1. Soil Microbiome of Native and Invasive Marsh Grasses in Blackbird Creek, Delaware Lathadevi K.Chintapenta 1#, Gulnihal Ozbay 1#, Venu Kalavacharla 1* Figure 5. Bacterial community ordination by marsh grass type, based on the relative abundance of taxonomic groups, by principal component analysis Acknowledgments We thank the NSF-EPSCoR program (EPS-1301765) and CIBER at DSU and USDA for their funding support and DNREC for their assistance with sampling. We also thank our undergraduate and graduate students and other lab members at Delaware State University for their constant support. Figure 1. Blackbird Creek sampling sites Introduction Blackbird creek is a dynamic saltmarsh ecosystem which provides a critical habitat for diverse flora and fauna. This reserve is considered to be pristine by Delaware National Estuarine Research Reserve (DNERR), but is experiencing problems such as loss of marsh habitat and intrusion of invasive species due to sea level rise and environmental changes (DNERR, 2013). This project studies the microbial community structure of the reserve as microbes are beneficial in reflecting the soil trophic status (Smit et al, 2001). Soil microbes play a critical role in nutrient cycling and retention with demonstrable effects on water quality and marsh management. In this study the diversity and abundance of microbes has been studied in sites with Spartina alterniflora (native marsh grass), Phragmites australis (invasive marsh grass) and mixed areas (Fig.1). Soil nutrient quality has been assessed in order to find the correspondence between microbial communities and nutrients. Results  The percent of total composition of major clades was estimated from 150 sequences. The marsh sites are dominated by Archaebacteria (16% of the clones), α- proteobacteria (12%), β-proteobacteria and δ- proteobacteria (14%).  Unknown bacteria accounted for 20% of the total clones. Less than 1% of all classified sequences include Bacteroidetes, Chloroflexi and Cyanobacteria  Shannon’s index (H’) calculated at a genetic distance of 3% from the Operational Taxonomic Units (OTU) and phylogenetic data indicated that mixed marsh areas had higher bacterial diversity (species richness). Methods Soil samples were collected by composite sampling. Soil pore water was analyzed for nutrients using GC-MS at University of Delaware. DNA was extracted with MO BIO Power Soil DNA Isolation kit and further amplified with 16S rDNA bacterial and archaeal primers. Fifty clones were prepared from each soil sample using TOPO TA cloning kit. The cloned DNA was then sequenced at DBI, University of Delaware. The DNA sequences identified were compared to NCBI BLAST database and sequences with more than 98% matches were accepted. # Aquatic and Environmental Sciences; * Plant Molecular Genetics-Centre for Integrated Biotechnology Research 1 Delaware State University, Department of Agriculture & Natural Resources, Dover, DE 19901 E-mail: kchintapenta@desu.edu Phylogenetic identities were determined by creating a phylogenetic tree (Fig. 3) of the clone sequences using parsimony analysis (1000 boot strap replicates) in MEGA 4.0. Figure 3. Phylogenetic tree of 16S rDNA sequences obtained from marsh soils Figure 4. Microbial diversity of the marsh areas Soil bacteria are categorized into groups on the basis of soil carbon availability. Betaproteobacteria are abundant in mixed sites (Fig.4) that are largely occupied by Spartina alterniflora, a C-4 plant. In aquatic environments, β-Proteobacteria are commonly present in areas that are rich in organic carbon (Fazi et al. 2005, Firer et al. 2007). Proteobacteria PC1 explains 73% of variance while PC2 explains additional 22% variance among samples. Results reveal shifts in the composition of bacterial communities and the abundance of specific taxonomic groups in relation to soil nutrients which reflects the shifts in their biogeochemical cycles. Soil nitrate concentration increased with soil pH. We observed shifts in the abundance of alpha and beta proteobacteria in mixed marsh areas (Fig.5). Green non-sulfur bacteria and Proteobacteria are high in rhizosphere soil whereas Archaeabacteria and Actinobacteria were more in bulk soil. Conclusions Study sites with invasive marsh grass (Phragmites australis) has less number of microbial species than the native marsh and the mixed marsh grasses. This infers that soil structure in the marsh ecosystem is being affected and thereby the microbial community structure is altered. This has a strong implication on function and health of marsh habitat. Current research Analysis of soil total carbon and total nitrogen. Continue collecting and studying bacterial community in these marsh sites. Figure 2a & 2b. Drawings of Phragmites australis and Spartina alterniflora (www.classicnatureprints.com) 2a 2b Proteobacteria percentagepercentage