Leaf spectroscopy of resistance to Ceratocystis wilt of 'Ohi'a.

Plant pathogens are increasingly compromising forest health, with impacts to the ecological, economic, and cultural goods and services these global forests provide. One response to these threats is the identification of disease resistance in host trees, which with conventional methods can take years or even decades to achieve. Remote sensing methods have accelerated host resistance identification in agricultural crops and for a select few forest tree species, but applications are rare. Ceratocystis wilt of 'ohi'a, caused by the fungal pathogen Ceratocystis lukuohia has been killing large numbers of the native Hawaiian tree, Metrosideros polymorpha or 'Ohi'a, Hawaii's most common native tree and a biocultural keystone species. Here, we assessed whether resistance to C. lukuohia is detectable in leaf-level reflectance spectra (400-2500 nm) and used chemometric conversion equations to understand changes in leaf chemical traits of the plants as indicators of wilt symptom progression. We collected leaf reflectance data prior to artificially inoculating 2-3-year-old M. polymorpha clones with C. lukuohia. Plants were rated 3x a week for foliar wilt symptom development and leaf spectra data collected at 2 to 4-day intervals for 120 days following inoculation. We applied principal component analysis (PCA) to the pre-inoculation spectra, with plants grouped according to site of origin and subtaxon, and two-way analysis of variance to assess whether each principal component separated individuals based on their disease severity ratings. We identified seven leaf traits that changed in susceptible plants following inoculation (tannins, chlorophyll a+b, NSC, total C, leaf water, phenols, and cellulose) and leaf chemistries that differed between resistant and early-stage susceptible plants, most notably chlorophyll a+b and cellulose. Further, disease resistance was found to be detectable in the reflectance data, indicating that remote sensing work could expedite Ceratocystis wilt of 'ohi'a resistance screenings.

Vacuum Steam Treatment of Metrosideros polymorpha Logs for Eradication of Ceratocystis huliohia and C. lukuohia.

A new and devastating disease, rapid ohia death (ROD), in Hawaii led to a state quarantine that regulates interisland transport of ohia wood and plant material to prevent spread of the causal pathogens. Heat treatments of ohia logs in commercial trade were considered for phytosanitary treatment. Vacuum steam (VS) was evaluated for its ability to eradicate the pathogens, Ceratocystis lukuohia and C. huliohia, in main stem logs from ROD-affected forest trees. Replicate loads of three debarked logs (24 to 43 cm in diameter, 1.7 to 2.0 m long) were VS treated at 56°C for 30 min (five loads) or 60°C for 60 min (four loads) at a sapwood depth equal to 70% of log radius. Percentage isolation of Ceratocystis from VS and ambient temperature logs before treatment and summarized by source tree ranged from 12 to 66% and 6 to 31% based on carrot baiting assays of tissue taken from outer and inner sapwood, respectively. No viable Ceratocystis was detected in sapwood locations for the 60°C/60 min schedule or inner locations for the 56°C/30 min schedule after treatment. Only one subsample (0.48%, n = 208) of the latter schedule treatment yielded Ceratocystis. Time needed for treatment ranged from 7.4 to 15 h for the 56°C/30 min schedule and from 8.6 to 19.2 h for the 60°C/60 min schedule. These results demonstrate that VS is an effective and efficient method for treating large-diameter ohia logs that mill owners and regulatory plant pathologists may consider for use in Hawaii.

Use of Semiochemicals for the Management of the Redbay Ambrosia Beetle.

This review highlights current advances in the management of the redbay ambrosia beetle, Xyleborus glabratus, a primary vector of the pathogenic fungus, Raffaelea lauricola, that causes laurel wilt. Laurel wilt has a detrimental effect on forest ecosystems of southeastern USA, with hundreds of millions of Lauraceae deaths. Currently, preventive measures mostly focus on infected-tree removal to potentially reduce local beetle populations and/or use of preventative fungicide applications in urban trees. Use of semiochemicals may offer an opportunity for the management of X. glabratus. Research on attractants has led to the development of α-copaene lures that are now the accepted standards for X. glabratus sampling. Research conducted on repellents first included methyl salicylate and verbenone and attained significant reduction in the number of X. glabratus captured on redbay and swamp bay trees treated with verbenone. However, the death rate of trees protected with verbenone, while lower compared to untreated trees, is still high. This work underscores the necessity of developing new control methods, including the integration of repellents and attractants into a single push-pull system.

Pathogenicity, Symptom Development, and Colonization of Metrosideros polymorpha by Ceratocystis lukuohia.

Extensive mortality of Metrosideros polymorpha (`ohi`a) trees has been associated with Ceratocystis spp. on Hawai`i Island and was named rapid `ohi`a death (ROD). Both C. lukuohia and C. huliohia have been associated with ROD, although C. lukuohia appears to be the more important pathogen. Crown observations and dissections of forest trees either wound-inoculated with, or naturally infected by, C. lukuohia were conducted to confirm pathogenicity and document patterns of host colonization. In pathogenicity trials, one of three and two of three trees inoculated with the fungus in February and August, respectively, exhibited crown wilt symptoms at 92 and 69 days after inoculation. Extensive, radial, black staining of the sapwood was found in main stems, while no crown wilt or xylem staining was found in control trees. Xylem staining, necrotic phloem, and fungus presence was noted in six trees inoculated in May to June and harvested 37 to 42 days later, and these observations were compared with those in two naturally infected trees felled in early August. Contiguous xylem staining was found in the main stems and into crowns of all diseased trees, while discontinuous streaks of xylem staining extended into the main forks and side branches. Necrotic phloem associated with xylem staining occurred on the lower stems of inoculated trees. Aside from the necrotic phloem and radial staining of the sapwood, symptom development in `ohi`a infected with C. lukuohia is similar to other systemic wilt diseases on hardwood trees. We propose Ceratocystis wilt of `ohi`a as the official name of the disease.

The Fungus Raffaelea lauricola Modifies Behavior of Its Symbiont and Vector, the Redbay Ambrosia Beetle (Xyleborus Glabratus), by Altering Host Plant Volatile Production.

The redbay ambrosia beetle Xyleborus glabratus is the vector of the symbiotic fungus, Raffaelea lauricola that causes laurel wilt, a highly lethal disease to members of the Lauraceae family. Pioneer X. glabratus beetles infect live trees with R. lauricola, and only when tree health starts declining more X. glabratus are attracted to the infected tree. Until now this sequence of events was not well understood. In this study, we investigated the temporal patterns of host volatiles and phytohormone production and vector attraction in relation to laurel wilt symptomology. Following inoculations with R. lauricola, volatile collections and behavioral tests were performed at different time points. Three days after infection (DAI), we found significant repellency of X. glabratus by leaf odors of infected swamp bay Persea palustris as compared with controls. However, at 10 and 20 DAI, X. glabratus were more attracted to leaf odors from infected than non-infected host plants. GC-MS analysis revealed an increase in methyl salicylate (MeSA) 3 DAI, whereas an increase of sesquiterpenes and leaf aldehydes was observed 10 and 20 DAI in leaf volatiles. MeSA was the only behaviorally active repellent of X. glabratus in laboratory bioassays. In contrast, X. glabratus did not prefer infected wood over healthy wood, and there was no associated significant difference in their volatile profiles. Analyses of phytohormone profiles revealed an initial increase in the amount of salicylic acid (SA) in leaf tissues following fungal infection, suggesting that the SA pathway was activated by R. lauricola infection, and this activation caused increased release of MeSA. Overall, our findings provide a better understanding of X. glabratus ecology and underline chemical interactions with its symbiotic fungus. Our work also demonstrates how the laurel wilt pathosystem alters host defenses to impact vector behavior and suggests manipulation of host odor by the fungus that attract more vectors.

Attraction of Redbay Ambrosia Beetle, Xyleborus Glabratus, To Leaf Volatiles of its Host Plants in North America.

The redbay ambrosia beetle, Xyleborus glabratus, is an important pest of redbay (Persea borbonia) and swamp bay (P. palustris) trees in forests of the southeastern USA. It is also a threat to commercially grown avocado. The beetle is attracted to host wood volatiles, particularly sesquiterpenes. Contrary to other ambrosia beetles that attack stressed, possibly pathogen-infected, and dying trees, X. glabratus readily attacks healthy trees. To date little is known about the role of leaf volatiles in the host selection behavior and ecology of X. glabratus. To address this question, an olfactometer bioassay was developed to test the behavioral response of X. glabratus to plant leaf volatiles. We found that X. glabratus was attracted to the leaf odors of their hosts, redbay and swamp bay, with no attraction to a non-host tree tested (live oak, Quercus virginiana), which served as a negative control. Gas chromatography-mass spectrometry (GS/MS) analysis of leaves revealed the absence of sesquiterpenes known to be attractive to X. glabratus and present in host wood, suggesting that additional leaf-derived semiochemicals may serve as attractants for this beetle. An artificial blend of chemicals was developed based on GC/MS analyses of leaf volatiles and behavioral assays. This blend was attractive to X. glabratus at a level that rivaled currently used lures for practical monitoring of this pest. This synthetic redbay leaf blend also was tested in the field. Baited traps captured more X. glabratus than unbaited controls and equivalently to manuka oil lures. We hypothesize that leaf volatiles may be used by X. glabratus as an additional cue for host location.