2013 NORTHEASTERN NATURALIST 20(1):49–68 Host Fishes and Conservation Status of Alasmidonta marginata (Bivalvia: Unionidae) in Minnesota Kylie H. Bloodsworth1,*, Ben R. Bosman1, Bernard E. Sietman1, and Mark C. Hove2 Abstract -We examined laboratory host suitability and assessed the distribution and status of Alasmidonta marginata (Elktoe) in Minnesota. Of the 85 fish species tested, glochidia metamorphosed on 27 species in 6 families (Cyprinidae, Catostomidae, Fundulidae, Poeciliidae, Gasterosteidae, and Cottidae). All catostomid species facilitated metamorphosis, and overall, Catostomidae produced more juvenile mussels per fish. This result, in combination with a previous finding of naturally infested fish, suggests that catostomids are an important host for A. marginata in nature. From extensive surveys, we found extant populations of A. marginata in the St. Croix River, Upper Mississippi River, and Minnesota River systems. Alasmidonta marginata is apparently extirpated or its range has decreased in several interior Minnesota watersheds and the Mississippi River main stem. Barrier waterfalls and habitat degradation have influenced A. marginata’s historic and recent distribution more so than the range of its hosts. Further study of naturally occurring and laboratory hosts for A. marginata and other Alasmidonta species is needed in order to improve conservation efforts and elucidate phylogenetic relationships for this group of mussels. Introduction North America contains the largest number of freshwater mussel (Unionidae) species in the world, yet the majority of taxa are recognized as endangered, threatened, or of special concern (Bogan 2008, Lydeard et al. 2004, Williams et al. 1993). Improving rare mussel conservation requires a better understanding of a species’ distribution and life history. Freshwater mussels have a specialized life cycle in which their larvae (glochidia) must temporarily parasitize the gills, fins, and other extremities of fish to complete development into free-living juveniles (Lefevre and Curtis 1910, Zale and Neves 1982a). Variation in host associations among unionid groups reveal patterns that are important for understanding evolutionary lineages (Barnhart et al. 2008). Species in the Tribe Anodontini, known for their terminally hooked, triangular-shaped glochidia (Clarke 1981a, Lefevre and Curtis 1910), have been shown to use a wide variety of fish hosts (Barnhart and Roberts 1997, Haag and Warren 1997, Trdan and Hoeh 1982, van Snik Gray et al. 2002). Suitable host relationships within the genus Alasmidonta, follow two general patterns: 1) species in the subgenera Alasmidonta and Decurambis with hosts in the fish families Cyprinidae (Minnows), Catostomidae (Suckers), 1Minnesota Department of Natural Resources, Division of Ecological and Water Resources, 500 Lafayette Road, Saint Paul, MN 55155. 2University of Minnesota, Department of Fisheries, Wildlife and Conservation Biology, 1980 Folwell Avenue, Saint Paul, MN 55108. *Corresponding author - Kbloodsworth@une.edu. 50 Northeastern Naturalist Vol. 20, No. 1 Poeciliidae (Live Bearers), Cottidae (Sculpins), Centrarchidae (Sunfishes), and Percidae (Perches), and 2) species in the subgenus Pressodonta with hosts in the families of Cottidae and Percidae (Darters) only (see Appendix 1). There has been no comprehensive phylogenetic analysis published for Alasmidonta; however, variation in host associations could reflect divergent lineages (Barnhart et al. 2008, Haag and Warren 2003), such as with genera in the Tribe Quadrulini (Fritts et al. 2012). Alasmidonta marginata (Elktoe) is the most widespread member of its genus, occurring broadly in the Mississippi, Ohio, and Great Lakes drainages (Clarke 1973, 1981a, 1981b). Potential host fishes for A. marginata include Catostomus commersoni (White Sucker), Hypentelium nigricans (Northern Hogsucker), Moxostoma macrolepidotum (Shorthead Redhorse), Ambloplites rupestris (Rock Bass), and Lepomis gulosus (Warmouth) (Howard and Anson 1922). These host identifications, however, were based solely on observations of naturally infested glochidia without confirmation of successful transformation to the juvenile stage. Such evidence can lead to erroneous host designations (Fritts et al. 2012, Haag and Warren 2003). While these hosts are similar to other Alasmidonta species within the subgenus Decurambis (see Appendix 1), laboratory testing is needed to assess host efficacy and validate unconfirmed host designation s. Adult mussels have limited vagility compared to fish. Larval dispersal, especially upstream, is thought to be the primary advantage of the parasitic phase of the unionid life cycle (Barnhart et al. 2008). Knowledge of host distribution and vagility can explain mussel colonization patterns, range limitations, and extirpations. Although broadly distributed in eastern North America, A. marginata is considered a species of special concern under national conservation assessment (Williams et al. 1993), with evidence that it has declined in parts of its range (Cummings and Mayer 1997, Williams et al. 2008). In Minnesota, it is currently listed as threatened (Natural Heritage and Nongame Research Program 1996), but there has been no recent comprehensive assessment of its distribution and status. Determining the host relationships and conservation status of A. marginata will help guide conservation efforts and provide informative characters for phylogenetic studies. Therefore, the objectives of this study were to 1) identify suitable glochidial hosts for A. marginata in laboratory trials, including species previously reported as potential natural hosts, and 2) evaluate the conservation status of A. marginata in Minnesota by determining its historical and recent distribution. Methods Host suitability Host suitability was examined using standard procedures of artificially inoculating fishes with glochidia and monitoring the success of these infections (Hove et al. 2000, Zale and Neves 1982a). We ran host trials at the University of Minnesota Wet Laboratory (UM) from 2005 to 2008, and the Minnesota Pollution 2013 K.H. Bloodsworth, B.R. Bosman, B.E. Sietman, and M.C. Hove 51 Control Agency Biomonitoring Laboratory (MPCA) from 2008 to 2010. Alasmidonta marginata is presumably bradytictic (long-term brooder), as it becomes gravid in the fall and retains glochidia into winter, similar to other anodontines (Baker 1928). We were unable to find gravid individuals in early spring after ice melt, suggesting glochidia are released sometime during winter or early spring. Therefore, we collected gravid female A. marginata during September and October from the St. Croix River, Washington County, MN (45.393658°N, 92.663634°W); the Root River, Fillmore County, MN (43.814835°N, 92.095656°W); the South Fork Zumbro River, Olmsted County, MN (44.020564°N, 92.456396°W); and the Chippewa River, Dunn County, WI (44.756486°N, 91.815162°W). We collected fishes used for host trials from rivers and lakes in Minnesota and southeast Missouri by seining, electrofishing, and angling. Other fishes were obtained from hatcheries in Wisconsin and Missouri, or the local aquarium trade. Host suitability can vary across river drainages; however, we assume that even though there could be some incompatibilities with mussels and fish collected from different watersheds, potential host(s) will at least exhibit some transformation (Riusech and Barnhart 2000, Rogers et al. 2001). All test fish were held in laboratory tanks at least two weeks prior to inoculating with glochidia or were inspected to ensure no pre-existing glochidia were present. Unionid nomenclature follows Turgeon et al. (1998), and fish nomenclature follows Nelson et al. (2004), except for taxonomic revisions in Blum et al. (2008), Strange and Mayden (2009), and Wood et al. (2002). To prepare for inoculations, we extracted glochidia from brooding female mussels in the laboratory by puncturing the inflated gill and flushing the contents with a syringe (Zale and Neves 1982a). Glochidia were considered mature if valves were fully formed and open, and tissue was present. We tested glochidia viability by adding NaCl to 15–30 glochidia from each female (Coker et al. 1921). If ≥70% of glochidia snapped shut quickly in response to salt exposure, the remaining glochidia were used for inoculation (Coker et al. 1921). Adult mussels were returned to their original collection site. We inoculated 85 fish species (17 families) and 1 amphibian species with glochidia. Fish were placed in a vigorously aerated water bath (3–7 L [1–2 gal]) containing several thousand glochidia. Depending on each species’ susceptibility to infestation, fish were exposed to glochidia for 30 seconds to 45 minutes until at least 10–20 glochidia had attached to gills and/or fins of fishes 2–10 cm in length, or 50–100 glochidia to fishes >10 cm in length (Hove et al. 2000). In some instances, we pipetted glochidia directly onto the gills if little or no attachment had occurred after 45 minutes. After exposure, we assessed glochidia attachment using a dissecting microscope while another person applied a gentle stream of water to separate the gill filaments. After sufficient glochidia attachment was observed, fish were placed in community holding tanks. Three to four days after inoculation, each fish was re-examined for encysted glochidia. If glochidia were no longer present on the gills or fins of any individuals, the trial for that fish species was ended. If encysted glochidia remained on the gills or fins after 3–4 days, all individual members of 52 Northeastern Naturalist Vol. 20, No. 1 that fish species were housed in a single monospecific aquarium at 22 ± 2 °C (MPCA) or 18 ± 3 °C (UM) for additional monitoring. Fish were fed daily. Each aquarium contained either a false bottom or suspended nets to prevent fish from consuming sloughed glochidia and juvenile mussels. Subsequently, water from the aquarium floor was siphoned every 3–4 days to monitor the presence of sloughed glochidia and transformed juveniles. Siphoned water was washed across two sieves with 1-mm and 125-μm mesh openings, respectively. We recorded the number of juveniles per tank for each siphoning event using a dissecting microscope. Transformed juveniles were distinguished from glochidia by valve movement, the presence or movement of a foot, growth beyond the glochidial valve, or the presence of two adductor muscles. We also counted empty shells of dead juveniles that showed obvious growth beyond the glochidial valve. Recovery of juveniles confirmed successful transformation had occurred, and a sample of transformed juveniles from each trial was preserved in 95% ethanol. A trial continued until 3 consecutive siphoning events revealed no glochidia or juveniles, or fish gills and fins were without glochidia upon visual inspection. Occasionally, a fish died at the beginning or during the juvenile release period. We inspected the dead fish’s gills under a dissecting microscope, and if glochidia were attached we clipped out the gills and put them in a 500-ml beaker with water and gentle aeration. Contents were sieved every 2–4 days until glochidia were no longer attached to gills. We recorded the number of juvenile mussels and combined them with the total count for their respective fish species. In 2009, trials with Notemigonis crysoleucas (Golden Shiner), Rhinichthys cataractae (Longnose Dace), Ictiobus bubalus (Smallmouth Buffalo), and Moxostoma anisurum (Silver Redhorse) produced some juveniles at the beginning of the transformation period (10 days post inoculation) that did not exhibit the standard juvenile-defining criteria i.e., obvious adductor muscles or valve movement. We defined these individuals as pre-juveniles on the date of collection. Consequently, we held these young mussels in a species-specific 500-ml beaker containing aerated water and re-examined them 2 and 5 days post-collection. By day 5, we recovered 33 juveniles exhibiting well-defined characteristics. In 2010, we recovered pre-juveniles from 8 species: Erimyzon oblongus (Creek Chubsucker), Hypentelium nigricans, Moxostoma erythrurum (Golden Redhorse), Fundulus diaphanus (Banded Killifish), Fundulus olivaceus (Blackspotted Topminnow), Gambusia affinis (Western Mosquitofish), Poecilia sphenops (Mexican Molly), and Xiphophorus maculatus (Southern Platyfish) 10–14 days post inoculation, and held and monitored them as before. We recovered 339 juveniles and added them to the total count for their respective fish species. Distribution and status We gathered information from several sources to describe the recent status and distribution of A. marginata in Minnesota. Most data came from the Minnesota Department of Natural Resources (MN DNR) statewide surveys from 1999 to 2011. Surveys were based on qualitative methods (i.e., timed searches) (Allen et al. 2007). Additionally, we compiled data from mussel surveys in the 2013 K.H. Bloodsworth, B.R. Bosman, B.E. Sietman, and M.C. Hove 53 Zumbro (Bright et al. 1988), Minnesota (Bright et al. 1990), Pomme de Terre and Chippewa (Bright et al. 1995), Cannon (Davis 1987), and St. Croix (Doolittle 1988, Hove and Hornbach 2004) rivers. All data used to evaluate the recent status of A. marginata in Minnesota were collected between 1985 and 201 1. To determine the historical distribution of A. marginata, we gathered data from several sources, including 1) relic shells collected in our recent surveys listed above, 2) museums records housed at the Illinois Natural History Survey Mollusk Collection (INHS), University of Minnesota’s James Ford Bell Museum of Natural History (JFBM), Milwaukee Public Museum, and Ohio State Museum of Biological Diversity, and 3) literature pertaining to the region (Baker 1928, Chelberg 1974, Dawley 1945, Fuller 1980, Grier 1922, Havlik 1983, Havlik and Stansbery 1978, Thiel 1981, van der Schalie and van der Schalie 1950). Results Host suitability Alasmidonta marginata glochidia successfully transformed on 27 fish species in 6 families (Appendix 2). Metamorphosis was confirmed for all species of Catostomidae (9 species in 6 genera), Cottidae (3 species), and Gasterosteidae (Sticklebacks) (1 species) tested. Among fish families that produced juvenile mussels, Catostomidae had the highest mean number of juveniles per fish (Fig. 1). Successful juvenile transformation was variable among species of Cyprinidae (8 out of 29 species) and Poeciliidae (3 out of 4 sp ecies), and results were inconsistent among trials for six cyprinid species and one fundulid species (Appendix 2). Distribution and status We compiled data from 3166 sample sites to describe the recent and historical distribution of A. marginata in Minnesota and bordering waters. Since 1985, 384 Figure 1. Juvenile production of Alasmidonta marginata among fish host families. Values (± SE) were calculated by averaging the mean number of juveniles/fish among trials for each species within a given family. n = number of fish species with successful transformation. 54 Northeastern Naturalist Vol. 20, No. 1 live individuals were found in the St. Croix, Minnesota, and Upper Mississippi (below St. Anthony Falls) river main stems and 8 tributaries (Table 1, Fig. 2). Over 75% were found in the St. Croix River drainage, with the main stem having the largest population in Minnesota. However, relative abundance was low (less than 2%) in all river systems (Table 1). Alasmidonta marginata is presumably extirpated from the Cannon and Cedar River drainages and nearly extirpated from the Minnesota River system, except for a small recruiting population in the Pomme de Terre River (Table 1, Fig. 2). The range of A. marginata has apparently declined in the Mississippi River main stem and Root River drainage, where it occurred at only 50% and 41% of historic sites, respectively (T able 1, Fig. 2). Discussion Host suitability Catostomidae have been viewed as poor hosts for unionids (Coker et al. 1921), however, all catostomid species we tested with A. marginata glochidia produced juveniles, and as a group, they produced significantly more juveniles per fish than other fish families that facilitated metamorphosis. Howard and Anson (1922) listed C. commersoni, H. nigricans, and M. macrolepidotum as hosts for A. marginata based on the identification of glochidia from naturally infested fishes. Our laboratory results confirmed these species as suitable hosts, indicating they are probable natural hosts for A. marginata. This finding suggests that contrary to the generalist host associations of other anodontines, A. marginata may specialize on catostomid hosts. This specialization may also be the case for other species in the subgenera Alasmidonta and Decurambis. For Table 1. Summary of recent population characteristics of Alasmidonta marginata in Minnesota. Individuals found in smaller tributaries are included in total for that drainage. Relative Freq. of individuals No. sites abundance No. at sites (%) Major drainage/river Year sampled sampled (%) live Live Live or dead St. Croix1 1987–2011 349 0.58 296 22.92 32.66 St. Croix main stem 1987–2011 222 0.54 177 22.97 36.04 Sunrise 2010–2011 16 0.40 17 18.75 31.25 Snake 1998–2009, 2011 58 0.70 59 29.31 32.76 Kettle 1998–2000, 2011 53 0.88 43 16.98 18.87 Minnesota 1989–2010 209 0.10 13 3.35 17.70 Minnesota main stem 1989–2008 123 0.02 1 0.81 20.33 Pomme de Terre 1990, 2007, 2010 63 0.15 11 7.94 15.87 Yellow Medicine 2000–2003 23 0.15 1 4.35 8.70 Upper Mississippi2 1987–2011 829 0.09 75 3.74 7.36 Mississippi main stem 2000–2011 530 0.02 11 1.89 3.77 Zumbro 1988–2002, 2010–2011 213 0.51 41 5.63 9.86 Root 1998–2003 63 1.21 19 11.11 26.98 Upper Iowa 1999 23 0.26 4 8.70 13.04 1Minnesota tributaries only. 2Mississippi River main stem and southeastern tributaries downstream from St. Anthony Falls within the MN state borders. 2013 K.H. Bloodsworth, B.R. Bosman, B.E. Sietman, and M.C. Hove 55 Figure 2. Historical and recent distribution of Alasmidonta marginata in Minnesota. Solid black circles = live occurrences (1985–2010), gray-filled circles = historic occurrences (Live and dead pre–1985 + dead post–1985), gray dots = sample sites. Capital letters represent river drainages; A = St. Croix, B = Mississippi headwaters, C = Minnesota, D = Cedar, E = Upper Mississippi. Black arrow denotes location of barrier falls; SAF = St. Anthony Falls. Voucher specimens held in the University of Minnesota and Illinois Natural History Survey mollusk collections: Mississippi River, Dakota County, JFBM14097, Wabasha County, INHS33422; St. Croix River, Chisago County, INHS36713, JFBM10999; Groundhouse River, Kanabec County, INHS 36370; Zumbro River (Middle Fork), Olmsted County, JFBM7729; Root River (North Branch), Olmsted County, JFBM14649; Turtle Creek (Cedar River Drainage), Mower County, JFBM10620; Upper Iowa River, Fillmore County, JFBM 10193; Minnesota River, Carver County, JFBM21551, Redwood County, INHS33757; Pomme de Terre River, Swift County, INHS36710. 56 Northeastern Naturalist Vol. 20, No. 1 example, Gordon and Layzer (1993) identified H. nigricans as the only suitable host for A. atropurpurea of 24 fish species tested. In other studies, however, H. nigricans was rejected as a host for A. raveneliana (Gordon and Moorman 2002), and C. commersoni was rejected as a host for A. heterodon (Michaelson and Neves 1995). Some species of Fundulidae (killifishes), Poeciliidae, and Cottidae had relatively high juvenile mussel production per fish, and using this criterion, would be candidates for future natural host studies. Fundulidae and Poeciliidae, however, are considered “universal hosts” for many unrelated mussel species (Haag and Warren 1997), and most likely would not be hosts for A. marginata in nature, due to their surface-feeding behavior and dissimilar habitat preferences (Haag and Warren 1997, Pflieger 1997). Cottidae and Gasterosteidae have universal host characteristics as well since both are suitable laboratory hosts for several distantly related mussel species (Allen et al. 2007, Barnhart and Roberts 1997, Bruenderman and Neves 1993, Rogers et al. 2001, Trdan and Hoeh 1982, van Snik Gray et al. 2002, Watters et al. 2005). Culaea inconstans (Brook Stickleback) is an unlikely natural host in our study region as their habitat is primarily restricted to small, clear, vegetated streams, or stream margins (Becker 1983). However, Cottidae have a benthic lifestyle (Pflieger 1997), which could expose them to glochidia, making them potential natural hosts for A. marginata in areas where they co-occur. Zale and Neves (1982b) documented such an occurrence by recovering A. viridis glochidia from naturally infested Cottus carolinae (Banded Sculpin) in Virginia. Additionally, Gordon and Moorman (2002) found that C. carolinae was the only suitable laboratory host for A. raveneliana of 18 species tested. Although our study found some minnows are suitable laboratory hosts for A. marginata, results were marginal with only 8 of 29 species facilitating transformation at low levels. These potential relationships should be examined further by identifying species that are hosts for A. marginata in nature. Although other species of Alasmidonta (A. varicosa, A. undulata) can transform on Centrarchidae (see Appendix 1), all 9 centrarchid species we tested, including Ambloplites rupestris, sloughed A. marginata glochidia. Therefore, Howard and Anson’s (1922) listing of A. rupestris, and likely L. gulosus, as suitable hosts is probably incorrect. Infested fish could have been caught before glochidia were sloughed, or glochidia were misidentified. This illustrates the importance of conducting laboratory host trials in conjunction with natural infestation studies, particularly if metamorphosis through the juvenile stage is not confirmed from naturally infested fishes. Proposed subgeneric groupings within Alasmidonta are based on a combination of morphological and reproductive characteristics. Clarke (1981a) placed A. marginata in the subgenus Decurambis along with A. raveneliana, A. atropurpurea, and A. varicosa (see Appendix 1). There is some overlap in reported laboratory hosts of A. atropurpurea, A. raveneliana, and A. varicosa with A. marginata; however, A. varicosa hosts include Centrarchidae and Percidae, which are more similar to those of A. undulata in the subgenus Alasmidonta. More broadly, Alasmidonta marginata differs from A. varicosa and A. undulata due to its apparent 2013 K.H. Bloodsworth, B.R. Bosman, B.E. Sietman, and M.C. Hove 57 inability to transform on centrarchids or percids. We suspect that comprehensive laboratory host studies with other members of the subgenus Decurambis would find suitable hosts similar to A. marginata. Clarke (1981a) placed A. viridis and A. heterodon in the subgenus Pressodonta, whose hosts appear to be restricted to Cottidae and Percidae (Darters) (J.P.E. Morrison reported in Clarke and Berg 1959, Michaelson and Neves 1995). The remaining species within the subgenera Alasmidonta and Decurambis have a broader range of potential hosts consisting of cyprinids, catostomids, poeciliids, cottids, centrarchids, or percids. Although comprehensive host studies are needed for more species, this observed variation in host associations could reflect divergent lineages within Alasmidonta. Distribution and status Alasmidonta marginata live in small, medium, and large sized, fast moving streams (van der Schalie 1938), a characteristic confirmed in our study (Fig. 2). The St. Croix River system currently holds the largest and most stable population of A. marginata in Minnesota, as well as several other rare mussel species (Hornbach 2001). The St. Croix is designated as a National Scenic Riverway and is among the highest quality watersheds in the upper Midwest due to its limited urban development, low nutrients, and minimal suspended sediments loads relative to other large Midwestern rivers (Fago and Hatch 1993). Although A. marginata’s abundance in Minnesota is relatively low and is a common characteristic of this species (Cummings and Mayer 1997, Strayer and Jirka 1997, Watters et al. 2009), its reduced range suggests it is sensitive to disturbance. Alasmidonta marginata has experienced its greatest decline in the Minnesota River watershed where it is in danger of extirpation due to intensive land-use-related impacts and pollution (Musser et al. 2009, Niemela and Feist 2002). Mussel populations in the Pomme de Terre River, including A. marginata, seem to be doing comparatively better than the Minnesota River watershed as a whole. Headwater tributaries such as the Chippewa, Pomme de Terre, and Lac Qui Parle, have generally experienced less biotic degradation than downstream tributaries such as the Blue Earth River (Schmidt and Proulx 2007, Sietman 2007). Sedimentation and nutrient loading has been most severe in the lower Minnesota River, most of which originates from the Blue Earth and Le Sueur watersheds (Belmont et al. 2011, Brezonik et al. 1999). The result has been an amplified loss of mussel species in the lower Minnesota River watershed (Sietman 2007). Likewise, the Upper Mississippi River main stem has been degraded from over a century of activities associated with river regulation, pollution, and more recently, Dreissena polymorpha Pallas (Zebra Mussel) introduction (Fremling 2005, Johnson and Aasen 1989), which are likely attributable to the decline of A. marginata. The distribution of A. marginata in Minnesota is limited to the St. Croix, Minnesota, and Upper Mississippi (below St. Anthony Falls) river drainages due to physical barriers rather than host distributions. St. Anthony Falls on the Mississippi River in Minneapolis, Hennepin County has been a post-glacial migration barrier to numerous mussel species since the late Pleistocene (Graf 1997). Although some mussels occur upstream of this barrier, no A. marginata 58 Northeastern Naturalist Vol. 20, No. 1 specimens have been recorded there. This finding suggests that A. marginata colonized this region of the Upper Mississippi River drainage after the barrier was established. The St. Croix River, Mississippi headwaters (above St. Anthony Falls), and the Lake Superior drainage were connected prior to the formation of St. Anthony Falls (Wright 1972). Consequently, many fish groups including suckers migrated into the Mississippi headwaters and are now present in both large and small streams throughout Minnesota (Phillips and Underhill 1971). Although it does not seem likely that the distribution of catostomids is a limiting factor, it is notable that I. bubalus, H. nigricans, and Carpiodes carpio (River Carpsucker) have ranges similar to A. marginata (K. Schmidt, MNDNR, St. Paul, MN, pers. comm.). Alasmidonta marginata was listed by the Minnesota Department of Natural Resources as a threatened species in 1996, due primarily to its reduced range. Findings from our study corroborate with this. With few reproducing populations outside the St. Croix River, we agree that A. marginata deserves its current “threatened” status in Minnesota. Conservation efforts aimed at improving and protecting aquatic habitat should focus on streams with reproducing A. marginata populations, i.e., St. Croix main stem and the Pomme de Terre, Root, and Zumbro rivers. This study provides important information on the life history and distribution of a sensitive mussel species that will aid conservation efforts. Future research should focus on recovering juvenile A. marginata from naturally infested fishes (e.g., Boyer et al. 2011). In addition, if habitat conditions improve in streams where A. marginata has been lost or populations are critically low, results from our study can be used to initiate propagation and reintroduction measures. By understanding host associations and current and historical geographical distributions, we can develop successful strategies for improving freshwater unionid conservation. Acknowledgments This study was supported in part by the Minnesota Environment and Natural Resources Trust Fund and the Federal Wildlife Conservation and Restoration Program, the University of Minnesota, and the Minnesota Department of Na tural Resources and US Fish and Wildlife Service who provided funds through Minnesota’s State Wildlife Grants Program. We thank Anne Kapuscinski (University of Minnesota) and the Minnesota Pollution Control Agency for use of laboratory space and equipment. Karen Baumann, Andrea Fritts, Ben Dickinson, Andy Edgecumbe, Traci Griffith, Angela Lager, Marta Lyons, Brendan O’Gorman, Kim Phillips, Madeline Pletta, Nissa Rudh, Andrea Stoneman, Jon Wagner, Nicole Ward, Kurtis Weber, and Cara Weggler assisted with host trials, fish and gravid mussel collection, and mussel surveys. Robert Hrabik, Brad Pobst, Michael Taylor, Konrad Schmidt, Nick Proulx, Matt Haworth, and Brett Nagle assisted with the collection and identification of fishes. Harold Wiegner, Joel Chirhart, Tom Klein, Jenny Kruckenberg, Sarah Wren, and Ann Kuitunen assisted with laboratory maintenance and fish care. We thank two anonymous reviewers for their helpful comments on the manuscript. 2013 K.H. Bloodsworth, B.R. Bosman, B.E. Sietman, and M.C. Hove 59 Literature Cited Allen, D.C., M.C. Hove, B.E. Sietman, J.M. Davis, D.E. Kelner, J.E. 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Alasmidonta Say species Fish hosts Source of information Subgenus Alasmidonta arcula Lea (Altamaha Arcmussel) Gambusia holbrooki Girard (Eastern Mosquitofish) (LT) O’Brien 2002 undulata Say (Triangle Floater) Campostoma anomalum Rafinesque (Central Stoneroller) (LT) Watters et al. 1998 Luxilus cornutus (LT) Nedeau et al. 2000, Watters et al. 1998 Notropis rubellus Agassiz (Rosyface Shiner) (LT) Watters et al. 1998 Rhinichthys obtusus (LT) Nedeau et al. 2000 Rhinichthys cataractae (LT) Nedeau et al. 2000 Semotilus corporalis Mitchill (Fallfish) (LT) Nedeau et al. 2000 Catostomus commersoni (LT) Nedeau et al. 2000 Hypentelium nigricans (LT) Watters et al. 1998 Lepomis gibbosus (LT) Nedeau et al. 2000, Watters et al. 1998 Micropterus salmoides (LT) Nedeau et al. 2000, Watters et al. 1998 Cottus cognatus (LT) Nedeau et al. 2000, Watters et al. 1998 Etheostoma flabellare (LT) Watters et al. 1998 Subgenus Decurambis atropurpurea Rafinesque (Cumberland Elktoe) Hypentelium nigricans (LT) Gordon and Layzer 1993 marginata Say (Elktoe) Catostomus commersoni (NI) Howard and Anson 1922 Hypentelium nigricans (NI) Howard and Anson 1922 Moxostoma macrolepidotum (NI) Howard and Anson 1922 Ambloplites rupestris (NI) Howard and Anson 1922 Lepomis gulosus Cuvier (Warmouth) (NI) Howard and Anson 1922 raveneliana Lea (Appalachian Elktoe) Cottus carolinae (LT) Moorman and Gordon 1993 varicosa Lamarck (Brook Floater) Notemigonus crysoleuca (LT) Nedeau et al. 2000 Rhinichthys obtusus (LT) Nedeau et al. 2000 Rhinichthys cataractae (LT) Nedeau et al. 2000 Noturus insignis Richardson (Margined Madtom) (LT) Nedeau et al. 2000 Cottus cognatus (NS) Nedeau et al. 2000 Lepomis gibbosus (LT) Nedeau et al. 2000 Perca flavescens (LT) Nedeau et al. 2000 2013 K.H. Bloodsworth, B.R. Bosman, B.E. Sietman, and M.C. Hove 65 Alasmidonta Say species Fish hosts Source of information Subgenus Pressodonta viridis Rafinesque (Slippershell Mussel) Cottus bairdii (NS) Clarke and Berg 1959 Cottus carolinae (NI) Zale and Neves 1982b Etheostoma nigrum (NS) Clarke and Berg 1959 heterodon Lea (Dwarf Wedgemussel) Cottus bairdii (LT) Michaelson and Neves 1995 Etheostoma nigrum (LT) Michaelson and Neves 1995 Etheostoma olmstedi Storer (Tessellated Darter) (LT) Michaelson and Neves 1995 Appendix 2. Fishes identified as suitable hosts for Alasmidonta marginata glochidia in the laboratory1. Recovery period is the number of days post-infection during which juvenile mussels were observed or, for trials that produced no juveniles, the number of days unt il individuals ceased to carry glochidial infec - tions. Mean no. juveniles/fish is sum of days for juveniles observed/per fish. Incomplete trials are denoted with an *, test subjects died before completion of study or study ended prematurely. Location of fish collections if other than Minnesota are: MO = Missouri, AT = aquarium trade, HR = hatchery raised, EX = exotic. Trial Water temp. No. fish infested Recovery No. juveniles Mean no. Species date range (°C) (survivors) period (d) recovered juveniles/ fish Cyprinidae Luxilus chrysocephalus Rafinesque (Striped Shiner) Oct07 15–17 5 (0) 21–25* 7* 1.4 Oct08 16–18 3 3 0 0.0 Luxilus cornutus Mitchill (Common Shiner) Oct05 18–20 6 22–37 6 1.0 Sept06 19–21 13 20 0 0.0 Oct08 24 6 16 0 0.0 Oct09 21–23 3 17 2 0.7 Luxilus zonatus Agassiz (Bleeding Shiner) (MO) Oct07 15–17 6 21–31 31 5.2 Oct08 24 10 9–11 6 0.6 Margariscus margarita Cope (Pearl Dace) Nov09 22–23 1 13–16 4 4.0 Nocomis biguttatus Kirtland (Hornyhead Chub) Sept06 19–21 2 4 0 0.0 Sept06 19–21 1 4 0 0.0 Nov09 22–23 2 13 1 0.5 66 Northeastern Naturalist Vol. 20, No. 1 Trial Water temp. No. fish infested Recovery No. juveniles Mean no. Species date range (°C) (survivors) period (d) recovered juveniles/ fish Notemigonis crysoleucas Mitchill (Golden Shiner) Oct05 18–20 2 6 0 0.0 Sept06 19–21 2 4 0 0.0 Sept06 19–21 3 4 0 0.0 Oct09 21–23 10 (8) 10–21 22 2.3 Rhinichthys cataractae Valenciennes (Longnose Dace) Oct05 18–20 1 4 0 0.0 Sept06 19–21 1 4 0 0.0 Oct08 16–18 3 3 0 0.0 Oct08 24 5 13–20 7 1.4 Oct09 21–23 6 10–17 3 0.5 Semotilus atromaculatus Mitchill (Creek Chub) Sept06 19–21 5 4 0 0.0 Sept06 19–21 1 4 0 0.0 Oct08 24 1 9–11 5 5.0 Catostomidae Carpiodes carpio Rafinesque (River Carpsucker) Oct07 15–17 1 22–35 109 109.0 Carpiodes cyprinus Lesueur (Quillback) Oct07 15–17 7 17–45 150 21.3 Oct09 21–23 8 (4) 10–21 58 7.5 Catostomus commersoni Lacepède (White Sucker) Sept06 19–21 5 23–26 4 0.8 Oct07 15–17 3 21–28 15 4.9 Oct08 16–18 2 22–39 180 91.0 Oct08 24 4 9–11 12 3.0 Oct10 23–24 6 (5) 10–31 854 147.3 Erimyzon oblongus Mitchell (Creek Chubsucker) (MO) Oct10 23–24 2 10–21 42 21.0 Hypentelium nigricans Lesueur (Northern Hogsucker) Oct07 15–17 2 (0) 17–25* 16* 8.0 Oct09 22–23 1 (0) 14–21* 43* 43.0 Oct10 23–24 6 10–31 1467 146.7 Ictiobus bubalus Rafinesque (Smallmouth Buffalo) Oct09 21–23 1 10–31 92 92.0 Moxostoma anisurum Rafinesque (Silver Redhorse) Sept06 19–21 1 26–35* 18* 18.0 Oct07 15–17 1 17–45 99 99.0 Oct09 21.5–23 2 (1) 10–28 104 104.0 2013 K.H. Bloodsworth, B.R. Bosman, B.E. Sietman, and M.C. Hove 67 Trial Water temp. No. fish infested Recovery No. juveniles Mean no. Species date range (°C) (survivors) period (d) recovered juveniles/ fish Moxostoma erythrurum Rafinesque (Golden Redhorse) Oct07 15–17 7 21–43 361 51.0 Oct10 23–24 5 10–24 547 109.4 Moxostoma macrolepidotum Lesueur (Shorthead Redhorse) Sept06 19–21 4 (3) 23–52 207 55.5 Oct07 15–17 2 (1) 25–47 358 179.2 Oct08 16–18 1 22–39 290 290.0 Oct08 24 5 9–27 61 12.2 Fundulidae Fundulus catenatus Storer (Northern Studfish) (MO) Oct07 15–17 3 (1) 25–40* 66* 32.0 Fundulus diaphanus Lesueur (Banded Killifish) Oct05 18–20 3 30–37 3 1.0 Sept06 19–21 2 19 0 0.0 Oct10 23–24 10 10–16 7 0.7 Fundulus olivaceus Storer (Blackspotted Topminnow) (MO) Oct07 15–17 3 17–45 151 50.6 Oct08 24 9 13 4 0.4 Oct10 23–24 2 10 3 1.5 Poeciliidae Gambusia affinis Baird and Girard (Western Mosquitofish) (MO) Oct08 24 10 13–18 19 1.9 Oct09 21–23 15 (13) 10–21 61 4.2 Oct10 23–24 2 10–21 29 14.5 Poecilia sphenops Valenciennes (Mexican Molly) (AT) Oct10 23–24 15 10-14 7 0.5 Xiphophorus maculatus Günther (Southern Platyfish) (AT) Oct10 23–24 12 14 1 0.1 Gasterosteidae Culaea inconstans Kirtland (Brook Stickleback) Oct10 23–24 2 17–28 13 6.5 Cottidae Cottus bairdii Girard (Mottled Sculpin) Oct10 23–24 9 11–27 49 5.4 Cottus carolinae Gill (Banded Sculpin) (MO) Oct08 16–18 1 27–39 21 21.0 Cottus cognatus Richardson (Slimy Sculpin) Oct10 24 2 (0) 10* 4* 4.0 1Fish species that did not facilitate glochidia metamorphosis (number of trials, total number of fish, range of last days to rejection): Acipenseridae (Sturgeons) - Acipenser fulvescens Rafinesque (Lake Sturgeon) (HR) (1, 7, 4); Lepisosteidae (Gars) - Lepisosteus osseus L. (Longnose Gar) (2, 5, 3); Cyprinidae (Minnows) - Carassius auratus L. (Goldfish) (EX) (1, 5, 3), Cyprinella spiloptera Cope (Spotfin Shiner) (3, 22, 3), Cyprinella venusta Girard (Blacktail 68 Northeastern Naturalist Vol. 20, No. 1 Shiner) (MO) (2, 10, 3–7), Cyprinus carpio L. (Common Carp) (EX) (2, 4, 2–3), Hybognathus nuchalis Agassiz (Mississippi Silvery Minnow) (1, 1, 7), Lythurus umbratilis Girard (Redfin Shiner) (1, 1, 4), Macrhybopsis storeriana Kirtland (Silver Chub) (2, 5, 4–16), Notropis atherinoides Rafinesque (Emerald Shiner) (2, 12, 3), Notropis blennius Girard (River Shiner) (1, 6, 4), Notropis dorsalis Agassiz (Bigmouth Shiner) (1, 12, 4), Notropis heterolepis Eigenmann and Eigenmann (Blacknose Shiner) (1, 8, 3), Notropis hudsonius Clinton (Spottail Shiner) (1, 6, 3), Notropis topeka Gilbert (Topeka Shiner) (HR) (2, 23, 3–4), Notropis volucellus Cope (Mimic Shiner) (1, 12, 3), Phenacobius mirabilis Girard (Suckermouth Minnow) (1, 9, 4), Chrosomus eos Cope (Northern Redbelly Dace) (2, 7, 5–7),Chrosomus erythrogaster Rafinesque (Southern Redbelly Dace) (1, 8, 9), Pimephales notatus Rafinesque (Bluntnose Minnow) (3, 14, 3–4), Pimephales promelas Rafinesque (Fathead Minnow) (2, 7, 3–4), Pimephales vigilax Baird and Girard (Bullhead Minnow) (1, 8, 6), Rhinichthys obtusus Hermann (Blacknose Dace) (3, 11, 3–6); Ictaluridae (Catfishes) - Ameiurus melas Rafinesque (Black Bullhead) (1, 7, 12), Ictalurus furcatus Lesueur (Blue Catfish) (HR) (1, 5, 17), Ictalurus punctatus Rafinesque (Channel Catfish) (3, 17, 2–9), Noturus exilis Nelson (Slender Madtom) (1, 3, 4), Noturus flavus Rafinesque (Stonecat) (2, 3, 4), Noturus gyrinus Mitchill (Tadpole Madtom) (2, 3, 3–9), Pylodictus olivaris Rafinesque (Flathead Catfish) (2, 4, 3); Esocidae (Pikes) - Esox lucius L. (Northern Pike) (1, 1, 17), Esox masquinongy Mitchill (Muskellunge) (1, 1, 3); Umbridae (Mudminnows) - Umbra limi Kirtland (Central Mudminnow) (1,1,20); Salmonidae (Trouts) - Salvelinus fontinalis Mitchell (Brook Trout) (1, 1, 3); Aphredoderidae (Pirate Perches) - Aphredoderus sayanus Gilliams (Pirate Perch) (MO) (1, 3, 9); Poeciliidae (Live Bearers) - Poecilia reticulata Peters (Guppy) (AT) (1, 5, 20); Moronidae (Temperate Basses) - Morone chrysops Rafinesque (White Bass) (2, 9, 7–23); Centrarchidae (Sunfishes) - Ambloplites rupestris Rafinesque (Rock Bass) (6, 30, 3–20), Lepomis cyanellus Rafinesque (Green Sunfish) (4, 23, 4–14), Lepomis gibbosus L. (Pumpkinseed) (5, 12, 3–14), Lepomis humilis Girard (Orangespotted Sunfish) (4, 14, 3–14), Lepomis macrochirus Rafinesque (Bluegill) (4, 34, 3–14), Micropterus dolomieui Lacepède (Smallmouth Bass) (4, 22, 5–12), Micropterus salmoides Lacepède (Largemouth Bass) (4, 20, 3–11), Pomoxis nigromaculatus Lesueur (Black Crappie) (2, 15, 14–17); Percidae (Perches) - Etheostoma caeruleum Storer (Rainbow Darter) (1, 14, 11), Etheostoma flabellare Rafinesque (Fantail Darter) (1, 6, 2), Etheostoma nigrum Rafinesque (Johnny Darter) (2, 5, 3–8), Etheostoma zonale Cope (Banded Darter) (1, 3, 3), Perca flavescens Mitchill (Yellow Perch) (2, 37, 9–20), Percina caprodes Rafinesque (Logperch) (3, 30, 3–9), Percina evides Jordan and Copeland (Gilt Darter) (1, 7, 4), Percina maculata Girard (Blackside Darter) (1, 6, 3), Percina phoxocephala Nelson (Slenderhead Darter) (1, 2, 4), Percina shumardi Girard (River Darter) (2, 8, 3–6), Sander canadensis Griffith and Smith (Sauger) (2, 20, 3–9), Sander vitreus Mitchill (Walleye) (2, 4, 13–24); Sciaenidae (Drums) - Aplodinotus grunniens Rafinesque (Freshwater Drum) (2, 2, 3–14); Proteidae (Mudpuppies) - Necturus maculosus Rafinesque (Common Mudpuppy) (1, 10, 3).