Zequanox® is a commercial formulation of the killed bacterium, Pseudomonas fluorescens (strain CL145A) that has USEPA approval for use in open water to kill dreissenid mussels. Previous nontarget studies demonstrated the safety and selectivity of the product, but the database is limited for macroinvertebrate taxa and exposure conditions. We evaluated the safety of the product to two high value macroinvertebrates, the amphipod Gammarus lacustris lacutris, and nymphs of the burrowing mayfly, Hexagenia spp., at the maximum approved concentration (100 mg/L A.I.) and exposure duration (8 h). Survival of animals was measured at the end of 8 h exposure and at 24 h post-exposure and 96 h post-exposure. Additionally, histological changes in the digestive tract of both species were compared with controls at 96 h post-exposure. The results showed no effect of Zequanox on survival of either species. Survival of G. lacustris exceeded 85% in all concentrations at all three sampling time points. Survival of Hexagenia spp. ranged from 71% (control) to 91% (50 mg/L) at 8 h, 89-93% at 24 h post-exposure and 70-73% at 96 post-exposure h across all treatments. We also saw no evidence of pathology in the visceral organs of treated animals. Our results indicate that application of Zequanox at the maximum approved concentration and exposure duration does not cause acute mortality or pathology to G. lacustris and Hexagenia spp.

The efficacy and subsurface application of Zequanox®, a commercially prepared spray-dried powder formulation of Pseudomonas fluorescens (strain CL145A), were evaluated for controlling zebra mussels (Dreissena polymorpha) within 27-m2 enclosures in Lake Minnetonka (Deephaven, Minnesota). Five treatments consisting of (1) two whole water column Zequanox applications, (2) two subsurface Zequanox applications, and (3) an untreated control were completed on each of three independent treatment days during September 2014. The two types of samplers used in the study were (1) custom built multi-plate samplers (type 1 samplers), with wood, perforated aluminum, and tile substrates that were placed into Robinson’s Bay in June of 2013 to allow for natural colonization by zebra mussels, and (2) samplers that were designed to contain zebra mussels (type 2 samplers) which consisted of zebra mussels adhering to perforated aluminum trays that were placed into mesh containment bags. One day prior to treatment, three individual samplers of each type were distributed to test enclosures and exposed to a randomly assigned treatment. Sampling to determine the living zebra mussel biomass adhering to type 1 samplers and the survival assessments for zebra mussels contained in type 2 samplers were completed ~40 days after exposure. The living zebra mussel biomass adhering to type 1 samplers and the survival of zebra mussels contained in type 2 samplers were significantly less in groups treated with the highest Zequanox concentrations and in groups that received whole water column applications compared to groups treated with lower Zequanox concentrations and subsurface applications. However, standardization biomass and survival results to the amount of Zequanox applied showed that the lower Zequanox concentrations and subsurface applications were more efficient at reducing zebra mussel biomass and inducing zebra mussel mortality. Although more efficient, biological significance and management goals should be evaluated prior to selecting subsurface application methods and lower treatment concentrations for Zequanox applications. Development and refinement of additional application techniques may improve the utility of the subsurface Zequanox applications.

The efficacy and subsurface application of Zequanox®, a commercially prepared spray-dried powder formulation of Pseudomonas fluorescens (strain CL145A), were evaluated for controlling zebra mussels (Dreissena polymorpha) within 27-m2 enclosures in Lake Minnetonka (Deephaven, Minnesota). Five treatments consisting of (1) two whole water column Zequanox applications, (2) two subsurface Zequanox applications, and (3) an untreated control were completed on each of three independent treatment days during September 2014. The two types of samplers used in the study were (1) custom built multi-plate samplers (type 1 samplers), with wood, perforated aluminum, and tile substrates that were placed into Robinson’s Bay in June of 2013 to allow for natural colonization by zebra mussels, and (2) samplers that were designed to contain zebra mussels (type 2 samplers) which consisted of zebra mussels adhering to perforated aluminum trays that were placed into mesh containment bags. One day prior to treatment, three individual samplers of each type were distributed to test enclosures and exposed to a randomly assigned treatment. Sampling to determine the living zebra mussel biomass adhering to type 1 samplers and the survival assessments for zebra mussels contained in type 2 samplers were completed ~40 days after exposure. The living zebra mussel biomass adhering to type 1 samplers and the survival of zebra mussels contained in type 2 samplers were significantly less in groups treated with the highest Zequanox concentrations and in groups that received whole water column applications compared to groups treated with lower Zequanox concentrations and subsurface applications. However, standardization biomass and survival results to the amount of Zequanox applied showed that the lower Zequanox concentrations and subsurface applications were more efficient at reducing zebra mussel biomass and inducing zebra mussel mortality. Although more efficient, biological significance and management goals should be evaluated prior to selecting subsurface application methods and lower treatment concentrations for Zequanox applications. Development and refinement of additional application techniques may improve the utility of the subsurface Zequanox applications.

Histopathological assessments of young-of-the-year smallmouth bass (YOY SMB) in the Susquehanna River drainage identified a high prevalence of the myxozoan Myxobolus inornatus. This myxozoan infects the connective tissue of the muscle below the skin but is sometimes observed in the esophagus and buccal cavity. In some instances, shallow infections cause breaks in the skin which could increase the chance of opportunistic bacterial infections. Several microbial pathogens including Flavobacterium columnare, Aeromonas spp. and largemouth bass virus (LMBV) have also been cultured from clinically diseased YOY. A multiplex fluorescence in situ hybridization (FISH) assay was developed to determine potential co-localization of M. inornatus, Flavobacterium spp., and Aeromonas spp. infections. With FISH, 75% of YOY SMB exhibited M. inornatus infections, 10% had Aeromonas spp. infections, and 5% had Flavobacterium spp. infections, while 3% had coinfections with both bacterial species and M. inornatus. The results of the multiplex FISH assay revealed a low occurrence of coinfections of Flavobacterium spp. and/or Aeromonas spp. with M. inornatus in randomly sampled individuals.

The survival of Pseudogymnoascus destructans (Pd) was evaluated at temperatures outside of its thermal range of growth on three different artificial growth media; Sabouraud dextrose agar (SD), brain-heart infusion agar (BHI), and brain-heart infusion agar supplemented with 10% sheep red blood cells (BHI+B). Pd was harvested from starting cultures grown of MEA agar at 7˚C for 60 days. Harvested conidia were diluted in Phosphate Buffered Saline + Tween20 and spread onto plates of a given medium. Plate were then incubated at either 24, 30 or 37˚C. Plates were incubated for 1, 5, 9, 15, 30, 60, 90, 120, or 150 days before being transferred to a 7˚C incubator for 50 days. Colony forming units (CFUs) of Pd were then enumerated, resulting in a time series of Pd survival on a given medium at a given temperature. As each medium was inoculated from a different starting culture of Pd, a control group for each medium was created by inoculating plates as above and then immediate incubation at 7˚C for 50 days. The number of CFUs on the control plates was used as a statistical offset factor which allowed for the fair comparison of Pd survival between different media. The number of conidia initially plated onto each plate varied between 100, 1000 , and 10,000, depending on the temperature-medium treatment combination. In order to ensure robust statistical analysis, all data was rescaled by an appropriate correction factor which is also presented in the datafile.

This data represents the number of positive and negative Pd (Pseudogymnoascus destructans) detections by county over the sampling period 2008-2012. Pd is the fungus that is the causative agent of white-nose syndrome.

Freshwater mussels of the order Unionida are among the most endangered animal groups globally, but the causes of population declines are often enigmatic with little known about the role of disease. In 2018, we collected wild adult pheasantshell (Actinonaias pectorosa) and mucket (Actinonaias ligamentina) during an epidemiologic survey investigating an ongoing mussel mass mortality event in the Clinch River, USA. Histopathology and transmission electron microscopy showed a novel microsporidian parasite primarily infecting the ovary of pheasantshell. Sequencing of the small subunit rRNA gene produced a 1333 bp sequence with greatest similarity to Pseudonosema cristatellae (AF484694.1; 86.36%; e-value = 0), a microsporidium infecting the freshwater bryozoan (Cristatella mucedo). Microsporidia were observed in 65% (17/26) of the examined female pheasantshell (A. pectorosa) and in no (0/2) female mucket (A. ligamentina), and occurred at mortality and non-mortality sites. Our findings indicate that a novel parasite, Microsporidium clinchi n. sp., is present in pheasantshell in the Clinch River, USA, and while likely not a cause of mass mortality, could reduce fecundity and recruitment in this declining population and threaten the success of reintroductions. Surveillance for M. clinchi n. sp. and evaluation of brood stock and their progeny for microsporidia would therefore be prudent.

Freshwater mussels of the order Unionida are among the most endangered animal groups globally, but the causes of population declines are often enigmatic with little known about the role of disease. In 2018, we collected wild adult pheasantshell (Actinonaias pectorosa) and mucket (Actinonaias ligamentina) during an epidemiologic survey investigating an ongoing mussel mass mortality event in the Clinch River, USA. Histopathology and transmission electron microscopy showed a novel microsporidian parasite primarily infecting the ovary of pheasantshell. Sequencing of the small subunit rRNA gene produced a 1333 bp sequence with greatest similarity to Pseudonosema cristatellae (AF484694.1; 86.36%; e-value = 0), a microsporidium infecting the freshwater bryozoan (Cristatella mucedo). Microsporidia were observed in 65% (17/26) of the examined female pheasantshell (A. pectorosa) and in no (0/2) female mucket (A. ligamentina), and occurred at mortality and non-mortality sites. Our findings indicate that a novel parasite, Microsporidium clinchi n. sp., is present in pheasantshell in the Clinch River, USA, and while likely not a cause of mass mortality, could reduce fecundity and recruitment in this declining population and threaten the success of reintroductions. Surveillance for M. clinchi n. sp. and evaluation of brood stock and their progeny for microsporidia would therefore be prudent.

This dataset includes data used to summarize trends and identify best-fit models to explain patterns in presence-absence and abundance of Pseudogymnoascus destructans (Pd) in environmental substrates and on bats within six bat hibernacula at different stages of white-nose syndrome (WNS). Data relating to environmental substrates include: dates and relative spatial locations of samples collected within study hibernacula, presence and quantity of Pd in samples based on qPCR analysis, and daily temperature parameters at each sample location on the days samples were collected. Data relating to bats include: dates and relative spatial locations of hibernating bats that were sampled, species, sex, weight(g), forearm length(mm), body mass index (weight/forearm), proportion of the wing with visible fungus or fluorescence characteristic of WNS under hand-held UVA light and presence and quantity of Pd in wing-skin swab samples based on qPCR analysis. Measures of time since first detection of WNS at each study hibernaculum are also included in the dataset.

This dataset includes data used to summarize trends and identify best-fit models to explain patterns in presence-absence and abundance of Pseudogymnoascus destructans (Pd) in environmental substrates and on bats within six bat hibernacula at different stages of white-nose syndrome (WNS). Data relating to environmental substrates include: dates and relative spatial locations of samples collected within study hibernacula, presence and quantity of Pd in samples based on qPCR analysis, and daily temperature parameters at each sample location on the days samples were collected. Data relating to bats include: dates and relative spatial locations of hibernating bats that were sampled, species, sex, weight(g), forearm length(mm), body mass index (weight/forearm), proportion of the wing with visible fungus or fluorescence characteristic of WNS under hand-held UVA light and presence and quantity of Pd in wing-skin swab samples based on qPCR analysis. Measures of time since first detection of WNS at each study hibernaculum are also included in the dataset.