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2007 Plant Management Network.
Accepted for publication 22 December 2006. Published 20 April 2007.


Phytophthora citricola Causes a Stem Canker in Black Walnut (Juglans nigra)


Janna Beckerman, Department of Botany Plant Pathology, and Gail Ruhl, Plant and Pest Diagnostic Laboratory, Purdue University, West Lafayette, IN 47907


Corresponding author: Janna Beckerman. jbeckerm@purdue.edu


Beckerman, J., and Ruhl, G. 2007. Phytophthora citricola causes a stem canker in black walnut (Juglans nigra). Online. Plant Health Progress doi:10.1094/PHP-2007-0420-01-BR.


Introduction

Black walnut (Juglans nigra) is considered a premier species in the lumber industry, despite fluctuating prices and lumber availability (3). This tree is relatively free of serious disease problems, except when the plants are intensively managed in a nursery-type or agroforestry operation (6). Intensively managed, high-density plantings, monoculture, and grafting practices predispose plants to pathogens not normally seen in the wild. Since the early 1980s, a canker of plantation-grown walnut has been regularly reported in the Midwest, and in 1989, a survey of 189 plantations revealed widespread distribution of this disease (2).

In spring 2006, samples of container-grown nursery stock of grafted black walnut were submitted to the Purdue Plant and Pest Diagnostic Laboratory. All affected samples, whether submitted to the Purdue Plant and Pest Diagnostic Laboratory or viewed on site at the nursery, were scions of several superior selections of black walnut grafted to one- to two-year-old seedlings raised in the nursery from harvested black walnut seed. All Purdue Plant and Pest Diagnostic Laboratory-submitted samples and all site-observed samples had cankers on the scion-wood portion of the graft. None of the observed, symptomatic plants had dieback that had progressed to the rootstock.

Both field-planted (Fig. 1) and containerized forest nursery stock samples showed symptoms of discrete cankers and gummosis. Cankers were often sunken, and tissue directly beneath the cankers was blackened. Canker expansion resulted in girdling of the main stem, concomitant in dieback, and death of portions distal to the canker. Cankers were often associated with leaf scars and branch stubs.


 

Fig. 1. An accompanying photo of black walnut submitted to the Plant and Pest Diagnostic Laboratory in spring 2006, showing dieback, death, and gummosis associated with black walnut canker.

 

The canker tissue was surface sterilized with 70% ethanol and plated onto P10VP medium. A Phytophthora sp. was consistently isolated from the symptomatic lesions, and these isolates produced prolific zoosporangia and oospores (Fig. 2). Morphological examination of the isolate is consistent with previously described features of P. citricola, including homothallism and the corresponding copious production of oospores; sporangia shape was somewhat variable and sporangia were not easily dislodged (Fig. 3) (1,4).


 

Fig. 2. An oospore with paragynous antheridia in V-8 juice agar.

 

Fig. 3. Semipapillate, noncaducous zoosporangium typical of P. citricola.


An axenic culture of the isolated Phytophthora was submitted to the Plant Pathogen Identification Laboratory at North Carolina State University for species identification. Blast analysis of the internal transcribed spacer region of ribosomal DNA (ITS-rDNA) sequence of the submitted sample identified 100% similarity with the sequences of two isolates of Phytophthora citricola Sawada (1927), previously submitted to GenBank.


Phytophthora citricola and Pathogenicity to Black Walnut

To date, P. citricola has been identified as the causal agent of root rot and trunk canker of more than 45 genera of plants (4). In black walnut, P. citricola was reported to cause a seedling root rot (5), but has never been reported to cause cankers. To test the pathogenicity of the Phytophthora isolate to cause cankers and dieback, five 2-year-old grafted black walnut scions were inoculated with a 5-mm plug from 4-day-old culture of P. citricola grown on 20% V-8 juice agar. Each plant was inoculated by: (i) wounding of old wood with a scalpel; (ii) wounding of new green growth with scalpel; (iii) inoculation of green growth without wounding; (iv) inoculation of detached petioles; (v) inoculation of bud scars, and (vi) inoculation of leaflets. Two 2-year-old grafted black walnut scions were used as control plants, and were subjected to the same six treatments as the P. citricola-inoculated plants above using 4-day old V-8 juice agar. All inoculations were wrapped with Parafilm M for 24 h. Plants were maintained in a 70F greenhouse and watered as needed. Foliage was kept dry to prevent dislodging of the inoculum.

Infection and canker development resulted within 36 h regardless of treatment (Fig. 4). No cankers developed in the control plants (Fig. 5). Symptomatic tissue was excised from the margin of the canker and plated on to P10VP agar. P citricola was re-isolated from all cankers in the manner outlined previously, fulfilling Kochs Postulates.



A

 
B

 
C
 

Fig. 4. Thirty-six hours post-inoculation, P. citricola-inoculated tissue shows lesion development, necrosis, and spread on both wounded (A) and unwounded (B) tissue. Gummosis was a common symptom (C).


 

Fig. 5. Thirty-six hours post-inoculation, necrosis is apparent on wounded control (left), but no visibly spreading lesions developed. Unwounded controls showed no symptom development (right).


P. citricola-inoculated and control black walnut grafts were maintained in a 70F greenhouse with regular watering for four months. By three months, all ten plants inoculated with P. citricola had suffered significant dieback, and seven of the grafts had died (Fig. 6); Scions on the remaining three died, but were producing rootstock suckers that did not yet show symptoms of disease. Control plants were asymptomatic and healthy.

To our knowledge, this is the first report of P. citricola causing a stem canker on black walnut in North America. Current work is underway to examine fungicide use and cultural management options to control this disease problem.


 

Fig. 6. Three months after inoculation, P. citricola inoculated black walnuts continued to decline, finally dying back to the root graft.

 

Literature Cited

1. Bush, E. A., Stromberg, E. L., Hong, C., Richardson, P. A., and Kong, P. 2006. Illustration of key morphological characteristics of Phytophthora species identified in Virginia nursery irrigation water. Online. Plant Health Progress doi:10.1094/PHP-2006-0621-01-RS.

2. Cummings-Carlson, J. 1994. Walnut Tips- Black Walnut Canker: Its Distribution and Management. Forestry Facts No. 72.

3. Cassens, D. L. 2004. Factors affecting the quality of walnut lumber and veneer. Pages 161-167 in: Black Walnut in a New Century: Proc. of the 6th Walnut Counc. Res. Symp., 2004 July 25-28, Lafayette, IN. Gen. Tech. Rep. NC-243. C. H. Michler, P. M. Pijut, J. W. Van Sambeek, M. V. Coggeshall, J. Seifert, K. Woeste, R. Overton, F. Ponder, Jr., eds. USDA Forest Serv., North Central Res. Stn., St. Paul, MN.

4. Erwin, D. C., and Ribeiro, O. K. 1996. Morphology and identification of Phytophthora species. Pages 99-144, 282-287 in: Phytophthora Diseases Worldwide. The American Phytopathological Society, St. Paul, MN.

5. Green, R. J., and Pratt, R.G. 1970. Root rot of black walnut seedling caused by Phytophthora citricola. Plant Dis. 54:583-585

6. Mielke, M., and Ostry, M. 2004. Diseases of Intensively Managed Black Walnut. Pages 110-113 in: Black Walnut in a New Century: Proc. of the 6th Walnut Counc. Res. Symp., 2004 July 25-28, Lafayette, IN. Gen. Tech. Rep. NC-243. C. H. Michler, P. M. Pijut, J. W. Van Sambeek, M. V. Coggeshall, J. Seifert, K. Woeste, R. Overton, F. Ponder, Jr., eds. USDA Forest Serv., North Central Res. Stn., St. Paul, MN.