Our Students and Their Research

Direct and enzymatic methods for urea determination were compared and analyzed for specificity. Organic nitrogen compounds (arginine and humics) were added to urea standards to measure interference in both methods. In the enzymatic method, urea concentration appeared to decrease with increasing arginine concentration. However, minor ammonia contamination in the arginine stock solution created uncertainty as to these results. However, the presence of arginine appeared to have no effect on the direct method. In the presence of humic material, the enzymatic method underestimated true urea concentrations, but the direct method was not as significantly affected. Pursuing a peripheral issue led to the conclusion that the enzymatic method underestimates urea concentrations under saline conditions. Urea standards and Pocomoke River samples were assayed using both methods. The enzymatic method underestimated urea concentrations relative to the direct method. Culture studies with the brown tide organism Aureococcus anophagefferens indicated that arginine is not an effective substrate for the organism's urease.

The calanoid copepod Eurytemora affinis is known to be highly adaptable in the dynamic conditions they encounter in the estuarine environments they inhabit. The geographic and seasonal distribution of the species is restricted, however, and could possibly be linked to osmotic limitations. These limitations involve both lethal and sublethal responses to salinity stress and are possibly related to salinity regime. Questions arise as to how osmotic demands created by different salinity regimes affect E.affinis individuals. These effects include sublethal responses such as reproduction and lethal responses involving maintenance costs. Possible salinity regimes populations encounter in estuaries are both pulsed and chronic and can have high, low, or optimal salinity levels. Under these parameters, E.affinis appears to have greater fecundity and survivorship in high salinity environments and in general, populations have relatively higher growth rates in varying vs. constant salinity regimes. Because salinity stress is often a major factor in many estuarine species' distribution, the response of E.affinis populations and individuals to different salinity regimes could be useful in explaining the factors limiting the distribution of the species in estuaries.

The accumulation of contaminants within the sediments of the Baltimore Harbor region has led to concern for their possible impact on aquatic ecosystems. In order to determine possible effects of contaminated sediment on three, co-occurring, indigenous species in the Chesapeake Bay we used 28d and 14d exposure tests. The three species studied were the benthic crustaceans, Palaemonetes pugio, Leptocheirus plumulosus, and Gammarus mucronatus. P. pugio and G. mucronatus were used in both 28d and 14d exposure tests, whereas L. plumulosus was only used in a 28d test. Organisms were exposed to diluted volumes of sediment from Colgate Creek (CC), a tributary in Baltimore Harbor. Contaminated sediments were mixed with the control substrate from Fishing Bay (FB) to make 25% CC and 50% CC concentrations. Reference sediments remained 100% FB. The responses of organisms, following the 28d and 14d exposures, were measured by quantifying changes in individual mass, mean reproduction, mean proportion survived, and standard metabolic rate for organisms in each of the three sediment treatments. The contaminated sediments showed no significant effects in measured response variables for P. pugio or L. plumulosus. Mean reproduction for G. mucronatus was significantly higher in the control sediments (FB) than in the contaminated sediments (CC). Standard metabolic rate values showed not significant differences between treatments implying that energy allocation for production was not altered. Effects on reproduction may have thus derived from mechanisms other than contaminant-driven energetic constraints. 14d exposures yielded similar survivorship results to that of the 28d exposure.

The Estuarine Turbidity Maximum (ETM), located at the head of many estuaries like the Chesapeake Bay, is a region of high turbidity and suspended particle concentrations. The ETM is a nursery area for larval fish and may concentrate zooplankton species. The goal of this study is to determine the spatial distribution of adult copepods and cladocera that are potential prey of fish larvae in and around the Chesapeake Bay ETM in both a wet (1998) and dry (1999) year. Plankton concentrations were analyzed from depth-stratified 1-m2 Tucker-trawl (280 micron mesh) samples that were taken at stations in the channel of the upper Chesapeake Bay. The study focused on the abundant calanoid copepods Eurytemora affinis and Acartia tonsa, and the cyclopoid parasitic copepod Ergasilus labricus. Cladocera analyzed were Bosmina longirostris and Daphnia pulex. Although total zooplankton concentrations were highest within and below the ETM zone in both 1998 and 1999, zooplankton distribution varied between species, year, and depth. Eurytemora and Ergasilus were concentrated in the ETM in 1998, but not in 1999. Acartia was concentrated below the ETM zone, and Cladocera were mostly found above the ETM. Vertical distribution varied among species, and between sexes of the same species. Changes in distribution and abundance of zooplankton in the ETM may have important consequences for higher trophic levels since the distribution of prey and parasitic copepods overlapped and was similar to that of larval striped bass and white perch.

The effects of diets consisting of the harmful algal bloom (HAB) species Pfiesteria piscicida (strain FDEPMDR23) and Prorocentrum minimum on the egg production and hatching success of the calanoid copepod Acartia tonsa were studied in laboratory experiments. These experiments were conducted in order to test the hypotheses that P. piscicida and P. minimum represent nutritionally deficient, or possibly toxic, diets to A. tonsa. A similar experiment was also conducted using the cryptophyte species Storeatula major since it was used as a food source to grow the P. piscicida. Each egg production and hatching experiment was run for 96h, which included a 48h acclimation period, a 24h experimental period, and an additional 24h period to allow eggs produced during the experimental period to hatch. Results obtained form the experiment using P. minimum as a food source show decreased egg production rates (expressed as eggs/female/day) and hatching success compared to diets consisting various diatoms and non-HAB forming dinoflagellates (information concerning these algal species was obtained from various sources cited within this paper). However, the P. minimum cultures used for this experiment were contaminated with S. major, so the results of this experiment cannot be directly linked to a diet consisting purely of P. minimum. Due to problems with copepod survival rates there is presently no data for the experiments involvingP. piscicida. or S. major. However, future experiments will be conducted to obtain this information. These results will also be combined with information on how these HAB diets affect the somatic growth of A. tonsa.

G. galatheanum and P. minimum are two small, bloom-forming dinoflagellates that have been associated with red tides in Chesapeake Bay. Algal blooms are the consequence of net growth, from increase division rates, or decreased mortality rates. In the past, most attention and research has been directed towards variables that increase growth rates, however, little is understood about the impact of predation, or zooplankton grazing, in terms of development and termination of toxic blooms. This study examines the distribution of the potential grazing pressure on G. galatheanum and P. minimum in the Chesapeake Bay and its tributaries. In situ grazing experiments were conducted in June and July, on (2) separate cruises. Dinoflagellate cultures were stained with CMFDA, a vital green fluorscent stain, and added to treatments with (<200 micron) and without (<1.2 micron) the natural microzooplankton assemblages collected at each station. In 58.6% of the experiments, the grazing coefficient was higher the maximum potential growth rate of G. galatheanum and P. minimum in culture, 0.94/d (Li et al., 1999) and 1.15/d (Grzebyk & Berland, 1996), respectively. Thus, grazing by microzooplankton has the potential to prevent net growth of these dinoflagellates. In June, there was a significant correlation between ciliate abundance (>20 micron) and potential grazing on P. minimum. Grazing pressure appeared to be higher in June than in July, but the difference was not statistically significant. In conclusion, microzooplankton have a high potential to control dinoflagellate growth, although the grazing pressure was rather variable throughout Chesapeake Bay and its tributaries.

The influence of sediment deposition on a meadow (Vallisneria americana) and a canopy (Myriophyllum spicatum) forming species of submerged aquatic vegetation was investigated at Blossom Point, MD. Two depths (5 and 15 cm) of sediment were deposited among these plants. After 7 days, productivity was determined by measuring photosynthesis vs. irradiance curves and biomass change over time. V. americana showed an increase in the rate of O₂ production (alpha) with increasing sediment levels however the maximum O₂ production (Pmax) decreased at the high treatment. M. spicatum's rate of production of O₂ (alpha) followed the light history pattern at the specific sites. The maximum amount of O₂ produced (Pmax) decreased with an increase in sediment deposition possibly because of the increase in phytotoxins at the base of the plant. Biomass data provided no conclusive data possibly due to the short time period of this experiment. In summary, sediment deposition affected the meadow and canopy species through different processes.

This study examined the uptake and assimilation of the trace metals cadmium, chromium, and zinc from suspended sediment by the estuarine copepod Eurytemora affinis. Adult females were exposed to suspended sediment labeled with radioisotopes of Cd, Cr, and Zn at four different concentrations for approximately 44 hours. Activity in the animals was then determined using a gamma counter before depuration and again after a four-hour depuration period. Cadmium uptake showed significant difference from controls at 300 mg/L sediment levels, as well as at 50 mg/L on one occasion. Zinc uptake was highly significant at all sediment levels. No significant differences were observed between non-depurated and depurated copepods, suggesting that perhaps material in guts is an insignificant contribution to total body load.

The hypothesis that humic substances stimulate the growth of dinoflagellate populations was tested in natural assemblages from the Chicamacomaco and Choptank Rivers. All humic additions were from a stock solution of Aldrich Humic Acid. Experiment 1 used samples from both rivers to see the effect of adding humics (HS), at a concentration of 5mg/L on heterotrophic and phototrophic dinoflagellate populations as well as on other parameters such as absorbance, pH, nutrient concentration, and Chl a concentration. Both trophic types of dinoflagellates decreased in density throughout the experiment, however, this decline was up to 50% lower in the HS treatments. This result was not consistent when the experiment was repeated. Nutrient Experiment 1 repeated this procedure for a Choptank sample, but added two treatments: one with nitrogen at 44μM and phosphorus at 1.8μM (NP), and one to which both nutrients and HS were added (HSNP). Heterotrophic dinoflagellates showed positive growth rates, however, they were 50% lower in the HSNP treatment then in the NP treatment. Phototrophs again showed a decline, however, the rate of decrease was slowed by 300% lower in the HSNP treatment than the NP treatment. Data, such as Chl a, indicate other parts of the community may be affected differently by the addition of HS.

We offer a quantitative synthesis of current research on in situ microbial community diversity. The lack of explicit quantitative statements on phylogenetic distribution, and the recent exponential climb in the application of 16S rRNA gene probes to natural systems prompted our research. In this review, studies in several natural systems, both broad and remote, were examined from some leading journals, including Applied and Environmental Microbiology, and FEMS Microbiology. Studies utilizing molecular techniques, such as polymerase chain reaction amplification (PCR) or fluorescence in situ hybridization (FISH), were synthesized according to the reported percentages of rRNA contribution by phylogenetic clades to the total community rRNA amount. Distinct shifts in community profile were observed across marine, freshwater, sediment, soil, and some extreme systems. Moreover, comparison of Eubacteria staining success observed a systematic variance in resolution across these natural systems. This observation, when taken with the small, unbalanced collection of in situ studies, suggests a need for further research to refine our understanding of the extent and function of microbial diversity.

The Atlantic Croaker, Micropogonius undulatus, is increasing in abundance in the Chesapeake Bay thus increasing its importance in the ecosystem. On three TIES (Trophic Interactions in Estuarine Systems) cruises per year from 1995 to 1999, stomach contents of Atlantic croaker were examined to obtain percentage and frequency of occurrence of prey. Over the summer of 1999, 323 croaker were examined and contents were categorized into polychaetes, fish, crustaceans and bivalves. Out of the factors examined during this study, there were three significant factors that play a role in croaker diet. The primary was ontogenetic size classes, with two other factors: one seasonal and one regional difference. There was also a noticeable difference in three fish caught by hook and line off of the pier at CBL. Such dietary changes bode well for the future of croaker, demonstrating high adaptability and an ability to manage environmental instability.

The purpose of this study was to determine the influence of nutrients on bacterial metabolism in salt marsh water with respect to the natural system and under conditions of experimental nutrient enrichments. Water samples were collected from the Monie Bay National Estuarine Reserve in Maryland during June and August 1999. Control and enriched samples were incubated over a 36 hour period in an environmental growth chamber at ambient temperature. Nitrogen and phosphorus were added to samples to achieve 80μM (NH₄) 2SO₄ and 5μM KH₂PO₄ concentrations. Bacterial production rates were measured using the 3H-leucine uptake method (Smith et al. 1992) and bacterial respiration was measured as changes in Oz concentrations using membrane inlet mass spectrometry (Kana et al. 1994), 4 gradient of nutrients was found in the natural system with a corresponding gradient of bacterial growth efficiency and bacterial production. Total nutrient concentrations (N and P) and average growth efficiencies were highest in Little Monie Creek, a tidal creek in close proximity to a chicken farm, and lowest in the Open Bay. Little Monie Creek was not limited by nutrients while the Open Bay waters and Little Monie Creek was not limited by nutrients while the Open Bay waters and Little Creek, a relatively pristine tidal creek were limited by nitrogen, phosphorus, or both. The experimental nutrient additions of nitrogen, phosphorus and nitrogen and phosphorus resulted in increased growth efficiencies for the Open Bay and Little Creek, but not for Little Monie Creek. The results suggest that bacterial growth efficiency is positively related to nutrient concentrations in Monie Bay National Estuarine Reserve.

Grazing preferences for controlled environment of the Calanoid copepod Acartia tonsa under a normal and an enriched nauplii concentration were determined using ambient Choptank River water. The sampling was done on two separate days in the MEERC lab at Horn Point. Two sets of tanks were sampled, 3 control and 3 nutrient tanks. Samples from the tanks were split into 12 1-L bottles, 6 containing copepods (experimental bottles) and 6 containing none (control bottles). Also, bottles 3,4,7,8,11 and 12 contained a 3x the normal enriched nauplii concentration. The results showed that when more nauplii were available, more nauplii and less phytoplankton were ingested. Copepods may switch their grazing preferences during changing environmental conditions and to regulate the population density of their environment.

The potential for ecological impacts, due to additional suspended sediment introduced to the water column, as a result of agriculture and construction, have become more widely recognized in recent years. However, the possible effects of increasing levels of suspended sediment on the general health and reproduction of copepods, a major food source for larval and commercial fishes are not well understood. In this study Acartia tonsa were added to l-L jars containing 0.5 ppm Thalassiosira weissflogii and varying levels of commercially available Kaolin corresponding to suspended sediment levels of 0, 49.5, 147.5, and 382 mg/L. The females were allowed to reproduce for 24hr, at which time the contents of the jars were screened and preserved for counting. Each sediment concentration was tested in quadruplicate. Results varied between trials. In the first trial, egg and nauplii production showed a significant decrease with the addition of sediment. In the second trial there was virtually no difference in reproduction at low levels of sediment, with a significant increase at the highest level. Possible explanations and implications for these results will be discussed.