Science-based restoration monitoring of coastal habitats, Volume One: A framework for monitoring plans under the Estuaries and Clean Waters Act of 2000 (Public Law 160-457)

Executive Summary: The Estuary Restoration Act of 2000 (ERA), Title I of the Estuaries and Clean Waters Act of 2000, was created to promote the restoration of habitats along the coast of the United States (including the US protectorates and the Great Lakes). The NOAA National Centers for Coastal Ocean Science was charged with the development of a guidance manual for monitoring plans under this Act. This guidance manual, titled Science-Based Restoration Monitoring of Coastal Habitats, is written in two volumes. It provides technical assistance, outlines necessary steps, and provides useful tools for the development and implementation of sound scientific monitoring of coastal restoration efforts. In addition, this manual offers a means to detect early warnings that the restoration is on track or not, to gauge how well a restoration site is functioning, to coordinate projects and efforts for consistent and successful restoration, and to evaluate the ecological health of specific coastal habitats both before and after project completion (Galatowitsch et al. 1998). The following habitats have been selected for discussion in this manual: water column, rock bottom, coral reefs, oyster reefs, soft bottom, kelp and other macroalgae, rocky shoreline, soft shoreline, submerged aquatic vegetation, marshes, mangrove swamps, deepwater swamps, and riverine forests. The classification of habitats used in this document is generally based on that of Cowardin et al. (1979) in their Classification of Wetlands and Deepwater Habitats of the United States, as called for in the ERA Estuary Habitat Restoration Strategy. This manual is not intended to be a restoration monitoring “cookbook” that provides templates of monitoring plans for specific habitats. The interdependence of a large number of site-specific factors causes habitat types to vary in physical and biological structure within and between regions and geographic locations (Kusler and Kentula 1990). Monitoring approaches used should be tailored to these differences. However, even with the diversity of habitats that may need to be restored and the extreme geographic range across which these habitats occur, there are consistent principles and approaches that form a common basis for effective monitoring. Volume One, titled A Framework for Monitoring Plans under the Estuaries and Clean Waters Act of 2000, begins with definitions and background information. Topics such as restoration, restoration monitoring, estuaries, and the role of socioeconomics in restoration are discussed. In addition, the habitats selected for discussion in this manual are briefly described. (PDF contains 116 pages

Transfection of eastern oyster (Crassotrea virginica) embryos

There is a need for research in disease resistance and microbial elimination in the eastern oyster Crassosostrea virginica. Gene transfer may lead to advances in this area, and a means of selecting transfected larvae would be useful. We transfected 3-hour-postfertilization embryos with the bacterial gene aminoglycoside phosphotransferase II (neonr), which confers resistance to neomycin and related antibiotics such as G418. The antibiotic G418 was examined as a potential selective agent. A neutral red assay was used to determine survival after 48 hours of exposure to various concentrations of G418 (0-4 mg/ml). We examined the effects of electroporation and chemically mediated transfection of 3-hour-postfertilization embryos on survival to straight-hinge larvae. DNA alone was found to have no effect on survival (P\u3e.05). For electroporation we found that increased voltage and pulse duration decreased survival (P\u3c.05). Chemically mediated transfection did not significantly affect survival (P=.5172). Transgenic larvae were identified after electroporation and chemically mediated transfection. These larvae were reared for 24 hours and exposed to G418 at 0.3 mg/ml for 48 hours. Significant differences in survival between transfected and nontransfected larvae were detected for electroporation (P=.0147) and chemically mediated transfection (P=.037). Gene transfer was also confirmed with polymerase chain reaction and observation of expression of green fluorescent protein. This study documents the first successful insertion and expression of foreign DNA in eastern oyster larvae

An improved procedure to count Perkinsus marinus in eastern oyster hemolymph

Perkinsus marinus infection intensity in Crassostrea virginica can be quantified without killing of oysters by determining parasite density in hemolymph samples incubated in fluid thioglycollate medium (FTM). The goal of this study was to improve existing protocols for counting of P. marinus in oyster hemolymph. Specifically, the objectives were to examine the effects on parasite number and diameter of: 1) adding supplements to FTM such as lipid and oyster extract; 2) incubating with various FTM preparations with and without agar or beef extract; 3) incubating with various hemocyte densities (105, 106, and 107 hemocytes/mL of FTM) in a constant FTM volume; 4) incubating with different volumes of FTM (0.2 mL, 1.0 mL, 5.0 mL, and 25.0 mL); and 5) sodium hydroxide digestion of cellular debris. From these results, an improved hemolymph protocol was developed. The diameters and numbers of enlarged parasites or hypnospores in hemolymph of 20 oysters measured by the improved protocol and the standard FTM hemolymph assay of Gauthier and Fisher were compared. Finally, the standard and improved protocols were compared with the FTM body burden assay. The diameter of hypnospores from samples processed with the improved protocol (26 ± 13 μm) was significantly greater than the diameters from samples processed with the standard protocol (10 ± 4 μm). The number of hypnospores in samples processed with the improved protocol (8.6 × 103 ± 3.3 × 103) was significantly greater than the numbers in samples processed with the standard protocol (1.9 × 103 ± 3.4 × 103). Results of the body burden assay were significantly correlated with results of the standard hemolymph assay and with results of the improved hemolymph assay. The coefficient of determination (r2 = 0.7602) and slope (0.91189) of the regression of the FTM body burden assay against the improved FTM hemolymph assay was improved from the coefficient of determination (0.5543) and slope (0.61257) of the regression of the FTM body burden assay against of the standard FTM hemolymph assay

Environmental significance of freshets in reducing Perkinsus marinus infection in eastern oysters Crassostrea virginica: Potential management applications

The effects of extreme freshwater events on Perkinsus marinus-Crassostrea virginica interactions remain unexplored. The effects of freshwater events on P. marinus infection in C. virginica and oyster survival were therefore examined in controlled laboratory experiments and a field study. For the laboratory experiments, oysters were collected in spring, summer and winter from an area in Louisiana where P. marinus is endemic. Oysters were placed in 2 recirculating water systems at a salinity and temperature similar to their collection site. They were subjected to 2 salinity treatments (freshet and control). Freshet events were simulated by reducing the water to salinities of 0 to 1 ppt over a 48 h period, and maintained for a 21 d period. Control oysters were maintained at the initial salinity. Thirty oysters were sampled prior to the freshet event, and 30 oysters per treatment group (freshet, control) were sampled on Days 7, 14 and 21 after initiation of the freshet event. Oyster mortality, P. marinus infection intensities, oyster condition index and oyster plasma osmolality were measured in weekly samples. All 3 simulated freshet events (i.e. spring, summer, winter) resulted in a significant reduction in P. marinus infection intensity, but failed to eliminate infection. The failure of the oyster plasma to reach very low osmolality (\u3c50 mOsm kg-1) provides a likely explanation for the lack of complete P. marinus elimination. The field study involved sampling oysters monthly in the Caloosahatchee estuary, Florida, from September 2000 to February 2002, and determining P. marinus weighted prevalence and condition index of wild oysters, and growth and survival of caged juvenile oysters. The data strongly support the contention that the numerous freshwater releases to the Caloosahatchee River kept P. marinus infection intensities in oysters at low levels, resulting in an overall low weighted prevalence, low oyster mortality and good growth. Data from our field study appear to support the hypothesis that repetitive and well-timed freshet events can prevent infection of oysters with P. marinus or at least maintain P. marinus infections at non-lethal intensities (e.g. \u3c106 parasites g-1 wet tissue) in oyster populations. The use of an adaptive management approach involving control of freshwater inflows could be invaluable to the oyster industry in areas close to freshwater diversion projects