1. The Effects of Beech Bark Disease on the Health of American Beech (Fagus grandifolia) Trees in the College Woods Natural Area, Durham, NH Kevin McDermott, Natural Resources Department, kjm82592@gmail.com Fall 2013 Abstract: This study attempts to investigate the effects of stress caused by beech bark disease on the health of beech leaves and the storage of starch in the twigs of beech trees. After detecting the spectral reflectance patterns of unaffected, moderately affected, and severely affected leaf samples, the red-edge inflection point values for the severely damaged samples were the lowest on average. Tests examining the starch content in the twigs also revealed higher amounts of starch present in severely affected samples. Introduction: The primary goal of this project is to examine the precise effects of beech bark disease (BBD) on the health of mature American beech (Fagus grandifolia) trees. Specifically, this study will examine the effects of BBD on the physiology of beech leaves and on the storage of starch in the twigs of this species. There are two main hypotheses to be tested in this study: 1.The spectral reflectance curves of beech leaves collected from severely affected trees will most likely reveal significantly lower red-edge inflection point (REIP) values and a lower near-infrared reflectance than that observed in the leaves of unaffected trees. 2.Starch content found in the twigs and buds of beech trees severely affected by beech bark disease will be significantly lower than that found in the twigs and buds of unaffected trees. A similar study comparing starch content in sugar maple (Acer saccharum) twigs between stressed and healthy trees found a significant reduction in starch within the stressed twigs. Cells located directly below the apical meristem in these stressed twigs contained almost no starch, while the same cells in the healthy twig contained abundant amounts of starch (Carlson, n.d.). Witter et al. (1983) also noted a significant pattern in the annual growth increment of trees affected by BBD. Infected beech trees displayed a reduction in radial growth by as much as 20-40% in comparison to unaffected trees on sites in New Hampshire and New York, while similar results found in studies in Michigan found a 19% growth reduction (Witter et al. 1983). By measuring and comparing the health of the leaves and carbohydrate storage in the buds and twigs of beech trees among various disease classes, we may find other effects resulting from BBD that would likely explain the annual growth increment decline observed in previous studies. Methods: Leaf and twig samples were collected from six individual beech trees in College Woods, Durham, NH. Samples were collected from two trees within each of the three BBD damage classes, including unaffected, moderately damaged and severely damaged, based on the presence of the scale insect and deformation or cankers on the bark. Each sample was collected from the lowest and most easily accessible branches, which typically occurred at between 20-30 feet from ground level on the mature beech trees. These samples were collected during the last week of September. Leaves from each sample were scanned using a VIRIS (Visible-Infrared Intelligent Spectrometer) to produce a representative spectral reflectance curve for each of the six samples. Values such as the red-edge inflection point (REIP) and ratios such as the normalized difference vegetation index (NDVI), TM band 4/3, TM band 5/4, and NIR 3/1 were calculated by the VIRIS software and averaged for each disease class. The VIRIS is a very useful tool for examining the spectral reflectance of vegetation, and has been used in previous studies for this purpose with a high degree of success. A paper written by Rock et al. (1986) highlights the advantages of using remote sensing technology such as the VIRIS for assessing forest damage. Higher reflectance values in the NIR plateau detected by the VIRIS were found to be an indicator of healthy leaf tissues in Picea rubens (red spruce) samples from a study of acid cloud damage on Camels Hump in Vermont (Rock et al., 1986). Twigs were selected from each sample and an analysis of starch content was conducted using two methods. The first method involved staining radial bisections of the twig and bud samples with an iodine solution to reveal relative amounts of starch contained in the pith. A sample representing each disease class was selected and photographed for comparison. A scanning electron microscope (SEM) was also used to quantify the amounts of starch contained within individual cells in each of the samples on a much larger scale. Results: Spectral reflectance curves produced by the VIRIS scans did not appear to indicate any significant patterns or correlation between stress level and relative amount of reflectance in the NIR plateau (Figure 1). While there was also no apparent difference in the NDVI or the TM band 4/3, 5/4, and NIR 3/1 ratios, there was an observable difference in the average REIP values (Table 1). The average REIP values ranged from 727 nm in the unaffected and moderately damaged samples to 724.65 nm in the severely damaged samples (Table 1). Results from the iodine test revealed significant differences in the amounts of starch contained in the twigs of the unaffected samples versus that contained in the moderate and severely affected samples. The severely damaged sample seemed to have much greater concentrations of starch in comparison to the moderate and unaffected samples, particularly in the region directly below the apical meristem (Figure 2). Unaffected samples tended to show very little to no starch content in this region of the stem. The results of the iodine staining test were confirmed by the images taken with the SEM (Figure 3). Figure 2. Comparison of starch content between an unaffected (A), moderately damaged (B), and severely damaged beech twig (C) following iodine staining. The severely damaged twig appears to contain a significantly higher amount of starch, particularly near the apical meristem. Figure 3. SEM images comparing abundance of individual starch grains between the unaffected (A, C) and severely damaged (B, D) twig samples. The portion of the stem closest to the apical meristem in the severely damaged sample (B) seems to show a greater number of starch grains in each cell when compared to this same region observed in the unaffected stem (A). A similar observation was made in the lower portion of the stem between the unaffected (C) and severely damaged (D) samples. The size of the parenchyma cells in the severely damaged sample also appear significantly larger, considering that both sets of imagery were taken at the same scale. Discussion: The results collected from the VIRIS spectral scans seem to confirm the hypothesis that REIP values would be lower in the leaf samples that experienced higher levels of damage from BBD. While the average REIP of the severely damaged leaf samples differed by only a few nanometers, this difference is enough to conclude that the stress caused by BBD may be causing a decline in the chlorophyll production of the beech tree (Table 1). The hypothesis that a significant decline in near- infrared reflectance would also be observed in the severely damaged samples could not be confirmed by the results obtained from the VIRIS scans. Based on the spectral reflectance curves produced by the VIRIS, there did not appear to be any observable difference in the relative levels of near-infrared reflectance between disease classes (Figure 1). The results of the iodine staining and SEM imagery contradict the second hypothesis, which assumed less starch content in the severely damaged samples. Starch content appeared to be significantly greater in the twig samples of severely infected trees, which contradicts the findings of the study by Carlson (n.d.) on starch in maple buds (Figure 2, 3). This may indicate some type of compensatory mechanism by the tree in order to counteract the damage caused by BBD. Higher amounts of calcium oxalate crystals observed in the severely damaged twig samples also indicate that the tree was undergoing greater levels of stress as a result of BBD. Conclusions: While the average REIP value was lower in the severely damaged samples indicating a potential decline in chlorophyll production, this value was still within the range considered to be healthy. BBD did not seem to have a significant impact on the NIR reflectance and thus the overall health of severely affected beech leaf samples. The stress caused by BBD may have stimulated increased production and storage of starch in the pith of severely damaged beech twigs in order to ensure adequate growth of new leaves next season. While these conclusions provide some interesting insights and raise new questions regarding the effects of BBD on the physiology of beech trees, further investigation will be needed to confirm these conclusions. Due to the small number of samples and potential for human error, the significance of these results cannot be fully tested or confirmed. The methods used for estimating the relative abundance of starch, such as the SEM imagery, were also fairly subjective and strictly based on observation. Further investigation of starch content and spectral reflectance patterns will be necessary to better understand how BBD impacts the health of beech trees on a cellular level. References: Carlson, M. n.d. A study of leaf abscission and twig starch. Natural Resources and Earth Systems Science Program, UNH, Durham, NH. Rock, B. N., J. E. Vogelmann, D. L. Williams, A. F. Vogelmann and T. Hoshizaki. “Remote Detection of Forest Damage.” BioScience, Vol. 36, No. 7, Ecology from Space (Jul. - Aug., 1986), pp. 439-445. Witter, J. A., J. L. Stoyenoff, H. A. Petrillo, J. L. Yocum, and J. I. Cohen. (1983). “Effects of Beech Bark Disease on Trees and Ecosystems.” University of Michigan, School of Natural Resources and Environment. Starch Grains AB C D A B C