Black 1

Effects of Schizachyrium scoparium coverage on soil and food web structure in coastal plain

Max Black1, Linda Deegan2, Chris Neill2, Marta De Giuli3

1. Franklin & Marshall College, Lancaster, PA 17603 2. Woods Hole Research Center, Falmouth, MA 02540 3. University of Chicago, Chicago, IL 60637, USA.

Black 2

Abstract

Coastal sandplain grasslands serve as hubs for and provide a unique habitat for rare and threatened . In recent years there have been major efforts to bring back grasslands since their decline caused by human development.

Management strategies, namely controlled burning, often focus on promoting warm season grasses such as Schizachyrium scoparium (little bluestem), a perennial bunch grass native to most of . The goal is to maintain the grasslands and prevent invasion trees and other late successional species. We are interested in examining the effects of increased little bluestem coverage on soil organic matter, invertebrate diversity and abundance, and determining if there is a shift in the plants at the base of the food web.

We studied a sandplain at Crane Wildlife Management Area in

Falmouth, Massachusetts, focusing on five sites that vary in little bluestem coverage.

Since little bluestem is the only major C4 plant in the grassland, we used the difference in carbon isotope fractionation to discriminate between C3 plant derived carbon and C4 plant derived carbon. We took 30 cm soil cores to analyze total soil organic matter and

ẟC13 by depth to determine the proportion of little bluestem derived soil carbon. We employed pitfall traps and berlese funnels to assess invertebrate diversity and abundance, and analyzed several species for ẟC13 to assess the impact of percent little bluestem coverage on their diets.

We found less soil organic matter in the areas with greater than 50% little bluestem coverage and greater ẟ13C values for areas with greater than 50% little bluestem coverage. There was less invertebrate diversity in plots with greater little

Black 3

bluestem coverage. There was an increase in invertebrate abundance as little bluestem coverage increased from 0-49% little bluestem coverage, and a decline in abundance as little bluestem coverage increased from 50-100%. The ẟ13C values of the sow bugs and woodlice (detritivores) increased as the little bluestem coverage increased. The

ẟ13C values of the (herbivores) remained relatively constant at around -

28 ppt among all of the cover types.

Little bluestem is contributing a significant proportion of soil carbon in areas with high bluestem coverage. Grasshoppers, an important direct herbivore in the food web, do not eat little bluestem; this could have implications higher up in the food web. There were less predators where there was higher little bluestem coverage, a sign of food web truncation. We observed a shift in ecosystem function, occurring at around 50% little bluestem coverage.

Key Words: coastal sandplain grasslands; biodiversity; ecosystem function; invertebrate; food web; soil organic matter; ẟC13

Introduction

Coastal plain grasslands, along with many open habitats, are historically maintained by natural disturbances such as fire, grazing, and salt spray (Dunwiddie et al, 1996). These disturbance cycles have been disrupted by increased development, causing a decline in coastal plain grasslands. In recent years, there have been major efforts to restore coastal plain grasslands, especially because of the ways that they promote biodiversity and are home to many rare and threatened species (Simmons and

Mazzei, 2014). One of the major ways wildlife organizations manage for grasslands is through the use of burning and other tactics to promote warm season grasses.

Black 4

Little bluestem is a warm season, perennial bunch grass that is native to most of

North America between and northern (Tober and Jensen, 2013). Often considered as a foundation species, it produces a large amount of seeds which provide nutritious meals for birds all across North America, and serves as an important food source for many grazing (Campbell, 2015). The clumped structure provides shelter for small mammals and ground nesting birds. Some animals that inhabit fields of little bluestem also feed on which feed on the grass’ inner shoots, or live within the grass. Little bluestem is more prevalent in upland open habitats like grasslands and shrub lands; habitats maintained by natural disturbances.

It’s widely understood that lower plant diversity is correlated with a decline in biodiversity and ecosystem services (SEP 2015). We wanted to assess the effects of increased little bluestem coverage on soil organic matter, invertebrate biodiversity and abundance, and if there is a shift in the plants at the base of the food web. Since little bluestem is the only major species using the C4 photosynthetic pathway in the grassland, we took advantage of the difference in carbon 13 isotope fractionation by using ẟC13 values as a proxy for carbon originating with little bluestem, or carbon originating with the C3 grassland species (O’Leary, 1988). Our research examined the soil, plant, and invertebrate composition of five sites of varying levels of little bluestem coverage within Crane Wildlife Management Area in Falmouth, Massachusetts.

Crane Wildlife Management Area is a large grassland in Massachusetts, with about 200 acres of open sandplain grassland; much of this area has high little bluestem coverage. Since the 1990’s, the Department of Fish and Wildlife has burned, mowed, and planted various native plants to expand and maintain the grassland ecosystem

Black 5

(Simmons and Mazzei, 2014). We examined five sites within the Crane WMA that vary in little bluestem coverage, to assess the impacts that higher coverage of little bluestem has on the soil and food web structure. We identified five 314 m2 sites with little bluestem coverage ranging from 4-15% to 75-100% coverage. We took soil cores from each site to assess the SOM and ẟC13 at four depths. Since little bluestem is the only major C4 plant in the ecosystem, ẟC13 will be used as a proxy for the amount of little bluestem derived carbon. Pitfall traps and berlese funnels will be deployed at each site to collect the invertebrates. We will assess invertebrate diversity and abundance, and determine the diets of several species using their ẟC13 values.

Methods

Field: We first visually identified five sites of varying little bluestem cover. We then marked each site with a surveyor’s flag and recorded the GPS coordinates. We used a tape measure and surveying tape to mark off a 10 meter radius circle using the flag as the center. Within these sites, we used a 3 x 3 meter PVC square to randomly select three quadrats where we estimated little bluestem coverage as well as percent coverage of other classes of plants, e.g. forbs, grasses, etc.

We took five soil cores to 30 cm within each site. We separated each core into four segments: 0-5 cm, 6-10 cm, 11-20 cm, and 21-30 cm and stored them in ziplock bags. These total to 100 soil samples. To deploy pitfall traps we placed 9 ounce plastic cups in the holes from which the soil cores were taken. We created a mix of 50 mL

Dawn Original dish detergent and 1 gallon of water, which we poured ~2 ounces of into each plastic cup. After 72 hours we retrieved the cups, replaced the lids, and returned them to the lab. To ensure that we found a sampling of invertebrates that was

Black 6

representative of the ecosystem, we collected five 10 x 10 cm quadrats of plant litter in ziplock bags to catch insects that reside on the plants and weren’t caught by falling into the pitfall traps; they will be processed via berlese funnels in the lab. Finally, we took clippings of little bluestem and several C3 plants to be used as baselines for C4 and C3

ẟC13 signatures in this specific grassland system.

Lab: We dried the clippings of Schyzarium scoparium, Rubus flagellaris,

Solidago canadensis, and Carex pensylvanica in a drying oven for 48 hours. We ground these clippings using a Wiglbug and analyzed them for ẟC13 (Otter, 2017). We dried the soil samples in a drying oven at 75℃ for 36 hours, removed roots and rocks and then ground them with a mortar and pestle to prepare them for loss on ignition and

ẟC13 isotope analysis. For all five cores from each site, we took subsamples from each depth and combined them according to depth, to make four aggregate samples per site.

This made a total of 20 soil samples for isotope analysis.

About 5 grams of dried and ground soil from each depth from each core were then placed in pre-ashed and pre-weighed aluminum tins. The initial weights of these samples were recorded. We then placed the tins in a muffle furnace set for 375℃ for 16 hours to burn off all organic material. After cooling for two hours, we reweighed the tins.

The difference between the initial weight and final weight was the weight of the organic matter in each sample (Robertson, 2011).

To isolate the soil surface invertebrates from the soapy water mixture, we poured the contents of each plastic cup through a mesh screen. After picking out large debris with tweezers, we rinsed the invertebrates with deionized water until no soapy water remained. We then rinsed with a small amount of 70% ethanol and then deposited the

Black 7

organisms into a glass jar with enough ethanol so that they were all covered. The organisms from all five pitfall traps for one site were combined into one jar, for a total of five jars. We identified the organisms using a microscope, several online references, and the help of Matt Pelikan, then recorded numbers of individuals as well as species for each site. Sow bugs, woodlice, and grasshoppers were abundant at all five of the sites; we chose to analyze these species for ẟC13. We dried the invertebrates in a drying oven for 24 hours and then ground them with a mortar and pestle.

To isolate the litter dwelling invertebrates, I built 10 berlese funnels (Figure 1)

These consisted of a plastic funnel lined with a 3 mm mesh screen to keep the litter from falling through the funnel. Underneath the funnel was a 1 L screw-cap bottle, with

~3 mL of 70% ethanol. We clipped a 65 watt incandescent light bulb directly above each funnel to provide a source of heat. The berlese funnels sat for 1 week with the lights constantly on and the ethanol replenished. The invertebrates collected through this method were identified in the same way as the pitfall trap invertebrates.

Results

We determined the average ẟ13C value of the three C3 plant species we collected to be -30 ppt, and the ẟ13C of little bluestem to be -12.2 ppt (Figure 2). We found there to be less soil organic matter in the areas with greater than 50% little bluestem coverage than in areas with less than 50% little bluestem coverage (Figure 3).

There were greater ẟ13C values for areas with greater than 50% little bluestem coverage (Figure 4). With greater soil depth, there was a slight shift towards higher

ẟ13C values for all five sites (Figure 5).

Black 8

There was less diversity in invertebrates in the plots with greater than 50% little bluestem coverage than the plots with less than 50% little bluestem coverage (Figure

6). There was a steady increase in invertebrate abundance as little bluestem coverage increased up to ~50% little bluestem coverage. As the sites increased in little bluestem coverage to greater than 50%, the invertebrate abundance steadily decreased (Figure

7). The ẟ13C values of the sow bugs and woodlice, which are detritivores, increased as the little bluestem coverage increased (Figure 8). The ẟ13C values of the grasshoppers remained relatively constant at around -28 ppt among all of the cover types (Figure 9).

Discussion

The ẟ13C values for C3 and C4 plants that we found were expected as compared to literature values (O’Leary, 1988) and used in further calculations in soil and invertebrate analysis. Combining the SOM values for the three lower coverage sites as one sample group, and the two higher coverage sites as another sample group, we ran a simple t-test and found a significant difference in total organic matter among these two groups. There was also a significant difference in ẟ13C values between these two sample groups. We averaged all of the ẟ13C values from all sites with <50% little bluestem coverage and sites with >50% little bluestem coverage and calculated the contributions of C3 carbon and C4 carbon based on the average C3 and C4 values from our plant isotope analysis. There was 16% little bluestem derived carbon in sites with less than 50% little bluestem coverage, and 35% little bluestem derived carbon in sites with more than 50% little bluestem. This is a significant difference in soil carbon contributions. Soil organic matter has a turnover time on the order of decades, and the little bluestem hasn’t present in this grassland for more than a few years. The

Black 9

difference in the contribution of C4 carbon is likely a function of the greater bluestem coverage, and a longer time scale. It is likely that the areas of higher little bluestem coverage have been managed and burned for longer periods of time, resulting in the cultivation of little bluestem.

The shift towards higher ẟ13C values in deeper soils is likely due to the very slight fractionation in carbon isotopes associated with trophic interactions (Post, 2002).

As soil microbes consume organic matter and respire, there is some discrimination between C12 and C13. The rooting depth of little bluestem can reach about four feet, resulting in a positive shift in ẟ13C values for areas of higher bluestem coverage at all depths in the 30 cm soil core (Weaver et al, 1935). The most organic matter is deposited at the soil surface, from above ground biomass, but the plant roots also contribute some organic matter to the soil (Weaver, 1935).

We determined the composition of the invertebrate community at each site in terms of percentage of predators, herbivores, detritivores, and carpenter . Only the invertebrates collected via the pitfall trap method were included, since we didn’t identify the invertebrates collected via Berlese funnels as closely, and knew less about them.

Predators include , , wasps, crickets, ants, centipedes, flies, and several parasitic species like mites, ticks, and fleas. Herbivores include grasshoppers, leaf- miner flies, honey bees, weevils, and skiff beetles. Detritivores include sow bugs, woodlice, millipedes, gastropods, cockroaches, and snails. Carpenter ants made up over 20% of the total species composition at all five sites. This could be attributed to the high availability of wood throughout the grassland, left over from the clearing of trees, and subsequent clearing of saplings that have continued to grow up throughout

Black 10

the years. There was a lower proportion of predators in areas of higher little bluestem coverage. This finding supports the idea that there is often a truncation of food webs in areas of decreased biodiversity (Ramus and Long, 2015). There were the greatest proportion of detritivores in the site with 25-49% little bluestem coverage; there wasn’t a major change in the number of detritivores between areas of higher and lower little bluestem coverage. There was not much of a difference in the proportion of herbivores across the gradient of bluestem coverage, although the sample size was quite small

(Figure 10).

Sow bugs and woodlice, also known as pill bugs, are detritivores, meaning that they primarily consume dead and decaying plant matter. Their ẟ13C values increased as little bluestem coverage increased; this indicated to us that the detritivores do not discriminate between eating little bluestem or forbs and other grasses. The detritivores ate more little bluestem when more little bluestem was available. The grasshoppers are direct herbivores. Their ẟ13C values remained relatively constant at around -28 ppt among all of the cover types which indicates that grasshoppers were not eating little bluestem- only the C3 forbs and grasses. While an important structural species for habitats, little bluestem isn’t a food source for some herbivorous species like grasshoppers. Grasshoppers are an important food source for birds, which many naturalists strive to protect. For this reason, nearing a monoculture of little bluestem or other C4 grass may mean a decline in available food for grasshoppers and perhaps other direct herbivores, causing further implications higher up in the food web.

Black 11

Conclusions

It is clear that at around 25-50% little bluestem coverage there is a change in the ecosystem function of a grassland. It would be beneficial to expand on this study to determine exactly when the change occurs, and what the implications are for biodiversity in terms of species richness, abundance, and evenness. Further insight into the optimal range of little bluestem coverage for grasslands will be crucial for future management of natural lands, to achieve peak biodiversity and ecosystem function.

Black 12

Acknowledgements

I would like to thank my mentors Linda Deegan and Chris Neill for their help designing and completing research on this project, as well as providing me with invaluable wisdom, resources, and experiences. Marshall Otter made this study possible by analyzing all of the isotope samples. I’d also like to thank Marta De Giuli for working closely in the field with me and helping to collect all of my samples. Matt

Pelikan and Michael Whittemore were both extremely helpful in assisting with species identification. Finally, my TA’s Richard McHorney, Jordan Stark, Em Stone, and Alana

Thurston for always being available and helpful answering questions and guiding me with lab work.

Black 13

Literature Cited

Campbell, Dana. September 22, 2015. Little Bluestem - Schizachyrium Scoparium - Brief Summary. Encyclopedia of Life.

Dunwiddie, Peter W.; Zaremba, Robert E.; Harper, Karen A. 1996. A classification of coastal heathlands and sandplain grasslands in Massachusetts. Rhodora. 98(894): 117-145.

O'Leary, Marion H. 1988. “Carbon Isotopes in Photosynthesis.” BioScience, vol. 38, no. 5, pp. 328–336. JSTOR

Otter, Marshall. 2017. Stable Isotope Lab. Marine Biological Laboratory. Woods Hole, Massachusetts 02543

Post, D. M. 2002. Using stable isotopes to estimate trophic position: Models, methods, and assumptions. Ecology, 83: 703–718.

Ramus, Aaron; Long, Zachary. 2015. Producer diversity enhances consumer stability in a benthic marine community. Journal of Ecology. 104. 572-579. 10.

Robertson, Sarah. June 27, 2011. Direct Estimation of Organic Matter by Loss on Ignition: Methods. SFU Soil Science Lab. Simon Frasier University. Burnaby, BC V5A 1S6, Canada

Science for Environment Policy May 2015. Ecosystem Services and the Environment. In-depth Report. Issue 11. Produced for the European Commission, DG Environment by the Science Communication Unit, UWE, Bristol.

Simmons, Time; Mazzei, Benjamin. March 2014. Biodiversity Initiative Site Plan, Sandplain Grassland & Pitch Pine/Scrub Oak Savannah Restoration. Frances Crane Wildlife Management Area North Falmouth, Massachusetts.

Black 14

Tober, D. and N. Jensen. 2013. Plant guide for little bluestem (Schizachyrium scoparium). USDA Natural Resources Conservation Service, Plant Materials Center, Bismarck, North Dakota 58501

Weaver, J. E.; Hougen, V. H.; and Weldon, M. D. 1935. "Relation of Root Distribution to Organic Matter in Prairie Soil". Agronomy & Horticulture -- Faculty Publications. 449.

Black 15

Figures and Tables

Figure 1. Diagram of a berlese funnel a picture of the 10 berlese funnels we set up in our lab.

Figure 2. ẟ13C values for Schyzarium scoparium, Rubus flagellaris, Solidago canadensis, and Carex pensylvanica.

Figure 3. Average percent soil organic matter for each of the five sites.

Figure 4. Average soil ẟ13C values for each of the five sites.

Figure 5. Soil ẟ13C values at four depths for each of the five sites.

Figure 6. Total invertebrate diversity in terms of number of species for each of the five sites.

Figure 7. Total invertebrate abundance in terms of number of individuals for each of the five sites.

Figure 8. ẟ13C values of sow bugs and woodlice for each of the five sites.

Figure 9. ẟ13C values of grasshoppers for the four sites at which they were present.

Figure 10. Species assemblage for each of the five sites as grouped into predators, herbivores, detritivores, and carpenter ants.

Table I. All species collected via pitfall traps, their common name, and number of individuals, grouped according to site.

Table II. All species collected via Berlese funnels, their common name, and number of individuals, grouped according to site.

Black 16

Figure 1. Diagram of a berlese funnel a picture of the 10 berlese funnels we set up in our lab.

Black 17

Figure 2. ẟ13C values for Schyzarium scoparium, Rubus flagellaris, Solidago canadensis, and Carex pensylvanica.

Black 18

Figure 3. Average percent soil organic matter for each of the five sites.

Black 19

Figure 4. Average soil ẟ13C values for each of the five sites.

Black 20

Figure 5. Soil ẟ13C values at four depths for each of the five sites.

Black 21

Figure 6. Total invertebrate diversity in terms of number of species for each of the five sites.

Black 22

Figure 7. Total invertebrate abundance in terms of number of individuals for each of the five sites.

Black 23

Figure 8. ẟ13C values of sow bugs and woodlice for each of the five sites.

Black 24

Figure 9. ẟ13C values of grasshoppers for the four sites at which they were present.

Black 25

Figure 10. Species assemblage for each of the five sites as grouped into predators, herbivores, detritivores, and carpenter ants.

Black 26

Table I. All species collected via pitfall traps, their common name, and number of individuals, grouped according to site.

Little bluestem Species Common name # Indivuduals coverage 4-15% Sowbug 4 4-15% vulgare 5 4-15% Cepaea sp. Snail 1 4-15% Allonemobius sp. Ground cricket 2 4-15% Napomyza gymnostoma Leaf miner fly 1 4-15% Alticini sp. 2 4-15% Aculeata sp. Wasp 3 4-15% Camponotus vicinus Carpenter 21 4-15% Staphylinidae sp. 2 Rove beetle larvae 7 4-15% Scutigeromorpha sp. Centipede 1 4-15% Staphylinidae sp. 3 Rove beetle 3 4-15% Blattodea sp. Cockroach 1 4-15% Curculionoidea Weevil or leaf notcher 2 4-15% Rabidosa sp. Wolf 1 4-15% Araneidae sp. 1 Ord weaver spider 6 4-15% Araneidae sp. 2 Spider 2 4-15% Araneidae sp. 3 Spider 1 4-15% Gastropod Slug 7 16-24% Camponotus vicinus Carpenter ant 29 16-24% Staphylinidae sp. 2 Rove beetle larvae 12 Red-legged 16-24% femurrubrum 2 16-24% Aphaenogaster sp. Ant 6 16-24% Cepaea sp. Snail 3 16-24% Achatininae Snail 1 16-24% Napomyza gymnostoma Leaf miner fly 1 16-24% Ceraphronoidea Wasp 1 16-24% Blattodea sp. Cockroach 3 16-24% Siphonaptera sp. Flea 1 16-24% Staphylinidae sp. 1 Rove beetle 1 16-24% Hydroscaphidae Skiff beetle 1 16-24% Unknown Clear bugs 13 16-24% Gastropod Slug 3 16-24% Lycosidae Spider 18 16-24% Porcellio scaber sowbug 1 16-24% woodlice 2 25-49% Porcellio scaber Sowbug 2 25-49% Armadillidium vulgare Woodlouse 49 25-49% Rabidosa rabida Rabid 1 25-49% Harpilini sp. Beetle 2 25-49% Julida sp. 1 Millipede 1 25-49% contractus 3 25-49% Staphylinidae sp. 1 Rove beetle 2 25-49% Siphonaptera sp. Flea 1 25-49% Homaeotarsus sp. Rove beetle 2 25-49% Camponotus vicinus Carpenter ant 30 25-49% Linepithema humile Ant 1 25-49% caespitum Pavement ant 1 25-49% Aphaenogaster sp. Ant 5 25-49% Trombidiina sp. Red velvet mite 2

Black 27

Bandwing 25-49% Arphiini sp. 1 grasshopper 25-49% Agelenopsis sp. Spider 3 25-49% Araneidae sp. 2 Spider 1 25-49% Clubionidae Spider 1 25-49% Unknown Clear bugs 44 25-49% Gastropod Slug 15 50-74% Napomyza krygeri Leaf miner fly 1 50-74% Euceros sp. Wasp 1 50-74% Porcellio scaber Sowbug 2 50-74% Armadillidium vulgare Woodlouse 2 Red-legged 50-74% Melanoplus femurrubrum 1 grasshopper 50-74% Gryllus pennsylvanicus Field cricket 1 50-74% Schizocosa sp. Wolf spider 1 50-74% Neoscona oaxacensis Orb weaver spider 1 50-74% Camponotus vicinus Carpenter ant 19 50-74% Hypoponera sp. Ant 1 50-74% Aphaenogaster sp. Ant 6 50-74% Gastropod Slug 13 75-100% Apis mellifera Honey bee 1 75-100% Armadillidium vulgare Woodlouse 6 75-100% Porcellio scaber Sowbug 1 Red-legged 75-100% Melanoplus femurrubrum 1 grasshopper 75-100% Camponotus vicinus Carpenter ant 23 75-100% Aphaenogaster sp. Ant 4 75-100% Tetramorium caespitum Ant 2 75-100% Araneidae sp. 4 Orb weaver spider 1 75-100% Grammonota sp. Spider 2 75-100% Unknown Clear bugs 13 75-100% Gastropod Slug 1

Black 28

Table II. All species collected via Berlese funnels, their common name, and number of individuals, grouped according to site.

Little # bluestem Species Common name Indivuduals coverage

4-15% Araneidae sp. 4 Spider 1 4-15% Coleoptera sp. 2 Beetle 7 4-15% sp. 1 Fly 1 4-15% Hymenoptera sp. 2 Fly 1 16-24% Coleoptera 4 Beetle 1 16-24% Araneidae sp. 4 Spider 3 16-24% Aphaenogaster sp. Ant 2 Rove beetle 16-24% Staphylinidae sp. 2 1 larva 16-24% Staphylinidae sp. 3 Rove beetle 1 16-24% Collembola sp. Springtail 3 25-49% Staphylinidae sp. 1 Rove beetle 1 Rove beetle 25-49% Staphylinidae sp. 2 2 larva 25-49% Staphylinidae sp. 3 Rove beetle 5 25-49% Coleoptera sp. 2 Beetle 29 25-49% Harpilini sp. Beetle 1 25-49% Coleoptera sp. 3 Beetle 1 25-49% Araneidae sp. 2 Spider 1 25-49% Siphonaptera sp. Flea 1 25-49% Ixodida sp. 1 Tick 4 25-49% Ixodida sp. 2 Tick 3 25-49% Staphylinidae sp. 4 Rove beetle 1 25-49% Acariformes Mite 1 50-74% Julida sp. 2 Millipede 1 50-74% Coleoptera sp. 2 Beetle 48 50-74% Staphylinidae sp. 4 Rove beetle 4 50-74% Coleoptera sp. 3 Beetle 1 50-74% Araneidae sp. Spider 1 75-100% Staphylinidae sp. 3 Rove beetle 5 Camponotus 75-100% Carpenter ant 1 vicinius 75-100% Coleoptera sp. 2 Beetle 10 75-100% Harpilini sp. Beetle 1 Rove beetle 75-100% Staphylinidae sp. 2 1 larva 75-100% Diptera sp. Fly juvenile? 1 Rove beetle 75-100% Staphylinidae sp. 5 1 larva