Pathogenicity experiments

Greenhouse experiments have demonstrated that Cylindrocarpon obtusisporum, Phaeoacremonium chlamydosporum, P. inflatipes, and P. aleophilum are aggressive pathogens of grape seedlings (fig. 1). Seedlings are used for bioassays because they can produce symptoms rapidly and are certain to be disease-free at the beginning of a trial.

One-month-old seedlings (Vitis vinifera “Carignane”) were inoculated by dipping their roots in a 106 spore/ml suspension for 30 minutes. After 60 days, 75% of seedlings inoculated with Cylindrocarpon obtusisporum, 50% with Phaeoacremonium aleophilum, 63% P. chlamydpsporum, and 56% with P. inflatipes, had died. The pathogens were easily recovered from both the roots and the stems of the moribund seedlings. Control plants treated with water remained healthy, and pathogens were never detected.

Field inoculations to confirm Phaeoacremonium as a pathogen of young vines were done at UC Davis. In August 1994, own-rooted grapevines (Vitis vinifera “Chardonnay”) were inoculated with a 106 spore/ml suspension and planted in 3.8L pots. Spores were introduced by drilling a small hole into the crown and filling the hole with spores or cutting the roots, then dipping the vine into the spore suspension for 30 minutes. Control plants were treated in the same manner but were inoculated with sterile water.

In February 1995, these vines were planted in the field. In October 1997, the test grapevines were removed from the field and destructively sampled. Both inoculation methods gave similar results, and the data were combined for analysis.

All inoculated grapevines were visibly stunted with small chlorotic leaves. The vascular elements and the pith near the crowns of inoculated vines were discolored. The caliper of the wood, measured at 2.5 cm above the soil line, was 28 mm for the control vines, compared to only 13 mm for inoculated vines, a reduction of more than 50%. (See Table III).

The dry weight of leaves and petioles was approximately three times less and the dry weight of wood was approximately four times less in the vines inoculated with P. chlamydosporum compared with water-inoculated controls.

Pathogen survival

We’ve demonstrated the ability of Phaeoacremonium inflatipes, P. chlamydosporum and P. aleophilum. to survive in nutrient-poor sand for over 28 days. It is possible that these pathogens are introduced into the grape rootstocks during the callusing stage of nursery production.

In the callusing process, cuttings are removed from cold storage and placed in heated bins of sand, sawdust, or peat moss for 25 to 48 days. The wounded end of the cutting forms a callus layer and initiates rooting. Small pathogen populations might be introduced into the callusing trays from infected cuttings. Once in the warm, damp media, they could spread through the callusing tray.

We have begun research with assistance of two commercial nurseries to determine whether these pathogens infect vines during commercial production of nursery stock.

New disease names

Fungi in the genera Cylindrocarpon and Phaeoacremonium have been consistently isolated from young grapevines showing decline symptoms. Cylindrocarpon spp. are generally recognized as relatively weak pathogens on vegetables, small fruits, tree fruits, forage, ornamentals, and conifers and therefore of limited economic importance. Production of chlamydospores allows the pathogen to survive in soil for extended lengths of time in the absence of a suitable host plant.

Cylindrocarpon destructans has been identified as causing a disease of young vines in France known as black-foot. One California isolate of Cylindrocarpon was identified as C. destructans in 1994. Since 1994, C. obtusisporum has been the most commonly isolated species.

However, because of the similarities in symptom expression caused by the two closely related species of Cylindrocarpon, we propose to maintain the name “black-foot disease” for decline caused by either species. We propose the name “Phaeoacremonium grapevine decline” to define the symptoms associated with infection by Phaeoacremonium spp.

Hence two new diseases of young grapevine have been identified in California: black-foot, caused by Cylindrocarpon obtusisporum or C. destructans; and Phaeoacremonium grapevine decline, caused by as many as three species of Phaeoacremonium.

Fungi in the genus Phaeoacremonium have also been implicated as the pathogens in “black measles” or “esca” disease of older grapevines (Vasquez and Gubler, unpublished data). Black measles is a destructive disease of woody tissues in grapes and has an extensive range of symptoms. These include stunted growth and dieback, leaf chlorosis and necrosis, raisining of fruit, plugged xylem vessels, and occasionally sudden wilting and/or death of vines. Black measles symptoms have been seen in some young vines infected with Phaeoacremonium.

These organisms have been in California for many years, and it is unclear why pathogens formerly associated only with older grapevines have recently emerged as a threat to young grapevines. Cultural practices or environmental conditions may have shifted in favor of the pathogens.

For example, at several sites, grapevines that were exposed to low temperatures in the winter of 1990/1991 have showed typical decline symptoms. Alternatively, perhaps new strains of these pathogens have evolved, or perhaps the huge planting boom has simply increased awareness of young vine decline.

Thus far, no obvious relationship exists between rootstock or scion varieties and the severity of either Cylindrocarpon black foot disease or Phaeoacremonium grapevine decline. Because of the phylloxera epidemic, AXR-1 rootstock, used almost exclusively for 20 years, has been replaced by other rootstock varieties. The new rootstocks may prove to be highly susceptible to these diseases, particularly if pre-disposed by devigorating conditions. Investigations into differences in susceptibility and symptom expression in rootstock varieties are in progress.

Due to the similarity of symptoms, diagnosis of Cylindrocarpon black foot disease or Phaeoacremonium grapevine decline should never be made solely on the basis of visual symptoms. Young grapevines can fail for many reasons, including poor nutrition, improper irrigation (too much or too little), poor planting techniques, virus-induced incompatibility, nematodes, and poor quality planting stock.

Vascular symptoms can also have many biotic and abiotic causes. Black or amber streaks seen in the vascular elements are the result of deposition of phenolic compounds in response to wounding, a general defense mechanism of the plant. Physical injuries to the grapevine due to root tearing, disbudding, or other trauma during production or planting can result in phenolic depositions and black discoloration, or even outright death of tissue.

Other terms have been used in the popular press to describe the visual symptoms of phenolic deposition and formation of gums and tyloses in the vascular tissues, including “black goo” and “black xylem disease.” These names do not encompass the full range of symptoms associated with Cylindrocarpon black foot disease or Phaeoacremonium grapevine decline nor do they provide a description of the problem adequate enough to be of value to plant pathology or to the viticulture industry.

Our lab at UC Davis is focusing on increasing our understanding of pathogen biology, disease epidemiology, and symptom development in Cylindrocarpon black foot disease and Phaeoacremonium vine decline. Studies are underway to determine how these pathogens survive and spread. Sources of pathogen inoculum will be identified. Eventually, with an increased understanding of the disease biology, effective control strategies can be developed.

References
Cylindrocarpon Black-Foot
1. Maluta, D.R. and P. Larignon 1991. “Pied-noir: Nieux vaut prevenir.” Viticulture 11: 71-72.
2. Grasso, S, and G.M. Lio 1975. “Infezioni de Cylindrocarpon obtusisporum su piante di vite in Sicilia.” Vitis 14: 1, 36-39.
3. Scheck, H.J., S.J. Vasquez, P. Larignon, D. Fogle, and W.G. Gubler, 1997. “Cylindrocarpon black-foot disease and Phaeoacremonium grapevine decline in California.” California Agriculture, in press.

Phaeoacremonium Vine Decline

1. Ferreira, J.H.S., P.S. Van Wyk, and E. Venter 1994. “Slow dieback of grapevine: Association of Phialophora parasitica. S. Afr. J. Enol. Vitic. 15: 9-11.
2. Crous, P.W., W. Gams, M.J. Wingfield, and P.S. van Wyk, 1996. “Phaeoacremonium gen. associated with wilt and decline diseases of woody hosts and human infections.” Mycologia 88: 786-796.