Andrew E Goodwin

Publications of Andrew E Goodwin

• Mortality and carrier status of bluegills exposed to viral hemorrhagic septicemia virus genotype IVb at different temperatures.

  Authors: Andrew E Goodwin, Gwenn E Merry

  Journal of aquatic animal health. 06/2011; 23(2):85-91.

  The emergence of the viral hemorrhagic septicemia virus (VHSV) genotype IVb (VHSV-IVb) in the Great Lakes of North America has led to concern that the virus might spread to natural fisheries andThe emergence of the viral hemorrhagic septicemia virus (VHSV) genotype IVb (VHSV-IVb) in the Great Lakes of North America has led to concern that the virus might spread to natural fisheries and aquaculture in the southern USA. We exposed bluegills Lepomis macrochirus to VHSV-IVb by intraperitoneal injection at six temperatures from 10 degrees C to 30 degrees C and followed the disease course by quantitative real-time reverse transcriptase polymerase chain reaction (qrt-RT-PCR). Mortality of injected fish was 90% at 10 degrees C, 35% at 14 degrees C, and 10% at 18 degrees C; no mortality attributable to VHSV was observed at temperatures of 22-30 degrees C. In survivors tested at 21 d postchallenge, viral copies and prevalence determined by qrt-RT-PCR were inversely related to temperature, and VHSV-IVb could not be detected in fish held at temperatures above 22 degrees C. Similar results were obtained for bluegills that were exposed by cohabitation with the intraperitoneally injected fish. Acclimation of the fish to 12 degrees C after 21 d at higher temperatures did not appear to cause a re-emergence of the virus. Based on our findings, the temperature range of VHSV-IVb appears to be the same as published values for VHSV genotype I, which has an optimum of 9-12 degrees C and an upper limit of 18-20 degrees C.

• Thelohanellus toyamai (syn. Myxobolus toyamai) infecting the gills of koi Cyprinus carpio in the eastern United States.

  Authors: Matt J Griffin, Andrew E Goodwin

  The Journal of parasitology. 06/2011; 97(3):493-502.

  A myxozoan resembling species of Thelohanellus was isolated from the gills of koi (Cyprinus carpio) cultured in North Carolina. Plasmodia measuring ∼200 µm in diameter contained tear-shapedA myxozoan resembling species of Thelohanellus was isolated from the gills of koi (Cyprinus carpio) cultured in North Carolina. Plasmodia measuring ∼200 µm in diameter contained tear-shaped myxospores containing a single pyriform polar capsule. The spore body was concave on one side, measuring 16.2 (14.7-16.8) µm long and 5.6 (4.5-6) µm wide. The polar capsule was 6.4 (5.8-7.2) µm long and 4.2 (3.4-4.6) µm wide, containing a polar filament coiled perpendicular to the longitudinal axis of the spore body making 8 turns. Occasionally, an oblong, irregularly shaped mass of protoplasm was observed between the polar capsule and spore capsule. Analysis of 18S small-subunit ribosomal DNA sequence demonstrated this isolate as a 99.9% match with Myxobolus toyamai from gills of C. carpio. However, the case isolate lacked the characteristic second polar capsule of Myxobolus, morphologically placing it within the Thelohanellus. Here we supplement genetic sequence data with histopathology, an amended morphological description of the agent, and a review of the original classification. For future reference, we suggest this organism be referred to as Thelohanellus toyamai Kudo, 1933, in accordance with the original classification and the nomial M. toyamai be avoided because it is at best outdated and, at worst, incorrect.

• Replication and persistence of VHSV IVb in freshwater turtles.

  Authors: Andrew E Goodwin, Gwenn E Merry

  Diseases of aquatic organisms. 05/2011; 94(3):173-7.

  With the emergence of viral hemorrhagic septicemia virus (VHSV) strain IVb in the Great Lakes of North America, hatchery managers have become concerned that this important pathogen could beWith the emergence of viral hemorrhagic septicemia virus (VHSV) strain IVb in the Great Lakes of North America, hatchery managers have become concerned that this important pathogen could be transmitted by animals other than fish. Turtles are likely candidates because they are poikilotherms that feed on dead fish, but there are very few reports of rhabdovirus infections in reptiles and no reports of the fish rhabdoviruses in animals other than teleosts. We injected common snapping turtles Chelydra serpentine and red-eared sliders Trachemys scripta elegans intraperitoneally with 10(4) median tissue culture infectious dose (TCID50) of VHSV-IVb and 21 d later were able to detect the virus by quantitative real-time reverse transcriptase PCR (qrt-RTPCR) in pools of kidney, liver, and spleen. In a second experiment, snapping turtles, red-eared sliders, yellow-bellied sliders T. scripta scripta, and northern map turtles Grapetemys geographica at 14 degrees C were allowed to feed on tissues from bluegill dying of VHSV IVb disease. Turtle kidney, spleen, and brain pools were not positive by qrt-RTPCR on Day 3 post feeding, but were positive on Days 10 and 20. Map turtles on Day 20 post-feeding were positive by both qrt-RTPCR and by cell culture. Our work shows that turtles that consume infected fish are a possible vector for VHSV IVb, and that the fish rhabdoviruses may have a broader host range than previously suspected.

• Detection and prevalence of the nonsyncytial American grass carp reovirus Aquareovirus G by quantitative reverse transcriptase polymerase chain reaction.

  Authors: Andrew E Goodwin, Gwenn E Merry, Houssam Attoui

  Journal of aquatic animal health. 03/2010; 22(1):8-13.

  The American grass carp reovirus (AGCRV) Aquareovirus G is not strongly associated with disease in fish, but it is often detected by cell culture during routine inspections of healthy fish. TheThe American grass carp reovirus (AGCRV) Aquareovirus G is not strongly associated with disease in fish, but it is often detected by cell culture during routine inspections of healthy fish. The cytopathic effect of AGCRV does not involve the typical syncytia associated with most aquareoviruses. Instead, the AGCRV produces a pattern of cell rounding that is very similar to that produced by rhabdoviruses, including those that are highly regulated. We have developed a quantitative polymerase chain reaction assay that can be used to identify AGCRV in cell cultures or directly on fish tissues. The assay detects as few as two copies of the plasmid template, has a coefficient of variation of 15% among assays performed on different days, and does not cross-react with any other aquareoviruses tested. Assays performed on tissues of cultured golden shiners Notemigonus crysoleucas and fathead minnow Pimephales promelas revealed a high prevalence of infection among healthy fish but no association with disease.

• The effects of largemouth bass virus on a quality largemouth bass population in arkansas.

  Authors: J Wesley Neal, Michael A Eggleton, Andrew E Goodwin

  Journal of wildlife diseases. 08/2009; 45(3):766-71.

  A 22.4-ha impoundment experienced an outbreak of Largemouth bass (Micropterus salmoides) virus (LMBV) disease in the summer of 2006. All dead or dying largemouth bass observed throughout the entireA 22.4-ha impoundment experienced an outbreak of Largemouth bass (Micropterus salmoides) virus (LMBV) disease in the summer of 2006. All dead or dying largemouth bass observed throughout the entire event were recorded and removed. In this study, we estimated mortality and examined size distribution, condition, and biomass following the outbreak. Boat-mounted electrofishing was used to collect largemouth bass for a mark-recapture population estimate and other population metrics. Fish samples were examined for evidence of LMBV, other infectious diseases, and physical abnormalities. Cell cultures inoculated with samples from moribund fish developed cytopathic effects typical of LMBV, and polymerase chain reaction (PCR) confirmed the presence of LMBV. The total number (N+/-95% confidence interval) of stock-size largemouth bass remaining was estimated to be 2,301+/-528 fish (103 bass/ha). The total observed mortality, including dead and dying individuals, during the LMBV outbreak was 176 largemouth bass (7% of the initial population). The total biomass remaining was estimated at 1,592 kg of stock-size bass and a relative biomass of 71.5 kg of stock-size largemouth bass per hectare. Largemouth bass size structure was dominated by quality and preferred (300-510 mm) size classes, with very few memorable-size or larger (>510 mm) fish, and the relative weight of largemouth bass was unusually variable. These results demonstrate that largemouth bass abundance and biomass in the reservoir remained very high despite mortalities attributed to a LMBV outbreak.

• Are all koi ulcer cases associated with infection by atypical Aeromonas salmonicida? Polymerase chain reaction assays of koi carp skin swabs submitted by hobbyists.

  Authors: Andrew E Goodwin, Gwenn E Merry

  Journal of aquatic animal health. 06/2009; 21(2):98-103.

  Infection by atypical Aeromonas salmonicida is regarded as the cause of ulcer disease (KUD) in koi carp Cyprinus carpio and goldfish Carassius auratus. However, other causes--including parasites,Infection by atypical Aeromonas salmonicida is regarded as the cause of ulcer disease (KUD) in koi carp Cyprinus carpio and goldfish Carassius auratus. However, other causes--including parasites, viral infection, and fungi--have been proposed. In our diagnostic work, we often fail to isolate A. salmonicida even when clear clinical signs of KUD are present. This failure may be because these fastidious and slow-growing bacteria are difficult to isolate in culture or because the bacteria are not actually present in the lesions. In this study, we used polymerase chain reaction (PCR) to detect A. salmonicida in DNA samples swabbed from koi carp ulcers. These alcohol-preserved samples were collected and submitted by hobbyists and included 40 separate cases from 12 different states. We identified atypical A. salmonicida by PCR in 52 of 62 samples submitted and in 33 of 40 unique cases. The negative findings for A. salmonicida by PCR could all be attributed to high water temperatures, prior antibiotic use, poor sample quality, or misdiagnosis of columnaris disease as KUD. Tests for Aphanomyces invadans by PCR were negative in every case. This work confirms that A. salmonicda is still the predominant cause of KUD and that our negative culture results were most likely due to technical failures rather than an absence of A. salmonicda in the ulcer lesions.

• Development of a polymerase chain reaction assay to detect cyprinid herpesvirus 2 in goldfish.

  Authors: Thomas B Waltzek, Tomofumi Kurobe, Andrew E Goodwin, Ronald P Hedrick

  Journal of aquatic animal health. 04/2009; 21(1):60-7.

  Cyprinid herpesvirus 2 (CyHV2) has been associated with epidemic mortality among cultured populations of goldfish Carassius auratus. As the principal target tissues are hematopoietic cells in theCyprinid herpesvirus 2 (CyHV2) has been associated with epidemic mortality among cultured populations of goldfish Carassius auratus. As the principal target tissues are hematopoietic cells in the kidney and spleen, the disease is designated herpesviral hematopoietic necrosis (HVHN). Originally described from Japan, the virus is present in at least five other countries and probably has a global distribution in goldfish. Preventing the further spread of the virus via control programs that exploit sensitive viral detection methods is critical. We developed a conventional polymerase chain reaction (PCR) test based on unique sequences found in the putative helicase gene of CyHV2 and completed initial steps toward the validation of this test. The helicase CyHV2 PCR has an analytic sensitivity of at least 78 copies of the target sequence per reaction in serially diluted plasmid and 84 copies/microg of DNA from the kidney and spleen of goldfish experimentally infected with CyHV2. The analytic specificity of the helicase CyHV2 PCR was demonstrated by the lack of amplification of genomic DNA from cyprinid herpesvirus 1, cyprinid herpesvirus 3, and ictalurid herpesvirus 1 (IcHV1). The helicase CyHV2 PCR effectively detected DNA from CyHV2 from goldfish over a broad geographic range, including Japan, California, Ohio, and Pennsylvania. The performance of the helicase CyHV2 PCR was compared with that of the previously described real-time TaqMan PCR for CyHV2 on a set of 37 samples of DNA from goldfish after experimental or natural exposure to CyHV2. The two tests had very strong agreement (kappa coefficient = 0.907) in classifying fish as positive or negative for CyHV2. The helicase CyHV2 PCR therefore complements the real-time PCR test as a conventional diagnostic method for preventing the further spread of CyHV2.

• Complete characterisation of the American grass carp reovirus genome (genus Aquareovirus: family Reoviridae) reveals an evolutionary link between aquareoviruses and coltiviruses.

  Authors: Fauziah Mohd Jaafar, Andrew E Goodwin, Mourad Belhouchet, Gwenn Merry, Qin Fang, Jean-François Cantaloube, Philippe Biagini, Philippe de Micco, Peter P C Mertens, Houssam Attoui

  Virology. 04/2008; 373(2):310-21.

  An aquareovirus was isolated from several fish species in the USA (including healthy golden shiners) that is not closely related to members of species Aquareovirus A, B and C. The virus, which isAn aquareovirus was isolated from several fish species in the USA (including healthy golden shiners) that is not closely related to members of species Aquareovirus A, B and C. The virus, which is atypical (does not cause syncytia in cell cultures at neutral pH), was implicated in a winter die-off of grass carp fingerlings and has therefore been called 'American grass carp reovirus' (AGCRV). Complete nucleotide sequence analysis of the AGCRV genome and comparisons to the other aquareoviruses showed that it is closely related to golden ide reovirus (GIRV) (>92% amino acid [aa] identity in VP5(NTPase) and VP2(Pol)). However, comparisons with grass carp reovirus (Aquareovirus C) and chum salmon reovirus (Aquareovirus A) showed only 22% to 76% aa identity in different viral proteins. These findings have formed the basis for the recognition of AGCRV and GIRV as members of a new Aquareovirus species 'Aquareovirus G' by ICTV. Further sequence comparisons to other members of the family Reoviridae suggest that there has been an 'evolutionary jump,' involving a change in the number of genome segments, between the aquareoviruses (11 segments) and coltiviruses (12 segments). Segment 7 of AGRCV encodes two proteins, from two distinct ORFs, which are homologues of two Coltivirus proteins encoded by genome segments 9 and 12. A similar model has previously been reported for the rotaviruses and seadornaviruses.

• Vertical transmission of Ovipleistophora ovariae (microspora) within the eggs of the golden shiner.

  Authors: Nicholas B D Phelps, Andrew E Goodwin

  Journal of aquatic animal health. 04/2008; 20(1):45-53.

  Fertilized eggs collected from broodfish infected by Ovipleistophora ovariae were tested by quantitative polymerase chain reaction (PCR) and found to be positive for the O. ovariae genome at 7.77 XFertilized eggs collected from broodfish infected by Ovipleistophora ovariae were tested by quantitative polymerase chain reaction (PCR) and found to be positive for the O. ovariae genome at 7.77 X 10(2) to 3.26 x 10(7) copies per microgram of host DNA. Fry hatched from these eggs contained from 1.37 X 10(2) to 9.89 X 10(6) copies of the O. ovariae genome per microgram of host DNA. Surface treatments of fertilized eggs with 150 mg formalin/L (used by farms as a fungicide) or a 1.5% solution of sodium sulfite (which removes the adhesive egg matrix) did not reduce vertical transmission to fry. Treatment of eggs with a 10% solution of bleach or a proprietary commercial DNA denaturant did not reduce the number of egg-associated copies of the O. ovariae genome. Histology of ovaries of infected fish demonstrated spores within the oocytes. However, no spores were observed by histology in positive fry hatched from infected eggs. The PCR and histological demonstration of the presence of O. ovariae spores in oocytes and fry, and the failure of strong DNA denaturants to reduce egg-associated copies, give evidence that O. ovariae is vertically transmitted within eggs.

• Validation of a quantitative PCR diagnostic method for detection of the microsporidian Ovipleistophora ovariae in the cyprinid fish Notemigonus crysoleucas.

  Authors: Nicholas B D Phelps, Andrew E Goodwin

  Diseases of aquatic organisms. 08/2007; 76(3):215-21.

  Microsporidian parasites are easily detected by light microscopy when infections are heavy and spores are present. However, early infections without spores, or light infections with low numbers ofMicrosporidian parasites are easily detected by light microscopy when infections are heavy and spores are present. However, early infections without spores, or light infections with low numbers of spores, are easily missed. This limitation has made it difficult to conduct investigations into microsporidian prevalence and transmission. In this study, we developed a quantitative TaqMan polymerase chain reaction assay to assess the presence of Ovipleistophora ovariae in the tissues of the cyprinid fish Notemigonus crysoleucas (golden shiner). The efficiency of the primer set was 100.8%, with a correlation coefficient of threshold position to copy number of 0.997 over 9 logs using a plasmid containing the cloned reaction product. No product was produced from other closely related microsporidian species (Nucleospora salmonis, Pseudoloma neurophila, Glugea stephani, Heterosporis sp., and O. mirandella). The coefficient of variation for replicate assays done on different days was 12.4%. The assay detects O. ovariae reliably at less than 10 genomic copies and 0.14 spores per reaction, but maximum sensitivity is only achieved when sonication is included as part of the DNA purification step. Using the assay, we found 4.44 x 10(1) to 7.91 x 10(6) copies microg(-1) host DNA in female golden shiners, with the spore density increasing during the spawning season. The parasite was also detected for the first time in the testes of male golden shiners at 2.60 x 10(1) to 8.62 x 102 copies microg(-1) host DNA.

• Complete characterisation of the American grass carp reovirus genome (genus Aquareovirus: family Reoviridae) reveals an evolutionary link between aquareoviruses and coltiviruses

  Authors: Fauziah Mohd Jaafar, Andrew E. Goodwin, Mourad Belhouchet, Gwenn Merry, Qin Fang, Jean-François Cantaloube, Philippe Biagini, Philippe de Micco, Peter P.C. Mertens, Houssam Attoui

  Virology.

  An aquareovirus was isolated from several fish species in the USA (including healthy golden shiners) that is not closely related to members of species Aquareovirus A, B and C. The virus, which isAn aquareovirus was isolated from several fish species in the USA (including healthy golden shiners) that is not closely related to members of species Aquareovirus A, B and C. The virus, which is atypical (does not cause syncytia in cell cultures at neutral pH), was implicated in a winter die-off of grass carp fingerlings and has therefore been called ‘American grass carp reovirus’ (AGCRV). Complete nucleotide sequence analysis of the AGCRV genome and comparisons to the other aquareoviruses showed that it is closely related to golden ide reovirus (GIRV) (> 92% amino acid [aa] identity in VP5(NTPase) and VP2(Pol)). However, comparisons with grass carp reovirus (Aquareovirus C) and chum salmon reovirus (Aquareovirus A) showed only 22% to 76% aa identity in different viral proteins. These findings have formed the basis for the recognition of AGCRV and GIRV as members of a new Aquareovirus species ‘Aquareovirus G’ by ICTV.Further sequence comparisons to other members of the family Reoviridae suggest that there has been an ‘evolutionary jump,’ involving a change in the number of genome segments, between the aquareoviruses (11 segments) and coltiviruses (12 segments). Segment 7 of AGRCV encodes two proteins, from two distinct ORFs, which are homologues of two Coltivirus proteins encoded by genome segments 9 and 12. A similar model has previously been reported for the rotaviruses and seadornaviruses.