Freshwater Mussel Viromes Increase Rapidly in Diversity and Abundance When Hosts Are Released from Captivity into the Wild.

Freshwater mussels (order: Unionida) are highly imperiled globally and are increasingly the focus of captive propagation efforts to protect and restore wild populations. The Upper Tennessee River Basin (UTRB) in Virginia is a freshwater biodiversity hotspot hosting at least 45 of North America's ~300 species of freshwater mussels, including 21 threatened and endangered species listed under the U.S. Endangered Species Act. Recent studies have documented that viruses and other microbes have contributed to freshwater mussel population declines in the UTRB. We conducted a multi-year longitudinal study of captive-reared hatchery mussels released to restoration sites throughout the UTRB to evaluate their viromes and compare them to captive hatchery environments. We documented 681 viruses from 27 families. The hatchery mussels had significantly less viruses than those deployed to wild sites, with only 20 viruses unique to the hatchery mussels. After the hatchery mussels were released into the wild, their number of viruses initially spiked and then increased steadily over time, with 451 viruses in total unique to the mussels in the wild. We found Clinch densovirus 1 (CDNV-1), a virus previously associated with mass mortality events in the Clinch River, in all samples, but the wild site mussels consistently had significantly higher CDNV-1 levels than those held in the hatchery. Our data document substantial differences between the viruses in the mussels in the hatchery and wild environments and rapid virome shifts after the mussels are released to the wild sites. These findings indicate that mussel release programs might benefit from acclimatization periods or other measures to mitigate the potential negative effects of rapid exposure to infectious agents found in natural environments.

Octomacrum spinum (Monogenoidea: Polyopisthocotylea: Octomacridae) from highland stoneroller, Campostoma spidaceum (Cypriniformes: Leuciscidae) from Arkansas and Oklahoma, USA, with a summary of the genus from North American and European fishes.

Octomacrum spinum Dansby & Shoemaker, 1973, is reported for the first time from 23 of 146 (16%) highland stonerollers, Campostoma spadiceum collected from central and western Arkansas and southeastern Oklahoma, USA. Analysis of molecular data revealed that the SSU rRNA gene of specimens of O. spinum from Arkansas was identical to a number of Octomacrum spp. and the LSU rRNA gene was 97.46% similar to Octomacrum europaeum. We also provide a summary of the species of Octomacrum reported from North American and European fishes.

A NOVEL SPECIES OF MYXOBOLUS (CNIDARIA: MYXOSPOREA: MYXOBOLIDAE) FROM THE GILL OF GREEN SUNFISH, LEPOMIS CYANELLUS (PERCIFORMES: CENTRARCHIDAE), FROM THE BLACK RIVER OF NORTHEASTERN ARKANSAS.

During March 2023, 7 green sunfish (Lepomis cyanellus) and 2 bluegill (Lepomis macrochirus) were collected from the Black River (White River drainage) in Lawrence County, Arkansas. In addition, during March 2023 and again in May-June 2023, 13 L. cyanellus and 6 L. macrochirus were taken from Butcherknife and Big Fork creeks (Ouachita River drainage), Polk County, Arkansas, 9 L. cyanellus were collected from the Caddo River, Montgomery County, Arkansas, and 5 green sunfish were taken from Clear Creek at Savoy, Washington County, Arkansas. All fish had their gill, gallbladder, fins, integument, musculature, and other major organs examined for myxozoans. The gill of 1 of 34 (3%) L. cyanellus was infected with a new myxozoan, Myxobolus fergusoni n. sp. Qualitative and quantitative morphological data were obtained from fresh myxospores, and molecular data consisted of a 1,933-base-pair sequence of the partial small subunit (SSU) ribosomal RNA (rRNA) gene. Phylogenetic analysis grouped M. fergusoni n. sp. with other centrarchid-infecting myxobolids from North America and placed this cluster in a larger clade comprising myxozoans that infect North American and European esocids, a North American aphredoderid, European percids, and a gasterosteid from Japan. Myxobolus fergusoni n. sp. infects the gill arches of L. cyanellus, similar to Myxobolus cartilaginis (Hoffman, Putz, and Dunbar, 1965), which was described from head cartilage, gill arches, and large fin rays of L. cyanellus. Another is Myxobolus mesentericusKudo, 1920, which was described from the viscera of green sunfish. A large polysporic plasmodium filled with myxospores was present in a basifilamental location associated with multiple gill filaments at their junction with the gill arch. The intact plasmodium replaced connective tissue within the arch but elicited only mild proliferation of overlying epithelium and a minimal host inflammatory response. This is the third time a myxozoan has been described from L. cyanellus, as well as being the first time it has been described from an Arkansas specimen.

Novel Aquareovirus isolated from channel catfish (Ictalurus punctatus) used in mussel restoration efforts in Wisconsin.

Channel catfish (Ictalurus punctatus) are a food fish extensively reared in aquaculture facilities throughout the world and are also among the most abundant wild catfish species in North America, making them a popular target of anglers. Furthermore, channel catfish are important members of aquatic ecosystems; for example, they serve as a glochidial host for the endangered winged mapleleaf mussel (Quadrula fragosa), making them critical for conserving this species through hatchery-based restoration efforts. During a routine health inspection, a novel aquareovirus was isolated from channel catfish used in mussel propagation efforts at a fish hatchery in Wisconsin. This virus was isolated on brown bullhead cells (ATCC CCL-59) and identified through metagenomic sequencing as a novel member of the family Spinareoviridae, genus Aquareovirus. The virus genome consists of 11 segments, as is typical of the aquareoviruses, with phylogenetic relationships based on RNA-dependent RNA polymerase and major outer capsid protein amino acid sequences showing it to be most closely related to golden shiner virus (aquareovirus C) and aquareovirus C/American grass carp reovirus (aquareovirus G) respectively. The potential of the new virus, which we name genictpun virus 1 (GNIPV-1), to cause disease in channel catfish or other species remains unknown.

A NEW SPECIES OF THELOHANELLUS (CNIDARIA: MYXOSPOREA: MYXOBOLIDAE) FROM THE GILL OF QUILLBACK, CARPIODES CYPRINUS (CYPRINIFORMES: CATOSTOMIDAE), FROM THE ARKANSAS RIVER DRAINAGE OF OKLAHOMA.

During May 2022 and again in March 2023, 5 quillbacks, Carpiodes cyprinus, were collected from the Verdigris River, Wagoner County, Oklahoma (n = 1), and the Black River, Lawrence County, Arkansas (n = 4), and their gill, gallbladder, fins, integument, musculature, and other major organs were macroscopically examined for myxozoans. Gill lamellae from the single quillback from the Verdigris River was infected with a new myxozoan, Thelohanellus oklahomaensis n. sp. Qualitative and quantitative morphological data were obtained from fresh and formalin-fixed preserved myxospores, and molecular data consisted of a 1,767 base pair sequence of the partial small subunit (SSU) ribosomal RNA gene. Phylogenetic analysis grouped T. oklahomaensis n. sp. with myxozoans known to infect North American catostomids and Eurasian cyprinids. Histological examination localized plasmodia to an intralamellar developmental site and revealed a possible vestige of a second polar capsule. Although plasmodia markedly expanded lamellae, there were no associated epithelial or inflammatory changes. Thelohanellus oklahomaensis n. sp. is the only member of the genus known to infect the gills of C. cyprinus.

Five Species of Wild Freshwater Sport Fish in Wisconsin, USA, Reveal Highly Diverse Viromes.

Studies of marine fish have revealed distant relatives of viruses important to global fish and animal health, but few such studies exist for freshwater fish. To investigate whether freshwater fish also host such viruses, we characterized the viromes of five wild species of freshwater fish in Wisconsin, USA: bluegill (Lepomis macrochirus), brown trout (Salmo trutta), lake sturgeon (Acipenser fulvescens), northern pike (Esox lucius), and walleye (Sander vitreus). We analyzed 103 blood serum samples collected during a state-wide survey from 2016 to 2020 and used a metagenomic approach for virus detection to identify known and previously uncharacterized virus sequences. We then characterized viruses phylogenetically and quantified prevalence, richness, and relative abundance for each virus. Within these viromes, we identified 19 viruses from 11 viral families: Amnoonviridae, Circoviridae, Coronaviridae, Hepadnaviridae, Peribunyaviridae, Picobirnaviridae, Picornaviridae, Matonaviridae, Narnaviridae, Nudnaviridae, and Spinareoviridae, 17 of which were previously undescribed. Among these viruses was the first fish-associated coronavirus from the Gammacoronavirus genus, which was present in 11/15 (73%) of S. vitreus. These results demonstrate that, similar to marine fish, freshwater fish also harbor diverse relatives of viruses important to the health of fish and other animals, although it currently remains unknown what effect, if any, the viruses we identified may have on fish health.

Mussel mass mortality in the Clinch River, USA: metabolomics detects affected pathways and biomarkers of stress.

Biologists monitoring freshwater mussel (order Unionida) populations rely on behavioral, often subjective, signs to identify moribund ("sick") or stressed mussels, such as gaping valves and slow response to probing, and they lack clinical indicators to support a diagnosis. As part of a multi-year study to investigate causes of reoccurring mortality of pheasantshell (Ortmanniana pectorosa; synonym Actinonaias pectorosa) in the Clinch River, Virginia and Tennessee, USA, we analyzed the hemolymph metabolome of a subset of mussels from the 2018 sampling period. Mussels at the mortality sites were diagnosed in the field as affected (case) or unaffected (control) based on behavioral and physical signs. Hemolymph was collected in the field by non-lethal methods from the anterior adductor muscle for analysis. We used ultra-high-performance liquid chromatography with quadrupole time-of-flight mass spectroscopy to detect targeted and untargeted metabolites in hemolymph and compared metabolomic profiles by field assessment of clinical status. Targeted biomarker analysis found 13 metabolites associated with field assessments of clinical status. Of these, increased gamma-linolenic acid and N-methyl-l-alanine were most indicative of case mussels, while adenine and inosine were the best indicators of control mussels. Five pathways in the targeted analysis differed by clinical status; two of these, purine metabolism and glycerophospholipid metabolism, were also indicated in the untargeted analysis. In the untargeted nalysis, 22 metabolic pathways were associated with clinical status. Many of the impacted pathways in the case group were catabolic processes, such as degradation of amino acids and fatty acids. Hierarchical clustering analysis matched clinical status in 72% (18 of 25) of mussels, with control mussels more frequently (5 of 16) not matching clinical status. Our study demonstrated that metabolomic analysis of hemolymph is suitable for assessing mussel condition and complements field-based indicators of health.

Hypersensitive response to interferon-stimulated gene (ISG)-inducing double-stranded RNA in American bullfrog tadpole fibroblasts.

American bullfrogs are thought to be carriers of ranaviruses and contribute to their global spread via trade. Bullfrog tadpoles succumb to ranaviral infection's more severe and deadly effects than bullfrog adults. Presently, little is known about bullfrog tadpoles' innate antiviral immunity, possible due to the lack of available bullfrog tadpole cell lines. In this study, we describe a novel bullfrog tadpole fibroblast cell line named BullTad-leg. Its general cellular attributes, gene expression and function of class-A scavenger receptors (SR-As), and responses to poly IC (a synthetic dsRNA mimicking viral dsRNAs and a potent inducer of the interferon (IFN)-mediated antiviral responses) are investigated. Its abundant expression of vimentin corroborated with the cells' fibroblast morphology. BullTad-leg cells expressed transcripts of four SR-A members: SR-AI, SCARA3, SCARA4, and SCARA5, but transcripts of MARCO, the fifth SR-A member, were not detected. BullTad-leg cells expressed functional SR-As and could bind AcLDL. BullTad-leg cells exhibited cytotoxicity in response to poly IC treatment via SR-As. Additionally, very low doses of poly IC were able to induce dose-dependent expressions of ISGs including Mx, PKR, ISG20, and IFI35. This research sheds new light on the innate immune response, particularly SR-A biology and dsRNA responsiveness, in bullfrog tadpoles.

Viruses of Freshwater Mussels during Mass Mortality Events in Oregon and Washington, USA.

Freshwater mussels (Unionida) are globally imperiled, in part due to largely unexplained mass mortality events (MMEs). While recent studies have begun to investigate the possibility that mussel MMEs in the Eastern USA may be caused by infectious diseases, mussels in the Western USA have received relatively little attention in this regard. We conducted a two-year epidemiologic investigation of the role of viruses in ongoing MMEs of the Western pearlshell (Margaritifera falcata) and the Western ridged mussel (Gonidea angulata) in the Chehalis River and Columbia River watersheds in the Western USA. We characterized viromes of mussel hemolymph from 5 locations in 2018 and 2020 using metagenomic methods and identified 557 viruses based on assembled contiguous sequences, most of which are novel. We also characterized the distribution and diversity of a previously identified mussel Gammarhabdovirus related to pathogenic finfish viruses. Overall, we found few consistent associations between viruses and mussel health status. Variation in mussel viromes was most strongly driven by location, with little influence from date, species, or health status, though these variables together only explained ~1/3 of variation in virome composition. Our results demonstrate that Western freshwater mussels host remarkably diverse viromes, but no single virus or combination of viruses appears to be associated with morbidity or mortality during MMEs. Our findings have implications for the conservation of imperiled freshwater mussels, including efforts to enhance natural populations through captive propagation.

Pathology and infectious agents of unionid mussels: A primer for pathologists in disease surveillance and investigation of mortality events.

Freshwater mussels are one of the most imperiled groups of organisms in the world, and more than 30 species have gone extinct in the last century. While habitat alteration and destruction have contributed to the declines, the role of disease in mortality events is unclear. In an effort to involve veterinary pathologists in disease surveillance and the investigation of freshwater mussel mortality events, we provide information on the conservation status of unionids, sample collection and processing techniques, and unique and confounding anatomical and physiological differences. We review the published accounts of pathology and infectious agents described in freshwater mussels including neoplasms, viruses, bacteria, fungi, fungal-like agents, ciliated protists, Aspidogastrea, Digenea, Nematoda, Acari, Diptera, and Odonata. Of the identified infectious agents, a single viral disease, Hyriopsis cumingii plague disease, that occurs only in cultured mussels is known to cause high mortality. Parasites including ciliates, trematodes, nematodes, mites, and insects may decrease host fitness, but are not known to cause mortality. Many of the published reports identify infectious agents at the light or ultrastructural microscopy level with no lesion or molecular characterization. Although metagenomic analyses provide sequence information for infectious agents, studies often fail to link the agents to tissue changes at the light or ultrastructural level or confirm their role in disease. Pathologists can bridge this gap between identification of infectious agents and confirmation of disease, participate in disease surveillance to ensure successful propagation programs necessary to restore decimated populations, and investigate mussel mortality events to document pathology and identify causality.

Plasticity, Paralogy, and Pseudogenization: Rhabdoviruses of Freshwater Mussels Elucidate Mechanisms of Viral Genome Diversification and the Evolution of the Finfish-Infecting Rhabdoviral Genera.

Viruses in the family Rhabdoviridae display remarkable genomic variation and ecological diversity. This plasticity occurs despite the fact that, as negative sense RNA viruses, rhabdoviruses rarely if ever recombine. Here, we describe nonrecombinatorial evolutionary processes leading to genomic diversification in the Rhabdoviridae inferred from two novel rhabdoviruses of freshwater mussels (Mollusca: Bivalvia: Unionida). Killamcar virus 1 (KILLV-1) from a plain pocketbook (Lampsilis cardium) is closely related phylogenetically and transcriptionally to finfish-infecting viruses in the subfamily Alpharhabdovirinae. KILLV-1 offers a novel example of glycoprotein gene duplication, differing from previous examples in that the paralogs overlap. Evolutionary analyses reveal a clear pattern of relaxed selection due to subfunctionalization in rhabdoviral glycoprotein paralogs, which has not previously been described in RNA viruses. Chemarfal virus 1 (CHMFV-1) from a western pearlshell (Margaritifera falcata) is closely related phylogenetically and transcriptionally to viruses in the genus Novirhabdovirus, the sole recognized genus in the subfamily Gammarhabdovirinae, representing the first known gammarhabdovirus of a host other than finfish. The CHMFV-1 G-L noncoding region contains a nontranscribed remnant gene of precisely the same length as the NV gene of most novirhabdoviruses, offering a compelling example of pseudogenization. The unique reproductive strategy of freshwater mussels involves an obligate parasitic stage in which larvae encyst in the tissues of finfish, offering a plausible ecological mechanism for viral host-switching. IMPORTANCE Viruses in the family Rhabdoviridae infect a variety of hosts, including vertebrates, invertebrates, plants and fungi, with important consequences for health and agriculture. This study describes two newly discovered viruses of freshwater mussels from the United States. One virus from a plain pocketbook (Lampsilis cardium) is closely related to fish-infecting viruses in the subfamily Alpharhabdovirinae. The other virus from a western pearlshell (Margaritifera falcata) is closely related to viruses in the subfamily Gammarhabdovirinae, which until now were only known to infect finfish. Genome features of both viruses provide new evidence of how rhabdoviruses evolved their extraordinary variability. Freshwater mussel larvae attach to fish and feed on tissues and blood, which may explain how rhabdoviruses originally jumped between mussels and fish. The significance of this research is that it improves our understanding of rhabdovirus ecology and evolution, shedding new light on these important viruses and the diseases they cause.

A Bacteriological Comparison of the Hemolymph from Healthy and Moribund Unionid Mussel Populations in the Upper Midwestern U.S.A. Prompts the Development of Diagnostic Assays to Detect Yokenella regensburgei.

Recent bacteriological investigations of freshwater mussel mortality events in the southeastern United States have identified a variety of bacteria and differences in bacterial communities between sick and healthy mussels. In particular, Yokenella regensburgei and Aeromonas spp. have been shown to be associated with moribund mussels, although it remains unclear whether these bacteria are causes or consequences of disease. To further understand the role of bacteria in mussel epizootics, we investigated mortality events that occurred in the upper Midwest in the Embarrass River (Wisconsin) and the Huron River (Michigan). For comparison, we also studied mussels from an unaffected population in the St. Croix River (Wisconsin). Diverse bacterial genera were identified from these sites, including Y. regensburgei from moribund mussels in the Embarrass River (Wisconsin). This bacterium has also been consistently isolated during ongoing mortality events in the Clinch River (Virginia). Subsequently, we developed and validated molecular assays for the detection of Yokenella to use in future investigations of mussel mortality events and to identify environmental reservoirs of this bacterium.

Efficacy of hydrogen peroxide to reduce Gyrodactylus species infestation density on four fish species.

OBJECTIVE: The ability to effectively treat parasitic infestations of fish is of high importance for fish culture facilities. However, tools or approved therapies for treating infestations on fish are limited. This paper summarizes results from four separate clinical field studies that evaluated the efficacy of hydrogen peroxide (H2 O2 ; 35% PEROX-AID) for reducing Gyrodactylus spp. infestation density. METHODS: Three species of Gyrodactylus were studied (G. salmonis, hosts: Brook Trout Salvelinus fontinalis and Lake Trout S. namaycush; G. freemani, host: Yellow Perch Perca flavescens; G. hoffmani, host: Fathead Minnow Pimephales promelas) before and after the application of immersion H2 O2 therapy. RESULT: Parasite density was significantly reduced for each parasite × host combination to which H2 O2 therapy was applied. Two clinical field studies in salmonids were found to demonstrate substantial effectiveness that enabled 35% PEROX-AID approval. CONCLUSION: Further assessments of Gyrodactylus spp. could expand the use of H2 O2 for controlling these parasites in aquaculture. Specifically, H2 O2 was effective at all levels tested (50 or 75 mg H2 O2 /L for 60 min for the Yellow Perch and Fathead Minnow clinical field studies; 100 or 150 mg H2 O2 /L for 30 min regardless of salt pre-treatment for the Brook Trout study; and 100 mg H2 O2 /L for 30 min or 50 mg H2 O2 /L for 60 min for the Lake Trout study).

A new Myxobolus (Cnidaria: Myxosporea: Myxobolidae) from the gills of the southern striped shiner, Luxilus chrysocephalus isolepis (Cypriniformes: Leuciscidae), from southwestern Arkansas, USA.

The southern striped shiner, Luxilus chrysocephalus isolepis (Hubbs & Brown) is a relatively large minnow belonging to the true minnow family Leuciscidae Bonaparte. Between May 2020 and January 2022, 55 L. c. isolepis were collected from watersheds in Montgomery (n = 6), Polk (n = 17) and Sevier (n = 32) counties, Arkansas, USA, and their gills, gallbladders, urinary bladders, fins, integument, other major organs, and musculature were examined for myxozoans. Gills of 11 (34%) individual southern striped shiners from Sevier County were infected with a new myxozoan, Myxobolus carlhubbsi n. sp. A qualitative and quantitative morphological description was based on formalin-fixed preserved myxospores, and molecular data consisted of a 1,970 base pair sequence of the partial small subunit rRNA gene from ethanol-preserved specimens. Histologically, plasmodia filled and expanded interlamellar troughs. Hyperplastic epithelial and goblet cells filled interlamellar troughs adjacent to plasmodia, but inflammatory response was limited to scattered lymphocytes. Phylogenetic analysis revealed that M. carlhubbsi n. sp. is a member of a clade of species with pyriform myxospores parasitizing North American Pogonichthyinae Girard and North American and Eurasian Leuciscinae Bonaparte. This is the first report of a myxozoan from L. c. isolepis. This article was registered in the Official Register of Zoological Nomenclature (ZooBank) as urn:lsid:zoobank.org:pub:D10D71C2-2C75-4A1C-80ED-B98FF36CB509.

Pseudomurraytrema fergusoni n. sp. (Monogenea: Dactylogyridae) from gills of Pealip Redhorse, Moxostoma pisolabrum (Catostomidae) from Arkansas, USA, with a description of gill pathology.

Pseudomurraytrema fergusoni n. sp. is described from the Pealip Redhorse, Moxostoma pisolabrum from the Black River (White River drainage), Lawrence County, Arkansas, USA. This represents the second monogenean described from M. pisolabrum as well as the second species of Pseudomurraytrema reported from an Arkansas catostomid. The description includes partial 18S rRNA and 28S rRNA gene sequences (732 bp and 851 bp, respectively), helping fill a void in sequence data from North American monogeneans, particularly those in the genus Pseudomurraytrema. In addition, histopathologic changes associated with the infection resulted in severe localized pathologic lesions in gills of the host, suggesting compromise of respiratory surfaces within affected areas adjacent to the worms.

Freshwater Mussels Show Elevated Viral Richness and Intensity during a Mortality Event.

Freshwater mussels (Unionida) are among the world's most imperiled taxa, but the relationship between freshwater mussel mortality events and infectious disease is largely unstudied. We surveyed viromes of a widespread and abundant species (mucket, Actinonaias ligamentina; syn: Ortmanniana ligamentina) experiencing a mortality event of unknown etiology in the Huron River, Michigan, in 2019-2020 and compared them to viromes from mucket in a healthy population in the St. Croix River, Wisconsin and a population from the Clinch River, Virginia and Tennessee, where a mortality event was affecting the congeneric pheasantshell (Actinonaias pectorosa; syn: Ortmanniana pectorosa) population. We identified 38 viruses, most of which were associated with mussels collected during the Huron River mortality event. Viral richness and cumulative viral read depths were significantly higher in moribund mussels from the Huron River than in healthy controls from each of the three populations. Our results demonstrate significant increases in the number and intensity of viral infections for freshwater mussels experiencing mortality events, whereas individuals from healthy populations have a substantially reduced virome comprising a limited number of species at low viral read depths.

A NEW SPECIES OF MYXOBOLUS (CNIDARIA: MYXOSPOREA: MYXOBOLIDAE) FROM THE GILLS OF CREEK CHUB, SEMOTILUS ATROMACULATUS (CYPRINIFORMES: LEUCISCIDAE: PLAGIOPTERINAE), FROM THE OUACHITA DRAINAGE OF ARKANSAS.

During October and November 2021, 33 creek chubs, Semotilus atromaculatus, were collected from 3 sites in Polk County, Arkansas (Ouachita River drainage), and their gills, gallbladder, fins, integument, musculature, and other major organs were examined for myxozoans. The gills of 9 (27%) were infected with a new myxozoan, Myxobolus fountainae n. sp. Qualitative and quantitative morphological data were from fresh and formalin-fixed preserved spores, while molecular data consisted of a 1918 base pair sequence of the partial small subunit ribosomal RNA gene. Phylogenetic analysis grouped M. fountainae n. sp. with the other leuciscid-infecting myxobolids from North America and within a larger clade of European myxozoans. In addition, histological information is provided on the infection. A previous record of Myxobolus muelleriBütschli, 1882, from the gills and ureters of S. atromaculatus is considered invalid and represents an unknown species. Myxobolus fountainae n. sp. is the only named myxozoan known to infect the gill filaments of S. atromaculatus, whereas Myxobolus pendula (Guilford, 1967) infects the gill arches.

A new species of Myxidium (Cnidaria: Myxosporea: Myxidiidae) from the gallbladder of pickerels, Esox spp. (Esociformes: Esocidae), from southwestern Arkansas, USA.

During April 2016 and again in November 2021, four Chain Pickerels, Esox niger were collected from Union (n = 3) and Nevada (n = 1) counties, Arkansas, USA, and 65 Grass Pickerels, Esox americanus vermiculatus were collected between January 2015 and December 2021 from four counties of Arkansas (n = 31) and McCurtain County, Oklahoma, USA (n = 34), and examined for myxozoans. Gallbladders of an individual E. niger from Nevada County, Arkansas, as well as a single individual of E. a. vermiculatus from Sevier County, Arkansas, were infected with a new myxozoan, Myxidium whippsi n. sp. Qualitative and quantitative morphological data were from formalin-fixed preserved myxospores while molecular data (SSU rRNA gene) consisted of 2031bp (host: E. niger) and 1723 bp (host: E. a. vermiculatus) partial sequences of the small subunit ribosomal RNA gene. Phylogenetic analysis placed M. whippsi n. sp. in a clade with two other myxozoans, Zschokkella nova and Myxidium truttae, previously reported from cyprinids and salmonids, respectively. We document the first report of a myxozoan from E. a. vermiculatus. This article was registered in the Official Register of Zoological Nomenclature (ZooBank) as urn:lsid:zoobank.org:pub:A50FCEB3-68C3-428E-A04E-37A16790F1EB.

Isolations of the Spring Viremia of Carp Virus in the Upper Mississippi River (USA), Including a New Host, the Quillback.

In July of 2018 and 2019, wild fish health surveys were conducted along the Wisconsin and Minnesota portions of the upper Mississippi River. Spring viremia of carp virus (SVCV) was isolated from Common Carp Cyprinus carpio as well as a newly identified host species, the Quillback Carpiodes cyprinus. Sanger sequencing of the gene encoding for the G protein revealed a high similarity of the Quillback isolate to various SVCV isolates identified from Common Carp that were collected during earlier wild fish health surveys and mortality events in the USA. Despite annual monitoring, this virus has been infrequently identified. The speculative role of native fish and invertebrates in allowing the virus to persist for long periods without detection is discussed.

Development of duplex qPCR targeting Carnobacterium maltaromaticum and Vagococcus salmoninarum.

In November 2018, Vagococcus salmoninarum was identified as the causative agent of a chronic coldwater streptococcosis epizootic in broodstock brook trout (Salvelinus fontinalis) at the Iron River National Fish Hatchery in Wisconsin, USA. By February 2019, the epizootic spread to adjacent raceways containing broodstock lake trout (Salvelinus namaycush), whereby fish were found to be coinfected with Carnobacterium maltaromaticum and V. salmoninarum. To differentiate these two pathogens and determine the primary cause of the lake trout morbidity, a quantitative real-time PCR (qPCR) was developed targeting the C. maltaromaticum phenylalanyl-tRNA synthase alpha subunit (pheS) gene. The qPCR was combined with a V. salmoninarum qPCR, creating a duplex qPCR assay that simultaneously quantitates C. maltaromaticum and V. salmoninarum concentrations in individual lake trout tissues, and screens presumptive isolates from hatchery inspections and wild fish from national fish hatchery source waters throughout the Great Lakes basin. Vagococcus salmoninarum and C. maltaromaticum were co-detected in broodstock brook trout from two tribal hatcheries and C. maltaromaticum was present in wild fish in source waters of several national fish hatcheries. This study provides a powerful new tool to differentiate and diagnose two emerging Gram-positive bacterial pathogens.