Hemlock woolly adelgid resistance E. Preisser, B. Maynard, R Hemlock woolly adelgid resistance E. Preisser, B. Maynard, R. Casagrande The hemlock woolly adelgid Adelges tsugae (HWA) attacks and kills both eastern hemlock (Tsuga canadensis) and Carolina hemlock (T. caroliniana). Both hemlock species are considered completely susceptible to this invasive insect, and there has been no report of HWA-resistant individuals of either species. No reported resistance in Tsuga canadensis or T. caroliniana

Western hemlock W. hemlocks resist eastern HWA Hemlocks CAN resist adelgids, however; for example, the western hemlock (found on the west coast of the US) is resistant to the western ‘strain’ of HWA and to the eastern ‘strain’ of HWA that kills eastern and Carolina hemlocks. Eastern hemlocks are also not hugely harmed by the western strain of HWA. W. hemlocks resist eastern HWA and E. hemlocks resist western HWA

Mountain hemlock Difficult to grow in northeast The mountain hemlock is also HWA-resistant; because it is difficult to grow in the northeast, however, it isn’t a suitable replacement for the native hemlock species. Difficult to grow in northeast

Chinese and Japanese hemlocks are HWA-resistant Same thing for Chinese and Japanese hemlocks: HWA-resistant but difficult to grow in the northeast. Also, they are very different in appearance from eastern hemlock and so often don’t work as a landscaping replacement for eastern hemlocks. Chinese and Japanese hemlocks are HWA-resistant (But poor choice for landscape or forest)

Are some eastern hemlocks resistant to HWA? Todd Caswell Richard Casagrande Laura Ingwell Brian Maynard Evan Preisser Laura Radville While most HWA researchers feel that adelgid resistance does not exist in either eastern or Carolina hemlock, a group of researchers at the University of Rhode Island has been working to experimentally test whether very rare individuals (1 in 10,000, say, or 1 in 100,000) might possess rare mutations that provide some degree of HWA resistance.

Preisser et al. (2008) sampled 142 hemlock stands in 2005 This work started in 2005, when researchers surveying HWA-devastated hemlock stands discovered some mature hemlock trees that appeared unaffected by HWA. These trees were growing in stands where >95% of the mature trees were dead (and the rest were rapidly dying); despite the obvious presence of HWA, however, these rare individuals didn’t have HWA and appeared healthy.

“Potentially resistant hemlock” In FOREST stands with ≥95% mortality Mature (>10 m) trees Appeared relatively healthy largely free of HWA and elongate hemlock scale INNATE OR ENVIRONMENTAL? After completing the 2005 survey, URI researchers were interested in following up on these observations… had the trees just gotten lucky, was it some environmental difference, or did the trees possess some degree of innate resistance? In order to address this question, researchers first had to decide how to determine whether a surviving tree might in fact be putatively resistant. They did this by establishing a series of criteria that candidate trees had to meet in order to be considered potentially resistant.

Evaluating hemlock resistance Propagate cuttings Grow under identical conditions Infest with HWA under controlled conditions Once a tree was identified as having the potential for HWA resistance, the degree of resistance had to be evaluated under controlled conditions. This meant taking cuttings (i.e., plant material that is genetically identical to the parent tree), growing these cuttings in the greenhouse along with cuttings from HWA-susceptible ‘control’ trees, and testing the cuttings from both putatively-resistant and control trees with HWA in order to see whether or not HWA survives on them.

CT sample sites The researchers identified three CT sites that had a few trees that fit the listed criteria.

Burnham Brook, CT There were two trees in Burnham Brook CT (note: these later turned out not to be resistant)

Lyme, CT Three trees from Lyme CT (these also turned out not to be resistant)

Madison, CT And several trees from a relatively wet site in Madison CT (they look like they’re growing in the open, but they’re not; it’s just that all of the hemlocks around them have died and new trees haven’t yet grown up). Trees from this site are the most promising… and the ‘mother’ trees at this site still look great. To see other trees that look this good, you’ve got to travel to northern MA/southern VT.

Rooting hemlocks Two variables: time of season and hormone treatments This was the first time the researchers had worked with hemlock rooting, so a masters student (Todd Caswell MS ‘07) took cuttings from the parent trees at different times of the year and treated them with different rooting treatments (i.e. different mixes of hormones).

Propagation bed Once the cuttings had been taken and treated with the rooting hormones, they were placed in a propagation bed at URI’s East Farm (equipped with misting jets and bottom heating) and allowed to grow.

January 2006 cuttings Cuttings were allowed to grow over the winter of 2006.

Spring 2006 root ratings 0-3 In spring 2006, all of the cuttings were removed from the mass-rooting bed and given a root rating of 0 (none) to 3 (abundant roots).

Results of 2006 winter cuttings (number rooted/number treated) SITE CODE HORMEX 45 1:1 DIP 'N GROW 1% KIBA HORMODIN 3 CONTROL AA 0/18 12/18 2/18 0/10 BB 1 11/17 14/17 6/17 8/17 1/17 BB 2 9/15 8/15 5/15 10/16 2/12 M 1 2/14 6/15 0/15 NA M 2 5/7 4/7 0/7 OL 1 3/7 1/6 2/6 0/6 OL 2 0/17 3/17 0/16 Totals 30/95 48/95 20/95 18/87 3/47 % Rooted 31.58% 50.53% 21.05% 20.69% 6.38% A 1:1 solution of Dip ‘N Grow and water gave the best rooting results.

2007 rooting results Same thing in 2007. Treatment 0 = Control Treatment 1 = 1:1 ratio of Dip ‘n Grow to water Treatment 2 = 1:2 ratio of Dip ‘n Grow to water

Rooted cutting in June Rooted cuttings were allowed to grow for a year; they initially put on growth belowground, but began later in the season to produce new foliage.

HWA inoculation Once the cuttings were well-established, we applied HWA-infested branches to each of them to assess whether cuttings from ‘resistant’ trees would have a lower HWA density than cuttings from control trees.

Inoculated plant A closeup of a cutting with a HWA-infested branch applied to it (this picture makes the cutting look a lot worse than it is; the basically healthy cutting is obscured by the extremely unhealthy HWA-infested branch).

2006 HWA results The results of the 2006 experiment were intriguing; even though we had low sample sizes, trees that had been identified as potentially HWA-resistant had a substantially lower density of settled HWA than did control trees (we also tested a tree from the Arnold Arboretum, far left, and two western hemlocks, far right). Numbers above each bar give the number of cuttings tested from that site. 3

Mark Mayer’s NJ “bulletproof stand” We talked about our findings at the 2007 HWA symposium, and invited people who thought that they might know of potentially-resistant trees to contact us. One person who did was Mark Mayer, a NJ state forester. He told us about a ‘bulletproof stand’ in NJ that still appeared healthy despite being surrounded by dead or dying hemlocks. This stand has been the source of some of our most promising cuttings. Courtesy of Mark Mayer

2007 sample sites 100 cuttings each from 6 CT Trees 5 NJ Trees (Mark Mayer) 2 PA Trees (Scott Stitzer) 1 MD Tree (Dave Mausel) 4 Control Sites (MA, NY) 18 Trees (1800 cuttings) In 2007, we did large-scale sampling of trees that seemed to be the most likely candidates for HWA resistance (we also sampled trees that were known to be HWA-susceptible as controls). We took 100 cuttings from 18 trees scattered across six states, treated them all with rooting hormone, and grew them at URI under controlled greenhouse conditions.

2007 rooted cuttings held for testing in spring 2009 Allow additional time to recover from rooting and outgrow field environmental effects Provide 12 months of growth under standardized conditions Improve timing of plant growth and HWA inoculation Because our initial experiment had been criticized for exposing the cuttings too quickly to HWA infestation, we grew the winter 2007 cuttings for two years until testing them in spring 2009. This allowed the cuttings to successfully root, put on new foliage, and acclimate to standardized greenhouse conditions.

HWA inoculation tests – 2009 Inoculate all rooted cuttings with HWA in early spring 3 groups: control, resistant, and cuttings from western hemlocks Assessed HWA settlement and survival In spring 2009, we repeated the 2006 experiment with much greater sample sizes and better control over experimental conditions. Cuttings from western hemlocks were included to confirm that the colonizing HWA performed poorly on them.

2009 HWA inoculation results We found that HWA both avoided colonizing and did poorly on western hemlocks. HWA settlement on the control and ‘resistant’ eastern hemlock cuttings did not differ, but HWA survival was much lower on the resistant cuttings than on the controls.

2010 HWA inoculation results 6 4 HWA crawlers/cm) We conducted an additional experiment in 2010 that found lower HWA settlement on resistant versus control cuttings. 2 Resistant Control

Progress towards providing nurseries with plants Planning third large-scale test of HWA resistance in spring 2011 Carried out large-scale grafting experiment in 2010 – results in summer 2011 Set up ‘hemlock hedge’ of resistant trees for cuttings and assessment of growth form and HWA resistance So, where to go from here? We’d like to do another large-scale test of HWA resistance in the cuttings we’ve made, see if grafted plants work as well as rooted cuttings, and set up a ‘hemlock hedge’ of resistant trees to assess the growth form of the different accessions and to confirm that, when grown outside, that naturally-colonizing HWA doesn’t settle on them. If you’re interested in this work and/or would like more information, please contact Evan Preisser (preisser@uri.edu) or Dick Casagrande (casa@uri.edu).