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Bennie Osburn - One of the best experts on this subject based on the ideXlab platform.
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a multiplex pcr for simultaneous detection and differentiation of north american serotypes of bluetongue and epizootic hemorrhagic disease viruses
Comparative Immunology Microbiology and Infectious Diseases, 2003Co-Authors: Imadeldin E Aradaib, Bennie Osburn, Wayne L Smith, James S CullorAbstract:Abstract In the present study, a multiplex RT-PCR-based assay for simultaneous detection and differentiation of North American serotypes of bluetongue (BT) virus (BTV) and epizootic hemorrhagic disease (EHD) virus (EHDV) in cell culture and clinical samples was developed. Two pairs of primers (B1 and B4) and (E1 and E4) were designed to hybridize to non-structural protein 1 (NS1) genomes of (BTV-11) and (EHDV-1), respectively. The multiplex PCR-based assay utilized a single tube-PCR amplification in which EHDV and BTV primers were used simultaneously in a multiplex format. The BTV primers generated a 790 base pair (bp) specific PCR product from RNA samples of North American BTV serotypes 2, 10, 11, 13 and 17; whereas EHDV serotypes 1 and 2 or total nucleic acid extract from non-infected baby hamster kidney (BHK) cells failed to demonstrate the 790 bp specific BTV PCR product. Likewise, the EHDV primers produced a 387 bp specific PCR product from RNA samples of EHDV serotypes 1 and 2, but not from BTV serotypes 2, 10, 11, 13, 17 or from total nucleic acid extract of BHK cell controls. Two pairs of nested primers (B2 and B3) and (E2 and E3), internal to the annealing sites of primers (B1and B4) and primers (E1 and E4), produced a 520 bp specific BTV and a 224 bp specific EHDV PCR product from BTV and EHDV first amplification products, respectively. These nested amplifications increased the sensitivity of the PCR assay and confirmed the specificity of the first amplified EHDV or BTV PCR products. The described multiplex RT-PCR-based assay could be used to facilitate rapid detection and differentiation of North American BTV and EHDV serotypes and to provide a valuable tool to study the epidemiology of these orbivirus infections in susceptible animal populations.
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evaluation of rt pcr for detection of sudanese serotypes of epizootic hemorrhagic disease virus serogroup
Veterinarski Arhiv, 2002Co-Authors: Mohammed Abdalla, Imadeldin E Aradaib, Bennie OsburnAbstract:Epizootic hemorrhagic disease (EHD) is an infectious non-contagious disease of deer and cattle. At least 2 serotypes of EHD virus, designated EHDV-4 and EHDV-318, are enzootic in the Sudan. To facilitate clinical disease investigation and control of the disease a rapid diagnostic assay is urgently needed. A reverse transcriptase (RT) polymerase chain reaction (RT-PCR) protocol, previously reported for detection of the United States EHDV serotypes 1 and 2 ribonucleic acid (RNA) in cell culture and clinical specimens, was evaluated for detection of the Sudanese EHDV serotypes. RNA from Sudanese isolates of EHDV-4 and EHDV-318, propagated in cell cultures, were detected by the described RT-PCR-based assay. The specific 387 bp PCR products were visualized on ethidium bromide-stained agarose gel. Amplification product was not detected when the EHDV RT-PCR-based assay was applied to RNA from Sudanese bluetongue virus (BTV) serotype 4 (BTV-4) or total nucleic acid extracts from uninfected BHK-21 cells. The scientific observations reported in this paper indicated that the previously described EHDV RT-PCR assay could be applied for detection of EHDV infection among the Sudanese susceptible animal populations.
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pcr detection of north american and central african isolates of epizootic hemorrhagic disease virus ehdv based on genome segment 10 of ehdv serotype 1
Journal of Clinical Microbiology, 1998Co-Authors: Imadeldin E Aradaib, William C Wilson, C E Schore, Mohammed E H Mohammed, Tilahun Yilma, James S Cullor, Bennie OsburnAbstract:PCR amplification technology for the detection of epizootic hemorrhagic disease virus (EHDV) ribonucleic acid in cell culture and clinical specimens was developed. With oligoribonucleotide primers selected from genome segment 10 of EHDV serotype 1 (EHDV-1), which codes for two nonstructural proteins (NS3 and NS3a), the PCR-based assay resulted in a 535-bp PCR product. RNAs from North American EHDV-1 prototype, EHDV-2 prototype, and a number of EHDV field isolates, including the Central African isolates of EHDV-5 and EHDV-318 propagated in cell cultures, were detected by this PCR-based assay. The specific 535-bp PCR products were visualized onto agarose gels, and the identity of the PCR products was confirmed by chemiluminescent hybridization with a 352-bp internal probe. The sensitivity of the EHDV PCR assay was increased by chemiluminescent hybridization; by this EHDV-NS3 PCR, 10 fg of EHDV RNA was detected (equivalent to 600 viral particles). Amplification product was not detected when the PCR-based assay was applied to RNAs from North American bluetongue virus prototype serotypes 2, 10, 11, 13, and 17; total nucleic acid extracts from uninfected BHK-21 cells; or unfractionated blood from calves and deer that were EHDV seronegative and virus isolation negative. The described EHDV PCR-based assay with primers derived from segment 10 of EHDV-1 resulted in detection of EHDV RNA from blood and tissues collected from calves and deer with natural and experimental EHDV infections and provides a valuable tool to study the epidemiology of EHDV infection in susceptible ruminants.
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serogrouping and topotyping of sudanese and united states strains of epizootic hemorrhagic disease virus using pcr
Comparative Immunology Microbiology and Infectious Diseases, 1997Co-Authors: Imadeldin E Aradaib, Mohammed E H Mohammed, Muawia Mohamed Mukhtar, H W Ghalib, Bennie OsburnAbstract:The potential use of the recently reported polymerase chain reaction (PCR) protocol for detection of United States epizootic hemorrhagic disease virus (EHDV) serotype 1 (EHDV-1) and serotype 2 (EHDV-2) ribonucleic acid in cell culture and clinical specimens was evaluated for detection of Sudanese EHDV strains. EHDV serotype 5 (EHDV-5) and EHDV, isolate 318 (untyped) designated (EHDV-318), recovered from sentinel calves at the Khartoum University farm (Sudan) were studied. RNA from EHDV-5 and EHDV-318 and a number of EHDV field isolates, propagated in cell cultures, were detected by the described PCR-based assay. The specific 387 bp PCR products were visualized on ethidium-bromide stained agarose gel. Specificity of the PCR products was confirmed by chemiluminescent hybridization with non-radiolabeled internal probe. Amplification product was not detected when the PCR-based assay was applied to RNA from blutongue virus (BTV) prototypes serotypes 2, 10, 11, 13, 16 and 17; total nucleic acid extracts from uninfected BHK-21 cells. The results of this study indicated that the previously described EHDV-PCR assay could be applied for detection of Sudanese as well as United States strains of EHDV serogroup. In addition, the described EHDV-PCR assay could be used as a supportive diagnostic assay to the current conventional virus isolation procedures used for detection of EHDV infection in susceptible ruminants.
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interaction of epizootic hemorrhagic disease virus with bovine erythrocytes in vitro electron microscope study
Comparative Immunology Microbiology and Infectious Diseases, 1997Co-Authors: Imadeldin E Aradaib, A W Brewer, Bennie OsburnAbstract:The interaction of epizootic hemorrhagic disease virus (EHDV) with bovine erythrocytes in t'itro has been studied, and the results are presented herein. EHDV, a member of the Orbit, irus genus in the family Reoviridae, causes fatal hemorrhagic disease in North American white-tailed deer (Odocoileus virginianus). Other captive and free ranging ruminants may also be infected with the disease [1, 4]. At least 10 serotypes of EHDV are recognised worldwide, but serotypes 1 and 2 are enzootic in the United States. In white-tailed deer, viremia is short and the disease usually follows a peracute course leading to death. In cattle, EHDV infection is common but typically asymptomatic, and viremia in EHDV infected cattle is usually short. However, infection of European breed of cattle with laboratory adapted strain of EHDV induced viremia which persisted for up to 50 days [3], suggesting that cattle may play a role in the epidemiology of the disease [2]. During the later course of EHDV-infection, the virus is principally associated with erythrocytes. Erythrocyte-associated viremia has been reported with a number of orbiviruses including bluetongue virus (BTV); however, there is no information available on the interaction of EHDV with bovine erythrocytes. The mechanism involved in prolonged viremia in EHDV-infected cattle, despite the presence of circulating neutralising antibodies, has yet to be explained. A plaque-purified EHDV serotype 1 (New Jersey strain) was used to inoculate suspended bovine erythrocytes, obtained from 4-month-old calves, at a multiplicity of infection (MOI) of 1, and the inoculated erythrocytes were incubated at 37~>C for 1 h. Uninoculated erythrocytes were used as controls. The details for collection of blood, preparation of erythrocytes, preparation of sections for transmission electron microscope were described previously [5]. EHDV particles were commonly observed in close association with erythrocyte cell membrane and were typically present in invaginations (Fig. 1). The viral particles measure about 50 70 nm in diameter and were recognized as EHDV particles by their morphological appearance. The viral particles were not observed in the erythrocytes from controls. The results of this study indicated that EHDV becomes associated with the erythrocytic cell membrane soon after in vitro infection of bovine erythrocytes, with the virus particles becoming sequestered in invaginations of the cell membrane. The infection of erythrocytes did not progress beyond the cell membrane invagination, as free virons
David E Stallknecht - One of the best experts on this subject based on the ideXlab platform.
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the first 10 years 2006 15 of epizootic hemorrhagic disease virus serotype 6 in the usa
Journal of Wildlife Diseases, 2017Co-Authors: Mark G Ruder, Andrew B. Allison, Donna J Johnson, Eileen N Ostlund, Clara Kienzle, Jamie E Phillips, Rebecca L Poulson, David E StallknechtAbstract:Abstract Epizootic hemorrhagic disease virus (EHDV) is a Culicoides biting midge–transmitted orbivirus (family Reoviridae) of wild and domestic ruminants and is an important pathogen of white-tailed deer (Odocoileus virginianus). Historically, only two serotypes, EHDV-1 and EHDV-2, have been known to be endemic in the US. However, in 2006, an exotic serotype (EHDV-6) was first detected in the US by a long-term passive surveillance system for EHDV and bluetongue viruses. Here we report EHDV-6 detections made through these passive surveillance efforts by the Southeastern Cooperative Wildlife Disease Study (University of Georgia, Athens, Georgia, USA) and the National Veterinary Services Laboratories (US Department of Agriculture, Ames, Iowa, USA) over a 10-yr period (2006–15). The results demonstrated that EHDV-6 was detected from ruminants every year since 2006 and was widespread in the central and eastern US, providing evidence that EHDV-6 is likely now established in the US.
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genetic characterization of epizootic hemorrhagic disease virus strains isolated from cattle in israel
Journal of General Virology, 2015Co-Authors: William C Wilson, Mark G Ruder, David E Stallknecht, Eyal Klement, Daniel G Mead, Hagai Yadin, Dane C Jasperson, Elizabeth W HowerthAbstract:Epizootic hemorrhagic disease virus (EHDV), a member of the genus Orbivirus not reported previously in Israel, was isolated from Israeli cattle during a ‘bluetongue-like’ disease outbreak in 2006. To ascertain the origin of this new virus, three isolates from the outbreak were fully sequenced and compared with available sequences. Whilst the L2 gene segment clustered with the Australian EHDV serotype 7 (EHDV-7) reference strain, most of the other segments were clustered with EHDV isolates of African/Middle East origin, specifically Bahrain, Nigeria and South Africa. The M6 gene had genetic relatedness to the Australian/Asian strains, but with the limited data available the significance of this relationship is unclear. Only one EHDV-7 L2 sequence was available, and as this gene encodes the serotype-specific epitope, the relationship of these EHDV-7 L2 genes to an Australian EHDV-7 reflects the serotype association, not necessarily the origin. The genetic data indicated that the strains affecting Israel in 2006 may have been related to similar outbreaks that occurred in North Africa in the same year. This finding also supports the hypothesis that EHDV entered Israel during 2006 and was not present there before this outbreak.
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vector competence of culicoides sonorensis diptera ceratopogonidae to epizootic hemorrhagic disease virus serotype 7
Parasites & Vectors, 2012Co-Authors: Andrew B. Allison, Mark G Ruder, David E Stallknecht, Eyal Klement, Elizabeth W Howerth, Barbara S Drolet, Daniel G MeadAbstract:Background: Culicoides sonorensis (Diptera: Ceratopogonidae) is a vector of epizootic hemorrhagic disease virus (EHDV) serotypes 1 and 2 in North America, where these viruses are well-known pathogens of white-tailed deer (WTD) and other wild ruminants. Although historically rare, reports of clinical EHDV infection in cattle have increased in some parts of the world over the past decade. In 2006, an EHDV-7 epizootic in cattle resulted in economic loss for the Israeli dairy industry. White-tailed deer are susceptible to EHDV-7 infection and disease; however, this serotype is exotic to the US and the susceptibility of C. sonorensis to this cattle-virulent EHDV is not known. The objective of the study was to determine if C. sonorensis is susceptible to EHDV-7 infection and is a competent vector.
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detection of a novel reassortant epizootic hemorrhagic disease virus ehdv in the usa containing rna segments derived from both exotic ehdv 6 and endemic ehdv 2 serotypes
Journal of General Virology, 2010Co-Authors: Andrew B. Allison, William C Wilson, Virginia H Goekjian, A C Potgieter, Donna J Johnson, Peter P C Mertens, David E StallknechtAbstract:Epizootic hemorrhagic disease virus (EHDV) is a Culicoides-transmitted orbivirus that infects domestic and wild ruminants and is provisionally thought to be distributed throughout Africa, North America, Australia, East Asia and the Middle East. Historically, of the seven proposed serotypes of EHDV, only EHDV-1 and EHDV-2 have been reported from North America. In 2006, EHDV isolates were recovered from moribund or dead white-tailed deer (Odocoileus virginianus) in Indiana and Illinois that could not be identified as either EHDV-1 or EHDV-2 by virus neutralization tests or by serotype-specific RT-PCR. Additional serological and genetic testing identified the isolates as EHDV-6, a serotype that, although originally described from Australia, has recently been recognized as an emerging pathogen of cattle in Morocco, Algeria and Turkey. In 2007 and 2008, EHDV-6 was isolated again from white-tailed deer, this time in Missouri, Kansas and Texas, suggesting that the virus is capable of overwintering and that it may become, or already is, endemic in a geographically widespread region of the USA. Genetic characterization of the virus indicates that it is a reassortant, such that the outer capsid proteins determining serotype specificity (VP2 and VP5) are derived from exotic EHDV-6, whilst the remaining structural and non-structural proteins are apparently obtained from indigenous EHDV-2 (Alberta).
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detection of all eight serotypes of epizootic hemorrhagic disease virus by real time reverse transcription polymerase chain reaction
Journal of Veterinary Diagnostic Investigation, 2009Co-Authors: William C Wilson, David E Stallknecht, Daniel G Mead, Emily S Ohearn, Christian Tellgrenroth, James O MechamAbstract:Epizootic hemorrhagic disease virus (EHDV) has been associated with bluetongue-like disease in cattle. Although U.S. EHDV strains have not been experimentally proven to cause disease in cattle, there is serologic evidence of infection. Differentiation of Bluetongue virus (BTV) and EHDV is necessary because diagnosis of infection caused by these viruses is often confused. The previously developed nested reverse transcription polymerase chain reaction (nRT-PCR) test for indigenous EHDV disease is sensitive and specific, but it is prone to contamination problems. Additionally, the EHDV nRT-PCR only detects 7 of the 8 serotypes. To develop an improved diagnostic test, sequence analysis was performed on 2 conserved target genes; one is highly expressed in infected mammalian cells, whereas the other is highly expressed in infected insect cells. This information was used to develop a rapid EHDV real-time PCR that detects all 8 EHDV serotypes. The EHDV assay did not cross-react with BTV strains and performed similarly to the nRT-PCR tests with archived clinical samples. In addition, it is superior to the nRT-PCR, not only because it is a closed system with fewer cross-contamination problems, but also because it detects all 8 serotypes and is less labor and time intensive.
Imadeldin E Aradaib - One of the best experts on this subject based on the ideXlab platform.
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a multiplex pcr for simultaneous detection and differentiation of north american serotypes of bluetongue and epizootic hemorrhagic disease viruses
Comparative Immunology Microbiology and Infectious Diseases, 2003Co-Authors: Imadeldin E Aradaib, Bennie Osburn, Wayne L Smith, James S CullorAbstract:Abstract In the present study, a multiplex RT-PCR-based assay for simultaneous detection and differentiation of North American serotypes of bluetongue (BT) virus (BTV) and epizootic hemorrhagic disease (EHD) virus (EHDV) in cell culture and clinical samples was developed. Two pairs of primers (B1 and B4) and (E1 and E4) were designed to hybridize to non-structural protein 1 (NS1) genomes of (BTV-11) and (EHDV-1), respectively. The multiplex PCR-based assay utilized a single tube-PCR amplification in which EHDV and BTV primers were used simultaneously in a multiplex format. The BTV primers generated a 790 base pair (bp) specific PCR product from RNA samples of North American BTV serotypes 2, 10, 11, 13 and 17; whereas EHDV serotypes 1 and 2 or total nucleic acid extract from non-infected baby hamster kidney (BHK) cells failed to demonstrate the 790 bp specific BTV PCR product. Likewise, the EHDV primers produced a 387 bp specific PCR product from RNA samples of EHDV serotypes 1 and 2, but not from BTV serotypes 2, 10, 11, 13, 17 or from total nucleic acid extract of BHK cell controls. Two pairs of nested primers (B2 and B3) and (E2 and E3), internal to the annealing sites of primers (B1and B4) and primers (E1 and E4), produced a 520 bp specific BTV and a 224 bp specific EHDV PCR product from BTV and EHDV first amplification products, respectively. These nested amplifications increased the sensitivity of the PCR assay and confirmed the specificity of the first amplified EHDV or BTV PCR products. The described multiplex RT-PCR-based assay could be used to facilitate rapid detection and differentiation of North American BTV and EHDV serotypes and to provide a valuable tool to study the epidemiology of these orbivirus infections in susceptible animal populations.
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detection of united states orbivirus serogroup using a multiplex rt pcr
Veterinarski Arhiv, 2003Co-Authors: Imadeldin E Aradaib, Mohammed Abdalla, Abdelrahim E Karrar, B I Osburn, M A AbdallaAbstract:A multiplex RT-PCR assay, for simultaneous detection and differentiation of United States serogroup of Orbiviruses, including bluetongue virus (BTV) and epizootic hemorrhagic disease virus (EHDV) in cell culture, was developed. Sets of primers were designed to hybridize to genome segment six of EHDV-2 and to genome segment 10 of BTV-10. The RT-PCR assay utilized a single-tube PCR amplification in which EHDV and BTV primers were used simultaneously in a multiplex format. The EHDV primers produced a 387 base pair (bp) specific PCR product from RNA samples of cell culture-adapted EHDV serotypes 1 and 2, but not from BTV serotypes 2, 10, 11, 13, 17; or from total nucleic acid extract of baby hamster kidney (BHK) cells controls. Likewise, the BTV primers generated a 251-bp amplicon from RNA samples of BTV serotypes 2, 10, 11, 13, and 17, whereas EHDV-1 and EHDV-2; and BHK21 cells total nucleic acid extract failed to demonstrate the 251-bp specific BTV PCR product. EHDV and BTV PCR amplification products were easily identified on the basis of size differences on ethidium bromide-stained agarose gels. This multiplex RT-PCR assay provides supportive diagnostic method for rapid detection of BTV and/or EHDV-infections among susceptible ruminants.
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evaluation of rt pcr for detection of sudanese serotypes of epizootic hemorrhagic disease virus serogroup
Veterinarski Arhiv, 2002Co-Authors: Mohammed Abdalla, Imadeldin E Aradaib, Bennie OsburnAbstract:Epizootic hemorrhagic disease (EHD) is an infectious non-contagious disease of deer and cattle. At least 2 serotypes of EHD virus, designated EHDV-4 and EHDV-318, are enzootic in the Sudan. To facilitate clinical disease investigation and control of the disease a rapid diagnostic assay is urgently needed. A reverse transcriptase (RT) polymerase chain reaction (RT-PCR) protocol, previously reported for detection of the United States EHDV serotypes 1 and 2 ribonucleic acid (RNA) in cell culture and clinical specimens, was evaluated for detection of the Sudanese EHDV serotypes. RNA from Sudanese isolates of EHDV-4 and EHDV-318, propagated in cell cultures, were detected by the described RT-PCR-based assay. The specific 387 bp PCR products were visualized on ethidium bromide-stained agarose gel. Amplification product was not detected when the EHDV RT-PCR-based assay was applied to RNA from Sudanese bluetongue virus (BTV) serotype 4 (BTV-4) or total nucleic acid extracts from uninfected BHK-21 cells. The scientific observations reported in this paper indicated that the previously described EHDV RT-PCR assay could be applied for detection of EHDV infection among the Sudanese susceptible animal populations.
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pcr detection of north american and central african isolates of epizootic hemorrhagic disease virus ehdv based on genome segment 10 of ehdv serotype 1
Journal of Clinical Microbiology, 1998Co-Authors: Imadeldin E Aradaib, William C Wilson, C E Schore, Mohammed E H Mohammed, Tilahun Yilma, James S Cullor, Bennie OsburnAbstract:PCR amplification technology for the detection of epizootic hemorrhagic disease virus (EHDV) ribonucleic acid in cell culture and clinical specimens was developed. With oligoribonucleotide primers selected from genome segment 10 of EHDV serotype 1 (EHDV-1), which codes for two nonstructural proteins (NS3 and NS3a), the PCR-based assay resulted in a 535-bp PCR product. RNAs from North American EHDV-1 prototype, EHDV-2 prototype, and a number of EHDV field isolates, including the Central African isolates of EHDV-5 and EHDV-318 propagated in cell cultures, were detected by this PCR-based assay. The specific 535-bp PCR products were visualized onto agarose gels, and the identity of the PCR products was confirmed by chemiluminescent hybridization with a 352-bp internal probe. The sensitivity of the EHDV PCR assay was increased by chemiluminescent hybridization; by this EHDV-NS3 PCR, 10 fg of EHDV RNA was detected (equivalent to 600 viral particles). Amplification product was not detected when the PCR-based assay was applied to RNAs from North American bluetongue virus prototype serotypes 2, 10, 11, 13, and 17; total nucleic acid extracts from uninfected BHK-21 cells; or unfractionated blood from calves and deer that were EHDV seronegative and virus isolation negative. The described EHDV PCR-based assay with primers derived from segment 10 of EHDV-1 resulted in detection of EHDV RNA from blood and tissues collected from calves and deer with natural and experimental EHDV infections and provides a valuable tool to study the epidemiology of EHDV infection in susceptible ruminants.
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serogrouping and topotyping of sudanese and united states strains of epizootic hemorrhagic disease virus using pcr
Comparative Immunology Microbiology and Infectious Diseases, 1997Co-Authors: Imadeldin E Aradaib, Mohammed E H Mohammed, Muawia Mohamed Mukhtar, H W Ghalib, Bennie OsburnAbstract:The potential use of the recently reported polymerase chain reaction (PCR) protocol for detection of United States epizootic hemorrhagic disease virus (EHDV) serotype 1 (EHDV-1) and serotype 2 (EHDV-2) ribonucleic acid in cell culture and clinical specimens was evaluated for detection of Sudanese EHDV strains. EHDV serotype 5 (EHDV-5) and EHDV, isolate 318 (untyped) designated (EHDV-318), recovered from sentinel calves at the Khartoum University farm (Sudan) were studied. RNA from EHDV-5 and EHDV-318 and a number of EHDV field isolates, propagated in cell cultures, were detected by the described PCR-based assay. The specific 387 bp PCR products were visualized on ethidium-bromide stained agarose gel. Specificity of the PCR products was confirmed by chemiluminescent hybridization with non-radiolabeled internal probe. Amplification product was not detected when the PCR-based assay was applied to RNA from blutongue virus (BTV) prototypes serotypes 2, 10, 11, 13, 16 and 17; total nucleic acid extracts from uninfected BHK-21 cells. The results of this study indicated that the previously described EHDV-PCR assay could be applied for detection of Sudanese as well as United States strains of EHDV serogroup. In addition, the described EHDV-PCR assay could be used as a supportive diagnostic assay to the current conventional virus isolation procedures used for detection of EHDV infection in susceptible ruminants.
William C Wilson - One of the best experts on this subject based on the ideXlab platform.
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genetic characterization of epizootic hemorrhagic disease virus strains isolated from cattle in israel
Journal of General Virology, 2015Co-Authors: William C Wilson, Mark G Ruder, David E Stallknecht, Eyal Klement, Daniel G Mead, Hagai Yadin, Dane C Jasperson, Elizabeth W HowerthAbstract:Epizootic hemorrhagic disease virus (EHDV), a member of the genus Orbivirus not reported previously in Israel, was isolated from Israeli cattle during a ‘bluetongue-like’ disease outbreak in 2006. To ascertain the origin of this new virus, three isolates from the outbreak were fully sequenced and compared with available sequences. Whilst the L2 gene segment clustered with the Australian EHDV serotype 7 (EHDV-7) reference strain, most of the other segments were clustered with EHDV isolates of African/Middle East origin, specifically Bahrain, Nigeria and South Africa. The M6 gene had genetic relatedness to the Australian/Asian strains, but with the limited data available the significance of this relationship is unclear. Only one EHDV-7 L2 sequence was available, and as this gene encodes the serotype-specific epitope, the relationship of these EHDV-7 L2 genes to an Australian EHDV-7 reflects the serotype association, not necessarily the origin. The genetic data indicated that the strains affecting Israel in 2006 may have been related to similar outbreaks that occurred in North Africa in the same year. This finding also supports the hypothesis that EHDV entered Israel during 2006 and was not present there before this outbreak.
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detection of a novel reassortant epizootic hemorrhagic disease virus ehdv in the usa containing rna segments derived from both exotic ehdv 6 and endemic ehdv 2 serotypes
Journal of General Virology, 2010Co-Authors: Andrew B. Allison, William C Wilson, Virginia H Goekjian, A C Potgieter, Donna J Johnson, Peter P C Mertens, David E StallknechtAbstract:Epizootic hemorrhagic disease virus (EHDV) is a Culicoides-transmitted orbivirus that infects domestic and wild ruminants and is provisionally thought to be distributed throughout Africa, North America, Australia, East Asia and the Middle East. Historically, of the seven proposed serotypes of EHDV, only EHDV-1 and EHDV-2 have been reported from North America. In 2006, EHDV isolates were recovered from moribund or dead white-tailed deer (Odocoileus virginianus) in Indiana and Illinois that could not be identified as either EHDV-1 or EHDV-2 by virus neutralization tests or by serotype-specific RT-PCR. Additional serological and genetic testing identified the isolates as EHDV-6, a serotype that, although originally described from Australia, has recently been recognized as an emerging pathogen of cattle in Morocco, Algeria and Turkey. In 2007 and 2008, EHDV-6 was isolated again from white-tailed deer, this time in Missouri, Kansas and Texas, suggesting that the virus is capable of overwintering and that it may become, or already is, endemic in a geographically widespread region of the USA. Genetic characterization of the virus indicates that it is a reassortant, such that the outer capsid proteins determining serotype specificity (VP2 and VP5) are derived from exotic EHDV-6, whilst the remaining structural and non-structural proteins are apparently obtained from indigenous EHDV-2 (Alberta).
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detection of all eight serotypes of epizootic hemorrhagic disease virus by real time reverse transcription polymerase chain reaction
Journal of Veterinary Diagnostic Investigation, 2009Co-Authors: William C Wilson, David E Stallknecht, Daniel G Mead, Emily S Ohearn, Christian Tellgrenroth, James O MechamAbstract:Epizootic hemorrhagic disease virus (EHDV) has been associated with bluetongue-like disease in cattle. Although U.S. EHDV strains have not been experimentally proven to cause disease in cattle, there is serologic evidence of infection. Differentiation of Bluetongue virus (BTV) and EHDV is necessary because diagnosis of infection caused by these viruses is often confused. The previously developed nested reverse transcription polymerase chain reaction (nRT-PCR) test for indigenous EHDV disease is sensitive and specific, but it is prone to contamination problems. Additionally, the EHDV nRT-PCR only detects 7 of the 8 serotypes. To develop an improved diagnostic test, sequence analysis was performed on 2 conserved target genes; one is highly expressed in infected mammalian cells, whereas the other is highly expressed in infected insect cells. This information was used to develop a rapid EHDV real-time PCR that detects all 8 EHDV serotypes. The EHDV assay did not cross-react with BTV strains and performed similarly to the nRT-PCR tests with archived clinical samples. In addition, it is superior to the nRT-PCR, not only because it is a closed system with fewer cross-contamination problems, but also because it detects all 8 serotypes and is less labor and time intensive.
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brief research reports detection of all eight serotypes of epizootic hemorrhagic disease virus by real time reverse transcription polymerase chain reaction
2009Co-Authors: William C Wilson, David E Stallknecht, Daniel G Mead, Christian Tellgrenroth, James O MechamAbstract:Epizootic hemorrhagic disease virus (EHDV) has been associated with bluetongue-like disease in cattle. Although U.S. EHDV strains have not been experimentally proven to cause disease in cattle, there is serologic evidence of infection. Differentiation of Bluetongue virus (BTV) and EHDV is necessary because diagnosis of infection caused by these viruses is often confused. The previously developed nested reverse transcription polymerase chain reaction (nRT-PCR) test for indigenous EHDV disease is sensitive and specific, but it is prone to contamination problems. Additionally, the EHDV nRT-PCR only detects 7 of the 8 serotypes. To develop an improved diagnostic test, sequence analysis was performed on 2 conserved target genes; one is highly expressed in infected mammalian cells, whereas the other is highly expressed in infected insect cells. This information was used to develop a rapid EHDV real-time PCR that detects all 8 EHDV serotypes. The EHDV assay did not cross-react with BTV strains and performed similarly to the nRT-PCR tests with archived clinical samples. In addition, it is superior to the nRT-PCR, not only because it is a closed system with fewer cross- contamination problems, but also because it detects all 8 serotypes and is less labor and time intensive.
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the s7 gene and vp7 protein are highly conserved among temporally and geographically distinct american isolates of epizootic hemorrhagic disease virus
Virus Research, 2003Co-Authors: James O Mecham, David E Stallknecht, William C WilsonAbstract:Complete sequences of genome segment 7 (S7) from six isolates of epizootic hemorrhagic disease virus serotype 1 (EHDV-1) and 37 isolates of serotype 2 (EHDV-2) were determined. These isolates were made between 1978 and 2001 from the southeast, mid-Atlantic, Midwest and intermountain United States. Analysis of the S7 sequence similarities showed 98.1% identity among the EHDV-1 isolates and 91.0% identity among the EHDV-2 isolates. Comparison of the deduced amino acid similarities showed an even greater degree of similarity among the isolates (100% among the EHDV-1 isolates and 98.9% identity among the EHDV-2 isolates). There was only 75.8% identity between the EHDV-1 and EHDV-2 isolates at the nucleic acid level; however, there was 93.7% identity between the two groups at the amino acid level. The ratio of non-synonymous to synonymous nucleotide indicates a strong selection for silent substitutions. There was no evidence for reassortment between EHDV-1 and EHDV-2 isolates. The high degree of conservation of S7 gene codons and the VP7 protein, suggests that little variation is allowed in preserving the function of this protein. The high degree of conservation also validates the use of diagnostic tests for EHDV based on S7 and VP7.
Eyal Klement - One of the best experts on this subject based on the ideXlab platform.
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genetic characterization of epizootic hemorrhagic disease virus strains isolated from cattle in israel
Journal of General Virology, 2015Co-Authors: William C Wilson, Mark G Ruder, David E Stallknecht, Eyal Klement, Daniel G Mead, Hagai Yadin, Dane C Jasperson, Elizabeth W HowerthAbstract:Epizootic hemorrhagic disease virus (EHDV), a member of the genus Orbivirus not reported previously in Israel, was isolated from Israeli cattle during a ‘bluetongue-like’ disease outbreak in 2006. To ascertain the origin of this new virus, three isolates from the outbreak were fully sequenced and compared with available sequences. Whilst the L2 gene segment clustered with the Australian EHDV serotype 7 (EHDV-7) reference strain, most of the other segments were clustered with EHDV isolates of African/Middle East origin, specifically Bahrain, Nigeria and South Africa. The M6 gene had genetic relatedness to the Australian/Asian strains, but with the limited data available the significance of this relationship is unclear. Only one EHDV-7 L2 sequence was available, and as this gene encodes the serotype-specific epitope, the relationship of these EHDV-7 L2 genes to an Australian EHDV-7 reflects the serotype association, not necessarily the origin. The genetic data indicated that the strains affecting Israel in 2006 may have been related to similar outbreaks that occurred in North Africa in the same year. This finding also supports the hypothesis that EHDV entered Israel during 2006 and was not present there before this outbreak.
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vector competence of culicoides sonorensis diptera ceratopogonidae to epizootic hemorrhagic disease virus serotype 7
Parasites & Vectors, 2012Co-Authors: Andrew B. Allison, Mark G Ruder, David E Stallknecht, Eyal Klement, Elizabeth W Howerth, Barbara S Drolet, Daniel G MeadAbstract:Background: Culicoides sonorensis (Diptera: Ceratopogonidae) is a vector of epizootic hemorrhagic disease virus (EHDV) serotypes 1 and 2 in North America, where these viruses are well-known pathogens of white-tailed deer (WTD) and other wild ruminants. Although historically rare, reports of clinical EHDV infection in cattle have increased in some parts of the world over the past decade. In 2006, an EHDV-7 epizootic in cattle resulted in economic loss for the Israeli dairy industry. White-tailed deer are susceptible to EHDV-7 infection and disease; however, this serotype is exotic to the US and the susceptibility of C. sonorensis to this cattle-virulent EHDV is not known. The objective of the study was to determine if C. sonorensis is susceptible to EHDV-7 infection and is a competent vector.
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vector competence of culicoides sonorensis diptera ceratopogonidae to epizootic hemorrhagic disease virus serotype 7
Parasites & Vectors, 2012Co-Authors: Eyal KlementAbstract:Culicoides sonorensis (Diptera: Ceratopogonidae) is a vector of epizootic hemorrhagic disease virus (EHDV) serotypes 1 and 2 in North America, where these viruses are well-known pathogens of white-tailed deer (WTD) and other wild ruminants. Although historically rare, reports of clinical EHDV infection in cattle have increased in some parts of the world over the past decade. In 2006, an EHDV-7 epizootic in cattle resulted in economic loss for the Israeli dairy industry. White-tailed deer are susceptible to EHDV-7 infection and disease; however, this serotype is exotic to the US and the susceptibility of C. sonorensis to this cattle-virulent EHDV is not known. The objective of the study was to determine if C. sonorensis is susceptible to EHDV-7 infection and is a competent vector. To evaluate the susceptibility of C. sonorensis, midges were fed on EHDV-7 infected WTD, held at 22 ± 1°C, and processed individually for virus isolation and titration on 4–16 days post feeding (dpf). Midges with a virus titer of ≥102.7 median tissue culture infective doses (TCID50)/midge were considered potentially competent. To determine if infected C. sonorensis were capable of transmitting EHDV-7 to a host, a susceptible WTD was then fed on by a group of 14–16 dpf midges. From 4–16 dpf, 45% (156/350) of midges that fed on WTD with high titer viremia (>107 TCID50/ml) were virus isolation-positive, and starting from 10–16 dpf, 32% (35/109) of these virus isolation-positive midges were potentially competent (≥102.7 TCID50/midge). Midges that fed on infected deer transmitted the virus to a susceptible WTD at 14–16 dpf. The WTD developed viremia and severe clinical disease. This study demonstrates that C. sonorensis is susceptible to EHDV-7 infection and can transmit the virus to susceptible WTD, thus, C. sonorensis should be considered a potential vector of EHDV-7. Together with previous work, this study demonstrates that North America has a susceptible ruminant and vector host for this exotic, cattle-virulent strain of EHDV-7.
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no evidence for involvement of sheep in the epidemiology of cattle virulent epizootic hemorrhagic disease virus
Veterinary Microbiology, 2011Co-Authors: Maor Kedmi, Carrie Batten, S Levi, Nadav Galon, V Bomborov, Hagai Yadin, Eyal KlementAbstract:Abstract Epizootic hemorrhagic disease virus (EHDV) is an Orbivirus. While not previously considered as an important disease in cattle, several EHDV serotypes (EHDV-6 and 7) have recently been implicated in disease outbreaks. The involvement of sheep in the epidemiology of EHDV is still not understood. In this study we compared the prevalence of antibodies to EHDV and bluetongue virus (BTV) in sheep to their prevalence in cattle after an outbreak of EHDV that occurred in Israel during 2006. Sixty-six sheep and lambs scattered in seven herds were compared to 114 cows and calves scattered in 13 dairy cattle herds, matched to the sheep herds by location. While antibody prevalence to EHDV was high in cattle (35.2% within the outbreak zone) no evidence of exposure to EHDV was found in sheep (p