Testing Procedures
for BMPs for Producing Clean Nursery Stock

3.3.  Individual plant sampling/baiting

Version 8/24/2017

Application:  Testing symptomatic or suspect individual plants.  Tests entire root system of individual plants for the presence of Phytophthora by detecting zoospores released after a flooding period.

Advantages:  Fairly sensitive test of entire root system, relatively easy to conduct, plant does not have to be unpotted.  Nondestructive and does not expose other plants to inoculum.  No false positives if bait spots are confirmed by culturing.  Culturing can identify Phytophthora species present. 

Limitations:  Positive reactions require at least several days to develop and can take longer to confirm depending on methods used.  Baiting conditions (temperatures, irrigation history) can affect test sensitivity.  Previous application of fungicides (systemic oomycete suppressive chemicals) to tested plants may interfere with test.  No single bait is optimal for all Phytophthora species.  Not suited to testing large numbers of plants.  This protocol is designed for plants that have been regularly irrigated up to the time of testing.  It may not be effective for plants that have been dry for an extended period (e.g., nonirrigated culls) because sporangia may not be present in such material. This method is easiest to use for containers up to about 1 gallon. For larger containers (e.g., 5 gal, 15 gal), we recommend using a single container version of the bench testing leachate test short duration protocol.

Methods:  Standard baiting uses green, nonwounded pears, which can be infected by a wide variety of Phytophthora species.  Leaf baits can also be used if you have arrangements for testing these baits. To minimize the possibility of cross-contamination between plants, wear waterproof gloves when handling containers and pear baits. Sanitize gloves before handling a different plant or other items assocaited with them.

Place the container of each plant to be tested in a heavy-duty plastic bag (e.g., 1 gallon Ziploc® freezer bags for 1 gallon pots or smaller stock).  If the container has enough room on top, use a clean waterproof glove to make a slight depression in the potting media in the top of the pot and place a rinsed, labeled, unwounded green pear in the depression (Figure 2A). Avoid scratching the pear if the potting media contains abrasive components such as perlite or lava rock by setting it lightly in the depression. If using a smaller container, place the pear in the bag alongside the container (Figure 2B). Leaf baits, if used, are added to the top of the container or the bag at this time. Irrigate the plant with clean water until water accumulates in the bag up to the depth of the top of the soil (Figure 2A,B).   Water temperature should be in the range of 18-24°C (65-75°F).  Do not expose the flooded plants to direct sun that could allow the water to heat up excessively. 

Leave plants flooded in the bags for one to two hours.  At the end of this period, remove the pot from the bag and allow the excess water to drain from the pot into the bag.  You will need to support the bag by placing it in a plastic container, such as a cut off 1 gallon plastic bottle (Figure 2C).  Collect drain water from the plant container until dripping has stopped or nearly stopped.  The total water volume from each pot will vary by pot size, but typically should be between 1 and 2.5 L (about 1-2.5 quarts).  If the pear bait was in the top of the container, transfer it to the water in the bag using a clean waterproof glove immediately after the pot is removed.  Wash, sanitize, and rinse your gloves before handling another test plant. 

This test can also be conducted by flooding the plants as described above and adding the pear bait at the end of the flooding period. However, in plants producing low amounts of inoculum (e.g., recently-infected plants), adding the pear at the end of the flooding period may reduce test sensitivity.

Incubate and evaluate pears as discussed above in section 3.2. Detection by baiting – general procedures.

Results:  Confirmed positive results provide proof that the tested plant is infected.  Adjacent plants should be considered to be at risk of infection.  They may not provide positive results if the plants have been infected only recently.  Negative results from a single test should be interpreted with caution. 

Greater confidence can be associated with negative tests from nurseries that are stringently following the procedures outlined in Best Management Practices (BMPs) for Producing Clean Nursery Stock.  If plants are targeted for use in sensitive or other non-infested sites, a second test conducted at least 1 week after the first, and testing with additional bait types, should be conducted to provide a higher level of confidence.

individual pot baiting 1 gal

Figure 2A. Baiting of individual flooded plants.  Water is added to pot until water level is at or slightly above the top of soil in the pot. Pears have been placed in each container before flooding.

individual pot baiting with pear in bag

Figure 2B. Baiting of individual plant by flooding. For small containers, the pear bait is added to the bag rather than placed into the container top.

pear bait in leachate

Phytophthora infected pear bait

Figure 2C.The bait is incubated in the water (left) until symptoms develop or for up to 3 days.  Pear bait on right was removed 2 days after being placed in water from an individual pot test of a Ceanothus and shows extensive symptom development.  Note that most of the of the infections are associated with lenticels in this pear.

Updated 8/24/2017 - added images, updated to show adding pears to pots/bags during flooding period