-rotting basidiomycetes Fomitiporia mediterranea, F. punctata and to a lesser extent, Stereum hirsutum (Larignon & Dubos 1997; Mugnai et al., 1999; Fischer 2002).  Young vines infected with Pa. chlamydospora and/or Phaeoacremonium species revealing Petri disease symptoms can later develop esca symptoms after the infection and colonisation of F. mediterranea, F. punctata or S. hirsutum. 

A monograph of Togninia and its Phaeoacremonium anamorphs
The genus Togninia (Fig. 2) was confirmed as the teleomorph of Phaeoacremonium (Fig. 3) by means of morphology, sexual compatibility, and DNA phylogeny (Mostert et al., 2003).  The genus Togninia was monographed along with its Phaeoacremonium anamorphs (Mostert et al., 2006b).  Ten species of Togninia and 22 species of Phaeoacremonium were treated.  Several new species of Togninia were described, namely T. argentinensis, T. austroafricana, T. krajdenii, T. parasitica, T. rubrigena and T. viticola.  Nine new Phaeoacremonium species, of which six were obtained as etiologic agents of human opportunistic infection, were identified by Mostert et al. (2005).  These species are Pm. alvesii, Pm. amstelodamense, Pm. australiense, Pm. griseorubrum, Pm. krajdenii, Pm. scolyti, Pm. subulatum, Pm. tardicrescens and Pm. venezuelense.  Additionally Mostert et al. (2006b) described six new species of Phaeoacremonium including Pm. argentinense, Pm. austroafricanum, Pm. novae-zealandiae, Pm. iranianum, Pm. sphinctrophorum and Pm. theobromatis. 

Species were identified based on their cultural and morphological characters, supported by DNA data derived from partial sequences of the actin and β-tubulin genes.  Phylogenies of the SSU and LSU rRNA genes were used to determine whether Togninia has more affinity with the Calosphaeriales or the Diaporthales.  These results confirmed that Togninia had a higher affinity to the Diaporthales than the Calosphaeriales.  Examination of type specimens obtained from various herbariums revealed that T. cornicola, T. vasculosa, T. rhododendri, T. minima var. timidula and T. villosa, were not members of Togninia.  New combinations such as Calosphaeria cornicola, Calosphaeria rhododendri, Calosphaeria transversa, Calosphaeria tumidula, Calosphaeria vasculosa and Jattaea villosa were therefore proposed to accommodate these species.

The mating strategy of several Togninia species was investigated with ascospores obtained from fertile perithecia produced in vitro (Mostert et al., 2006b).  Togninia argentinensis and T. novae-zealandiae have homothallic mating systems, whereas T. austroafricana, T. krajdenii, T. parasitica, T. rubrigena and T. viticola are heterothallic.  The species predominantly isolated from diseased grapevines are Pm. aleophilum, Pm. parasiticum and Pm. viticola.  Perithecia of two of these species, T. minima and T. viticola (Rooney-Latham et al., 2005; Eskalen et al., 2005), have been found on grapevines in the field, indicating that these species recombine in vineyards. 
Thirteen species of Phaeoacremonium have been isolated from grapevines of which eight have been found on diseased grapevines in South Africa. These species include Pm. aleophilum, Pm. austroafricanum, Pm. krajdenii, Pm. parasiticum, Pm. scolyti, Pm. subulatum, Pm. venezuelense and Pm. viticola.  Of the 42 Phaeoacremonium isolates investigated from grapevines in South Africa, 52% were confirmed as Pm. aleophilum, 21% Pm. parasiticum and 10% Pm. austroafricanum.  

Detection tools
The correct and rapid identification of Phaeoacremonium species is important to facilitate the understanding of their involvement in plant as well as human disease.  A rapid identification method was developed for the 22 species of Phaeoacremonium currently recognised (Mostert et al., 2005; 2006b).  It involved the use of 23 species-specific primers, including 20 primers targeting the β-tubulin gene and three targeting the actin gene.  These primers can be used in 14 multiplex reactions.  With the multiple-entry electronic key phenotypic and sequence data can be used to identify the different Phaeoacremonium species.  Separate dichotomous keys were provided for the identification of the Togninia and Phaeoacremonium species based on phenotypic characters.  Additionally, keys for the identification of Phaeoacremonium-like fungi and the genera related to Togninia were also provided. 
 
Genetic diversity among isolates of Phaeomoniella chlamydospora
The genetic diversity among isolates of Pa. chlamydospora on grapevines was investigated by means of Amplified Fragment Length Polymorphisms (AFLPs) (Mostert et al., 2006a).  Phaeomoniella chlamydospora isolates were collected from different positions on the same vine, different vines within a vineyard and different vineyards within South Africa.  Selected isolates from various grape producing countries, namely Australia, France, Italy, Iran, New Zealand, Slovenia and U.S.A. were also included.  The overall low level of genetic variation confirmed asexual reproduction to be dominant in the field.  Different genotypes were found among isolates of Pa. chlamydospora within the same grapevine, suggesting multiple infections from different inoculum sources.  Isolates from different countries had a high percentage of similarity and clustered together, indicating the absence of genotype-geographic structure.  The presence of the same genotype in different vineyards and production areas suggests that long-range dispersal through aerial inoculum or infected plant material played an important role in genotype distribution. 

Control of Petri disease and esca
Disease prevention seems to be the most effective means in managing Petri disease and esca.  In the nurseries, infections in rootstock mother blocks can best be prevented by sanitation and pruning wound protection. Hot water treatment of rootstock cuttings prior to grafting for 30 min at 50°C proved to be the most effective means of reducing the infections of  propagation material (Crous et al., 2001; Edwards et al., 2004; Fourie and Halleen, 2004).  In the vineyard, stress factors in young vines that might predispose it to Petri disease and esca should be avoided.  Sanitation practices, such as the removal of infected plants, plant parts and/or pruning debris, will lead to lower inoculum levels in vineyards.  Additionally, pruning wound protection in vineyards will limit infection by these pathogens.  

In conclusion
Seven species of Togninia and 15 species of Phaeoacremonium were newly described.  The data published in the monograph, as well as the newly developed dichotomous and online polyphasic keys will enable scientists to correctly identify all the known species and also provide a reference database to which new species can be added in future.  The newly reported host ranges and distributions, together with pathogenicity data, will enable scientists to identify species of possible quarantine concern.  Rapid identification of these species based on the data generated in this thesis, would prevent the accidental import of infected grapevine material into countries where these species presently do not occur.

References

1. Crous PW, Swart L, Coertze S. (2001). The effect of hot-water treatment on fungi occurring in apparently healthy grapevine cuttings. Phytopathologia Mediterranea 40: S464-S466.
2. Edwards J, Pascoe I, Salib S, Laukart N. (2004). Hot water treatment of grapevine cuttings reduces incidence of Phaeomoniella chlamydospora in young vines. Phytopathologia Mediterranea 43: 158-159 (abstract).
3. Eskalen A, Rooney-Latham S, Gubler WD (2005). First report of perithecia of Phaeoacremonium viticola on grapevine (Vitis vinifera) and ash tree (Fraxinus latifolia) in California. Plant Disease 89: 686.
4. Fischer M. (2002). A new wood-decaying basidiomycete species associated with esca of grapevine: Fomitiporia mediterranea (Hymenochaetales). Mycological Progress 1: 315-324.
5. Fourie PH, Halleen F. (2004). Proactive control of Petri disease of grapevine through treatment of propagation material. Plant Disease 88: 1241-1245.
6. Groenewald M, Kang J-C, Crous PW, Gams W. (2001). ITS and beta-tubulin phylogeny of Phaeoacremonium and Phaeomoniella species. Mycological Research 105: 651-657.
7. Larignon P, Dubos B. (1997). Fungi associated with esca disease in grapevine. European Journal of Plant Pathology 103: 147-157.
8. Mostert L, Abeln ECA, Halleen F, Crous PW. (2006a). Genetic diversity among isolates of Phaeomoniella chlamydospora on grapevines. Australasian Journal of Plant Pathology 35: 453-460.
9. Mostert L, Groenewald JZ, Summerbell RC, Gams W, Crous PW.  (2006b). Taxonomy and pathology of Togninia (Diaporthales) and its Phaeoacremonium anamorphs.  Studies in Mycology 54.
10. Mostert L, Groenewald JZ, Summerbell RC, Robert V, Sutton DA, Padhye AA, Crous PW.  (2005). Species of Phaeoacremonium associated with human infections and environmental reservoirs in infected woody plants.  Journal of Clinical Microbiology 43:1752-1767.
11. Mostert L, Crous PW, Groenewald JZ, Gams W, Summerbell R. (2003).  Togninia (Calosphaeriales) is confirmed as teleomorph of Phaeoacremonium by means of morphology, sexual compatibility, and DNA phylogeny.   Mycologia 95: 646-659.
12. Mugnai L, Graniti A, Surico G. (1999). Esca (black measles) and brown wood-streaking: two old and elusive diseases of grapevines. Plant Disease 83: 404-416.
13. Rooney-Latham S, Eskalen A, Gubler WD. (2005). Occurrence of Togninia minima perithecia in esca-affected vineyards in California. Plant Disease 89: 867-871.
14. Scheck HJ, Vasquez SJ, Gubler WD. (1998). First report of three Phaeoacremonium spp. causing young grapevine decline in California. Plant Disease 82: 590.