Effects of Non-native Riparian Tree Species on Soil Microbial Community Activity

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1 Effects of Non-native Riparian Tree Species on Soil Microbial Community Activity Russian olive (Elaeagnus angustifolia) on the right and Plains cottonwoods (Populus deltoids) on the left co-existing at Riverfront Park, Billings, MT. Ksenia Lynch Dan Albrecht Rocky Mountain College Rocky Mountain College Yellowstone Research Center Professor of Biology 1511 Poly Drive 1511 Poly Drive Billings, MT Billings, MT Please cite this report as: Lynch, Ksenia and D. Albrecht Effects of non-native riparian tree species on soil microbial community activity. Report YRRC Submitted to RMC SEED Fund and the Yellowstone River Research Center, Rocky Mountain College, Billings, MT. Pp P a g e

2 Introduction Non-native species are of great concern because of the variety of ways they impact the ecosystems they invade. Non-natives can decrease agricultural yields, pose significant public health risks, and influence ecosystems by physically altering habitats, hybridizing with native species, and displacing native species (Livingston and Osteen 2008). When native species are displaced, additional indirect effects can then spread throughout the system, influencing nutrient cycling and altering ecosystem stability. This variety of effects results in billions of dollars spent annually to control the spread of non-natives (Moline and Poff 2008). In Montana, Russian olive (Elaeagnus angustifolia) is a non-native species of special concern. Introduced into the region only about 140 years ago, it is now the third most common riparian tree species and is widely credited both with causing the decline in native cottonwoods (Populus spp.) and altering plant and animal communities in riparian forests (Friedman et al. 2005). Studies of invasive species revolve around two main questions. First, how do invasive species affect the systems they invade? Second, how do invasives become established so successfully and so quickly? This study combines these two issues in an investigation of a possible trait that could have broader effects on the native systems, allowing for successful invasion. As mentioned above, Russian olive affects the systems it invades through directly competing with and displacing the native cottonwood species. Additionally, Russian olive has indirect effects that spread throughout the system. For example, Kinkade (pers. comm) observed the Russian olive leaf packs attract a greater abundance of macroinvertebrates and decay more quickly than cottonwood leaves. Additionally, Obritschkewitsch (pers. comm) observed increased CO 2 emission from soils beneath Russian olive trees. Finally, Albrecht (pers comm) had noted that Russian olive leaves suffer less damage from herbivores than cottonwood leaves. These observations are all consistent with the idea that Russian olive has both direct and indirect effects on the systems it occupies. The second important question with invasive species is how they become established so successfully and so quickly. Russian olive possesses a number of traits that contribute to its success: thorns, shade tolerance, nitrogen fixing, high lignin, and high cellulose (Katz and Shafroth 2003). There is also the possibility that Russian olive is successful because it produces allelochemicals. Allelochemicals are substances secreted by a plant as a defense mechanism against herbivores or competing plants. In two previous projects, it was observed that simple extracts from Russian olive leaves can inhibit germination of other plant species that could compete with Russian olive as well deterring invertebrate herbivores that may damage Russian olive (Albrecht pers comm). This project investigates the possibility that allelochemicals produced by Russian olive may alter the activity of the soil microbial community and in turn the nutrient cycling in the invaded ecosystem. Soil microbes (producers, consumers, and decomposers) are an important part of ecosystem function. A new species of plant in the community of producers might influence ecosystem function (Donald and William 2003) if its allelechemicals alter the activity of the community of soil microbes. Two hypotheses were examined to explain how Russian olive might influence the activity of microbial community in the soils it occupies. As mentioned above, Russian olive is a nitrogen fixer, and its leaves contain 2-3X more nitrogen than cottonwood leaves (Moline and Poff 2008). H1: The higher nitrogen content of Russian olive leaves might stimulate higher rates of soil microbial activity. Alternatively, H2: Russian olive produces allelochemicals and these chemicals might inhibit soil microbial activity. Page 2

3 Methods Area Sampled Soil samples were collected at Riverfront Park, Billings, MT ( , ) and Kindsfater Wetlands, Laurel MT ( , ) where Russian olives and plains cottonwoods (Populus deltoids) occur (Figure 1). Samples were collected from the base of neighboring trees that did not have overlapping crowns, but with the rim of canopies never further than five meters away. At each site, measurements for soil temperature, soil ph, and tree DBH were recorded. Using a soil core an estimated 10 grams of soil was acquired in the morning and transported back to the lab within the hour. To determine the soil moisture content for each sample, four grams of sample soil were removed and dried at 60 C, After 24 hours the soil was reweighed to determine the gravimetric water content. Figure 1. Map of two study areas. Another 4 grams of soil from each sample was mixed with 32ml of sterile 0.85% NaCl. After, the solution was placed on a shaker for three hours in order to dispense as many microbes into solution as possible. The solution was then centrifuged at 4400 rpm for three minutes. The supernatant was used to inoculate the Biolog Ecoplates with 135μl per well. The plates were incubated at 20 C and read with a 96-well Microplate Manager 6 Reader at 595nm every 12 hours for three days. Biolog Ecoplates A single plate is comprised of 96 wells, with each well containing a specific carbon source (31 carbon sources, repeated 3 times, including 23 chemical sensitivity assays and 3 deionized water controls) and tetrazolium dye (Figure 2). The dye turns purple in the presence of NADH, which is a product of metabolism. This color change indicates the extent to which the carbon source is metabolized directly relating to the level of microbial activity in that well. Page 3

4 Microplate Manager 6 produced a spreadsheet of the productivity numbers. To determine standard productivity of any given Biolog plate, all the wells were averaged on that Biolog plate, and then the three control wells were subtracted (Figure 2). Figure 2. A basic layout of the Biolog Plate pictured in the top right corner. Bottom half illustrates an example of the Excel spreadsheet, showing deionized water controls indicated by the red arrows and the Ecoplate productivity number circled in red. Results The average productivity number for the Russian olive sites at 12 hours was 0.02±0.007 and a slightly higher average productivity number of 0.03±0.011 for cottonwood sites. For the next 60 hours the average productivity numbers typically increased 0.125/12hrs for Russian olive sites and 0.119/12hrs for cottonwood sites (Figures 3). These results show no statistical difference in microbial activity between Russian olive (Elaeagnus angustifolia) and native cottonwoods (Populus spp.) soils. Page 4

5 Productivity # Figure 3. An average microbe productivity number of soils exposed to Russian olive and cottonwoods trees separated by 12 hours periods Time (hrs) RO CW Although there was no variance between the two species, the two zones sampled were notably different from each other (Figure 4). Within the Russian olive sites, the average productivity numbers from Riverfront park sites had higher values (by on average) versus those from Kindsfater wetland (Figure 5) p= Figure 4. Average microbe productivity number for soils exposed to Russian olive and cottonwoods trees separated by 12-hour periods and location. There was a similar trend with the cottonwood sites: Riverfront productivity numbers were on average higher than the ones from the Kindsfater wetland location with a p= A complete list of p-values can be found in Table 1. Page 5

6 Productivity # Figure 5. Average productivity number comparison of soils exposed to Russian olive and cottonwood at Riverfront Park and Kindsfater wetland in Billings, MT RO CW Riverfront Park Sample #'s Kindsfater Wetland Table 1. P-values for comparing the Riverfront Park and Kindsfater wetland locations, separated by time periods and tree species. Highlighted in yellow difference significant at 5%, and in green at 10%. Time RO CW 12hrs hrs hrs E-06 48hrs E-05 60hrs hrs Discussion The preliminary results from this study show no statistically significant difference in soil microbial activity between soils of Russian olive and native cottonwood species thus H1 and H2 were not supported. H1: Russian olive might stimulate higher rates of soil microbial activity due to the higher nitrogen content in its leaves. H2: Russian olive might inhibit soil microbial activity through the production of allelochemicals. It is important to note that this study only examined two areas, 11 miles apart and both approximately 26 acres. River Front Park in Billings, MT has an established riparian community for both Russian olive and cottonwood trees. It also contains a few substantial water sources: the Yellowstone River, Lake Josephine and a number of small creeks. Kindsfater wetland in Laurel, MT, the second area sampled, lacks large bodies of water. It served as a gravel mining operation until 1987, and in 2002 identified as a potential wetland restoration site with work to develop the site as a wetland bank under progress. The tree communities at Kindsfater are thus only recently established and the soil composition may differ greatly since topsoil was removed to access the gravel. Page 6

7 An attempt was made to evaluate some of these variables by examining average microbial activity numbers in relation to soil moisture, temperature, ph, and DBH, but no positive correlation was found. However, the significant difference between the two sites should not be ignored. One way of interpreting these results is to speculate that the greater nitrogen content did not stimulate and the allelochemicals did not inhibit the soil microbial activity. If one mechanism stimulates and the other inhibits, the two factors may result in negligible changes in overall microbial activity. Kruse et al (2000) found evidence that K. angustifolia exhibits mechanisms of germination inhibition of native riparian species by suppressing of nitrogenfixing bacteria. The interaction between plant chemicals and soil nitrogen-fixing microorganisms could potentially be an interdependent mechanism keeping the soil environment at equilibrium. In this context, although the overall microbial activity would remain unaffected, the composition of soil microorganisms could vary drastically, as the nitrogen-fixing bacteria could be displacing the native microbes for those soils. Finally, there is the issue of soil microbial community composition. Russian olive s potential allelopathic mechanism works through releasing chemicals into the surrounding soil environment by leaching, root exudation, volatilization, residue decomposition, and other processes (Ehrenfeld et al. 2003). In recent years, work has been done on another allelopathic shrub, Polygonella myriophylla. The chemicals this plant secretes are hydroquinone and gallic acid. Microorganisms in scrub soils have been shown to convert these compounds to forms they are able to utilize (Weidenhamer and Romeo 2004). It is possible that the soil samples examined in this study were only similar in the total microbial activity levels because a certain species of microorganism was thriving, while potentially many other species were absent. To assess this new hypothesis future studies should attempt to initially examine the already existing Biolog data, focusing on the metabolic patterns and eventually conducting new experiments evaluating the microbial community composition in greater detail. Acknowledgements These experiments were carried out with support from the Montana Space Grant Consortium, Yellowstone River Research Center, and Rocky Mountain College SEED Grant. The results were presented at the Murdock College Science Research Program Conference, Annual Yellowstone River Research Center Student Forum and the Montana Space Grant Consortium Student Research Symposium. Page 7

8 Literature Cited Ehrenfeld, J. G Effects of exotic plant invasions on soil nutrient cycling processes. Ecosystems 6 (6): doi: /s Friedman, J., G. Auble, P. Shafroth,, M. Scott, M. Merigliano, M. Freehling and N. Griffin Biological invasions 7.4: Katz, L. and P. Shafroth Biology, ecology and management of (Elaeagnus angustifolia L). Russian olive in western North America. Wetlands 23(4):763. Kaur, H., R. Kaur, S. Kaur, I. Baldwin and T. Inderjit Taking ecological function seriously: soil microbial communities can obviate allelopathic effects of released metabolites. PLoS ONE 4:e4700. Kruse, M., M. Standberg and B. Strandberg Ecological effects of allelopathic plants a review. National Environmental Research Institute. NERI Technical Report No. 315, Silkeborg, P66. Moline, A.B. and N.L. Poff Growth of an invertebrate shredder on native (Populus) and non-native (Tamarix, Elaeagnus) leaf litter. Freshwater Biology 53: Weidenhamer, J.D. and J.T. Romeo Allelochemicals of (Polygonella myriophylla) chemistry and soil degradation. J. Chem. Ecol. 30: Zak, D., W. Holmes, D. White, A. Peacock and D. Tilman Plant Diversity, Soil Microbial Communities, and Ecosystem Functions: Are There Any Links? Ecology 84: Page 8