Patrick Biber of the University of Southern Mississippi Gulf Coast Research Lab, Tim Sherman of University of South Alabama, and Allison Walker of Acadia University, will study the incidence and abundance of the seagrass parasite Plasmodiophora diplantherae in the North-Central Gulf of Mexico. The project will collect seagrass samples to determine coverage and infection rate. This project will lay the groundwork for future studies of this marine plant parasite by mapping the incidence and abundance of the parasite. Scientists also will obtain DNA sequences that can be used in future tools to detect the parasite in seagrass plants and coastal sediments.
Objectives
The objectives of the proposed study are to map the incidence and relative abundance of the seagrass parasite P. diplantherae in the North-Central Gulf of Mexico (LA, MS, AL, FL) and to obtain genetic material (DNA sequences) to place this parasite phylogenetically. Successful DNA extraction and amplification will aid in future development of rapid molecular diagnostic tools for the detection of this parasite in seagrass plants and coastal sediments.
Methodology
Field work will be done at seven locations in the northern Gulf of Mexico, focusing on MS and AL seagrass beds. Proposed locations were chosen based on where this parasite has been recorded previously and include in Grand Bay, Horn Island, Ship Island, Cat Island in MS; the northern Chandeleur Islands, Pointe aux Pines (AL) and Johnson Beach at Perdido Bay. At each location, 2-3 sites will be visited for intense sampling including obtaining seagrass cores and transect-based observations for infected H. wrightii plants. Three seagrass cores (15cm diameter, 30cm deep) will be collected at 10m intervals along each transect (total 6 cores/site). In addition to the seagrass core, water depth, seagrass percent cover in a 0.25m2 quadrat, and prevalence of H. wrightii in mixed species beds will be noted for that station. Each core will be transferred to a floating sieve 850 μm (size 20) mesh to remove the sediments and the cleaned shoots placed in labelled sterile plastic bags and placed on ice.
Core samples will be processed in the lab within 24 hours of collection. All shoots in a core will be counted and the number of infected shoots will be recorded to calculate the percent infection. Only infected shoots will be frozen for later DNA analysis. Infected shoot samples will be shipped on ice to Dr. Walker in Ottawa, Canada for DNA extraction and gene sequencing. DNA will be extracted from infected H. wrightii tissue using an UltraClean Microbial DNA Isolation Kit. The 18S region of ribosomal DNA will be amplified using Illustra Ready-To-Go™ PCR Beads with Plasmodiophorid-specific primers.18S rDNA PCR amplification will be confirmed using gel electrophoresis and forward and reverse sequences will be obtained by the dideoxy chain-terminating method using Plasmodiophorid-specific sequencing primers and ABI PRISM 3100 or ABI PRISM 3130xl automated DNA sequencers. Sequences will be edited and aligned using Geneious 6.0.4 software. Consensus sequences will searched against the reference NCBI sequence database GenBank using the BLAST search algorithm, as well as other relevant sequence databases. The top sequence matches will be subsequently downloaded and a maximum likelihood analysis will be performed in PHYML to place this species in a phylogenetic framework.
Rationale
The seagrass Halodule wrightii is infected by the protistan parasite Plasmodiophora diplantherae in the Gulf of Mexico, but how it may contribute to current seagrass losses is unknown. This parasite causes plant stunting and seagrass uprooting, which can be detrimental to seagrass ecosystem health and coastal seagrass restoration projects. It was first reported from the north-central Gulf of Mexico (LA and MS) in 2009; however its wider distribution and abundance in this economically import region are currently unknown and may have expanded since the Deepwater Horizon incident in 2010.
Infections with Plasmodiophora spp. can decrease host fitness by reducing the growth of the host plant, reducing the formation of inflorescences, and by altering host metabolism and/or reproductive success. Plasmodiophorid parasites cause increased uprooting of their angiosperm hosts as a consequence of reduced root growth. In areas where seagrass-restoration projects are ongoing, this uprooting can subsequently damage seagrass beds and there is also a considerable risk of floating plants spreading this pathogen to adjacent populations of seagrasses.