Publications by authors named "Thomas B Waltzek"

107 Publications

In-Frame 12-Nucleotide Deletion within Open Reading Frame 3a in a SARS-CoV-2 Strain Isolated from a Patient Hospitalized with COVID-19.

Microbiol Resour Announc 2021 Feb 25;10(8). Epub 2021 Feb 25.

Emerging Pathogens Institute, University of Florida, Gainesville, Florida, USA.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) strain UF-8, with an in-frame 12-nucleotide deletion within open reading frame 3a (ORF3a), was isolated from a 78-year-old COVID-19 patient in March 2020.
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http://dx.doi.org/10.1128/MRA.00137-21DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7909084PMC
February 2021

Emerging Pathogens and a Current-Use Pesticide: Potential Impacts on Eastern Hellbenders.

J Aquat Anim Health 2021 Mar 16;33(1):24-32. Epub 2021 Feb 16.

Center for Wildlife Health, University of Tennessee, Knoxville, Tennessee, 37996, USA.

Populations of the eastern hellbender Cryptobranchus alleganiensis alleganiensis have been declining for decades, and emerging pathogens and pesticides are hypothesized to be contributing factors. However, few empirical studies have attempted to test the potential effects of these factors on hellbenders. We simultaneously exposed subadult hellbenders to environmentally relevant concentrations of either Batrachochytrium dendrobatidis (Bd) or a frog virus 3-like ranavirus (RV), a combination of the pathogens, or each pathogen following exposure to a glyphosate herbicide (Roundup). Additionally, we measured the ability of the skin mucosome to inactivate Bd and RV in growth assays. We found that mucosome significantly inactivated RV by an average of 40% but had no negative effects on Bd growth. All treatments that included RV exposure experienced reduced survival compared to controls, and the combination of RV and herbicide resulted in 100% mortality. Histopathology verified RV as the cause of mortality in all RV-exposed treatments. No animals were infected with Bd or died in the Bd-only treatment. Our results suggest that RV exposure may be a significant threat to the survival of subadult hellbenders and that Roundup exposure may potentially exacerbate this threat.
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http://dx.doi.org/10.1002/aah.10117DOI Listing
March 2021

Earliest detection to date of SARS-CoV-2 in Florida: Identification together with influenza virus on the main entry door of a university building, February 2020.

PLoS One 2021 13;16(1):e0245352. Epub 2021 Jan 13.

Emerging Pathogens Institute, University of Florida, Gainesville, Florida, United States of America.

In February and March, 2020, environmental surface swab samples were collected from the handle of the main entry door of a major university building in Florida, as part of a pilot surveillance project screening for influenza. Samples were taken at the end of regular classroom hours, between the dates of February 1-5 and February 19-March 4, 2020. Influenza A(H1N1)pdm09 virus was isolated from the door handle on four of the 19 days sampled. Both SARS-CoV-2 and A(H1N1)pdm09 virus were detected in a sample collected on February 21, 2020. Based on sequence analysis, the Florida SARS-CoV-2 strain (designated UF-11) was identical to strains being identified in Washington state during the same time period, while the earliest similar sequences were sampled in China/Hubei between Dec 30th 2019 and Jan 5th 2020. The first human case of COVID-19 was not officially reported in Florida until March 1st. In an analysis of sequences from COVID-19 patients in this region of Florida, there was only limited evidence of subsequent dissemination of the UF-11 strain. Identical or highly similar strains, possibly related through a common transmission chain, were detected with increasing frequency in Washington state between end of February and beginning of March. Our data provide further documentation of the rapid early spread of SARS-CoV-2 and underscore the likelihood that closely related strains were cryptically circulating in multiple U.S. communities before the first "official" cases were recognized.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0245352PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7806172PMC
January 2021

Characterisation and comparison of the mucosa-associated bacterial communities across the gastrointestinal tract of stranded green turtles, .

AIMS Microbiol 2020 19;6(4):361-378. Epub 2020 Oct 19.

College of Public Health, Medical and Veterinary Sciences, James Cook University, Townsville, 4811, Qld, Australia.

are primarily herbivorous long-distance migratory sea turtles that contribute to marine ecosystems. Extensive research has been conducted to restore the populations of green turtles. Little is known about their gut microbiota which plays a vital role in their health. We investigated the mucosa-associated bacterial communities across the gastrointestinal (GI) tract of a total four (3, juvenile and 1, adult) stranded green turtles. Samples taken from four GI regions including oesophagus, stomach, small intestine and large intestine were analysed by high-throughput sequencing targeting hypervariable V1-V3 regions of the bacterial 16S rRNA gene. Bacterial diversity and richness decreased longitudinally along the GI tract from oesophagus to the small intestine of stranded turtles. The large intestine showed a higher bacterial diversity and richness compared to small intestine. The bacterial community of green turtles' GI tract was largely dominated by Firmicutes, Proteobacteria, Actinobacteria, Bacteroidetes and Fusobacteria. Aerobic and facultative anaerobic bacteria prevailed primarily in the oesophagus while anaerobes (, and ) constituted the bulk of large intestinal microbiota. Firmicutes dominated the GI tract except within the small intestine where Proteobacteria prevailed. At the OTU level, six percent of the total OTUs (>1% relative abundance) were common in all GI regions. This is a comprehensive characterisation of bacterial microbiota across the GI tract in green turtles which will provide a reference for future studies on turtle gut microbiome and their metabolism to improve their health and nutrition during rehabilitation.
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http://dx.doi.org/10.3934/microbiol.2020022DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7755585PMC
October 2020

Phaeohyphomycosis due to Veronaea botryosa in cultured white sturgeon (Acipenser transmontanus Richardson) from California USA during 2006 to 2015.

J Fish Dis 2020 Dec 17. Epub 2020 Dec 17.

Department of Infectious Diseases and Immunology, College of Veterinary Medicine, University of Florida, Gainesville, FL, USA.

Infection with Veronaea botryosa can result in rare cutaneous or disseminated, granulomatous to pyogranulomatous phaeohyphomycosis in humans, although disease due to the fungus has also been reported in non-mammalian vertebrates. This report documents disease due to V. botryosa in captive, juvenile to subadult or young adult white sturgeon (Acipenser transmontanus Richardson) from California USA and complements a previous report of the disease in captive Siberian sturgeon (Acipenser baerii) from Florida USA. Pathological examinations revealed granulomatous to pyogranulomatous inflammation of multiple organs. Isolates of the fungal agent were phenotypically consistent with V. botryosa, and molecular analyses of the D1/D2 region of the fungal 28S rRNA gene and the internal transcribed spacer (ITS) region located between the fungal 18S and 28S rRNA genes confirmed the aetiologic agent as V. botryosa. The disease in captive sturgeon results in a considerable economic encumbrance to the producer due to the loss of the cumulative financial resources invested in the production of older subadult to young adult sturgeon.
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http://dx.doi.org/10.1111/jfd.13308DOI Listing
December 2020

Genome Sequence of a Ranavirus Isolated from a Red-Eared Slider (Trachemys scripta elegans) in Poland.

Microbiol Resour Announc 2020 Nov 19;9(47). Epub 2020 Nov 19.

Department of Infectious Diseases and Immunology, College of Veterinary Medicine, University of Florida, Gainesville, Florida, USA

The red-eared slider (RES) ranavirus (RESRV) was isolated from a free-ranging RES turtle that died with evidence of respiratory disease. The RESRV genome sequence (106,878 bp) was determined, and phylogenetic analysis revealed that it is a common midwife toad virus (CMTV) strain. This study is the first report of CMTV in RES.
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http://dx.doi.org/10.1128/MRA.00781-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7679084PMC
November 2020

Genomic Characterization of Picornaviruses Isolated From Ribbon () and Harbor () Seals.

Front Vet Sci 2020 30;7:554716. Epub 2020 Oct 30.

Department of Infectious Diseases & Immunology, College of Veterinary Medicine, University of Florida, Gainesville, FL, United States.

The seal picornavirus 1, species , is currently the only recognized member of the genus within the family . The bear picornavirus 1 was recently proposed as the second species in the genus under the name aquamavirus B. Herein, we determined the complete genomes of two novel pinniped picornaviruses, the harbor seal picornavirus (HsPV) and the ribbon seal picornavirus (RsPV). The HsPV and the RsPV were isolated in Vero.DogSLAMtag cells from samples collected from stranded harbor () and ribbon () seals. RsPV-infected Vero.DogSLAMtag cells displaying extensive cytopathic effects were processed for transmission electron microscopy and revealed non-enveloped viral particles aggregated into paracrystalline arrays in the cytoplasm. A next-generation sequencing approach was used to recover the complete genomes of the HsPV and the RsPV (6,709 and 6,683 bp, respectively). Phylogenetic and genetic analyses supported the HsPV and the RsPV as members of the genus. Based on these results, RsPV represents a novel strain of , while the HsPV is a novel strain of the proposed species aquamavirus B. These discoveries provide information on the evolutionary relationships and ultrastructure of aquamaviruses and expands the known host range of those viruses. Our results underscore the importance of the application of classical virology and pathology techniques coupled with high-throughput sequencing technologies for the discovery and characterization of pathogens in wild marine mammals.
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http://dx.doi.org/10.3389/fvets.2020.554716DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7661754PMC
October 2020

Characterization of an alphavirus isolated from a stranded harbor porpoise (Phocoena phocoena) from Alaska.

Virus Res 2021 01 16;291:198187. Epub 2020 Oct 16.

Department of Infectious Diseases and Immunology, University of Florida, 2187 Mowry Road, 32611, Gainesville, FL, USA.

The family Togaviridae comprises several significant human and veterinary mosquito-borne pathogens. Two togaviruses (genus Alphavirus) have been previously identified in association with marine mammals, the southern elephant seal virus (SESV) and Eastern equine encephalitis virus (EEEV) from a fatal captive harbor seal infection. Herein we report the ultrastructural and phylogenomic characterization of a novel marine togavirus, the first isolated from a cetacean, an Alaskan harbor porpoise (Phocoena phocoena) displaying ulcerative dermatitis. A skin sample was processed for virus isolation on Vero.DogSLAMtag cells and cytopathic effects (CPE) were observed on primary isolation approximately 20 days post-infection. Transmission electron microscopy of the infected Vero.DogSLAMtag cells revealed typical alphavirus particles budding from both plasma and vacuolar membranes of infected cells. A next-generation sequencing approach was used to determine the near complete genome of the Alaskan harbor porpoise alphavirus (AHPV). Phylogenetic analysis supported the AHPV as the sister species to the SESV, forming a marine mammal alphavirus clade separate from the recognized alphavirus antigenic complexes. Genetic comparison of the protein coding sequence of the AHPV to other alphaviruses demonstrated amino acid identities ranging from 42.1-67.1%, with the highest identity to the SESV. Based on its genetic divergence, we propose the AHPV represents a novel alphavirus species, pending formal proposal to and ratification by the International Committee on Taxonomy of Viruses. The ecological and genetic characteristics of the AHPV and the SESV also suggest they represent a novel antigenic complex within the genus Alphavirus, which we propose to be named the Marine Mammal Virus Complex. The role of the AHPV in the associated harbor porpoise cutaneous pathology, if any, remains unclear. Further research is needed to determine AHPV's route(s) of transmission and potential vectors, host range, prevalence, and pathogenicity in cetaceans including harbour porpoises.
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http://dx.doi.org/10.1016/j.virusres.2020.198187DOI Listing
January 2021

Cytochrome oxidase gene sequencing reveals channel catfish ovary cell line is contaminated with brown bullhead cells.

J Fish Dis 2021 Jan 17;44(1):119-122. Epub 2020 Oct 17.

Department of Basic Sciences, College of Veterinary Medicine, Mississippi State University, Starkville, MS, USA.

The channel catfish (Ictalurus punctatus, Rafinesque) ovary (CCO) cell line is the standard cell line used for channel catfish diagnostics. Next-gen sequencing studies of a virus cultured in the CCO cells revealed mitochondrial sequences matching those of brown bullhead (Ameiurus nebulosus, Lesueur). Therefore, we systematically performed partial cytochrome oxidase 1 gene sequencing of several sources of the CCO cell line and all matched the brown bullhead and not the channel catfish.
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http://dx.doi.org/10.1111/jfd.13278DOI Listing
January 2021

Viable SARS-CoV-2 in the air of a hospital room with COVID-19 patients.

Int J Infect Dis 2020 Nov 16;100:476-482. Epub 2020 Sep 16.

Department of Environmental Engineering Sciences, College of Engineering, University of Florida, USA.

Objectives: Because the detection of SARS-CoV-2 RNA in aerosols but failure to isolate viable (infectious) virus are commonly reported, there is substantial controversy whether severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) can be transmitted through aerosols. This conundrum occurs because common air samplers can inactivate virions through their harsh collection processes. We sought to resolve the question whether viable SARS-CoV-2 can occur in aerosols using VIVAS air samplers that operate on a gentle water vapor condensation principle.

Methods: Air samples collected in the hospital room of two coronavirus disease-2019 (COVID-19) patients, one ready for discharge and the other newly admitted, were subjected to RT-qPCR and virus culture. The genomes of the SARS-CoV-2 collected from the air and isolated in cell culture were sequenced.

Results: Viable SARS-CoV-2 was isolated from air samples collected 2 to 4.8 m away from the patients. The genome sequence of the SARS-CoV-2 strain isolated from the material collected by the air samplers was identical to that isolated from the newly admitted patient. Estimates of viable viral concentrations ranged from 6 to 74 TCID units/L of air.

Conclusions: Patients with respiratory manifestations of COVID-19 produce aerosols in the absence of aerosol-generating procedures that contain viable SARS-CoV-2, and these aerosols may serve as a source of transmission of the virus.
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http://dx.doi.org/10.1016/j.ijid.2020.09.025DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7493737PMC
November 2020

Tilapia develop protective immunity including a humoral response following exposure to tilapia lake virus.

Fish Shellfish Immunol 2020 Nov 25;106:666-674. Epub 2020 Aug 25.

Department of Veterinary Microbiology and Immunology, Faculty of Veterinary Medicine, Kasetsart University. Thailand; Center for Advanced Studies for Agriculture and Food, Kasetsart University, Institute for Advanced Studies, Kasetsart University, Bangkok, 10900, Thailand (CASAF, NRU-KU), Thailand. Electronic address:

Tilapia lake virus (TiLV) is an emerging virus associated with high mortality in cultured tilapia. Since the first report of tilapia lake virus, it has been detected in diseased tilapia in sixteen countries around the world. Thus, there is an urgent need to develop an efficacious vaccine to prevent TiLV disease (TiLVD) and reduce its global economic impact. Understanding the role of the adaptive immune response following exposure of tilapia to TiLV is a critical step in the development of such a vaccine. In this study, we challenged red hybrid tilapia by cohabitation or intraperitoneal injection and demonstrated that surviving fish develop a protective immunity. We also demonstrated that tilapia that survived experimental infections possess significant antibodies against the protein encoded by the TiLV segment 4. We then developed a TiLV indirect ELISA to determine the antibody response in tilapia. The ELISA revealed high antibody levels in survivors of experimental challenges and following outbreaks on farms. The ELISA effectively distinguished TiLV-exposed from unexposed tilapia and was used to monitor anti-TiLV antibody kinetics following infection. During the primary infection, tilapia developed an antibody response as early as 7 days post TiLV challenge (dpc), peaked at 15 dpc, showed a gradual decline up until about 42 dpc, but persisted in some fish up until day 110 dpc. Upon re-infection, an increased antibody response occurred within 7-14 days, demonstrating that tilapia that survive TiLV infections develop humoral memory. In conclusion, our results demonstrated that tilapia mount antibody responses against TiLV that supports protective immunity to subsequent TiLV disease. The persistence of anti-TiLV antibodies in survivors following a single exposure suggests a single vaccination might be adequate to protect tilapia during the entire grow-out period. This study provides important information about the immune response of tilapia following exposure to TiLV as a first step in the development of an efficacious vaccine against this emerging and economically important viral disease.
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http://dx.doi.org/10.1016/j.fsi.2020.08.031DOI Listing
November 2020

Viable SARS-CoV-2 in the air of a hospital room with COVID-19 patients.

medRxiv 2020 Aug 4. Epub 2020 Aug 4.

Department of Environmental Engineering Sciences, College of Engineering, University of Florida, USA.

Background - There currently is substantial controversy about the role played by SARS-CoV-2 in aerosols in disease transmission, due in part to detections of viral RNA but failures to isolate viable virus from clinically generated aerosols. Methods - Air samples were collected in the room of two COVID-19 patients, one of whom had an active respiratory infection with a nasopharyngeal (NP) swab positive for SARS-CoV-2 by RT-qPCR. By using VIVAS air samplers that operate on a gentle water-vapor condensation principle, material was collected from room air and subjected to RT-qPCR and virus culture. The genomes of the SARS-CoV-2 collected from the air and of virus isolated in cell culture from air sampling and from a NP swab from a newly admitted patient in the room were sequenced. Findings - Viable virus was isolated from air samples collected 2 to 4.8m away from the patients. The genome sequence of the SARS-CoV-2 strain isolated from the material collected by the air samplers was identical to that isolated from the NP swab from the patient with an active infection. Estimates of viable viral concentrations ranged from 6 to 74 TCID50 units/L of air. Interpretation - Patients with respiratory manifestations of COVID-19 produce aerosols in the absence of aerosol-generating procedures that contain viable SARS-CoV-2, and these aerosols may serve as a source of transmission of the virus.
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http://dx.doi.org/10.1101/2020.08.03.20167395DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7418726PMC
August 2020

Genome Sequence of a CHeRI Orbivirus 3 Strain Isolated from a Dead White-Tailed Deer (Odocoileus virginianus) in Florida, USA.

Microbiol Resour Announc 2020 Jun 25;9(26). Epub 2020 Jun 25.

Department of Infectious Diseases and Immunology, College of Veterinary Medicine, University of Florida, Gainesville, Florida, USA

We report the genome sequence of an orbivirus isolated from a dead farmed white-tailed deer in Florida. The deer was coinfected with epizootic hemorrhagic disease virus type 2. Phylogenetic and genetic analyses supported the virus as the fourth strain of the CHeRI orbivirus 3 species.
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http://dx.doi.org/10.1128/MRA.00523-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7317105PMC
June 2020

Characterization of the bacterial microbiome among free-ranging bottlenose dolphins ().

Heliyon 2020 Jun 18;6(6):e03944. Epub 2020 Jun 18.

Department of Infectious Diseases and Immunology, College of Veterinary Medicine, University of Florida, Gainesville, Florida, USA.

Marine animals represent a dynamic and complex habitat for diverse microbial communities. The microbiota associated with bottlenose dolphins () are believed to influence their health status, but it remains poorly understood. We therefore characterized and compared the bacterial microbiome of bottlenose dolphins from six different anatomical sites that represent four different body systems (respiratory, digestive, reproductive, and integumentary). In this study, a total of 14 free-ranging bottlenose dolphins were sampled during the 2015 Sarasota Bay Dolphin Health Assessment. Bacterial diversity and abundance were assessed by PCR amplification of the hypervariable V3-V4 regions of the bacterial 16S rRNA gene for each sample, followed by sequencing on an Illumina MiSeq platform. Analysis showed that bottlenose dolphins harbor diverse bacterial communities with a unique microbial community at each body system. Additionally, the bottlenose dolphin bacterial microbiome was clearly distinct to the aquatic microbiome from their surrounding habitat. These results are in close agreement with other cetacean microbiome studies, while our study is the first to explore what was found to be a diverse bottlenose dolphin genital microbiome. The core bacterial communities identified in this study in apparently healthy animals might be informative for future health monitoring of bottlenose dolphins.
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http://dx.doi.org/10.1016/j.heliyon.2020.e03944DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7305398PMC
June 2020

Characterization of a novel picornavirus isolated from moribund aquacultured clownfish.

J Gen Virol 2020 07;101(7):735-745

Department of Infectious Diseases and Immunology, College of Veterinary Medicine, University of Florida, Gainesville, FL, USA.

Over the last decade, a number of USA aquaculture facilities have experienced periodic mortality events of unknown aetiology in their clownfish (). Clinical signs of affected individuals included lethargy, altered body coloration, reduced body condition, tachypnea, and abnormal positioning in the water column. Samples from outbreaks were processed for routine parasitological, bacteriological, and virological diagnostic testing, but no consistent parasitic or bacterial infections were observed. Histopathological evaluation revealed individual cell necrosis and mononuclear cell inflammation in the branchial cavity, pharynx, oesophagus and/or stomach of four examined clownfish, and large basophilic inclusions within the pharyngeal mucosal epithelium of one fish. Homogenates from pooled external and internal tissues from these outbreaks were inoculated onto striped snakehead (SSN-1) cells for virus isolation and cytopathic effects were observed, resulting in monolayer lysis in the initial inoculation and upon repassage. Transmission electron microscopy of infected SSN-1 cells revealed small round particles (mean diameter=20.0-21.7 nm) within the cytoplasm, consistent with the ultrastructure of a picornavirus. Full-genome sequencing of the purified virus revealed a novel picornavirus most closely related to the bluegill picornavirus and other members of the genus . Additionally, pairwise protein alignments between the clownfish picornavirus (CFPV) and other known members of the genus yielded results in accordance with the current International Committee on Taxonomy of Viruses criteria for members of the same genus. Thus, CFPV represents a proposed new limnipivirus species. Future experimental challenge studies are needed to determine the role of CFPV in disease.
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http://dx.doi.org/10.1099/jgv.0.001421DOI Listing
July 2020

Genomic Characterization of Tilapia Lake Virus Isolates Recovered from Moribund Nile Tilapia () on a Farm in the United States.

Microbiol Resour Announc 2020 Jan 23;9(4). Epub 2020 Jan 23.

Department of Infectious Diseases and Immunology, College of Veterinary Medicine, University of Florida, Gainesville, Florida, USA

Here, we present the complete coding sequences of two tilapia lake virus (TiLV) isolates recovered during an investigation of a mortality event in farmed Nile tilapia in the United States. Phylogenetic analysis supported the isolates as each other's closest relatives and members of a clade of Thai TiLV strains.
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http://dx.doi.org/10.1128/MRA.01368-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6979302PMC
January 2020

The effect of diet on the gastrointestinal microbiome of juvenile rehabilitating green turtles (Chelonia mydas).

PLoS One 2020 15;15(1):e0227060. Epub 2020 Jan 15.

Georgia Sea Turtle Center, Jekyll Island Authority, Jekyll Island, Georgia, United States of America.

Threatened and endangered green turtles (Chelonia mydas) are unique because as juveniles they recruit from pelagic to near-shore waters and shift from an omnivorous to primarily herbivorous diet (i.e. seagrass and algae). Nevertheless, when injured and ill animals are admitted to rehabilitation, animal protein (e.g. seafood) is often offered to combat poor appetite and emaciation. We examined how the fecal microbiome of juvenile green turtles changed in response to a dietary shift during rehabilitation. We collected fecal samples from January 2014 -January 2016 from turtles (N = 17) in rehabilitation at the Georgia Sea Turtle Center and used next generation sequencing to analyze bacterial community composition. Samples were collected at admission, mid-rehabilitation, and recovery, which entailed a shift from a mixed seafood-vegetable diet at admission to a primarily herbivorous diet at recovery. The dominant phyla changed over time, from primarily Firmicutes (55.0%) with less Bacteroidetes (11.4%) at admission, to primarily Bacteroidetes (38.4%) and less Firmicutes (31.8%) at recovery. While the microbiome likely shifts with the changing health status of individuals, this consistent inversion of Bacteroidetes and Firmicutes among individuals likely reflects the increased need for protein digestion, for which Bacteroidetes are important. Firmicutes are significant in metabolizing plant polysaccharides; thus, fewer Firmicutes may result in underutilization of wild diet items in released individuals. This study demonstrates the importance of transitioning rehabilitating green turtles to an herbivorous diet as soon as possible to afford them the best probability of survival.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0227060PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6961862PMC
April 2020

A New Family of DNA Viruses Causing Disease in Crustaceans from Diverse Aquatic Biomes.

mBio 2020 01 14;11(1). Epub 2020 Jan 14.

Department of Infectious Diseases and Immunology, College of Veterinary Medicine, University of Florida, Gainesville, Florida, USA

Panulirus argus virus 1 (PaV1) is the only known virus infecting the Caribbean spiny lobster () from the Caribbean Sea. Recently, related viruses, Dikerogammarus haemobaphes virus 1 (DhV1) and Carcinus maenas virus 1 (CmV1), have been detected in the demon shrimp () and the European shore crab (), respectively, from sites in the United Kingdom. The virion morphology of these crustacean viruses is similar to that of iridoviruses. However, unlike iridoviruses and other nucleocytoplasmic large DNA viruses (NCLDVs), these viruses complete their morphogenesis in the host cell nucleus rather than in the cytoplasm. To date, these crustacean viruses have remained unclassified due to a lack of genomic data. Using an Illumina MiSeq sequencer, we sequenced the complete genomes of PaV1, CmV1, and DhV1. Comparative genome analysis shows that these crustacean virus genomes encode the 10 hallmark proteins previously described for the NCLDVs of eukaryotes, strongly suggesting that they are members of this group. With a size range of 70 to 74 kb, these are the smallest NCLDV genomes identified to date. Extensive gene loss, divergence of gene sequences, and the accumulation of low-complexity sequences reflect the extreme degradation of the genomes of these "minimal" NCLDVs rather than any direct relationship with the NCLDV ancestor. Phylogenomic analysis supports the classification of these crustacean viruses as a distinct family, "Mininucleoviridae," within the pitho-irido-Marseille branch of the NCLDVs. Recent genomic and metagenomic studies have led to a dramatic expansion of the known diversity of nucleocytoplasmic large DNA viruses (NCLDVs) of eukaryotes, which include giant viruses of protists and important pathogens of vertebrates, such as poxviruses. However, the characterization of viruses from nonmodel hosts still lags behind. We sequenced the complete genomes of three viruses infecting crustaceans, the Caribbean spiny lobster, demon shrimp, and European shore crab. These viruses have the smallest genomes among the known NCLDVs, with losses of many core genes, some of which are shared with iridoviruses. The deterioration of the transcription apparatus is compatible with microscopic and ultrastructural observations indicating that these viruses replicate in the nucleus of infected cells rather than in the cytoplasm. Phylogenomic analysis indicates that these viruses are sufficiently distinct from all other NCLDVs to justify the creation of a separate family, for which we propose the name "Mininucleoviridae" (i.e., small viruses reproducing in the cell nucleus).
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http://dx.doi.org/10.1128/mBio.02938-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6960288PMC
January 2020

Genome characterization of cetaceanpox virus from a managed Indo-Pacific bottlenose dolphin (Tursiops aduncus).

Virus Res 2020 03 7;278:197861. Epub 2020 Jan 7.

Department of Infectious Diseases and Immunology, College of Veterinary Medicine, University of Florida, 2187 Mowry Road, 32611 Gainesville, Florida, USA. Electronic address:

Cetaceanpox viruses (CePVs) are associated with a cutaneous disease in cetaceans often referred to as "tattoo" lesions. To date, only partial genomic data are available for CePVs, and thus, they remain unclassified members of the subfamily Chordopoxvirinae within the family Poxviridae. Herein, we describe the first complete CePV genome sequenced from the tattoo lesion of a managed Indo-Pacific bottlenose dolphin (Tursiops aduncus), using next-generation sequencing. The T. aduncus CePV genome (CePV-TA) was determined to encode 120 proteins, including eight genes unique to the CePV-TA and five genes predicted to function as immune-evasion genes. The results of CePV-TA genetic analyses supported the creation of a new chordopoxvirus genus for CePVs. The complete sequencing of a CePV represents an important first step in unraveling the evolutionary relationship and taxonomy of CePVs, and significantly increases our understanding of the genomic characteristics of these chordopoxviruses.
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http://dx.doi.org/10.1016/j.virusres.2020.197861DOI Listing
March 2020

Detection and Preliminary Characterization of Phocine Distemper Virus in a Stranded Harp Seal () from the Gulf of St. Lawrence, Canada.

J Wildl Dis 2020 07 9;56(3):646-650. Epub 2020 Jan 9.

Department of Fisheries and Oceans Canada, 501 University Crescent, Winnipeg, Manitoba R3T 2N6, Canada.

A lethargic juvenile male harp seal () in poor nutritional condition was found on the beach on the north shore of Prince Edward Island, Canada, in June 2017. Microscopic examination revealed a severe nonsuppurative encephalitis positive for morbillivirus antigen on immunohistochemistry. Virus isolation attempts were negative. However, phocine distemper virus (PDV) was detected in brain tissue RNA extracts by a seminested reverse transcription PCR that targeted the paramyxovirus RNA-dependent RNA polymerase () gene. Comparison of the resulting partial PDV nucleotide sequence revealed it was nearly identical to PDV strains isolated from eastern Atlantic harbor seals () during a 1988 epizootic in the Wadden and Irish seas, and a western Atlantic harbor seal () that stranded in Maine, US, in 2006. Our study confirmed that closely related PDV strains are circulating in multiple seal species along the coastlines of North America and Europe.
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http://dx.doi.org/10.7589/2019-10-267DOI Listing
July 2020

Characterization of ranaviruses isolated from lumpfish L. in the North Atlantic area: proposal for a new ranavirus species (European North Atlantic Ranavirus).

J Gen Virol 2020 02 20;101(2):198-207. Epub 2019 Dec 20.

Technical University of Denmark, National Institute of Aquatic Resources, Aquatic Animal Health Unit, Kgs. Lyngby, Denmark.

The commercial production of lumpfish L. is expanding with the increased demand for their use as cleaner fish, to control sea-lice numbers, at marine Atlantic salmon L. aquaculture sites throughout Northern Europe. A new ranavirus has been isolated from lumpfish at multiple locations in the North Atlantic area. First isolated in 2014 in the Faroe Islands, the virus has subsequently been found in lumpfish from Iceland in 2015 and from Scotland and Ireland in 2016. The Icelandic lumpfish ranavirus has been characterized by immunofluorescent antibody test, optimal growth conditions and transmission electron microscopy. Partial sequences of the major capsid protein gene from 12 isolates showed 99.79-100% nt identity between the lumpfish ranaviruses. Complete genome sequencing from three of the isolates and phylogenetic analysis based on the concatenated 26 iridovirus core genes suggest these lumpfish ranavirus isolates form a distinct clade with ranaviruses from cod L. and turbot L. isolated in Denmark in 1979 and 1999, respectively. These data suggest that these viruses should be grouped together as a new ranavirus species, European North Atlantic Ranavirus, which encompasses ranaviruses isolated from marine fishes in European North Atlantic waters.
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http://dx.doi.org/10.1099/jgv.0.001377DOI Listing
February 2020

Three New Species Isolated from Farmed White-Tailed Deer () in the United States.

Viruses 2019 12 20;12(1). Epub 2019 Dec 20.

Emerging Pathogens Institute, University of Florida, Gainesville, FL 32610, USA.

We report the detection and gene coding sequences of three novel species found in six dead farmed white-tailed deer in the United States. Phylogenetic analyses indicate that the new orbiviruses are genetically closely related to the Guangxi, Mobuck, Peruvian horse sickness, and Yunnan orbiviruses, which are thought to be solely borne by mosquitos. However, four of the six viruses analyzed in this work were found as co-infecting agents along with a known cervid pathogen, epizootic hemorrhagic disease virus-2 (EHDV-2), raising questions as to whether the new viruses are primary pathogens or secondary pathogens that exacerbate EHDV-2 infections. Moreover, EHDV-2 is known to be a -borne virus, raising additional questions as to whether species can also serve as vectors for the novel orbiviruses, if mosquitoes can vector EHDV-2, or whether the deer were infected through separate bites by the insects. Our findings expand knowledge of the possible viral pathogens of deer in the United States. Moreover, due to the close genetic relatedness of the three new orbiviruses to viruses that are primary pathogens of cattle and horses, our findings also underscore a crucial need for additional research on the potential role of the three new orbiviruses as pathogens of other animals.
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http://dx.doi.org/10.3390/v12010013DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7019857PMC
December 2019

Genomic characterization of a novel circovirus from a stranded Longman's beaked whale (Indopacetus pacificus).

Virus Res 2020 02 29;277:197826. Epub 2019 Nov 29.

Department of Infectious Diseases and Immunology, College of Veterinary Medicine, University of Florida, Gainesville, FL, United States. Electronic address:

Tissues from a juvenile Longman's beaked whale that stranded in Hawaii in 2010 were screened for viruses using a Next-Generation Sequencing (NGS) approach. From the NGS data, the full genome (1,849 bp) of a novel beaked whale circovirus (BWCV) was determined. Two open reading frames (ORF) were annotated, including ORF1 that encodes the capsid gene, ORF2 that encodes the replication-associated gene, and a 9-bp conserved nonamer on the apex of the open loop found in all circoviruses. Independent phylogenetic analyses based on amino acid sequence alignments of the two CV proteins supported the BWCV as a member of the genus Circovirus, branching as the sister species to the recently discovered canine circovirus. A sequence identity matrix generated from complete genome alignments revealed the BWCV displays between from 51.1 to 56.7% nucleotide identity to other circoviruses, which is lower than the 80% threshold proposed for species demarcation. Considering the genetic and phylogenetic analyses, we propose the formal species designation of beaked whale circovirus. An endpoint PCR assay targeting the BWCV genome confirmed the presence of the BWCV DNA in every tissue from which DNA was extracted, including spleen, muscle, left ventricle, left adrenal gland, liver, lung, cerebrum, cerebellum, and lymph node. An automated in situ hybridization assay utilizing RNAscope® technology and targeting the replication-associated gene resulted in labeling of individual cells morphologically resembling mononuclear leukocytes and cells of blood vessels in diaphragm, liver, lymph nodes, lung, pericardium, oral mucosa and tongue, adrenal gland, testis, aorta, intestine, stomach and heart. The clinical or pathologic significance of BWCV is undetermined, as are its host range, prevalence, and pathogenicity in cetaceans of Hawaiian waters and elsewhere.
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http://dx.doi.org/10.1016/j.virusres.2019.197826DOI Listing
February 2020

Characterization of a peribunyavirus isolated from largemouth bass (Micropterus salmoides).

Virus Res 2019 11 17;273:197761. Epub 2019 Sep 17.

University of Wisconsin-La Crosse, 54601, La Crosse, WI, USA.

We report the complete genome sequencing of the first fish peribunyavirus determined using a next-generation sequencing approach. The virus was isolated during a routine health assessment of wild largemouth bass (Micropterus salmoides) in Wisconsin in April of 2009. Further research is needed to determine the epidemiology and pathogenicity of the largemouth bass bunyavirus.
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http://dx.doi.org/10.1016/j.virusres.2019.197761DOI Listing
November 2019

Genomic and developmental characterisation of a novel bunyavirus infecting the crustacean Carcinus maenas.

Sci Rep 2019 09 10;9(1):12957. Epub 2019 Sep 10.

Fisheries and Aquatic Science, University of Florida, Gainesville, Florida, 32653, USA.

Carcinus maenas is in the top 100 globally invasive species and harbours a wide diversity of pathogens, including viruses. We provide a detailed description for a novel bunyavirus (Carcinus maenas Portunibunyavirus 1) infecting C. maenas from its native range in the Faroe Islands. The virus genome is tripartite, including large (L) (6766 bp), medium (M) (3244 bp) and small (S) (1608 bp) negative sense, single-stranded RNA segments. Individual genomic segments are flanked by 4 bp regions of similarity (CCUG). The segments encode an RNA-dependent RNA-polymerase, glycoprotein, non-structural protein with a Zinc-Finger domain and a nucleoprotein. Most show highest identity to the 'Wenling Crustacean Virus 9' from an unidentified crustacean host. Phylogenomics of crustacean-infecting bunyaviruses place them across multiple bunyavirus families. We discuss the diversity of crustacean bunyaviruses and provide an overview of how these viruses may affect the health and survival of crustacean hosts, including those inhabiting niches outside of their native range.
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http://dx.doi.org/10.1038/s41598-019-49260-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6736955PMC
September 2019

Characterization of a novel rhabdovirus isolated from a stranded harbour porpoise (Phocoena phocoena).

Virus Res 2019 11 6;273:197742. Epub 2019 Sep 6.

University of Florida, 2187 Mowry Road, 32611, Gainesville, Florida, USA. Electronic address:

An adult male harbour porpoise (Phocoena phocoena) stranded off the coast of Alaska displaying poor body condition, scattered mild ulcerative dermatitis, and necrotizing balanoposthitis. Necropsy findings included severe verminous panniculitis, pneumonia, hepatitis, and enteritis. Histopathological examination of skin lesions revealed a pustular epidermitis and dermatitis, with ballooning degeneration of keratinocytes and occasional amphophilic intracytoplasmic inclusion bodies. A swab sample collected from the ulcerative penile lesions was processed for virus isolation resulting in cytopathic effects observed in primary beluga whale kidney (BWK) cells. Transmission electron microscopy revealed bullet-shaped virions budding from the cell surface of infected BWK cells consistent with a rhabdovirus. A cDNA library was prepared using RNA extracted from infected cell culture supernatant and sequenced on an Illumina MiSeq sequencer. The near-complete genome of a novel rhabdovirus was recovered. Genetic and phylogenetic analyses based on the complete L gene supported the harbour porpoise rhabdovirus (HPRV) as a new species. HPRV clustered phylogenetically with dolphin rhabdovirus (DRV) and this cetacean rhabdovirus clade was found to be the sister group to members of the genus Perhabdovirus that infect fish. A specific nested RT-PCR assay detected HPRV RNA in the epaxial musculature of the harbour porpoise. Our results are consistent with a previous hypothesis that cetacean rhabdoviruses may have arisen following a host jump from fish and suggest that DRV and HPRV represent separate species belonging in a new genus within the family Rhabdoviridae. Further research is needed to determine the health impact of HPRV in harbour porpoise populations, its prevalence, and route of transmission.
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http://dx.doi.org/10.1016/j.virusres.2019.197742DOI Listing
November 2019

A novel host-adapted strain of Salmonella Typhimurium causes renal disease in olive ridley turtles (Lepidochelys olivacea) in the Pacific.

Sci Rep 2019 06 27;9(1):9313. Epub 2019 Jun 27.

Department of Infectious Diseases and Immunology, College of Veterinary Medicine, University of Florida, Gainesville, Florida, 32603, United States of America.

Salmonella spp. are frequently shed by wildlife including turtles, but S. enterica subsp. enterica serovar Typhimurium or lesions associated with Salmonella are rare in turtles. Between 1996 and 2016, we necropsied 127 apparently healthy pelagic olive ridley turtles (Lepidochelys olivacea) that died from drowning bycatch in fisheries and 44 live or freshly dead stranded turtles from the west coast of North and Central America and Hawaii. Seven percent (9/127) of pelagic and 47% (21/44) of stranded turtles had renal granulomas associated with S. Typhimurium. Stranded animals were 12 times more likely than pelagic animals to have Salmonella-induced nephritis suggesting that Salmonella may have been a contributing cause of stranding. S. Typhimurium was the only Salmonella serovar detected in L. olivacea, and phylogenetic analysis from whole genome sequencing showed that the isolates from L. olivacea formed a single clade distinct from other S. Typhimurium. Molecular clock analysis revealed that this novel clade may have originated as recently as a few decades ago. The phylogenetic lineage leading to this group is enriched for non-synonymous changes within the genomic area of Salmonella pathogenicity island 1 suggesting that these genes are important for host adaptation.
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http://dx.doi.org/10.1038/s41598-019-45752-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6597722PMC
June 2019

Molecular characterization of a novel reassortment Mammalian orthoreovirus type 2 isolated from a Florida white-tailed deer fawn.

Virus Res 2019 09 19;270:197642. Epub 2019 Jun 19.

Department of Infectious Diseases and Immunology, College of Veterinary Medicine, University of Florida, Gainesville, FL, USA; Emerging Pathogens Institute, University of Florida, Gainesville, FL, USA. Electronic address:

Mammalian orthoreovirus (MRV) is the type species of the genus Orthoreovirus and causes a range of significant respiratory, nervous or enteric diseases in humans and animals. In 2016 a farmed white-tailed deer (Odocoileus virginianus) fawn became ill, displaying clinical signs of lethargy, dehydration, and profuse foul-smelling diarrhea. A necropsy was performed after the three-week-old fawn died and various tissue samples were submitted to the University of Florida's Cervidae Health Research Initiative for diagnostic evaluation. Aliquots of homogenized heart, liver, and spleen tissues were inoculated onto Vero E6 cells. After virus-specific cytopathic effects (CPE) were detected in Vero cells inoculated with spleen homogenate, infected cells were fixed in glutaraldehyde and analyzed by transmission electron microscopy (TEM), which revealed icosahedral virus particles approximately 75 nm in diameter with morphologies consistent with those of reoviruses within the cytoplasm of the infected cells. RNA extracted from virions in the spent media of infected cells with advanced CPE was used to prepare a cDNA library, which was sequenced using an Illumina MiSeq sequencer. Complete coding sequences for ten separate reovirus segments were attained, and these indicated the isolated agent was a MRV. Genetic and phylogenetic analyses based on the outer capsid sigma-1 (σ1) protein gene sequences supported the Florida white-tailed fawn isolate as a type 2 MRV that branched as the sister group to a MRV-2 strain previously characterized from the urine of a moribund lion (Panthera leo) in Japan. However, analyses based on 7/10 genes (L1-L2, M2-M3, S2-S4) supported the white-tailed deer MRV as the closest relative to a type 3 MRV strain isolated from a dead mink in China. These data suggest the white-tailed deer MRV may have resulted from the natural reassortment of MRVs originating from multiple wildlife species. To our knowledge, this is the first detection of MRV-2 infection in a white-tailed deer. Continued surveillance efforts are needed to determine whether this MRV-2 strain poses a health threat to farmed white-tailed deer populations.
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http://dx.doi.org/10.1016/j.virusres.2019.197642DOI Listing
September 2019

Characterization of a Novel Megalocytivirus Isolated from European Chub ().

Viruses 2019 05 15;11(5). Epub 2019 May 15.

Department of Infectious Diseases and Immunology, College of Veterinary Medicine, University of Florida, Gainesville, FL 32611, USA.

A novel virus from moribund European chub () was isolated on (EPC) cells. Transmission electron microscopic examination revealed abundant non-enveloped, hexagonal virus particles in the cytoplasm of infected EPC cells consistent with an iridovirus. Illumina MiSeq sequence data enabled the assembly and annotation of the full genome (128,216 bp encoding 108 open reading frames) of the suspected iridovirus. Maximum Likelihood phylogenetic analyses based on 25 iridovirus core genes supported the European chub iridovirus (ECIV) as being the sister species to the recently-discovered scale drop disease virus (SDDV), which together form the most basal megalocytivirus clade. Genetic analyses of the ECIV major capsid protein and ATPase genes revealed the greatest nucleotide identity to members of the genus including SDDV. These data support ECIV as a novel member within the genus . Experimental challenge studies are needed to fulfill River's postulates and determine whether ECIV induces the pathognomonic microscopic lesions (i.e., megalocytes with basophilic cytoplasmic inclusions) observed in megalocytivirus infections.
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http://dx.doi.org/10.3390/v11050440DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6563503PMC
May 2019

Genome Sequence of Tilapia Lake Virus Associated with Syncytial Hepatitis of Tilapia in an Ecuadorian Aquaculture Facility.

Microbiol Resour Announc 2019 May 2;8(18). Epub 2019 May 2.

Department of Infectious Diseases and Immunology, College of Veterinary Medicine, University of Florida, Gainesville, Florida, USA

Here, we report the complete coding sequences of tilapia lake virus (TiLV) associated with syncytial hepatitis of tilapia (SHT). The TiLV strain was sequenced from the liver RNA extract of a moribund Nile tilapia () fingerling from an Ecuadorian aquaculture facility in 2012.
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http://dx.doi.org/10.1128/MRA.00084-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6498224PMC
May 2019