1. Phytophthora ramorum A Short Course Presented by the California Oak Mortality Task Force Part 2 - Symptom Recognition, Diagnosis, and Sampling Background Photo Credits: Karl Buermeyer, UC Cooperative Extension (forest scene) Stephen Eales, Plant Health and Seed Inspectorate, DEFRA, UK (mountain laurel) Jan Hedberg, Oregon Department of Agriculture (viburnums in nursery) Steve Tjosvold, UC Cooperative Extension (rhododendron)

2. Course Outline Part 2 - Part 2 - Symptom Recognition, Diagnosis, and Sampling Bark Symptoms on Trees Foliar Symptoms on Wildland and Nursery Plants Scouting and Sampling Symptomatic Plants Review Questions Part 1 - Introduction to Phytophthora ramorum and Sudden Oak Death Part 3 - Regulations and Management of Phytophthora ramorum

3. Part 2 - Symptom Recognition, Diagnosis, and Sampling Bark Symptoms on Trees When Phytophthora ramorum spores germinate on the bark surface of a host tree, hyphae penetrate the bark. The pathogen then grows in the inner bark and outer xylem. In response, the tree “bleeds,” exuding a sticky reddish ooze. This is the only outward symptom of the disease until the tree turns brown and dies. In true oaks, these symptoms are only found within about 8 feet of the ground. Since other diseases cause similar symptoms, bleeding alone is not sufficient for diagnosis. If the outer bark of an infected tree is scraped away, cankers can be seen. Generally, red healthy bark tissue will be separated from brown, dull, diseased tissue by a distinct black zone line marking the progression of the disease. David Chambers, UC Cooperative Extension Karl Buermeyer, UC Cooperative Extension

4. Once infected by Phytophthora ramorum, trees are often attacked by secondary organisms. Ambrosia and oak bark beetles are attracted to the infected area of the trunk, and can be detected by small piles of boring dust (frass) around entry and exit holes in the tree trunk. Ambrosia beetles bore deeply into the xylem tissue, resulting in light colored frass, while oak bark beetles stay in the outer bark, and can be detected by reddish frass. Ambrosia Beetle Monarthrum species Oak Bark Beetle Pseudopityopthorus pubippenis A common condition in oak trees, known as “bacterial wetwood,” also causes bleeding symptoms which can be mistaken for Phytophthora ramorum. Associated with an injury or crack, wetwood bleeding is watery, foul smelling, and usually very copious, unlike the sticky, spotty oozing caused by Phytophthora ramorum. Karl Buermeyer, UC Cooperative Extension

5. Boring beetles and Hypoxylon can rapidly weaken trees, causing them to fail. These failures often occur where the cankers infected the trunk - the area to which the beetles are attracted. Ambrosia beetle galleries can be seen in the picture to the left where this tanoak failed. Karl Buermeyer, UC Cooperative Extension Another secondary organism commonly associated with Phytophthora ramorum is the fungus Hypoxylon thouarsianum. It produces hard, black, round fruiting bodies on the bark surface, and can be found where there is dead wood in a tree. Although not unique to Phytophthora ramorum-infested areas, it is often quite noticeable due to the large amount of dead wood available.

6. As mentioned in Part 1 of this course, tanoak is both a bark and foliar host. This may explain why it is the most vulnerable to mortality. Bark infections may or may not cause the oozing symptoms described above (slide 4). Tanoaks of all ages can be killed by the pathogen, whereas true oaks don’t seem to be at risk until they are about 6” in diameter. Symptoms on Foliar Hosts Symptoms present in tanoak that are not found in true oaks include the wilting of new spring growth due to disruption of water transport (shown above), and necrotic lesions on the twigs, resulting in the death of the leaves as the lesions progress along the twig (photo to the left). Tanoak (Lithocarpus densiflora) Pavel Svihra, UC Cooperative Extension Karl Buermeyer, UC Cooperative Extension

7. In California’s wildlands, the most important foliar host of Phytophthora ramorum is California bay laurel. The pathogen sporulates abundantly on the leaf surface. When it rains, the built up inoculum is washed or blown off the leaves onto the soil, other leaves, and the boles of other trees. Steve Tjosvold, UC Cooperative Extension California bay laurel symptoms include a brown leaf lesion with an irregular margin, sometimes with a yellow chlorotic band in the live portion of the leaf, and often separated by a uneven black zone line. Symptoms usually appear on the lowest part of the leaf, where water accumulates. Due to abundant sporulation, and the ease of culturing from these leaves, this tree is used as an early indicator and risk factor for the disease entering an area. Although infection can sometimes kill small twigs, Phytophthora ramorum does not significantly damage California bay laurel. California bay laurel (Umbellularia californica)

8. Evergreen huckleberry Vaccinium ovatum California buckeye Aesculus californica A number of other native broad-leaf species harbor Phytophthora ramorum in California and Oregon (See the complete list in Part 1.). Little is known about the role of these species in the life cycle and spread of the disease. The pathogen is difficult to culture from many of these species, and is difficult to diagnose because of the presence of other foliar diseases. Toyon Heteromeles arbutifolia Bigleaf maple Acer macrophyllum California honeysuckle Lonicera hispidula Pacific madrone Arbutus menziesii Wood rose Rosa gymnocarpa All photos: Garbelotto Lab, UC Berkeley

9. Rhododendrons are a significant Phytophthora ramorum nursery host, and in European gardens are suspected of spreading the pathogen to nearby trees that were growing amongst them. Rhododendrons may also play an important role in spreading the disease in wildland situations in Oregon. Rhododendrons (Rhododendron spp.) Ornamental Nursery Hosts Symptoms on rhododendrons include necrotic areas with diffused margins that are similar in appearance on both sides of the leaf. Steve Tjosvold, UC Cooperative Extension Bruce Moltzan, Missouri Department of Conservation

10. Stem infections can also occur in rhododendrons. Infection begins either in the leaf or at bud scars on the stems, and can travel either from the leaves through the midveins to the petioles and stems, or vice versa. In the two-week sequence above right, an artificial infection at point (A) travels down the leaf to infect the stem (B), and then travels throughout the stem and into other leaves (C). A B C (2nd artificial infection point) Steve Tjosvold, UC Cooperative Extension BBA Germany Alan Kanaskie, Oregon Dept. of Forestry

11. First confirmed as a host of Phytophthora ramorum in Europe, 2003, camellias are perhaps the most commonly detected host in US nurseries. Numerous cultivars of Camellia japonica, C. sasanqua, C. reticulata, and C. x williamsii have been found to be infected. Camellias (Camellia spp.) Camellia symptoms are limited to leaves. Lesions have diffused edges, and appear primarily at leaf tips and margins. No twig or branch infections have been observed in camellias, perhaps because infected leaves abscise readily when infected. When checking camellias, look at dropped leaves around the base of the plant, and for plants that have thin crowns, as shown to the right. Cheryl Blomquist, CDFA

12. In pieris, Phytophthora ramorum causes leaf necrosis and leaf abscision similar to that on Camellia. Branch tip dieback is also found. Pieris or Andromeda (Pieris spp.) Jan Hedberg, Oregon Dept. of Agriculture Central Science Laboratory, DEFRA, UK (Crown Copyright)

13. In viburnums, Phytophthora ramorum can cause leaf necrosis, tip dieback, and cankers. Viburnums can react similarly to bark hosts, in that the whole plant can die at once. Viburnum (Viburnum spp.) Cheryl Blomquist, CDFA Central Science Laboratory, DEFRA, UK (Crown Copyright) Jan Hedberg, Oregon Dept. of Agriculture

14. A number of other ornamental plants have been found to be susceptible to Phytophthora ramorum. Most of these have been found in the UK. Mountain laurel (Kalmia spp.) is of particular concern in the US due to its wide distribution in eastern forests. Other Ornamental Hosts Mountain Laurel Kalmia latifolia Drooping leuthocoe Leuthocoe fontanesiana Lilac Syringa vulgaris

15. Phytophthora ramorum symptoms vary greatly between and within host species, so there is no universal symptom indicating that a plant has the disease. Therefore, when surveying for the presence of this pathogen, it is best to be cautious and sample any suspicious host leaf or stem lesion. Conditions that may be Confused with Phytophthora ramorum on Ornamentals However, when sampling, rule out plants that only have sun scald symptoms. This condition only appears on one side of the plant (facing the sun), and unlike Phytophthora ramorum, it tends to avoid the leaf midvein. The photos to the right show sunscald on camellia and rhododendron. Tomas Pastalka, CDFA Steve Tjosvold, UC Cooperative Extension

16. Whether looking for Phytophthora ramorum in wildlands or nurseries, it is important to be familiar with host species symptoms. Diagnostic guides to assist in identification can be found at www.suddenoakdeath.org. At this site, review the document “Sudden Oak Death and Associated Diseases Caused by Phytophthora ramorum,” by Davidson et al., for information on native species where Phytophthora ramorum is present in west coast wildland habitats. It also gives a general overview of the disease and briefly covers ornamental hosts. For a more detailed guide to nursery host identification and management, access “Nursery Guide for Diseases of Phytophthora ramorum on Ornamentals: Diagnosis and Management,” by Tjosvold et al. Scouting and Sampling for Phytophthora ramorum

17. When surveying for Phytophthora ramorum, a 100% survey or strip survey can be used, checking all host species for symptoms. For larger properties, a 20% strip survey can be laid out by spacing transects 100m apart, and checking all host plants within 10m of the line (20m total width). Spacing the lines closer together will result in progressively more intensive surveys. Since the objective is to detect the disease, a suspicious plant just outside the transect border should be sampled. Lines should be perpendicular to the general contour of the property (uphill and downhill) so that different slope positions and draws will not be missed. Note the location of sampled plants on a map or with GPS so that they can be relocated if found to be positive. Surveying in Wildland Areas If surveys are to be done on a state or county level, 1/10 ha plots can be randomly distributed in areas that are likely to have host species, as identified by vegetation maps.

18. A nursery layout map that includes the locations of targeted species is useful to get oriented and develop a survey strategy. A scouting map includes species and cultivar names, locations, approximate quantity, and sources of targeted plants in scouted areas. During the walk-through, record the date, observations, and sampling information directly onto the scouting map. The recorded information should be reviewed and used to develop an efficient strategy each time the nursery is scouted. Scouting in Nurseries Begin the inspection with an overview of the area from the crop perimeter or with a quick walk-through. Examine any symptomatic plants more closely in an attempt to identify the problem. If symptoms are not apparent, start by walking a systematic path through the crop. Pay particular attention to low areas that may collect water, as well as the perimeter of the nursery if there are host species in the surrounding landscape.

19. Although it is easier to identify, sample, and culture Phytophthora ramorum on the leaves of some foliar hosts, it is often necessary to sample from the bark of potentially infected host trees. While the technique is described here, it is highly recommended that only trained individuals sample from bark. Bole Sampling for Phytophthora ramorum Upon locating symptomatic tissue… the canker is exposed with a hatchet or chisel.

20. Bole Sampling for Phytophthora ramorum and removed with tweezers. A small section of the margin tissue is separated...

21. Bole Sampling for Phytophthora ramorum The sample is imbedded in a PARP plate and sent to the lab for culturing and DNA testing. Photo series by Karl Buermeyer, UC Cooperative Extension

22. Leaf Sampling for Phytophthora ramorum Select a fresh, representative sample of symptomatic plant parts, including some associated leaves and stems. Be sure to include some healthy tissue along with the diseased. The sample should be placed in a plastic bag and labeled with the date, genus, species, cultivar (if appropriate), and location. The sample should be kept cool, away from direct sunlight, and delivered to the diagnostic laboratory within 24 hours. Send samples to your state’s plant pathology diagnostics laboratory, an approved private laboratory, or contact your local Agricultural Commissioner’s office.

23. Review Questions 1. What is the outward manifestation of Phytophthora ramorum on bark hosts? A reddish, sticky ooze from otherwise intact and healthy looking bark 2. In bark cankers, where is the disease active, and from where on the tree should someone take a sample to send in for laboratory testing? The black zone line surrounding the diseased tissue (found under the bark) 3. Name two organisms that are secondary invaders on infected trees, and hasten their decline. Hypoxylon fungi and ambrosia beetles

24. 4. Where are symptoms of Phytophthora ramorum most likely to occur on California bay laurel leaves, and why ? Review Questions, continued On the lowest most portion of the leaf tissue, as it hangs, where water accumulates 5. Name two reasons why some hosts of Phytophthora ramorum are not good for sampling or indicating the presence of the disease. It can be difficult to isolate the organism from leaves; Some hosts get several other leaf diseases 6. What is the single most distinguishing characteristic of Phytophthora ramorum lesions on the leaves of most ornamental plants? Diffused margins of necrotic areas

25. 7. What genus of ornamental plant can be infected near the base, so that the whole plant dies at once? Review Questions, continued Viburnum 8. Why might it be difficult to find symptomatic leaves on a camellia or pieris, even though it is infected? Infected leaves often abscise quickly 9. Name two high risk areas for Phytophthora ramorum in a nursery. Low spots where water collects; Along the perimeter if there are infected hosts outside the nursery