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METHOD DEVELOPMENT FOR RAPID STREAMSIDE ASSESSMENT OF FRESHWATER MUSSEL (ORDER UNIONIDA) HEMOCYTE CONCENTRATION AND MORPHOMETRICS
Madison E. Mueller, W. Gregory Cope, Christopher B. Eads, Loretta M. Lutackas, and Dorian J. Hayes

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ABSTRACT
Contemporary freshwater mussel population declines often lack discriminant explanations, and diagnoses are commonly post hoc, subjective, expensive, and/or lethal. To avoid further compromising imperiled populations, we need sublethal metrics and biomarkers that identify early warning signs of infection, stress responses, and deviations from physiological homeostasis or potential resilience to environmental pressures. Immune modulators can be differentiated by quantifying multiple physiological endpoints based on hemocyte form and function. Spontaneous hemocyte aggregation, cell instability, travel times from mussel collection sites, and a lack of access to costly instrumentation currently limit immune status assessments. Widespread capacity to understand immune responses to disease, toxicants, and other stressors relies on effective hemolymph handling and preservation techniques. In this study we optimized methods to measure total hemocyte concentration and stability across five antiaggregants (Alsever’s Solution, K2EDTA and sodium citrate Microtainers, Isoton II, and Leibovitz’s L15) over time using a Beckman Coulter Multisizer 4e (a laboratory benchtop unit) to validate results from aMillipore Scepter (a portable, field-ready hand-held unit). K2EDTA Microtainers held on ice for up to 6 h preserved hemolymph most effectively based on consistency in total hemocyte concentration and morphometrics over time. There was a highly significant agreement (R2 ¼ 0.9156, P , 0.0001) in total hemocyte concentration between the Multisizer 4e and Millipore Scepter. Portable and compact cell counters provide a less expensive, advantageous alternative to benchtop instruments for instantaneous evaluation at mussel collection sites when timely quantification would otherwise require extensive logistical planning. Our results lay the foundation for accessible streamside assessment, which can expedite the establishment of reference ranges among species and populations and identify how anticipated stressors such as infection and starvation alter the immune response. This approach also may be useful for quantifying freshwater mussel health among propagation facilities and in stable or declining ecosystems.


GENETIC VARIATION IN AN INVASIVE CLAM (FORM B: CORBICULA LARGILLIERTII) ACROSS ILLINOIS, USA
Brenden Mattfolk, Julia W. Bondi Nicholas J. Iacaruso, Jeremy Tiemann, Mark A. Davis, and Joel B. Corush

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ABSTRACT
Population genetics is an important tool for understanding the source, establishment, and subsequent spread of invasive species. It is particularly pertinent for invasive species that go unrecognized or are understudied due to their limited detection, cryptic diversity, and/or overall lack of interest. One genus of invasive clams, Corbicula, has been a particularly successful invader, both globally and in the USA, but has received less attention than other aquatic invaders (e.g., invasive carp or dreissenid mussels). Herein, we address the genetic diversity of one of the three invasive Corbicula species—Form B: Corbicula largillierti— by using the 28S nuclear and cytochrome c oxidase subunit 1 (COI) mitochondrial gene in 74 individuals from six localities across multiple watersheds in Illinois, USA.We identified spatial structure in both genes and revealed an overall high level of genetic diversity and differentiation among localities. This information can be useful for addressing questions pertaining to propagule pressure, local spread, and overall genetic variation of this otherwise understudied invasive species.


CHARACTERIZATION OF THE NATIVE FRESHWATER MUSSEL ASSEMBLAGE ACROSS A REACH OF THE RAPPAHANNOCK RIVER, VIRGINIA
Traci P. DuBose1, Jason Rohweder, Catherine Gatenby, Brian Watson, and Teresa J. Newton

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ABSTRACT
Native freshwater mussels occur in rivers of all sizes, yet much of the literature focuses on small- to medium-sized rivers, likely because of logistic challenges of sampling deep, fast, and turbid large rivers. As large-river surveys become more common, the information resulting from a systematic sampling design can provide baseline data on population sizes to inform decision making by resource managers. We collected baseline data onmussel resources in the Rappahannock River, Virginia, in August 2024. This Atlantic coast river, which drains into the Chesapeake Bay, is ~320 km long, up to 1.6 km wide, and up to 35 m deep.We conducted a systematic quantitative survey across the 75-km reach of the river from Fredericksburg to Leedstown, Virginia. Divers obtained 187 whole substrate samples to estimate the species richness, density, population size, size distribution, community composition, and longitudinal spatial distribution of the native mussel assemblage. Six species were found, and the community was dominated by Eastern Elliptio (Elliptio complanata).Mean reach density (6 1 standard error) and population sizewas 1.25 (0.21)musselsm22 and 19.3 (3.3) million mussels, respectively. The presence of few fresh dead mussels indicated low to moderate natural mortality. Shell length ranged from 32 to 122 mm, and multiple size classes were observed in E. complanata. Species richness and density had similar longitudinal patterns, with the highest values in themiddle of the sampled reach (river kilometers 60–95). These data provide resource managers with baseline information on the spatial distribution, density, length demography, diversity, and evenness of native freshwater mussels across this reach of the Rappahannock River. Identi!cation of mussel occurrence, density, and evidence of reproduction can be used by managers to aid conservation and management decisions such as prioritizing areas for reintroductions and habitat restoration.


UPPER THERMAL TOLERANCE OF THE WESTERN RIDGED MUSSEL (GONIDEA ANGULATA), WESTERN PEARLSHELL (MARGARITIFERA FALCATA), AND WESTERN SCULPIN SPECIES (COTTUS SPP.)
Rachael Valeria1, Alexa Maine, Christine Parent, and James J. Nagler

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ABSTRACT
Freshwater mussels are ecologically and culturally important and globally have the highest extinction and imperilment rates of any animal. The Western Ridged Mussel (Gonidea angulata) and the Western Pearlshell (Margaritifera falcata) are experiencing die-offs across their native ranges, and increasing water temperature is a possible contributing factor. To host their larval life stage, G. angulata uses the gills of sculpins (Cottus spp.) and M. falcata uses the gills of salmonids (Oncorhynchus spp.). The purpose of this laboratory study was to determine the upper thermal tolerances of G. angulata and M. falcata and two sculpin species, Margined Sculpin (Cottus marginatus) and Paiute Sculpin (Cottus beldingi), by using sublethal acute and chronic survival tests. The acute test consisted of increasing the water temperature until critical thermal maximum (CTmax) temperature behaviors were observed. The mean CTmax temperature was 34.1°C for G. angulata and 33.2°C for M. falcata, and the mean for the two Cottus spp. was 28.9°C. Based on the acute test results, G. angulata and M. falcata have a higher CTmax temperature than their host fish. The chronic test gradually increased the water temperature surrounding the test organism to 2°C below the CTmax temperature for each species. This temperature was maintained for 7 d, during which survival was recorded. All mussels perished, whereas 58.3% of the sculpins survived the chronic test. Survival curves from the chronic test showed that G. angulata survived longer and are more tolerant to prolonged elevated temperatures than M. falcata. These thermal tolerance tests demonstrate that a thermal separation between mussel and host fish is possible, thus limiting mussel reproduction as climate change leads to warming surface waters. In addition, tolerance to prolonged elevated temperature varies between these western mussel species.