CICEET Progress Report for the period 9/01/01 through 3/01/02

Project Title: Density-Dependent Effects on Grazing and Success for Seed Generated Seagrass (Zostera marina L.) Plants.
Principal Investigator(s): Scott W. Nixon, Stephen Granger, Brian Maynard, and Malia Schwartz

Accomplishments
Scheduled Tasks:
The focus of this reporting period has been largely one of transition. We were fortunate to receive a second award from CICEET, which began in September 2001. Part of the details from this progress report will meld into those included in the first progress report for the new project.

The tasks for this reporting period fall into three categories. First, to modify the design of the seeding machine and conduct large scale testing. Second, to search for and test other commercially available gelling agents that may increase the efficiency of mechanized seeding. Third, to finalize and publish a pamphlet on seed collection techniques.

During this past summer our lab was contracted by the US Navy to conduct a large-scale (0.25 acre) eelgrass restoration using the seeding machine developed with CICEET funding. To make a restoration of this size possible the existing prototype design (see previous progress report) needed to be scaled-up from the existing 4-tine prototype to an 8-tine model. A larger pump provided an additional volume of gel needed to support the increase in tines. Finally, approximately 1 million seeds needed to be collected to make planting possible. A goal of collecting 1.5 million seeds was set, approximately 3 times that collected in any of our previous efforts.

As mentioned in previous reports, germination rates observed from our initial trials of the seeding machine were lower than expected. We believe that the gelatin (Knox) used in the initial plantings placed additional respiratory demand on the sediments which lowered seed germination. We proposed to investigate new gelling agents that may work as alternatives to Knox gelatin.

Finally, we wanted to finalize and publish a pamphlet entitled "Growing Eelgrass from Seed- A Practical Guide to Collection and Storage of Seeds for the Propagation and Restoration of Zostera marina L.". By the end of the last reporting period a draft of the manual had been completed and editorial revisions, design, and printing were still to be conducted.

Progress on Tasks
During August and September 2001 our research group was successful in collecting over 1.75 million eelgrass seeds. This exceeded our original goal of 1.5 million and has positive implications towards making this technology a possibility for even larger efforts. We were successful in building an 8 tine model of the sled and, with advice from technicians at Edhard Corporation, were able to adapt the larger pump to our new sled design. In October and November, 2001 we conducted a large-scale seeding (400m²) operation at two locations (800m² total area planted) in Narragansett Bay. The seeding sled operated exceptionally well and after only a few modifications to the seed to gel ratio and winching system, we were able to plant a 400m² area in approximately 4 hours. We look forward to monitoring the success of this effort in April 2002. We have taken our experiences from this year's seeding machine trials to Design Machine Corporation (South County, RI) who is currently working on modifications which will minimize tine clogging, one of the problems encountered during this years plantings. We have submitted an application for a patent with the United States Patent Office to copyright the technology.

We have performed an extensive search of both the agricultural and chemical industries for various gelling agents that may work with our unique application. Through this search we have come up with 3 alternative gel types Cab-o-sil, Sodium Bentonite, and Agri-gel (See Table 1). In November 2001 seeds were suspended in each gel type and planted by hand in sediment filled plastic pots (10.2cm x 10.2cm) x 3 replicates. After planting, the pots were placed in flowing seawater tanks and kept at ambient temperature, salinity and irradiance.We will follow these treatments throughout the spring and summer monitoring seed germination and seedling survival.

We have completed the first draft of our seed collection pamphlet including text, illustrations and photographs. We have negotiated and contracted a local printer who will produce 2000 copies when finalized. The draft is now under editorial review by the Sea Grant Communications Office and when the final edits are finished will go to designer for the final layout. The editor, Malia Schwartz, has just returned from a six-month hiatus and has promised to finish editorial changes by the middle of February. The final pamphlet layout will be approved and sent to the printer by April 1st. We expect delivery from the printer by the middle of May 2002 with distribution occurring shortly there after.

Difficulties Encountered
One of the difficulties encountered during this year's plantings was tine clogging. As mentioned in our previous progress report (August 2001), the lower germination seen in the gel planted seed, was thought to be attributed to the increased respiratory demand added to the sediments by the Knox gelatin. To reduce the effect of the gel on the sediments, we worked to reduce the amount of gel needed for planting. The reduced gel to seed ratio resulted in clogging of tines during planting. The current design incorporates a 90° bend in the injection needle, which was the point at which most clogs occurred. We have met with Duane Sherman, the president of Design Machine Corporation (South County, RI) to discuss ways to redesign the layout of the manifolds and injection needles so as to eliminate these bends and minimize future clogs. This will, in turn, facilitate the reduction in the amount of gel needed for future plantings. As a second measure, we are working with alternative gel types with lower organic content in the hopes that they will have less impact on the sediment respiratory demand and increased germination will result.

Anticipated Success in Meeting Project Objectives in Scheduled Project Period
We do not anticipate any difficulties in meeting our project objectives during this reporting period.

Because of the difficulties encountered, a no-cost time extension to June 30, 2003 is requested for this project.

Preliminary Results
On January 28, 2002 a preliminary count of seedling emergence in the various gel plantings was performed. The results of this count are shown in (See Table 2). These initial figures show promise for other gelling substances. Sodium Bentonite showed significantly higher germination (p<0.05) than all other treatments including that of the no gel control. These results may indicate that Sodium Bentonite is having a positive effect on seed germination. At the time of this count all treatments had significantly higher germination (p<0.05) than that of the Knox gelatin treatment. It should be noted that these results are preliminary, final germination success will be realized in April when the period of germination slows and lateral shoot production begins.

Tasks and activities for next reporting period

Tasks for the next reporting period
Our primary task for the spring of 2002 will be to continue work on the seeding machine. Through the large-scale test of the machine this past fall; we have gained valuable experience with the machine and what improvements need to be made. As mentioned earlier clogging was one issue that will need to be addressed. Other issues include redesigning the sled to make it lighter and easier to handle, and modifying the pump head to work with a less expensive motor. We will test the effects of the redesigned tines this spring in preparation for another seeding to occur in the fall of 2002.

A second task for the spring and summer of 2002 will be to continue monitoring the germination and seedling success of seeds planted in our trials of alternate gel types. The preliminary results presented earlier are very promising and we will continue to monitor the emergent seedlings through one growing season observing the effects, if any, each gel may have on seed germination and seedling growth. We will also begin to run tests of the seeding machine using the alternate gels proven to be the most effective in terms of germination and seedling success. Previous plantings with the machine have all been accomplished utilizing Knox gelatin as the suspension media. The new alternate gel types will have different viscosities; there fore modifications will have to be made to the pumping system.

Finally, we will finalize and publish the pamphlet on seed collection techniques with a deadline for distribution set for May 2002. With the return of our editor we will begin the editorial review of our pamphlet and have final editorial changes finished by February 2002. To ensure that the pamphlet is easy to understand and factual we have made arrangements for personnel at Save the Bay and the Rhode Island Division of Environmental Management (RIDEM) to comment on the text before it goes out to press. In April 2002 we will send off the final draft for layout and printing.

Work plan to accomplish tasks
At the time of submission of this report we have met twice with Design Machine Corporation to discuss modifications to the pump and manifold systems as well as to the sled design. We have prioritized modifications and started with the reengineering of the manifold and tines. We have delivered all the necessary parts to make the modification and are awaiting the completion of the first order repairs, expected by March 15th 2002. We will test the new modifications in March and April 2002. With satisfactory results, we will begin modifications to the pump system and sled construction. We anticipate making test runs of the sled with all modifications by June 2002 in preparation for a second large scale planting to occur during the fall of 2002. This second planting is included in the funding of the 2001-2003 CICEET award.

In November 2001 we planted eelgrass seeds in marine sediment, using the four differing gels. From our previous experiments, we know that April is the time during which germination slows and lateral shoot production begins. As a result, we will make a count of shoot emergence and calculate a germination success at that time. We will continue to monitor the emergent seedlings through one growing season and by late July 2002 we will count the number of lateral shoots produced by each seedling, and measuring above ground and below ground biomass in our experimental treatments. This will indicate whether any of the gels were advantageous to seedling growth. Those gels that perform well in these initial plantings will then be tested for application with the mechanized delivery system.

Finally, we will work with Malia Swartz, to make final editorial changes to the seed collection pamphlet, complete the final layout, and have it printed. As mentioned earlier, we anticipate the time of delivery and distribution to be May, 2002.

Concerns or difficulties
We have currently seeded our largest restoration effort to date with the seeding machine. As mentioned in previous reports we are concerned with low germination success we have seen from the Knox gelatin plantings. However, we are encouraged by the initial results of our plantings using alternative gels (Table 2). We are confident that one of these alternatives will be optimal for future plantings. We anticipate using one of these alternative gel types when planting during the fall 2002.

Expenditures
We have had no difficulties staying within budget for this reporting period.