Discipline: Ecology Environmental and Earth Sciences
Subcategory: Geosciences and Earth Sciences
Session: 4
Room: Private Dining
Thaddisha James - Grambling State University
Co-Author(s): Lescia Valmond, Grambling State University, Grambling, LA; John Thomas, Grambling State University, Grambling, LA
Metagenomics, or sequencing of genetic material extracted directly from environmental sources, is typically used to study microorganisms including those found in soil and water. This metagenomic analysis of water and sediment from the Grambling pond and wastewater from Grambling State University and the city of Grambling sewersheds will deepen our understanding of the local biodiversity and uncover any antibiotic resistance genes that are present. Based on our previous knowledge of microbial ecology, we hypothesized that the profile of microorganisms identified in these various settings would be very different from one another. Conical tubes were used to collect water and sediment samples from the Grambling pond and a 24-hour composite wastewater sample was collected from the wastewater lift stations at Grambling State University and the city of Grambling. All water samples were centrifuged to obtain a pellet and DNA was extracted using the ZymoBIOMICS DNA Miniprep Kit. DNA extraction from the soil samples was carried out using the DNeasy PowerLyzer PowerSoil Kit. DNA quality was assessed by UV spectroscopy and DNA concentration was measured using a Qubit high sensitivity dsDNA assay. Libraries for sequencing were prepared using the Oxford Nanopore Rapid Barcoding Kit. Using the EPI2ME What’s In My Pot bioinformatics pipeline, 867,885 total reads were examined. Bacteria accounted for 94% of reads, another 5% were classified as Eukaryota, and fewer than 1% were classified as Archaea or viruses. In overall taxonomic abundance, the top ten species identified were Escherichia coli (111,351 reads), Homo sapiens (17,189), Cloacibacterium normanense (17,128), Sphaerotilus natans (6,700), Escherichia marmotae (4,851), Phocaeicola vulgatus (3,525), Aquaspirillum sp. LM1 (2,919), Salmonella enterica (2,439), Faecalibacterium prausnitzii (1,835), and Blautia sp. SC05B48 (1,741).Many of these are enteric bacteria and therefore not surprisingly found in the wastewater, but some were also detected in the pond water and sediment. The EPI2ME Antimicrobial Resistance workflow produced 1,344 alignments to 185 genes in the Comprehensive Antibiotic Resistance Database. Future studies will focus on the impact of these microorganisms and their antibiotic resistance on water quality and public health. References: Coughlan LM, Cotter PD, Hill C and Alvarez-Ordóñez A (2015) “Biotechnological applications of functional metagenomics in the food and pharmaceutical industries” www.frontiersin.org Sissel Juul, Fernando Izquierdo, Adam Hurst, Xiaoguang Dai, Amber Wright, Eugene Kulesha, Roger Pettett, Daniel J. Turner (2015) “What’s in my pot? Real-time species identification on the MinION” www.biorxiv.orgJulie Segre, (2022) “Metagenomics” www.genome.gov
Funder Acknowledgement(s): National Science Foundation Award #1818697
Faculty Advisor: Paul Kim Ph.D., Department of Biological Sciences, Grambling State University; Audrey Kim D.V.M., Ph.D., Department of Biological Sciences, Grambling State University, kimp@gram.edu
Role: In this research, I was responsible for: - Collection- DNA Extraction- Library Preparation- DNA Sequencing- Data Analysis