Modeling the risk of aquatic species invasion spread through boater movements and river connections.

Aquatic invasive species (AIS) are one of the greatest threats to the functioning of aquatic ecosystems worldwide. Once an invasive species has been introduced to a new region, many governments develop management strategies to reduce further spread. Nevertheless, managing AIS in a new region is challenging because of the vast areas that need protection and limited resources. Spatial heterogeneity in invasion risk is driven by environmental suitability and propagule pressure, which can be used to prioritize locations for surveillance and intervention activities. To better understand invasion risk across aquatic landscapes, we developed a simulation model to estimate the likelihood of a waterbody becoming invaded with an AIS. The model included waterbodies connected via a multilayer network that included boater movements and hydrological connections. In a case study of Minnesota, we used zebra mussels (Dreissena polymorpha) and starry stonewort (Nitellopsis obtusa) as model species. We simulated the impacts of management scenarios developed by stakeholders and created a decision-support tool available through an online application provided as part of the AIS Explorer dashboard. Our baseline model revealed that 89% of new zebra mussel invasions and 84% of new starry stonewort invasions occurred through boater movements, establishing it as a primary pathway of spread and offering insights beyond risk estimates generated by traditional environmental suitability models alone. Our results highlight the critical role of interventions applied to boater movements to reduce AIS dispersal.

Modelo del riesgo de la invasión de especies acuáticas dispersadas por movimiento de botes y conexiones entre ríos Resumen Las especies acuáticas invasoras (EAI) son una de las principales amenazas para el funcionamiento de los ecosistemas acuáticos a nivel mundial. Una vez que una especie invasora ha sido introducida a una nueva región, muchos gobiernos desarrollan estrategias de manejo para disminuir la dispersión. Sin embargo, el manejo de las especies acuáticas invasoras en una nueva región se complica debido a las amplias áreas que necesitan protección y los recursos limitados. La heterogeneidad espacial de un riesgo de invasión es causada por la idoneidad ambiental y la presión de propágulo, que puede usarse para priorizar la ubicación de las actividades de vigilancia e intervención. Desarrollamos una simulación para estimar la probabilidad de que un cuerpo de agua sea invadido por EAI para tener un mejor entendimiento del riesgo de invasión en los paisajes acuáticos. El modelo incluyó cuencas conectadas a través de una red multicapa que incluía movimiento de botes y conexiones hidrológicas. Usamos como especies modelo a Dreissena polymorpha y a Nitellopsis obtusa en un estudio de caso en Minnesota. Simulamos el impacto de los escenarios de manejo desarrollado por los actores y creamos una herramienta de decisiones por medio de una aplicación en línea proporcionada como parte del tablero del Explorer de EAI. Nuestro modelo de línea base reveló que el 89% de las invasiones nuevas de D. polymorpha y el 84% de las de N. obtusa ocurrieron debido al movimiento de los botes, lo que lo estableció como una vía primaria de dispersión y nos proporcionó información más allá de las estimaciones de riesgo generadas por los modelos tradicionales de idoneidad ambiental. Nuestros resultados resaltan el papel crítico de las intervenciones aplicadas al movimiento de los botes para reducir la dispersión de especies acuáticas invasoras.

Disease ecology and host range of Cyprinid herpesvirus 3 (CyHV-3) in CyHV-3 endemic lakes of North America.

Cyprinid herpesvirus-3 (CyHV-3) is an important pathogen of common carp (Cyprinus carpio, carp) causing significant economic and ecological impacts worldwide. The recent emergence of CyHV-3 in the Upper Midwest region of the United States has raised questions related to the disease ecology and host specificity of CyHV-3 in wild carp populations. To determine the prevalence of CyHV-3 in wild populations of fishes in Minnesota, we surveyed five lakes in 2019 in which the virus was known to have caused mass mortality events in carp from 2017 to 2018. A total of 28 species (n = 756 total fish) of native fishes and 730 carp were screened for the presence of CyHV-3 DNA using specific qPCR. None of the native fish tissues tested positive for CyHV-3 although the prevalence of CyHV-3 in carp was 10%-50% in the five lakes. A single lake (Lake Elysian) with a 50% DNA detection rate and evidence of ongoing transmission and CyHV-3-associated mortality was surveyed again in 2020 from April to September. During this period, none of the tissues from 24 species (n = 607 total fish) tested positive for CyHV-3 though CyHV-3 DNA and mRNA (indicating viral replication) was detected in carp tissues during the sampling period. CyHV-3 DNA was detected most often in brain samples without evidence of replication, potentially indicating that brain tissue is a site for CyHV-3 latency. Paired qPCR and ELISA testing for Lake Elysian in 2019-2020 identified young carp (especially males) to be the primary group impacted by CyHV-3-associated mortality and acute infections, but with no positive detections in juvenile carp. Seroprevalence of carp from Lake Elysian was 57% in 2019, 92% in April of 2020 and 97% in September 2020. These results further corroborate the host specificity of CyHV-3 to carp in mixed wild populations of fishes in Minnesota and provide additional insights into the ecological niche of CyHV-3 in shallow lake populations of carp in North America.

A bi-level model for state and county aquatic invasive species prevention decisions.

Recreational boats are important vectors of spread of aquatic invasive species (AIS) among waterbodies of the United States. To limit AIS spread, state and county agencies fund watercraft inspection and decontamination stations at lake access points. We present a bi-level model for determining how a state planner can efficiently allocate inspection resources to county managers, who independently decide where to locate inspection stations. In our formulation, each county manager determines a set of optimal plans for the locations of inspection stations under various resource constraints. Each plan maximizes inspections of risky boats that may carry AIS from infested to uninfested lakes within the county. Then, the state planner selects the set of county plans (i.e., one plan for each county) that maximizes the number of risky boats inspected throughout the state subject to a statewide resource constraint. We apply the model using information from Minnesota, USA, including the infestation status of 9182 lakes and estimates of annual numbers of boat movements from infested to uninfested lakes. Comparison of solutions of the bi-level model with solutions of a state-level model where a state planner selects lakes for inspection stations statewide shows that when state and county objectives are not aligned, the loss in efficiency at the state-level can be substantial.

AIS explorer: Prioritization for watercraft inspections-A decision-support tool for aquatic invasive species management.

Invasions of aquatic invasive species have caused significant economic and ecological damage to global aquatic ecosystems. Once an invasive population has established in a new habitat, eradication can be financially and logistically impossible, motivating management strategies to rely heavily upon prevention measures to reduce the introduction and spread. To be productive, on-the-ground management of aquatic invasive species requires effective decision-making surrounding the allocation of limited resources. Watercraft inspections play an important role in managing aquatic invasive species by preventing the overland transport of invasive species between waterbodies and providing education to boaters. In this study, we developed and tested an interactive web-based decision-support tool, AIS Explorer: Prioritization for Watercraft Inspections, to guide AIS managers in developing efficient watercraft inspection plans. The decision-support tool is informed by a network-based algorithm that maximized the number of inspected watercraft that move from AIS infested to uninfested lakes within and between counties in Minnesota, USA. It was iteratively built with stakeholder feedback, including consultations with county managers, beta-testing of the web-based application, and workshops to educate and train end-users. The co-development and implementation of data-driven decision support tools demonstrate how interdisciplinary methods can be used to connect science and management to support decision-making. The AIS Explorer: Prioritization for Watercraft Inspections application makes optimized research outputs accessible in multiple dynamic forms that maintain pace with discovery of new infestations and local needs. In addition, the decision support tool has supported improved and closer communication between AIS managers and researchers on this topic.

Influence of Environmental Stressors on the Microbiota of Zebra Mussels (Dreissena polymorpha).

Host-associated microbiota play a critical role in host fitness by providing nutrition, enhancing digestion capabilities, and by providing protection from pathogens. Here, we investigated the effects of two environmental stressors, temperature, and salinity, on the microbiota associated with zebra mussels (ZMs), a highly invasive bivalve in North America. To examine this in detail, lake-collected ZMs were acclimated to laboratory conditions, and subjected to temperature and salinity stress conditions. The impact of these stressors on the diversity, composition, and dynamics of ZM-associated microbiota were assessed by using amplicon- and shotgun-based sequencing, and qPCR-based approaches. Elevated temperature was found to be the primary driver of ZM mortality, although salinity alone also increased its likelihood. Stressor-induced ZM mortality, which ranged between 53 and 100%, was concomitant with significant increases in the relative abundance of several genera of putative opportunistic pathogens including Aeromonas. These genera were only present in low relative abundance in ZMs obtained from the control tank with 0% mortality. Shotgun sequencing and qPCR analyses indicated that the relative and absolute abundances of pathogenic Aeromonas species (particularly A. veronii) were significantly greater in temperature-induced dead ZMs. Taken together, our results show that environmental stress, especially elevated temperature (> 25 °C), is associated with the rapid mortality of ZMs as well as the proliferation of putative opportunistic bacterial pathogens.

Field and Laboratory Evaluation of the Microsporidian Parasite Heterosporis sutherlandae: Prevalence, Severity, and Transmission.

Heterosporis sutherlandae is an invasive microsporidian parasite in the Great Lakes region of North America that infects the skeletal muscle of numerous fish species, rendering the fillet unfit for human consumption. Although H. sutherlandae has been identified as a pathogen of concern by state management agencies, there is little information to inform regulation and intervention. We sampled fishes over 1 year from three lakes in northern Minnesota with known infected populations to determine the importance of host demographic and environmental variables for influencing H. sutherlandae infection prevalence. Heterosporis sutherlandae was present during all sampling periods, ranging in prevalence from 1% to 11%. The prevalence of H. sutherlandae among Yellow Perch Perca flavescens varied significantly according to season, with winter having the lowest prevalence (1%) and summer having the highest prevalence (11%). For other fish species, the prevalence of H. sutherlandae also varied significantly with season: the lowest prevalence occurred during spring (1%) and the highest prevalence occurred in fall (9%). Rates of pathogen transmission were estimated by exposing Fathead Minnows Pimephales promelas in the laboratory. Transmission rates were 23% when naïve fish were fed infected tissues and only 2% when naïve fish were held in cohabitation with tissue-fed fish. Exposure method and exposure duration (d) increased the probability that a fish was infected with H. sutherlandae. These findings suggest that H. sutherlandae transmission is greater when a susceptible host consumes infected tissue than when the fish is exposed to spores present in the water column. The current rates of infection in wild fishes are in stark contrast to the prevalence documented in 2004 (28%), suggesting a reduction in H. sutherlandae prevalence within at least one Yellow Perch population in the Laurentian Great Lakes region since the early 2000s.

Koi herpesvirus and carp oedema virus: Infections and coinfections during mortality events of wild common carp in the United States.

Koi herpesvirus (KHV; cyprinid herpesvirus-3) and carp oedema virus (CEV) are important viruses of common and koi carp (Cyprinus carpio); however, the distribution of these viruses in wild common carp in North America is largely unknown. During the summers of 2017 and 2018, 27 mass mortalities of common carp were reported from four states in the USA (Minnesota, Iowa, Pennsylvania and Wisconsin), the majority of which were distributed across eight major watersheds in southern Minnesota. Samples from 22 of these mortality events and from five clinically healthy nearby carp populations were screened for KHV, CEV and SVCV using real-time polymerase chain reaction (qPCR). KHV was confirmed in 13 mortality events, CEV in two mortality events and coinfections of KHV/CEV in four mortality events. Nucleotide sequence analysis revealed that the KHV and CEV detected here are closely related to European lineages of these viruses. While molecular detection alone cannot conclusively link either virus with disease, the cases described here expand the known range of two important viruses. This is also the first reported detection of KHV and CEV coinfections in wild carp populations.

Retrospective and Predictive Investigation of Fish Kill Events.

Fish kill investigations are critical to understanding threats to aquatic ecosystems and can serve as a measure of environmental disruption as well as an early indicator of emerging disease. The goal of this study was to analyze historical data related to such events among wild fish populations in Minnesota in order to assess the quality and completeness of the data and potential trends in fish kills. After excluding events with incomplete data (e.g., in which the location was not reported), we analyzed 225 unique fish kills from 2003 to 2013 that were recorded in two Minnesota Department of Natural Resources databases. The most reported fish kills occurred during 2007 (n = 41) and during the month of June (n = 81) across all years. Centrarchid species were present in the most fish kills (138), followed by cyprinid and ictalurid species, which were present in 53 and 40 events, respectively. Environmental factors were the most common cause of death reported. Models of environmental factors revealed that the maximum nighttime land surface temperature was the most critical factor in fish mortality, followed by changes in primary productivity and human disturbances. During the course of this study, data gaps were identified, including underreporting, inconsistent investigation, and the lack of definitive diagnoses, making interpretation of our results challenging. Even so, understanding these historical trends and data gaps can be useful in generating hypotheses and advancing data collection systems for investigating future fish kills. Our study is a primer investigation of fish kills providing information on the plausible areas, seasons, and fish groups at risk that can guide active environmental monitoring and epidemiological surveillance of fishes.

Genomic characterization of a novel calicivirus, FHMCV-2012, from baitfish in the USA.

During regulatory sampling of fathead minnows (Pimephales promelas), a novel calicivirus was isolated from homogenates of kidney and spleen inoculated into bluegill fry (BF-2) cells. Infected cell cultures exhibiting cytopathic effects were screened by PCR-based methods for selected fish viral pathogens. Illumina HiSeq next generation sequencing of the total RNA revealed a novel calicivirus genome that showed limited protein sequence similarity to known homologs in a BLASTp search. The complete genome of this fathead minnow calicivirus (FHMCV) is 6564 nt long, encoding a polyprotein of 2114 aa in length. The complete polyprotein shared only 21% identity with Atlantic salmon calicivirus,followed by 11% to 14% identity with mammalian caliciviruses. A molecular detection assay (RT-PCR) was designed from this sequence for screening of field samples for FHMCV in the future. This virus likely represents a prototype species of a novel genus in the family Caliciviridae, tentatively named "Minovirus".

Detection and molecular characterization of a novel piscine-myocarditis-like virus from baitfish in the USA.

During a survey of apparently healthy baitfish from the state of Minnesota, a novel piscine-myocarditis-like virus (PMCLV) was detected in golden shiners (Notemigonus crysoleucas). The nearly complete genome sequence is 5819 nt long, including a partial 5' untranslated region (UTR) of 100 nt. The sequence is divided into three ORFs: the complete ORF1 and ORF2, encoding proteins of 818 and 831 amino acids, respectively, and a partial ORF3 encoding 248 amino acids of the corresponding protein. This novel virus sequence was most closely related to piscine myocarditis virus (PMCV), with a 49.0 % and 58.2 % amino acid identity in the ORF1 (YP005481249)- and ORF2 (YP004581250)-encoded proteins, respectively. Six of 56 retail outlets (e.g., bait shops) were positive during the 2014-2015 survey, indicating a 10.7 % prevalence of the novel virus in this population of golden shiners. Currently, there is no disease that is known to be associated with this virus.

Detection and surveillance of viral hemorrhagic septicemia virus using real-time RT-PCR. I. Initial comparison of four protocols.

Eight laboratories worked collectively to evaluate 4 real-time RT-PCR (rRT-PCR) protocols targeting viral hemorrhagic septicemia virus (VHSV) being considered for deployment to a USA laboratory testing network. The protocols utilized previously published primers and probe sets developed for detection and surveillance of VHSV. All participating laboratories received and followed a standard operating protocol for extraction and for each of the rRT-PCR assays. Performance measures specifically evaluated included limit of detection (defined as the smallest amount of analyte in which 95% of the samples are classified as positive), analytical specificity, assay efficiency across genotype representatives, within- and between-plate variation within a laboratory, and variation between laboratories using the same platform, between platforms, and between software versions. This evaluation clearly demonstrated that the TaqMan®-based assay developed by Jonstrup et al. (2013; J Fish Dis 36:9-23) produced the most consistent analytical performance characteristics for detecting all genotypes of VHSV across the 8 participating laboratories.

Detection and surveillance of viral hemorrhagic septicemia virus using real-time RT-PCR. II. Diagnostic evaluation of two protocols.

Two real-time reverse transcription polymerase chain reaction (rRT-PCR) assays under consideration for deployment to multiple testing laboratories across the USA were evaluated for diagnostic sensitivity and specificity on tissue homogenates obtained from natural and experimental viral hemorrhagic septicemia (VHS)-infected fish. Estimates for diagnostic specificity using virus isolation as the reference method were similar between laboratories regardless of the assay. Diagnostic sensitivity estimates of 0.96 (95% CI: 0.95, 0.97) for Jonstrup et al. (2013)'s assay (J Fish Dis 36:9-23) exceeded the diagnostic sensitivity of 0.85 (95% CI: 0.83, 0.87) for Phelps et al. (2012)'s assay (J Aquat Anim Health 24:238-243). The Jonstrup rRT-PCR assay is robust as demonstrated by high sensitivity and specificity estimates across laboratories and can be used as a valuable tool for targeted surveillance and for testing of suspect VHSV samples.

Health of wild fish exposed to contaminants of emerging concern in freshwater ecosystems utilized by a Minnesota Tribal community.

Fish serve as indicators of exposure to contaminants of emerging concern (CECs)-chemicals such as pharmaceuticals, hormones, and personal care products-which are often designed to impact vertebrates. To investigate fish health and CECs in situ, we evaluated the health of wild fish exposed to CECs in waterbodies across northeastern Minnesota with varying anthropogenic pressures and CEC exposures: waterbodies with no human development along their shorelines, those with development, and those directly receiving treated wastewater effluent. Then, we compared three approaches to evaluate the health of fish exposed to CECs in their natural environment: a refined fish health assessment index, a histopathological index, and high-throughput (ToxCast) in vitro assays. Lastly, we mapped adverse outcome pathways (AOPs) associated with identified ToxCast assays to determine potential impacts across levels of biological organization within the aquatic system. These approaches were applied to subsistence fish collected from the Grand Portage Indian Reservation and 1854 Ceded Territory in 2017 and 2019. Overall, 24 CECs were detected in fish tissues, with all but one of the sites having at least one detection. The combined implementation of these tools revealed that subsistence fish exposed to CECs had histological and macroscopic tissue and organ abnormalities, although a direct causal link could not be established. The health of fish in undeveloped sites was as poor, or sometimes poorer, than fish in developed and wastewater effluent-impacted sites based on gross and histologic tissue lesions. Adverse outcome pathways revealed potential hazardous pathways of individual CECs to fish. A better understanding of how the health of wild fish harvested for consumption is affected by CECs may help prioritize risk management research efforts and can ultimately be used to guide fishery management and public health decisions. Integr Environ Assess Manag 2024;20:846-863. © 2023 The Authors. Integrated Environmental Assessment and Management published by Wiley Periodicals LLC on behalf of Society of Environmental Toxicology & Chemistry (SETAC).

Comparison of treatments to inactivate viral hemorrhagic septicemia virus (VHSV-IVb) in frozen baitfish.

Current US state and federal fish health regulations target the spread of viral hemorrhagic septicemia virus-IVb (VHSV-IVb) through movement restrictions of live fish; however, they largely ignore the potential for the virus to be spread through commercial distribution and use of frozen baitfish from VHSV-IVb-positive regions. Some state laws do require treatment of frozen baitfish to inactivate VHSV, and additional methods have been proposed, but few scientific studies have examined the efficacy of these treatments. In this study, bluegills Lepomis macrochirus were challenged with VHSV-IVb and frozen to represent standard industry methods, disinfected by various treatments, and tested for infectious VHSV-IVb using virus isolation. The virus was isolated from 70% of fish subjected to 3 freeze/thaw cycles. All other treatment methods were effective in inactivating the virus, including treatment with isopropyl alcohol, mineral oil, salt and borax, and dehydration. Dehydration followed by rehydration is rapid and effective, and therefore, seems to be the best option for inactivating VHSV-IVb present in frozen baitfish while maintaining their usefulness as bait.

Release of live baitfish by recreational anglers drives fish pathogen introduction risk.

Emerging diseases of wildlife are an existential threat to biodiversity, and human-mediated movements of live animals are a primary vector of their spread. Wildlife disease risk analyses offer an appealing alternative to precautionary approaches because they allow for explicit quantification of uncertainties and consideration of tradeoffs. Such considerations become particularly important in high-frequency invasion pathways with hundreds of thousands of individual vectors, where even low pathogen prevalence can lead to substantial risk. The purpose of this study was to examine the landscape-level dynamics of human behavior-mediated pathogen introduction risk in the context of a high-frequency invasion pathway. One such pathway is the use and release of live fish used as bait by recreational anglers. We used a stochastic risk assessment model parameterized by angler survey data from Minnesota, USA, to simulate one year of fishing in Minnesota and estimate the total number of risky trips for each of three pathogens: viral hemorrhagic septicemia virus, the microsporidian parasite Ovipleistophora ovariae, and the Asian fish tapeworm Schizocotyle acheilognathi. We assessed the number of introductions under four scenarios: current/baseline conditions, outbreak conditions (increased pathogen prevalence), source-focused control measures (decreased pathogen prevalence), and angler-focused control measures (decreased rates of release). We found that hundreds of thousands of introduction events can occur per year, even for regulated pathogens at low pathogen prevalence. Reducing the rate of illegal baitfish release had significant impact on risky trips in scenarios where a high number of anglers were involved, but was less impactful in circumstances with limited outbreaks and fewer affected anglers. In contrast, reducing pathogen prevalence in the source populations of baitfish had relatively little impact. In order to make meaningful changes in pathogen introduction risk, managers should focus efforts on containing local outbreaks and reducing illegal baitfish release to reduce pathogen introduction risk. Our study also demonstrates the risk associated with high-frequency invasion pathways and the importance of incorporating human behaviors into wildlife disease models and risk assessments.

Open water dreissenid mussel control projects: lessons learned from a retrospective analysis.

Dreissenid mussels are one of the most problematic aquatic invasive species (AIS) in North America, causing substantial ecological and economic effects. To date, dreissenid mussel control efforts in open water have included physical, biological, and chemical methods. The feasibility of successful dreissenid mussel management or eradication in lakes is relatively undocumented in the freshwater management literature. This review presents information on 33 open water dreissenid mussel control projects in 23 North America lakes. We reviewed data from past dreissenid mussel control projects and identified patterns and knowledge gaps to help inform adaptive management strategies. The three key lessons learned include (1) pre- and post-treatment survey methods that are designed to meet management objectives are beneficial, e.g., by sampling for all life stages and taking into account that no survey method is completely comprehensive; (2) defining the treatment area-particularly ensuring it is sufficiently large to capture all life stages present-is critical to meeting management objectives; and (3) control projects provide an opportunity to collect water chemistry, effects on non-target organisms, and other efficacy-related data that can inform safe and effective adaptive management.

An expert-based risk ranking framework for assessing potential pathogens in the live baitfish trade.

As global trade of live animals expands, there is increasing need to assess the risks of invasive organisms, including pathogens, that can accompany these translocations. The movement and release of live baitfish by recreational anglers has been identified as a particularly high-risk pathway for the spread of aquatic diseases in the United States. To provide risk-based decision support for preventing and managing disease invasions from baitfish release, we developed a hazard identification and ranking tool to identify the pathogens that pose the highest risk to wild fish via this pathway. We created a screening protocol and semi-quantitative stochastic risk ranking framework, combining published data with expert elicitation (n = 25) and applied the framework to identify high-priority pathogens for the bait supply in Minnesota, USA. Normalized scores were developed for seven risk criteria (likelihood of transfer, prevalence in bait supply, likelihood of colonization, current distribution, economic impact if established, ecological impact if established and host species) to characterize a pathogen's ability to persist in the bait supply and cause impacts to wild fish species of concern. The generalist macroparasite Schizocotyle acheilognathi was identified as presenting highest overall threat, followed by the microsporidian Ovipleistophora ovariae, and viral haemorrhagic septicaemia virus. Our findings provide risk-based decision support for managers charged with maintaining both the recreational fishing industry and sustainable, healthy natural resources. Particularly, the identification of several high-risk but currently unregulated pathogens suggests that focusing risk management on pathogens of concern in all potential host species could reduce disease introduction risk. The ranking process, implemented here for a single state case study, provides a conceptual framework for integrating expert opinion and sparse available data that could be scaled up and applied across jurisdictions to inform risk-based management of the live baitfish pathway.

Susceptibility of Pimephales promelas and Carassius auratus to a strain of koi herpesvirus isolated from wild Cyprinus carpio in North America.

Cyprinid herpesvirus-3 (CyHV-3, syn. koi herpesvirus) is an important pathogen worldwide and a common cause of mass mortality events of wild common carp (Cyprinus carpio) in North America, however, reference strains and genomes obtained from wild carp are not available. Additionally, it is unclear if fishes in North America are susceptible to CyHV-3 infection due to incomplete susceptibility testing. Here we present the first North American type strain and whole-genome sequence of CyHV-3 isolated from wild carp collected from a lake with a history and recent incidence of carp mortality. Additionally, the strain was used in an in-vivo infection model to test the susceptibility of a common native minnow (Pimephales promelas) and goldfish (Carrasius auratus) which is invasive in North America. Detection of CyHV-3 DNA was confirmed in the tissues of a single fathead minnow but the same tissues were negative for CyHV-3 mRNA and samples from exposed fathead minnows were negative on cell culture. There was no detection of CyHV-3 DNA or mRNA in goldfish throughout the experiment. CyHV-3 DNA in carp tissues was reproducibly accompanied by the detection of CyHV-3 mRNA and isolation on cell culture. Additionally, environmental CyHV-3 DNA was detected on all tank filters during the study. These findings suggest that fathead minnows and goldfish are not susceptible to CyHV-3 infection and that detection of CyHV-3 DNA alone in host susceptibility trials should be interpreted with caution.

Host Range of Carp Edema Virus (CEV) during a Natural Mortality Event in a Minnesota Lake and Update of CEV Associated Mortality Events in the USA.

Mass mortality events of common carp (Cyprinus carpio, carp) associated with carp edema virus (CEV) alone or in coinfections with koi herpesvirus (KHV), is an emerging issue. Despite recent outbreaks of CEV in wild carp populations, the host range of North American species has not been well studied. To that end, we intensively sampled carp (n = 106) and co-habiting native fish species (n = 5 species; n = 156 total fish) from a CEV-suspect mass-mortality event of carp in a small Minnesota lake (Lake Swartout). Additionally, fecal and regurgitant samples (n = 73 each) from double-crested cormorants (Phalacrocorax auritus, DCCO) were sampled to test the potential of DCCO to act as a vector for virus transmission. CEV was confirmed to be widespread in the Lake Swartout carp population during the outbreak with high viral loads and histological confirmation, suggesting that CEV was the cause of the mortality event. There were no detections of CEV in any native fish species; however, DCCO regurgitant and fecal samples were positive for CEV DNA. In addition, three CEV-positive and one CEV + KHV-positive mortality events were confirmed with no observed mortality or morbidity of non-carp species in other lakes. This study provides evidence that CEV infection and disease may be specific to carp during mortality events with mixed-species populations, identifies DCCO as a potential vector for CEV, and further expands the known range of CEV, as well as coinfections with KHV, in North America.