Tumours in white suckers from Lake Michigan tributaries: pathology and prevalence.

The prevalence and histopathology of neoplastic lesions were assessed in white sucker Catostomus commersonii captured at two Lake Michigan Areas of Concern (AOCs), the Sheboygan River and Milwaukee Estuary. Findings were compared to those observed at two non-AOC sites, the Root and Kewaunee rivers. At each site, approximately 200 adult suckers were collected during their spawning migration. Raised skin lesions were observed at all sites and included discrete white spots, mucoid plaques on the body surface and fins and large papillomatous lesions on lips and body. Microscopically, hyperplasia, papilloma and squamous cell carcinoma were documented. Liver neoplasms were also observed at all sites and included both hepatocellular and biliary tumours. Based on land use, the Kewaunee River was the site least impacted by human activities previously associated with fish tumours and had significantly fewer liver neoplasms when compared to the other sites. The proportion of white suckers with liver tumours followed the same patterns as the proportion of urban land use in the watershed: the Milwaukee Estuary had the highest prevalence, followed by the Root, Sheboygan and Kewaunee rivers. The overall skin neoplasm (papilloma and carcinoma) prevalence did not follow the same pattern, although the percentage of white suckers with squamous cell carcinoma exhibited a similar relationship to land use. Testicular tumours (seminoma) were observed at both AOC sites but not at the non-AOC sites. Both skin and liver tumours were significantly and positively associated with age but not sex.

Evidence of estrogenic endocrine disruption in smallmouth and largemouth bass inhabiting Northeast U.S. national wildlife refuge waters: A reconnaissance study.

Intersex as the manifestation of testicular oocytes (TO) in male gonochoristic fishes has been used as an indicator of estrogenic exposure. Here we evaluated largemouth bass (Micropterus salmoides) or smallmouth bass (Micropterus dolomieu) form 19 National Wildlife Refuges (NWRs) in the Northeast U.S. inhabiting waters on or near NWR lands for evidence of estrogenic endocrine disruption. Waterbodies sampled included rivers, lakes, impoundments, ponds, and reservoirs. Here we focus on evidence of endocrine disruption in male bass evidenced by gonad histopathology including intersex or abnormal plasma vitellogenin (Vtg) concentrations. During the fall seasons of 2008-2010, we collected male smallmouth bass (n=118) from 12 sites and largemouth bass (n=173) from 27 sites. Intersex in male smallmouth bass was observed at all sites and ranged from 60% to 100%; in male largemouth bass the range was 0-100%. Estrogenicity, as measured using a bioluminescent yeast reporter, was detected above the probable no effects concentration (0.73ng/L) in ambient water samples from 79% of the NWR sites. Additionally, the presence of androgen receptor and glucocorticoid receptor ligands were noted as measured via novel nuclear receptor translocation assays. Mean plasma Vtg was elevated (>0.2mg/ml) in male smallmouth bass at four sites and in male largemouth bass at one site. This is the first reconnaissance survey of this scope conducted on US National Wildlife Refuges. The baseline data collected here provide a necessary benchmark for future monitoring and justify more comprehensive NWR-specific studies.

Effects of urban land-use on largescale stonerollers in the Mobile River Basin, Birmingham, AL.

During the spring and fall of 2001 and the spring of 2002 a study was conducted to evaluate the health of the largescale stoneroller (Campostoma oligolepis) populations in streams along an urban land-use gradient. Sites were selected from a pool of naturally similar sub-basins (eco-region, basin size, and geology) of the Mobile River basin (MRB), using an index of urban intensity derived from infrastructure, socioeconomic, and land-use data. This urban land-use gradient (ULUG) is a multimetric indicator of urban intensity, ranging from 0 (background) to 100 (intense urbanization). Campostoma sp. have been used previously as indicators of stream health and are common species found in all sites within the MRB. Endpoints used to determine the effects of urban land-use on the largescale stoneroller included total glutathione, histology, hepatic apoptosis, condition factor and external lesions. Liver glutathione levels were positively associated with increasing urban land-use (r(2) = 0.94). Histopathological examination determined that some abnormalities and lesions were correlated with the ULUG and generally increased in prevalence or severity with increasing urbanization. Liver macrophage aggregates were positively correlated to the ULUG. The occurrence of nucleosomal ladders (indicating apoptotic cell death) did not correspond with urban intensity in a linear fashion. Apoptosis, as well as prevalence and severity of a myxozoan parasite, appeared to have a hormetic dose-response relationship. The majority of the biomarkers suggested fish health was compromised in areas where the ULUG ≥ 36.

Health of white sucker within the St. Louis River area of concern associated with habitat usage as assessed using stable isotopes.

In Spring 2011, 200 adult white sucker were collected in four areas of the St. Louis River area of concern (AOC), located in Minnesota and Wisconsin, USA. The areas included the upper AOC as a reference area, the upper estuary, St. Louis Bay and Superior Bay. Grossly visible abnormalities were documented and preserved for microscopic analyses, as were five to eight representative pieces of liver tissue. A piece of dorsal muscle was preserved for stable isotope analyses and otoliths removed for age determination. The incidence of raised skin lesions (mucoid plaques) was high (31 %), however, microscopically only 4.5 % of the white suckers had neoplasia (papillomas). The remaining lesions were epidermal hyperplasia. Superior Bay had the lowest percentage of skin/lip lesions (10 %), while St. Louis Bay had the highest (44 %). St. Louis Bay also had the highest incidence of skin neoplasms (12 %). No hepatocellular neoplasms were documented, however bile duct tumors were observed in 4.5 % of the suckers. Foci of cellular alteration were observed in fish from all sites except the upper AOC. Stable isotope data indicated that most of the suckers relied on the St. Louis River AOC for the majority (>75 %) of their diet, indicating they were resident within the AOC and not in Lake Superior. The amount of diet obtained from the upper estuary was a significant predictor of skin lesion incidence. Hence, habitat use within the AOC appears to be an important risk factor for skin and possibly, liver lesions.

Reproductive health indicators of fishes from Pennsylvania watersheds: association with chemicals of emerging concern.

Fishes were collected at 16 sites within the three major river drainages (Delaware, Susquehanna, and Ohio) of Pennsylvania. Three species were evaluated for biomarkers of estrogenic/antiandrogenic exposure, including plasma vitellogenin and testicular oocytes in male fishes. Smallmouth bass Micropterus dolomieu, white sucker Catostomus commersonii, and redhorse sucker Moxostoma species were collected in the summer, a period of low flow and low reproductive activity. Smallmouth bass were the only species in which testicular oocytes were observed; however, measurable concentrations of plasma vitellogenin were found in male bass and white sucker. The percentage of male bass with testicular oocytes ranged from 10 to 100%, with the highest prevalence and severity in bass collected in the Susquehanna drainage. The percentage of males with plasma vitellogenin ranged from 0 to 100% in both bass and sucker. Biological findings were compared with chemical analyses of discrete water samples collected at the time of fish collections. Estrone concentrations correlated with testicular oocytes prevalence and severity and with the percentage of male bass with vitellogenin. No correlations were noted with the percentage of male sucker with vitellogenin and water chemical concentrations. The prevalence and severity of testicular oocytes in bass also correlated with the percent of agricultural land use in the watershed above a site. Two sites within the Susquehanna drainage and one in the Delaware were immediately downstream of wastewater treatment plants to compare results with upstream fish. The percentage of male bass with testicular oocytes was not consistently higher downstream; however, severity did tend to increase downstream.

Temporal evaluation of estrogenic endocrine disruption markers in smallmouth bass (Micropterus dolomieu) reveals seasonal variability in intersex.

A reconnaissance project completed in 2009 identified intersex and elevated plasma vitellogenin in male smallmouth bass inhabiting the Missisquoi River, VT. In an attempt to identify the presence and seasonality of putative endocrine disrupting chemicals or other factors associated with these observations, a comprehensive reevaluation was conducted between September 2012 and June 2014. Here, we collected smallmouth bass from three physically partitioned reaches along the river to measure biomarkers of estrogenic endocrine disruption in smallmouth bass. In addition, polar organic chemical integrative samples (POCIS) were deployed to identify specific chemicals associated with biological observations. We did not observe biological differences across reaches indicating the absence of clear point source contributions to the observation of intersex. Interestingly, intersex prevalence and severity decreased in a stepwise manner over the timespan of the project. Intersex decreased from 92.8% to 28.1%. The only significant predictor of intersex prevalence was year of capture, based on logistic regression analysis. The mixed model of fish length and year-of-capture best predicted intersex severity. Intersex severity was also significantly different across late summer and early spring collections indicating seasonal changes in this metric. Plasma vitellogenin and liver vitellogenin Aa transcript abundance in males did not indicate exposure to estrogenic endocrine disrupting chemicals at any of the four sample collections. Analysis of chemicals captured by the POCIS as well as results of screening discrete water samples or POCIS extracts did not indicate the contribution of appreciable estrogenic chemicals. It is possible that unreported changes in land-use activity have ameliorated the problem, and our observations indicate recovery. Regardless, this work clearly emphasizes that single, snap shot sampling for intersex may not yield representative data given that the manifestation of this condition within a population can change dramatically over time.

Flow cytometric method for measuring chromatin fragmentation in fixed sperm from yellow perch (Perca flavescens).

Declining harvests of yellow perch, Perca flavescens, in urbanized watersheds of Chesapeake Bay have prompted investigations of their reproductive fitness. The purpose of this study was to establish a flow cytometric technique for DNA analysis of fixed samples sent from the field to provide reliable gamete quality measurements. Similar to the sperm chromatin structure assay, measures were made on the susceptibility of nuclear DNA to acid-induced denaturation, but used fixed rather than live or thawed cells. Nuclei were best exposed to the acid treatment for 1 minute at 37 °C followed by the addition of cold (4 °C) propidium iodide staining solution before flow cytometry. The rationale for protocol development is presented graphically through cytograms. Field results collected in 2008 and 2009 revealed DNA fragmentation up to 14.5%. In 2008, DNA fragmentation from the more urbanized watersheds was significantly greater than from reference sites (P = 0.026) and in 2009, higher percentages of haploid testicular cells were noted from the less urbanized watersheds (P = 0.032) indicating better reproductive condition at sites with less urbanization. For both years, total and progressive live sperm motilities by computer-assisted sperm motion analysis ranged from 19.1% to 76.5%, being significantly higher at the less urbanized sites (P < 0.05). This flow cytometric method takes advantage of the propensity of fragmented DNA to be denatured under standard conditions, or 1 minute at 37 °C with 10% buffered formalin-fixed cells. The study of fixed sperm makes possible the restrospective investigation of germplasm fragmentation, spermatogenic ploidy patterns, and chromatin compaction levels from samples translocated over distance and time. The protocol provides an approach that can be modified for other species across taxa.

The cellular immune response of rainbow trout (Salmo gairdneri Richardson) to sheep red blood cells.

The cellular immune response of rainbow trout, Salmo gairdneri, to sheep erythrocytes was investigated. Both the primary and secondary responses were measured using the migration inhibition factor (MIF), antigen-binding (ABC), and plaque-forming cell (PFC) assays. These immune function assays provide measures of both T and B cell activity. The kinetics of these three responses at 16 degrees C were determined by sampling fish over an 18 day period for the primary response and a ten day period for the secondary response. The peak MIF response occurred two days after injection, while the primary peak PFC response was observed 14 days post-injection. Two ABC peaks were observed in the primary response, one at four days and one at ten days after injection. In the secondary response the peak ABC response was observed four days and the peak PFC response six days post-inoculation. The possible interrelationships of the various cell populations are discussed.

Dietary influences on disease resistance factors in channel catfish.

The effects of diet on selected disease resistance factors were studied in channel catfish (Ictalurus punctatus). Two commercial feeds and a "control," laboratory-prepared diet were compared. Macrophage function (phagocytosis and intracellular killing) was used to assess nonspecific disease resistance and serum antibody was measured as an indication of specific immunity. We measured the immune response to Edwardsiella ictaluri, a common bacterial pathogen of catfish, as well as the phagocytosis and killing of the bacteria. In the initial experiment fish were maintained on the experimental diets for 116 days, vaccinated and responses assayed 14 days later. Significant differences among the groups were observed in the phagocytic index as well as in circulating antibody. An additional study showed that even when fed the experimental diets for only 42 days there were significant differences in the ability of macrophages from both immunized and nonimmunized fish to kill E. ictaluri.

Quantitative polymerase chain reaction for transforming growth factor-beta applied to a field study of fish health in Chesapeake Bay tributaries.

Fish morbidity and mortality events in Chesapeake Bay tributaries have aroused concern over the health of this important aquatic ecosystem. We applied a recently described method for quantifying mRNA of an immunosuppressive cytokine, transforming growth factor-beta (TGF-beta), by reverse transcription quantitative-competitive polymerase chain reaction to a field study of fish health in the Chesapeake Basin, and compared the results to those of a traditional cellular immunoassay macrophage bactericidal activity. We selected the white perch (Morone americana) as the sentinel fish species because of its abundance at all of the collection sites. White perch were sampled from Chesapeake Bay tributaries in June, August, and October 1998. Splenic mononuclear cell TGF-beta mRNA levels increased and anterior kidney macrophage bactericidal activity decreased, particularly in eastern shore tributaries, from June to August and October. The results of the two assays correlated inversely (Kendall's [Tau] b = -0.600; p = 0.0102). The results indicated both temporal and spatial modulation of white perch immune systems in the Chesapeake Basin, and demonstrated the utility of quantitative PCR for TGF-beta as a molecular biomarker for field assessment of teleost fish immune status.

Immunoregulation of fish nonspecific cytotoxic cell activity by retinolacetate but not poly I:C.

Nonspecific cytotoxic cells (NCC) from fish (Ictalurus punctatus) were treated with different concentrations of retinolacetate and poly I:C. Both in vitro and in vivo experiments were conducted. Retinolacetate significantly increased NCC activity against chromium-51 labeled human B-cell lymphoma target cells (NC-37). Preincubation (treatment prior to adding the labeled target cells) of NCC for 4 to 8 h in 10(-3) to 10(-12) M concentration of retinolacetate produced significant increases in NCC activity compared to treatment during the killing assay only. Similar experiments with different concentrations of poly I:C had no NCC augmenting effects when tested by adding poly I:C either during preincubation periods or during the cytotoxicity assay. Retinolacetate probably produces positive modulation of cytotoxicity by increasing the killing effectiveness of individual NCC, rather than recruiting larger numbers of cytolytic cells. In vitro studies were also conducted by injecting catfish (i.p.) with 1 X, 3X and 5X the daily recommended vitamin A dosages and determining NCC activity after 24, 48 and 72 h treatment. The 1X dose significantly increased NCC activity at 72 h. This increase was not transient because NCC activity after 33-37 days' treatment was significantly higher than controls in the 1X, 2X and 3X groups. Intraperitoneal injections of fish with poly I:C produced no significant increases in NCC activity at 24 or 72 h post-inoculation.

Tissue disposition and excretion of 14C-labelled aflatoxin B1 after oral administration in channel catfish.

The pharmacokinetics, tissue distribution and excretion of 14C-labelled aflatoxin B1 (AFB1) were examined after oral administration (250 micrograms/kg body weight) in channel catfish (Ictalurus punctatus). Plasma concentrations of parent AFB1 were best described by a one-compartment pharmacokinetic model, in which peak plasma concentration (503 ppb) occurred at 4.1 hr after dosing. The absorption and elimination half-lives were 1.5 and 3.7 hr, respectively. AFB1 was highly bound (95%) to plasma proteins. Concentrations of 14C (in AFB1 equivalents) measured in the tissues were highest at 4 hr, ranging from 596 ppb in the plasma to 40 ppb in the muscle. AFB1 residues were rapidly depleted; at 24 hr the concentrations in the plasma and muscle were 32 and less than 5 ppb, respectively. Concentrations in the bile exceeded 2000 ppb (at 24 hr), whereas the highest concentration in the urine was 51 ppb (4-6-hr collection interval). Renal and biliary excretion accounted for less than 5% of the administered dose, indicating incomplete absorption. Pharmacokinetic modelling and tissue data demonstrate a very low potential for the accumulation of AFB1 and its metabolites in the edible flesh of channel catfish through the consumption of AFB1-contaminated feed.

Comparative susceptibility of Atlantic salmon, lake trout and rainbow trout to Myxobolus cerebralis in controlled laboratory exposures.

The susceptibility of lake trout Salvelinus namaycush, rainbow trout Oncorhynchus mykiss and Atlantic salmon Salmo salar to Myxobolus cerebralis, the causative agent of whirling disease, was compared in controlled laboratory exposures. A total of 450 (225 for each dose) fry for each species were exposed to a low (200 spores per fish) or high (2000 spores per fish) dose of the infective triactinomyxon. At 22 wk post-exposure, 60 fish from each group, as well as controls for each species, were examined for clinical signs (whirling behavior, blacktail, deformed heads and skeletal deformities), microscopic lesions, and presence of spores. Rainbow trout were highly susceptible to infection, with 100% being positive for spores and with microscopic pathological changes in both exposure groups. Rainbow trout were the only species to show whirling behavior and blacktail. Atlantic salmon were less susceptible, with only 44 and 61% being positive for spores, respectively, in the low and high dose groups, while 68 and 75%, respectively, had microscopic pathology associated with cartilage damage. Rainbow trout heads contained mean spore concentrations of 2.2 (low dose) or 4.0 (high dose) x 10(6) spores g tissue(-1). The means for positive Atlantic salmon (not including zero values) were 1.7 (low) and 7.4 (high) x 10(4) spores g tissue(-1). Lake trout showed no clinical signs of infection, were negative for spores in both groups and showed no histopathological signs of M. cerebralis infection.

Myxosporean plasmodial infection associated with ulcerative lesions in young-of-the-year Atlantic menhaden in a tributary of the Chesapeake Bay, and possible links to Kudoa clupeidae.

Ulcers in Atlantic menhaden Brevoortia tyrannus (Latrobe) (Clupeidae), observed along the USA east coast, have been attributed to diverse etiologies including bacterial, fungal and, recently, harmful algal blooms. To understand the early pathogenesis of these lesions, we examined juvenile Atlantic menhaden collected during their seasonal presence in Chesapeake Bay tributaries from April to October 1999 and from March to August 2000. We conducted histopathological examinations of young-of-the-year fish from the Pocomoke River tributary, which has a history of fish mortalities and high lesion prevalence. Kudoa clupeidae (Myxozoa: Myxosporea) spores were present in the muscles of fish collected in both years. Of the fish assessed by histology in April, 5 to 14% were infected, while in May 90 to 96% were infected. Infection rates remained high during the summer. Mature spores were primarily located within myomeres and caused little or no observable pathological changes. Ultrastructure showed spores with capsulogenic cells bearing filamentous projections, and a basal crescentic nucleus with mottled nucleoplasm containing cleaved, condensed chromatin. Also, a highly invasive plasmodial stage of a myxozoan was found in the lesions of juvenile Atlantic menhaden. The plasmodia were observed in fish collected between May and July, with the maximum occurrence in late June 1999 and late May 2000. Plasmodia penetrated and surrounded muscle bundles, causing grossly observable raised lesions in 73% of all fish infected with this invasive stage. Plasmodia were also detected in the visceral organs, branchial arches, and interocular muscles of some fish. Some of the invasive extrasporogonic plasmodial lesions were associated with ulcers and chronic inflammatory infiltrates. The plasmodial stage appeared to slough out of the tissue with subsequent evidence of wound healing. Ultrastructure showed plasmodia with an elaborate irregular surface, divided into distinct ectoplasm and endoplasm; the latter contained numerous spherical vegetative nuclei, secondary generative cells, and occasional cell doublets. Our ultrastructural studies indicate that the plasmodial organisms, which are important in the etiology of the skin lesions, are myxozoans, and they may represent early stages of K. clupeidae.

Cartilage proliferation in response to metacercarial infections of fish gills.

The pathology observed in a metacercarial (species not yet identified) infection of pond-raised, aquarium fish is described. In natural infections, there was extensive proliferation of cartilage resulting in loss of respiratory epithelium and occasionally in fusion of primary lamellae. Grossly, the most consistent sign of infection was flared opercula with protruding gills. Although haemorrhage was produced in early infections as the cercariae migrated, it was not macroscopically evident. A few heavily infected fish did have grossly observable signs of haemorrhage but this was not a consistent finding. Cercariae were collected from snails (Pleurocera sp.) and fish were experimentally infected to study the response. Over a 6-day-period, a progression from fibroblast to chondroblast occurred, indicating that piscine fibroblasts are able to differentiate into chondroblasts. This differentiation may be dependent on environmental conditions or may be stimulated by the parasite itself.

Effects of whirling disease on selected hematological parameters in rainbow trout.

Hematological responses to whirling disease in rainbow trout (Oncorhynchus mykiss) were investigated. Two-mo-old fingerling rainbow trout were exposed to cultured triactinomyxon spores of Myxobolus cerebralis at 9,000 spores/fish in December, 1997. Twenty-four wks post-exposure, fish were taken from infected and uninfected groups for peripheral blood and cranial tissue sampling. Histological observations on cranial tissues confirmed M. cerebralis infection in all exposed fish. Differences in hematological parameters between the two groups included significantly lower total leukocyte and small lymphocyte counts for the infected fish. No effects on hematocrit, plasma protein concentration, or other differential leukocyte counts were noted.

Characterization of a new myxozoan species (Myxozoa: Myxobolidae: Myxosporea) in largescale stonerollers (Campostoma oligolepis) from the Mobile River Basin (Alabama).

Myxobolus stanlii sp. n. was described from largescale stonerollers ( Campostoma oligolepis ) from the Mobile River Basin in Alabama. The parasite was described using critical identifying morphological features, and the 18S small subunit ribosomal RNA (SSU rRNA) gene sequence. The spore body was ovoid, 10.03 ± 0.7 (7.5-11.0) µm long and 8.8 ± 1.5 (6.3-11.3) µm wide in frontal view. Spore thickness was 6.3 ± 2.7 (6.2-8.6) µm in sutural view. Polar capsules were pyriform, of equal size, and oriented in plane with the sutural ridge. Polar capsules were 2.45 ± 1.5 (range 2.1-4.3) µm in width and 4.6 ± 2.7 (range 4.5-6.9) µm in length. Based on the SSU rRNA gene sequence of Myxobolus stanlii sp. n. is most closely related to M. pseudodispar.

A redescription of Myxobolus inornatus from young-of-the-year smallmouth bass (Micropterus dolomieu).

During investigations of young-of-the year smallmouth bass ( Micropterus dolomieu ) mortalities in the Susquehanna River, Pennsylvania, U.S.A. and affected tributaries, raised areas were noted in the muscle in the vicinity of the caudal peduncle. The raised areas were caused by plasmodia of a myxozoan parasite. Spores found within plasmodia were similar to those of Myxobolus inornatus previously described from the caudal peduncle of fingerling largemouth bass ( Micropterus salmoides ) in Montana. Here, M. inornatus is redescribed based on histologic comparisons and spore measurements. The addition of spore photographs, line drawings, a voucher specimen, and partial small-subunit ribosomal (rSSU) DNA gene sequence are new in this study. This is also the first description of M. inornatus from smallmouth bass. The plasmodia of M. inornatus were grossly observed at the base of the caudal and dorsal fins and were 280.3 ± 33.5 (range 77.1-920.3) µm long and 320.6 ± 41.0 (range 74.85-898.4) µm wide. In some instances, plasmodia of M. inornatus were large enough to rupture the epidermis or were associated with misaligned vertebrae. The slightly pyriform spores were 11.3 ± 0.2 (range 8.6-17.4) µm in length and 8.6 ± 0.2 (range 7.1-13.7) µm wide with an iodinophilous vacuole and a sutural ridge with 8 to 10 sutural folds. The SSU rDNA gene sequence places M. inornatus in a sister group with Myxobolus osburni .

Mortality of centrarchid fishes in the Potomac drainage: survey results and overview of potential contributing factors.

Skin lesions and spring mortality events of smallmouth bass Micropterus dolomieu and selected other species were first noted in the South Branch of the Potomac River in 2002. Since that year morbidity and mortality have also been observed in the Shenandoah and Monocacy rivers. Despite much research, no single pathogen, parasite, or chemical cause for the lesions and mortality has been identified. Numerous parasites, most commonly trematode metacercariae and myxozoans; the bacterial pathogens Aeromonas hydrophila, Aeromonas salmonicida, and Flavobacterium columnare; and largemouth bass virus have all been observed. None have been consistently isolated or observed at all sites, however, nor has any consistent microscopic pathology of the lesions been observed. A variety of histological changes associated with exposure to environmental contaminants or stressors, including intersex (testicular oocytes), high numbers of macrophage aggregates, oxidative damage, gill lesions, and epidermal papillomas, were observed. The findings indicate that selected sensitive species may be stressed by multiple factors and constantly close to the threshold between a sustainable (healthy) and nonsustainable (unhealthy) condition. Fish health is often used as an indicator of aquatic ecosystem health, and these findings raise concerns about environmental degradation within the Potomac River drainage. Unfortunately, while much information has been gained from the studies conducted to date, due to the multiple state jurisdictions involved, competing interests, and other issues, there has been no coordinated approach to identifying and mitigating the stressors. This synthesis emphasizes the need for multiyear, interdisciplinary, integrative research to identify the underlying stressors and possible management actions to enhance ecosystem health.