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Sample records for distemper virus detection

  1. Detection of Arctic and European cluster of canine distemper virus in north and center of Iran

    PubMed Central

    Namroodi, Somayeh; Rostami, Amir; Majidzadeh-Ardebili, Keyvan; Ghalyanchi Langroudi, Arash; Morovvati, Abbas

    2015-01-01

    Canine distemper virus (CDV) creates a very contagious viral multi-systemic canine distemper (CD) disease that affects most species of Carnivora order. The virus is genetically heterogeneous, particularly in section of the hemagglutinin (H) gene. Sequence analysis of the H gene can be useful to investigate distinction of various lineages related to geographical distribution and CDV molecular epidemiology. Since vaccination program is conducted only in large cities of Iran, CD still remains as one of the major causes of death in dogs in this country. In order to monitor H gene, CDV has been detected in 14 out of 19 sampled dogs through the amplification of nucleoprotein (NP) gene in nested-PCR assay. In the next step 665 bp of H gene was amplified in 9 out of 14 NP-gene positive dogs. Phylogenetic analysis distinguished two distinct CDV genotypes in Iran. JN941238 has been embedded in European cluster and JN941239 has been embedded in Arctic cluster. Nucleic analysis has been shown high difference among both Iranian CDV lineages with CDV vaccine strains. PMID:26893808

  2. Sandwich-dot enzyme-linked immunosorbent assay for the detection of canine distemper virus.

    PubMed

    Li, Zhi; Zhang, Yanlong; Wang, Huiguo; Jin, Jinhua; Li, Wenzhe

    2013-10-01

    A sandwich-dot enzyme-linked immunosorbent assay (dot ELISA) was developed for the detection of canine distemper virus (CDV). In 56 dogs suspected to have CD the rates of detection of CDV antigen in samples of blood lymphocytes and palpebral conjunctiva by dot ELISA and ELISA were, respectively, 91% (49/54) and 81% (44/54) for the lymphocyte samples and 88% (28/32) and 75% (24/32) for the conjunctival samples. The CDV detection limits were 10 ng/50 μL for dot ELISA and 40 ng/50 μL for ELISA. The reliability of dot ELISA relative to electron microscopy was 96% with 22 samples: all 21 samples in which CDV particles were observed by electron microscopy yielded positive results with dot ELISA; the single sample in which particles were not observed yielded false-positive results with dot ELISA. The results indicate that the dot ELISA developed can serve as a reliable rapid diagnostic test in suspected cases of CD and also be useful for epidemiologic surveillance of the disease.

  3. Detection of canine distemper virus serum neutralizing antibodies in captive U.S. phocids.

    PubMed

    Clancy, Meredith M; Gamble, Kathryn C; Travis, Dominic A

    2013-03-01

    Antibodies to morbilliviruses have been documented in free-ranging pinnipeds throughout populations in the Atlantic and Arctic Oceans, but not from the Pacific Ocean. As a symbolic geographic barrier between the exposed Atlantic and naive Pacific populations, the captive phocid population in North America had undocumented serologic status. In this study, canine distemper virus (CDV) serum neutralization assays were used to assess the prevalence of antibodies in this population with participation of 25 U.S. institutions from grey seals (Halichoerus grypus, n = 6) and harbor seals (Phoca vitulina, n = 108). Historic and environmental risk factors associated with the epidemiology of distemper virus were collected by survey. Based on antibodies to canine distemper virus, the prevalence of exposure in this population was 25.5%, with 28 seals (grey, n = 2; harbor, n = 26) demonstrating antibody titers > or = 1:16, and positive titers ranged from 1:4 to 1:1,536. By survey analysis, strong associations with seropositive status were identified for captive origin (P = 0.013) and movement among institutions (P = 0.024). Size of population has positive correlation with likelihood of seropositive seals at an institution (P = 0.020). However, no major husbandry or enclosure-based risk factors were identified in institutions with seropositive seals, and no interaction between individual or institutional risk factors was identified. Previously undocumented prior to this study, CDV antibodies were measured in harbor seals (n = 2) recently stranded from the Pacific coast. PMID:23505705

  4. Detection and differentiation of wild-type and vaccine strains of canine distemper virus by a duplex reverse transcription polymerase chain reaction.

    PubMed

    Dong, X Y; Li, W H; Zhu, J L; Liu, W J; Zhao, M Q; Luo, Y W; Chen, J D

    2015-01-01

    Canine distemper virus (CDV) is the cause of canine distemper (CD) which is a severe and highly contagious disease in dogs. In the present study, a duplex reverse transcription polymerase chain reaction (RT-PCR) method was developed for the detection and differentiation of wild-type and vaccine strains of CDV. Four primers were designed to detect and discriminate the two viruses by generating 638- and 781-bp cDNA products, respectively. Furthermore, the duplex RT-PCR method was used to detect 67 field samples suspected of CD from Guangdong province in China. Results showed that, 33 samples were to be wild-type-like. The duplex RT-PCR method exhibited high specificity and sensitivity which could be used to effectively detect and differentiate wild-type and vaccine CDV, indicating its use for clinical detection and epidemiological surveillance. PMID:27175171

  5. Detection and differentiation of wild-type and vaccine strains of canine distemper virus by a duplex reverse transcription polymerase chain reaction

    PubMed Central

    Dong, X. Y.; Li, W. H.; Zhu, J. L.; Liu, W. J.; Zhao, M. Q.; Luo, Y. W.; Chen, J. D.

    2015-01-01

    Canine distemper virus (CDV) is the cause of canine distemper (CD) which is a severe and highly contagious disease in dogs. In the present study, a duplex reverse transcription polymerase chain reaction (RT-PCR) method was developed for the detection and differentiation of wild-type and vaccine strains of CDV. Four primers were designed to detect and discriminate the two viruses by generating 638- and 781-bp cDNA products, respectively. Furthermore, the duplex RT-PCR method was used to detect 67 field samples suspected of CD from Guangdong province in China. Results showed that, 33 samples were to be wild-type-like. The duplex RT-PCR method exhibited high specificity and sensitivity which could be used to effectively detect and differentiate wild-type and vaccine CDV, indicating its use for clinical detection and epidemiological surveillance. PMID:27175171

  6. Seroprevalence of Canine Distemper Virus in Cats

    PubMed Central

    Ikeda, Yasuhiro; Nakamura, Kazuya; Miyazawa, Takayuki; Chen, Ming-Chu; Kuo, Tzong-Fu; Lin, James A.; Mikami, Takeshi; Kai, Chieko; Takahashi, Eiji

    2001-01-01

    A seroepidemiological survey of canine distemper virus (CDV) infection in Asian felids revealed that the prevalence of antibodies varied depending on region and, in some cases, exposure to dogs. The serologic pattern in cats with antibodies indicated that they had likely been exposed to field strains rather than typical CDV vaccine strains. PMID:11329473

  7. Serological detection of infection with canine distemper virus, canine parvovirus and canine adenovirus in communal dogs from Zimbabwe.

    PubMed

    McRee, Anna; Wilkes, Rebecca P; Dawson, Jessica; Parry, Roger; Foggin, Chris; Adams, Hayley; Odoi, Agricola; Kennedy, Melissa A

    2014-01-01

    Domestic dogs are common amongst communities in sub-Saharan Africa and may serve as important reservoirs for infectious agents that may cause diseases in wildlife. Two agents of concern are canine parvovirus (CPV) and canine distemper virus (CDV), which may infect and cause disease in large carnivore species such as African wild dogs and African lions, respectively. The impact of domestic dogs and their diseases on wildlife conservation is increasing in Zimbabwe, necessitating thorough assessment and implementation of control measures. In this study, domestic dogs in north-western Zimbabwe were evaluated for antibodies to CDV, CPV, and canine adenovirus (CAV). These dogs were communal and had no vaccination history. Two hundred and twenty-five blood samples were collected and tested using a commercial enzyme-linked immunosorbent assay (ELISA) for antibodies to CPV, CDV, and CAV. Of these dogs, 75 (34%) had detectable antibodies to CDV, whilst 191 (84%) had antibodies to CPV. Antibodies to canine adenovirus were present in 28 (13%) dogs. Canine parvovirus had high prevalence in all six geographic areas tested. These results indicate that CPV is circulating widely amongst domestic dogs in the region. In addition, CDV is present at high levels. Both pathogens can infect wildlife species. Efforts for conservation of large carnivores in Zimbabwe must address the role of domestic dogs in disease transmission. PMID:25686382

  8. Serological detection of infection with canine distemper virus, canine parvovirus and canine adenovirus in communal dogs from Zimbabwe.

    PubMed

    McRee, Anna; Wilkes, Rebecca P; Dawson, Jessica; Parry, Roger; Foggin, Chris; Adams, Hayley; Odoi, Agricola; Kennedy, Melissa A

    2014-01-01

    Domestic dogs are common amongst communities in sub-Saharan Africa and may serve as important reservoirs for infectious agents that may cause diseases in wildlife. Two agents of concern are canine parvovirus (CPV) and canine distemper virus (CDV), which may infect and cause disease in large carnivore species such as African wild dogs and African lions, respectively. The impact of domestic dogs and their diseases on wildlife conservation is increasing in Zimbabwe, necessitating thorough assessment and implementation of control measures. In this study, domestic dogs in north-western Zimbabwe were evaluated for antibodies to CDV, CPV, and canine adenovirus (CAV). These dogs were communal and had no vaccination history. Two hundred and twenty-five blood samples were collected and tested using a commercial enzyme-linked immunosorbent assay (ELISA) for antibodies to CPV, CDV, and CAV. Of these dogs, 75 (34%) had detectable antibodies to CDV, whilst 191 (84%) had antibodies to CPV. Antibodies to canine adenovirus were present in 28 (13%) dogs. Canine parvovirus had high prevalence in all six geographic areas tested. These results indicate that CPV is circulating widely amongst domestic dogs in the region. In addition, CDV is present at high levels. Both pathogens can infect wildlife species. Efforts for conservation of large carnivores in Zimbabwe must address the role of domestic dogs in disease transmission.

  9. Phylogenetic analysis of the haemagglutinin gene of canine distemper virus strains detected from giant panda and raccoon dogs in China

    PubMed Central

    2013-01-01

    Background Canine distemper virus (CDV) infects a variety of carnivores, including wild and domestic Canidae. In this study, we sequenced and phylogenetic analyses of the hemagglutinin (H) genes from eight canine distemper virus (CDV) isolates obtained from seven raccoon dogs (Nyctereutes procyonoides) and a giant panda (Ailuropoda melanoleuca) in China. Results Phylogenetic analysis of the partial hemagglutinin gene sequences showed close clustering for geographic lineages, clearly distinct from vaccine strains and other wild-type foreign CDV strains, all the CDV strains were characterized as Asia-1 genotype and were highly similar to each other (91.5-99.8% nt and 94.4-99.8% aa). The giant panda and raccoon dogs all were 549Y on the HA protein in this study, irrespective of the host species. Conclusions These findings enhance our knowledge of the genetic characteristics of Chinese CDV isolates, and may facilitate the development of effective strategies for monitoring and controlling CDV for wild canids and non-cainds in China. PMID:23566727

  10. Establishment of reverse transcription loop-mediated isothermal amplification for rapid detection and differentiation of canine distemper virus infected and vaccinated animals.

    PubMed

    Liu, Da-Fei; Liu, Chun-Guo; Tian, Jin; Jiang, Yi-Tong; Zhang, Xiao-Zhan; Chai, Hong-Liang; Yang, Tian-Kuo; Yin, Xiu-Chen; Zhang, Hong-Ying; Liu, Ming; Hua, Yu-Ping; Qu, Lian-Dong

    2015-06-01

    Although widespread vaccination against canine distemper virus (CDV) has been conducted for many decades, several canine distemper outbreaks in vaccinated animals have been reported frequently. In order to detect and differentiate the wild-type and vaccine strains of the CDV from the vaccinated animals, a novel reverse transcription loop-mediated isothermal amplification (RT-LAMP) method was developed. A set of four primers-two internal and two external-were designed to target the H gene for the specific detection of wild-type CDV variants. The CDV-H RT-LAMP assay rapidly amplified the target gene, within 60 min, using a water bath held at a constant temperature of 65°C. The assay was 100-fold more sensitive than conventional RT-PCR, with a detection limit of 10(-1)TCID50ml(-1). The system showed a preference for wild-type CDV, and exhibited less sensitivity to canine parvovirus, canine adenovirus type 1 and type 2, canine coronavirus, and canine parainfluenza virus. The assay was validated using 102 clinical samples obtained from vaccinated dog farms, and the results were comparable to a multiplex nested RT-PCR assay. The specific CDV-H RT-LAMP assay provides a simple, rapid, and sensitive tool for the detection of canines infected with wild-type CDV from canines vaccinated with attenuated vaccine.

  11. Antiserum raised in pigs against canine distemper virus and its utility in diagnostic procedures for morbillivirus infections (canine distemper, phocine distemper, rinderpest).

    PubMed

    Zaghawa, A; Liess, B; Frey, H R

    1990-07-01

    Antiserum against canine distemper virus (CDV) was raised in pigs by intranasal inoculation with CDV strains CND65 and ROCKBORN. Immunoglobulin fractions were conjugated with horseradish peroxidase. Peroxidase-conjugated anti-CDV immunoglobulin preparations were used for the detection and titration of CDV, seal-derived (phocine) distemper virus (PDV) and rinderpest virus (RPV) in Vero cell cultures. For the detection and titration of corresponding neutralizing antibodies a direct neutralizing peroxidase-linked antibody (NPLA) assay was established. The results were compared with those obtained with the conventional microtitre neutralization test (MNT) based on CPE reading. In addition the sensitivity of an indirect peroxidase-linked antibody (PLA) assay was tested in parallel with that of the NPLA assay using sera obtained from CDV-immunized pigs.

  12. Real-time reverse transcription polymerase chain reaction method for detection of Canine distemper virus modified live vaccine shedding for differentiation from infection with wild-type strains.

    PubMed

    Wilkes, Rebecca P; Sanchez, Elena; Riley, Matthew C; Kennedy, Melissa A

    2014-01-01

    Canine distemper virus (CDV) remains a common cause of infectious disease in dogs, particularly in high-density housing situations such as shelters. Vaccination of all dogs against CDV is recommended at the time of admission to animal shelters and many use a modified live virus (MLV) vaccine. From a diagnostic standpoint for dogs with suspected CDV infection, this is problematic because highly sensitive diagnostic real-time reverse transcription polymerase chain reaction (RT-PCR) tests are able to detect MLV virus in clinical samples. Real-time PCR can be used to quantitate amount of virus shedding and can differentiate vaccine strains from wild-type strains when shedding is high. However, differentiation by quantitation is not possible in vaccinated animals during acute infection, when shedding is low and could be mistaken for low level vaccine virus shedding. While there are gel-based RT-PCR assays for differentiation of vaccine strains from field strains based on sequence differences, the sensitivity of these assays is unable to match that of the real-time RT-PCR assay currently used in the authors' laboratory. Therefore, a real-time RT-PCR assay was developed that detects CDV MLV vaccine strains and distinguishes them from wild-type strains based on nucleotide sequence differences, rather than the amount of viral RNA in the sample. The test is highly sensitive, with detection of as few as 5 virus genomic copies (corresponding to 10(-1) TCID(50)). Sequencing of the DNA real-time products also allows phylogenetic differentiation of the wild-type strains. This test will aid diagnosis during outbreaks of CDV in recently vaccinated animals.

  13. Detection of IgM antibodies against a recombinant nucleocapsid protein of canine distemper virus in dog sera using a dot-blot assay.

    PubMed

    Barben, G; Stettler, M; Jaggy, A; Vandevelde, M; Zurbriggen, A

    1999-03-01

    A dot-blot assay for the detection of IgM antibodies (ABs) against canine distemper virus (CDV) in canine serum is described. The diagnostic potential of this technique was evaluated by analysing sera from three test groups: (i) specific pathogen-free (SPF) beagle dogs experimentally infected with virulent CDV; (ii) SPF dogs immunized with a combined vaccine containing CDV, and (iii) SPF dogs immunized with a CDV-free vaccine. As antigen for the dot-blot assay we used the recombinant nucleocapsid protein (N protein) of the virulent A75/17 CDV strain. All 12 dogs of group 1, infected with virulent CDV, showed detectable CDV-specific IgM levels in their serum. All dogs of group 2 were also positive for anti-CDV IgM after the first immunization with the CDV-containing vaccine. The four dogs immunized with a CDV-free vaccine (group iii) remained negative throughout the course of the experiment. From these results, we conclude that the IgM detection test, which requires only a single serum sample, is a useful method for diagnosing current or recent CDV infection in CDV-infected or CDV-immunized dogs under experimental conditions. PMID:10216448

  14. Detection by RT-PCR and genetic characterization of canine distemper virus from vaccinated and non-vaccinated dogs in Argentina.

    PubMed

    Calderon, Marina Gallo; Remorini, Patricia; Periolo, Osvaldo; Iglesias, Marcela; Mattion, Nora; La Torre, José

    2007-12-15

    RT-PCR was used to detect canine distemper virus (CDV) RNA in clotted blood from Argentine domestic dogs. The NP gene was detected in 73 out of 99 blood samples analyzed. The deduced amino acid sequence of these gene fragments showed 100% identity with the sequence of other wild-type and vaccine strains. A fragment of the hemagglutinin gene was amplified from 24 (32.9%) of the NP-RNA-positive clinical specimens. These H fragments were further analyzed by restriction fragment length polymorphism (RFLP) and sequencing. A single NdeI site was detected in all 24 wild-type strains but was absent in the vaccine strains. Phylogenetic analysis of the partial hemagglutinin amino acid sequences showed close clustering for local strains, clearly distinct from vaccine strains and other wild-type foreign CDV strains. One of the local strains, Arg 23, branched out of the root of the Argentine clade, close to the European strains, suggesting that two different pathogenic CDV genotypes are currently circulating in Argentina, one of them clearly predominant. PMID:17628358

  15. Recombinant canine distemper virus serves as bivalent live vaccine against rabies and canine distemper.

    PubMed

    Wang, Xijun; Feng, Na; Ge, Jinying; Shuai, Lei; Peng, Liyan; Gao, Yuwei; Yang, Songtao; Xia, Xianzhu; Bu, Zhigao

    2012-07-20

    Effective, safe, and affordable rabies vaccines are still being sought. Attenuated live vaccine has been widely used to protect carnivores from canine distemper. In this study, we generated a recombinant canine distemper virus (CDV) vaccine strain, rCDV-RVG, expressing the rabies virus glycoprotein (RVG) by using reverse genetics. The recombinant virus rCDV-RVG retained growth properties similar to those of vector CDV in Vero cell culture. Animal studies demonstrated that rCDV-RVG was safe in mice and dogs. Mice inoculated intracerebrally or intramuscularly with rCDV-RVG showed no apparent signs of disease and developed a strong rabies virus (RABV) neutralizing antibody response, which completely protected mice from challenge with a lethal dose of street virus. Canine studies showed that vaccination with rCDV-RVG induced strong and long-lasting virus neutralizing antibody responses to RABV and CDV. This is the first study demonstrating that recombinant CDV has the potential to serve as bivalent live vaccine against rabies and canine distemper in animals. PMID:22698451

  16. Recombinant rabies virus expressing the H protein of canine distemper virus protects dogs from the lethal distemper challenge.

    PubMed

    Wang, Feng-Xue; Zhang, Shu-Qin; Zhu, Hong-Wei; Yang, Yong; Sun, Na; Tan, Bin; Li, Zhen-Guang; Cheng, Shi-Peng; Fu, Zhen F; Wen, Yong-Jun

    2014-12-01

    The rabies virus (RV) vector LBNSE expressing foreign antigens have shown considerable promise as vaccines against viral and bacteria diseases, which is effective and safe. We produced a new RV-based vaccine vehicle expressing 1.824 kb hemagglutinin (H) gene of the canine distemper virus (CDV) by reverse genetics technology. The recombinant virus LBNSE-CDV-H retained growth properties similar to those of vector LBNSE both in BSR and mNA cell culture. The H gene of CDV was expressed and detected by immunostaining. To compare the immunogenicity of LBNSE-CDV-H, dogs were immunized with each of these recombinant viruses by intramuscular (i.m.). The dogs were bled at third weeks after the immunization for the measurement of virus neutralizing antibody (VNA) and then challenged with virulent virus (ZJ 7) at fourth weeks. The parent virus (LBNSE) without expression of any foreign molecules was included for comparison. Dogs inoculated with LBNSE-CDV-H showed no any signs of disease and exhibited seroconversion against both RV and CDV H protein. The LBNSE-CDV-H did not cause disease in dogs and conferred protection from challenge with a lethal wild type CDV strain, demonstrating its potential value for wildlife conservation efforts. Together, these studies suggest that recombinant RV expressing H protein from CDV stimulated high levels of adaptive immune responses (VNA), and protected all dogs challenge infection. PMID:25465178

  17. Canine distemper virus (CDV) in another big cat: should CDV be renamed carnivore distemper virus?

    PubMed

    Terio, Karen A; Craft, Meggan E

    2013-09-17

    One of the greatest threats to the conservation of wild cat populations may be dogs or, at least, one of their viruses. Canine distemper virus (CDV), a single-stranded RNA virus in the Paramyxoviridae family and genus Morbillivirus, infects and causes disease in a variety of species, not just canids. An outbreak of CDV in wild lions in the Serengeti, Tanzania, in 1994 was a wake-up call for conservationists, as it demonstrated that an infectious disease could swiftly impact a previously healthy felid population. To understand how this virus causes disease in noncanid hosts, researchers have focused on specific mutations in the binding site of the CDV hemagglutinin gene. Now, Seimon et al. provide information on CDV in its latest feline victim, the endangered wild Amur tiger (Panthera tigris altaica) [T. A. Seimon et al., mBio 4(4):e00410-13, 2013, doi:10.1128/mBio.00410-13]. Their findings of CDV strains infecting tigers, in combination with recent information from other felids, paints a different picture, one in which CDV strains from a variety of geographic lineages and with a variety of amino acid residues in the hemagglutinin gene binding site can infect cats and cause disease. Although CDV has been known as a multihost disease since its discovery in domestic dogs in 1905, perhaps it is time to reconsider whether these noncanid species are not just incidental or "spillover" hosts but, rather, a normal part of the complex ecology of this infectious disease.

  18. Immunopathogenic and Neurological Mechanisms of Canine Distemper Virus

    PubMed Central

    Carvalho, Otávio Valério; Botelho, Clarisse Vieira; Ferreira, Caroline Gracielle Torres; Scherer, Paulo Oldemar; Soares-Martins, Jamária Adriana Pinheiro; Almeida, Márcia Rogéria; Silva Júnior, Abelardo

    2012-01-01

    Canine distemper is a highly contagious viral disease caused by the canine distemper virus (CDV), which is a member of the Morbillivirus genus, Paramyxoviridae family. Animals that most commonly suffer from this disease belong to the Canidae family; however, the spectrum of natural hosts for CDV also includes several other families of the order Carnivora. The infectious disease presents worldwide distribution and maintains a high incidence and high levels of lethality, despite the availability of effective vaccines, and no specific treatment. CDV infection in dogs is characterized by the presentation of systemic and/or neurological courses, and viral persistence in some organs, including the central nervous system (CNS) and lymphoid tissues. An elucidation of the pathogenic mechanisms involved in canine distemper disease will lead to a better understanding of the injuries and clinical manifestations caused by CDV. Ultimately, further insight about this disease will enable the improvement of diagnostic methods as well as therapeutic studies. PMID:23193403

  19. Canine distemper virus (CDV) in another big cat: should CDV be renamed carnivore distemper virus?

    PubMed

    Terio, Karen A; Craft, Meggan E

    2013-01-01

    One of the greatest threats to the conservation of wild cat populations may be dogs or, at least, one of their viruses. Canine distemper virus (CDV), a single-stranded RNA virus in the Paramyxoviridae family and genus Morbillivirus, infects and causes disease in a variety of species, not just canids. An outbreak of CDV in wild lions in the Serengeti, Tanzania, in 1994 was a wake-up call for conservationists, as it demonstrated that an infectious disease could swiftly impact a previously healthy felid population. To understand how this virus causes disease in noncanid hosts, researchers have focused on specific mutations in the binding site of the CDV hemagglutinin gene. Now, Seimon et al. provide information on CDV in its latest feline victim, the endangered wild Amur tiger (Panthera tigris altaica) [T. A. Seimon et al., mBio 4(4):e00410-13, 2013, doi:10.1128/mBio.00410-13]. Their findings of CDV strains infecting tigers, in combination with recent information from other felids, paints a different picture, one in which CDV strains from a variety of geographic lineages and with a variety of amino acid residues in the hemagglutinin gene binding site can infect cats and cause disease. Although CDV has been known as a multihost disease since its discovery in domestic dogs in 1905, perhaps it is time to reconsider whether these noncanid species are not just incidental or "spillover" hosts but, rather, a normal part of the complex ecology of this infectious disease. PMID:24045642

  20. Canine Distemper Virus (CDV) in Another Big Cat: Should CDV Be Renamed Carnivore Distemper Virus?

    PubMed Central

    Terio, Karen A.; Craft, Meggan E.

    2013-01-01

    ABSTRACT One of the greatest threats to the conservation of wild cat populations may be dogs or, at least, one of their viruses. Canine distemper virus (CDV), a single-stranded RNA virus in the Paramyxoviridae family and genus Morbillivirus, infects and causes disease in a variety of species, not just canids. An outbreak of CDV in wild lions in the Serengeti, Tanzania, in 1994 was a wake-up call for conservationists, as it demonstrated that an infectious disease could swiftly impact a previously healthy felid population. To understand how this virus causes disease in noncanid hosts, researchers have focused on specific mutations in the binding site of the CDV hemagglutinin gene. Now, Seimon et al. provide information on CDV in its latest feline victim, the endangered wild Amur tiger (Panthera tigris altaica) [T. A. Seimon et al., mBio 4(4):e00410-13, 2013, doi:10.1128/mBio.00410-13]. Their findings of CDV strains infecting tigers, in combination with recent information from other felids, paints a different picture, one in which CDV strains from a variety of geographic lineages and with a variety of amino acid residues in the hemagglutinin gene binding site can infect cats and cause disease. Although CDV has been known as a multihost disease since its discovery in domestic dogs in 1905, perhaps it is time to reconsider whether these noncanid species are not just incidental or “spillover” hosts but, rather, a normal part of the complex ecology of this infectious disease. PMID:24045642

  1. Selective spread and reduced virus release leads to canine distemper virus persistence in the nervous system.

    PubMed

    Zurbriggen, A; Graber, H U; Vandevelde, M

    1995-05-01

    In primary dog brain cell cultures (DBCC) the attenuated canine distemper virus (CDV) is cytolytic, whereas virulent CDV is not. Thus, the question why cytolysis does or does not occur appears to be intimately associated with the mechanism of persistence. Persistence is most likely related to the way these viruses replicate. In the present study we used morphological and immunocytochemical approaches to compare several aspects of virus replication between a cytolytic and a virulent CDV strain using DBCC to study both strains. Quantitative measurements did not detect a difference in the rate of virus protein synthesis at the level of the single cell, between the two types of infection. Electron microscopical results and virus titration experiments showed marked differences in virus spread and virus release. Immunocytochemical studies showed differences in the distribution of the nucleocapsid and matrix proteins between the two infections. Budding and cytolysis are strongly limited in the virulent A75/17-CDV infection as compared to attenuated viruses. This is probably due to structural changes in the virus proteins leading to modifications of virus assembly. Thus the present study supports a mechanism of CDV persistence through a type of virus maturation and spread by which very little virus is released outside of the cell.

  2. Serologic survey for antibodies to canine distemper virus in collared peccary (Tayassu tajacu) populations in Arizona.

    PubMed

    Noon, Ted H; Heffelfinger, James R; Olding, Ronald J; Wesche, Shannon Lynn; Reggiardo, Carlos

    2003-01-01

    In 1989, a disease outbreak was observed among collared peccaries (javelina, Tayassu tajacu) in southern Arizona (USA) and canine distemper virus (CDV) was isolated from affected animals. Subsequently, 364 sera were collected from hunter-harvested javelina over a 4 yr period (1993-96) and were tested for antibody to CDV. Neutralizing antibody to CDV was detected in 58% of the serum samples suggesting that CDV infection is probably enzootic in the collared peccary populations of southern Arizona.

  3. Vaccination of mice against canine distemper virus-induced encephalitis with vaccinia virus recombinants encoding measles or canine distemper virus antigens.

    PubMed

    Wild, T F; Bernard, A; Spehner, D; Villeval, D; Drillien, R

    1993-01-01

    Measles and canine distemper are caused by serologically related viruses. Although dogs immunized with measles virus (MV) do not elicit canine distemper virus (CDV) neutralizing antibodies, they are protected against the fatal disease. To investigate the potential role of the MV antigens in protection against CDV, we have immunized mice with vaccinia virus (VV) recombinants expressing the MV haemagglutinin (HA), fusion (F), nucleoprotein (NP) and matrix (M) antigens and challenged them with CDV. A partial protection was observed with the VV recombinants expressing the F, NP and M antigens, but not the HA. In contrast, immunization with a VV recombinant expressing the CDV F protein completely protected mice from CDV. PMID:8470428

  4. Chronic encephalomyelitis caused by canine distemper virus in a Bengal tiger.

    PubMed

    Blythe, L L; Schmitz, J A; Roelke, M; Skinner, S

    1983-12-01

    A chronic progressive neurologic disease was observed and monitored for 18 months in a young, tamed Bengal tiger. Clinical, serologic, and neuropathologic evidence of canine distemper virus infection was seen. Clinical signs included convulsions, myoclonus, and slowly progressive ataxia. Marked increases in neutralizing antibodies against canine distemper virus were seen in the serum and cerebrospinal fluid. Neuropathologic findings were nonsuppurative meningoencephalomyelitis, with perivascular cuffing, demyelination, and inclusion bodies typical of canine distemper virus. It was concluded that, in light of this case and an earlier report of canine distemper in lion cubs, vaccination of this subgroup of carnivores with a killed vaccine may be beneficial if exposure to other animals susceptible to canine distemper is anticipated.

  5. Chronic encephalomyelitis caused by canine distemper virus in a Bengal tiger.

    PubMed

    Blythe, L L; Schmitz, J A; Roelke, M; Skinner, S

    1983-12-01

    A chronic progressive neurologic disease was observed and monitored for 18 months in a young, tamed Bengal tiger. Clinical, serologic, and neuropathologic evidence of canine distemper virus infection was seen. Clinical signs included convulsions, myoclonus, and slowly progressive ataxia. Marked increases in neutralizing antibodies against canine distemper virus were seen in the serum and cerebrospinal fluid. Neuropathologic findings were nonsuppurative meningoencephalomyelitis, with perivascular cuffing, demyelination, and inclusion bodies typical of canine distemper virus. It was concluded that, in light of this case and an earlier report of canine distemper in lion cubs, vaccination of this subgroup of carnivores with a killed vaccine may be beneficial if exposure to other animals susceptible to canine distemper is anticipated. PMID:6685717

  6. Canine distemper virus persistence in the nervous system is associated with noncytolytic selective virus spread.

    PubMed

    Zurbriggen, A; Graber, H U; Wagner, A; Vandevelde, M

    1995-03-01

    Canine distemper virus (CDV), a negative-strand RNA morbillivirus, causes a progressive demyelinating disease in which virus persistence plays an essential role. The antiviral immune response leads to virus clearance in the inflammatory lesions. However, CDV can replicate and persist outside these inflammatory lesions within the brain. How CDV is capable of persisting in the presence of an effective antiviral immune response is poorly understood. In the present investigation, we studied several aspects of virus replication in primary dog brain cell cultures (DBCC), comparing an attenuated CDV strain and a virulent CDV strain. Confluent DBCC were infected with either virulent A75/17-CDV or attenuated Onderstepoort-CDV and monitored for 60 days. Persistence was not associated with defective virus production, because all mRNAs and corresponding proteins were continuously expressed in the noncytolytic infection. Quantitative measurements did not detect a difference between the two types of infection in the rate of virus transcription and protein synthesis at the level of the single cell. However, electron microscopy and virus titration experiments showed that in the persistent CDV infection virus budding is strongly limited compared with that of the attenuated virus. Morphometry and immunocytochemistry showed profound differences in the way the two viruses spread in the culture. The attenuated CDV spread randomly to immediately adjacent cells, whereas persistent CDV spread selectively to more-distant cells by way of cell processes. In conclusion, the present study supports a mechanism of CDV persistence through selective spread by way of cell processes, enabling virulent CDV to invade the central nervous system without the need of releasing much virus into the extracellular space.

  7. Canine distemper virus persistence in the nervous system is associated with noncytolytic selective virus spread.

    PubMed Central

    Zurbriggen, A; Graber, H U; Wagner, A; Vandevelde, M

    1995-01-01

    Canine distemper virus (CDV), a negative-strand RNA morbillivirus, causes a progressive demyelinating disease in which virus persistence plays an essential role. The antiviral immune response leads to virus clearance in the inflammatory lesions. However, CDV can replicate and persist outside these inflammatory lesions within the brain. How CDV is capable of persisting in the presence of an effective antiviral immune response is poorly understood. In the present investigation, we studied several aspects of virus replication in primary dog brain cell cultures (DBCC), comparing an attenuated CDV strain and a virulent CDV strain. Confluent DBCC were infected with either virulent A75/17-CDV or attenuated Onderstepoort-CDV and monitored for 60 days. Persistence was not associated with defective virus production, because all mRNAs and corresponding proteins were continuously expressed in the noncytolytic infection. Quantitative measurements did not detect a difference between the two types of infection in the rate of virus transcription and protein synthesis at the level of the single cell. However, electron microscopy and virus titration experiments showed that in the persistent CDV infection virus budding is strongly limited compared with that of the attenuated virus. Morphometry and immunocytochemistry showed profound differences in the way the two viruses spread in the culture. The attenuated CDV spread randomly to immediately adjacent cells, whereas persistent CDV spread selectively to more-distant cells by way of cell processes. In conclusion, the present study supports a mechanism of CDV persistence through selective spread by way of cell processes, enabling virulent CDV to invade the central nervous system without the need of releasing much virus into the extracellular space. PMID:7853504

  8. Canine Distemper Virus Antigen Detection in External Epithelia of Recently Vaccinated, Sick Dogs by Fluorescence Microscopy Is a Valuable Prognostic Indicator

    PubMed Central

    Neel, Tina

    2014-01-01

    Currently, there are no reliable predictors of the clinical outcomes of domesticated dogs that have been recently vaccinated against canine distemper virus (CDV) and develop respiratory disease. In this study, vaccinated dogs from Oklahoma City that were showing clinical signs of respiratory disease were evaluated for CDV antigen using a direct fluorescent antibody test (FAT). Clinical outcomes after standard symptomatic therapy for respiratory disease were recorded, and a statistical analysis of the results was performed. We present our study showing that CDV FAT results were predictive of clinical recovery (prognostic indicator, prospects of clinical recovery) among vaccinated dogs showing clinical signs of respiratory disease. Negative CDV FAT results equated to 80% chances of recovery after symptomatic therapy, compared to 55% chances of recovery when the CDV FAT results were positive. Based on the results of this study, we show that veterinarians can make better informed decisions about the clinical outcomes of suspected CDV cases, with 2-h turnaround times, by using the CDV FAT. Thus, antemortem examination with the CDV FAT on external epithelia of recently vaccinated, sick dogs is a clinically useful diagnostic test and valuable prognostic indicator for veterinarians. Application of the CDV FAT to these samples avoids unnecessary euthanasia of dogs with suspected CDV. PMID:25428156

  9. Rapid and sensitive detection of immunoglobulin M (IgM) and IgG antibodies against canine distemper virus by a new recombinant nucleocapsid protein-based enzyme-linked immunosorbent assay.

    PubMed

    von Messling, V; Harder, T C; Moennig, V; Rautenberg, P; Nolte, I; Haas, L

    1999-04-01

    Canine distemper morbillivirus (CDV) infection causes a frequently fatal systemic disease in a broad range of carnivore species, including domestic dogs. In CDV infection, classical serology provides data of diagnostic and prognostic values (kinetics of seroconversion) and is also used to predict the optimal vaccination age of pups. Routine CDV serology is still based on time- and cost-intensive virus neutralization assays (V-NA). Here, we describe a new capture-sandwich enzyme-linked immunosorbent assay (ELISA) that uses recombinant baculovirus-expressed nucleocapsid (N) protein of a recent CDV wild-type isolate (2544/Han95) for the detection of CDV-specific antibodies in canine sera. Recombinant antigen was produced with high efficacy in Heliothis virescens larvae. The capture-sandwich ELISA enabled a clear-cut qualitative evaluation of the CDV-specific immunoglobulin G (IgG) and IgM serostatuses of 196 and 35 dog sera, respectively. Inter-rater agreement analysis (kappa = 0.988) indicated that the ELISA can be used unrestrictedly as a substitute for the V-NA for the qualitative determination of CDV-specific IgG serostatus. In an attempt to semiquantify N-specific antibodies, a one-step-dilution (alpha method) IgG-specific ELISA was implemented. Alpha values of >/=50% showed very good inter-rater agreement (kappa = 0.968) with V-NA titers of >/=1/100 50% neutralizing dose (ND50) as measured against the central European CDV wild-type isolate 2544/Han95 in canine sera originating from northern Germany. An ND50 titer of 1/100 is considered a threshold, and titers of >/=1/100 indicate a resilient, protective immunity. CDV N-specific antibodies of the IgM class were detected by the newly developed ELISA in 9 of 15 sera obtained from dogs with symptoms of acute distemper. In leucocytes of 5 of the 15 dogs (all of which were also IgM positive) CDV RNA was detected by reverse transcription (RT)-PCR. The recombinant capture-sandwich ELISA detecting N-specific antibodies

  10. Rapid and Sensitive Detection of Immunoglobulin M (IgM) and IgG Antibodies against Canine Distemper Virus by a New Recombinant Nucleocapsid Protein-Based Enzyme-Linked Immunosorbent Assay

    PubMed Central

    von Messling, Veronika; Harder, Timm C.; Moennig, Volker; Rautenberg, Peter; Nolte, Ingo; Haas, Ludwig

    1999-01-01

    Canine distemper morbillivirus (CDV) infection causes a frequently fatal systemic disease in a broad range of carnivore species, including domestic dogs. In CDV infection, classical serology provides data of diagnostic and prognostic values (kinetics of seroconversion) and is also used to predict the optimal vaccination age of pups. Routine CDV serology is still based on time- and cost-intensive virus neutralization assays (V-NA). Here, we describe a new capture-sandwich enzyme-linked immunosorbent assay (ELISA) that uses recombinant baculovirus-expressed nucleocapsid (N) protein of a recent CDV wild-type isolate (2544/Han95) for the detection of CDV-specific antibodies in canine sera. Recombinant antigen was produced with high efficacy in Heliothis virescens larvae. The capture-sandwich ELISA enabled a clear-cut qualitative evaluation of the CDV-specific immunoglobulin G (IgG) and IgM serostatuses of 196 and 35 dog sera, respectively. Inter-rater agreement analysis (κ = 0.988) indicated that the ELISA can be used unrestrictedly as a substitute for the V-NA for the qualitative determination of CDV-specific IgG serostatus. In an attempt to semiquantify N-specific antibodies, a one-step-dilution (alpha method) IgG-specific ELISA was implemented. Alpha values of ≥50% showed very good inter-rater agreement (κ = 0.968) with V-NA titers of ≥1/100 50% neutralizing dose (ND50) as measured against the central European CDV wild-type isolate 2544/Han95 in canine sera originating from northern Germany. An ND50 titer of 1/100 is considered a threshold, and titers of ≥1/100 indicate a resilient, protective immunity. CDV N-specific antibodies of the IgM class were detected by the newly developed ELISA in 9 of 15 sera obtained from dogs with symptoms of acute distemper. In leucocytes of 5 of the 15 dogs (all of which were also IgM positive) CDV RNA was detected by reverse transcription (RT)-PCR. The recombinant capture-sandwich ELISA detecting N-specific antibodies of the

  11. Canine Distemper Virus Strains Circulating among North American Dogs▿

    PubMed Central

    Kapil, Sanjay; Allison, Robin W.; Johnston, Larry; Murray, Brandy L.; Holland, Steven; Meinkoth, Jim; Johnson, Bill

    2008-01-01

    Canine distemper virus (CDV) is a highly contagious virus that causes multisystemic disease in dogs. We received seven samples from dogs with CD from the United States during 2007. CDV isolates from these samples formed large, multinucleated syncytia in a Vero cell line expressing canine signaling lymphocyte activation molecule (SLAM). Based on the hemagglutinin gene sequences, the CDV isolates from three states (California, Missouri, and Oklahoma) formed two CDV genetic groups: group I (major; six of seven isolates) consisted of CDV isolates closely related to the European wildlife lineage of CDV, and group II (minor; one of seven isolates) was genetically related to the Arctic-like lineage of CDV. However, both CDV groups were genetically different from the current vaccine strains that belong to the American-1 lineage of the old (1930 to 1950) CDV isolates. PMID:18256210

  12. Phylogenetic analysis of Austrian canine distemper virus strains from clinical samples from dogs and wild carnivores.

    PubMed

    Benetka, V; Leschnik, M; Affenzeller, N; Möstl, K

    2011-04-01

    Austrian field cases of canine distemper (14 dogs, one badger [Meles meles] and one stone marten [Martes foina]) from 2002 to 2007 were investigated and the case histories were summarised briefly. Phylogenetic analysis of fusion (F) and haemagglutinin (H) gene sequences revealed different canine distemper virus (CDV) lineages circulating in Austria. The majority of CDV strains detected from 2002 to 2004 were well embedded in the European lineage. One Austrian canine sample detected in 2003, with a high similarity to Hungarian sequences from 2005 to 2006, could be assigned to the Arctic group (phocine distemper virus type 2-like). The two canine sequences from 2007 formed a clearly distinct group flanked by sequences detected previously in China and the USA on an intermediate position between the European wildlife and the Asia-1 cluster. The Austrian wildlife strains (2006 and 2007) could be assigned to the European wildlife group and were most closely related to, yet clearly different from, the 2007 canine samples. To elucidate the epidemiological role of Austrian wildlife in the transmission of the disease to dogs and vice versa, H protein residues related to receptor and host specificity (residues 530 and 549) were analysed. All samples showed the amino acids expected for their host of origin, with the exception of a canine sequence from 2007, which had an intermediate position between wildlife and canine viral strains. In the period investigated, canine strains circulating in Austria could be assigned to four different lineages reflecting both a high diversity and probably different origins of virus introduction to Austria in different years.

  13. Establishment of a Rescue System for Canine Distemper Virus

    PubMed Central

    Gassen, Uta; Collins, Fergal M.; Duprex, W. Paul; Rima, Bert K.

    2000-01-01

    Canine distemper virus (CDV) has been rescued from a full-length cDNA clone. Besides Measles virus (MV) and Rinderpest virus, a third morbillivirus is now available for genetic analysis using reverse genetics. A plasmid p(+)CDV was constructed by sequential cloning using the Onderstepoort vaccine strain large-plaque-forming variant. The presence of a T7 promoter allowed transcription of full-length antigenomic RNA by a T7 RNA polymerase, which was provided by a host range mutant of vaccinia virus (MVA-T7). Plasmids expressing the nucleocapsid protein, the phosphoprotein, and the viral RNA-dependent RNA polymerase, also under control of a T7 promoter, have been generated. Infection of HeLa cells with MVA-T7 and subsequent transfection of p(+)CDV plus the helper plasmids led to syncytium formation and release of infectious recombinant (r) CDV. Comparison of the rescued virus with the parental virus revealed no major differences in the progression of infection or in the shape and size of syncytia. A genetic tag, consisting of two nucleotide changes within the coding region of the L protein, has been identified in the rCDV genome. Expression by rCDV of all the major viral structural proteins has been demonstrated by immunofluorescence. PMID:11044118

  14. Fatal canine distemper virus infection of giant pandas in China

    PubMed Central

    Feng, Na; Yu, Yicong; Wang, Tiecheng; Wilker, Peter; Wang, Jianzhong; Li, Yuanguo; Sun, Zhe; Gao, Yuwei; Xia, Xianzhu

    2016-01-01

    We report an outbreak of canine distemper virus (CDV) infection among endangered giant pandas (Ailuropoda melanoleuca). Five of six CDV infected giant pandas died. The surviving giant panda was previously vaccinated against CDV. Genomic sequencing of CDV isolated from one of the infected pandas (giant panda/SX/2014) suggests it belongs to the Asia-1 cluster. The hemagglutinin protein of the isolated virus and virus sequenced from lung samples originating from deceased giant pandas all possessed the substitutions V26M, T213A, K281R, S300N, P340Q, and Y549H. The presence of the Y549H substitution is notable as it is found at the signaling lymphocytic activation molecule (SLAM) receptor-binding site and has been implicated in the emergence of highly pathogenic CDV and host switching. These findings demonstrate that giant pandas are susceptible to CDV and suggest that surveillance and vaccination among all captive giant pandas are warranted to support conservation efforts for this endangered species. PMID:27310722

  15. Phocine Distemper Virus: Current Knowledge and Future Directions

    PubMed Central

    Duignan, Pádraig J.; Van Bressem, Marie-Françoise; Baker, Jason D.; Barbieri, Michelle; Colegrove, Kathleen M.; De Guise, Sylvain; de Swart, Rik L.; Di Guardo, Giovanni; Dobson, Andrew; Duprex, W. Paul; Early, Greg; Fauquier, Deborah; Goldstein, Tracey; Goodman, Simon J.; Grenfell, Bryan; Groch, Kátia R.; Gulland, Frances; Hall, Ailsa; Jensen, Brenda A.; Lamy, Karina; Matassa, Keith; Mazzariol, Sandro; Morris, Sinead E.; Nielsen, Ole; Rotstein, David; Rowles, Teresa K.; Saliki, Jeremy T.; Siebert, Ursula; Waltzek, Thomas; Wellehan, James F.X.

    2014-01-01

    Phocine distemper virus (PDV) was first recognized in 1988 following a massive epidemic in harbor and grey seals in north-western Europe. Since then, the epidemiology of infection in North Atlantic and Arctic pinnipeds has been investigated. In the western North Atlantic endemic infection in harp and grey seals predates the European epidemic, with relatively small, localized mortality events occurring primarily in harbor seals. By contrast, PDV seems not to have become established in European harbor seals following the 1988 epidemic and a second event of similar magnitude and extent occurred in 2002. PDV is a distinct species within the Morbillivirus genus with minor sequence variation between outbreaks over time. There is now mounting evidence of PDV-like viruses in the North Pacific/Western Arctic with serological and molecular evidence of infection in pinnipeds and sea otters. However, despite the absence of associated mortality in the region, there is concern that the virus may infect the large Pacific harbor seal and northern elephant seal populations or the endangered Hawaiian monk seals. Here, we review the current state of knowledge on PDV with particular focus on developments in diagnostics, pathogenesis, immune response, vaccine development, phylogenetics and modeling over the past 20 years. PMID:25533658

  16. Fatal canine distemper virus infection of giant pandas in China.

    PubMed

    Feng, Na; Yu, Yicong; Wang, Tiecheng; Wilker, Peter; Wang, Jianzhong; Li, Yuanguo; Sun, Zhe; Gao, Yuwei; Xia, Xianzhu

    2016-01-01

    We report an outbreak of canine distemper virus (CDV) infection among endangered giant pandas (Ailuropoda melanoleuca). Five of six CDV infected giant pandas died. The surviving giant panda was previously vaccinated against CDV. Genomic sequencing of CDV isolated from one of the infected pandas (giant panda/SX/2014) suggests it belongs to the Asia-1 cluster. The hemagglutinin protein of the isolated virus and virus sequenced from lung samples originating from deceased giant pandas all possessed the substitutions V26M, T213A, K281R, S300N, P340Q, and Y549H. The presence of the Y549H substitution is notable as it is found at the signaling lymphocytic activation molecule (SLAM) receptor-binding site and has been implicated in the emergence of highly pathogenic CDV and host switching. These findings demonstrate that giant pandas are susceptible to CDV and suggest that surveillance and vaccination among all captive giant pandas are warranted to support conservation efforts for this endangered species. PMID:27310722

  17. Phocine distemper virus: current knowledge and future directions.

    PubMed

    Duignan, Pádraig J; Van Bressem, Marie-Françoise; Baker, Jason D; Barbieri, Michelle; Colegrove, Kathleen M; De Guise, Sylvain; de Swart, Rik L; Di Guardo, Giovanni; Dobson, Andrew; Duprex, W Paul; Early, Greg; Fauquier, Deborah; Goldstein, Tracey; Goodman, Simon J; Grenfell, Bryan; Groch, Kátia R; Gulland, Frances; Hall, Ailsa; Jensen, Brenda A; Lamy, Karina; Matassa, Keith; Mazzariol, Sandro; Morris, Sinead E; Nielsen, Ole; Rotstein, David; Rowles, Teresa K; Saliki, Jeremy T; Siebert, Ursula; Waltzek, Thomas; Wellehan, James F X

    2014-12-01

    Phocine distemper virus (PDV) was first recognized in 1988 following a massive epidemic in harbor and grey seals in north-western Europe. Since then, the epidemiology of infection in North Atlantic and Arctic pinnipeds has been investigated. In the western North Atlantic endemic infection in harp and grey seals predates the European epidemic, with relatively small, localized mortality events occurring primarily in harbor seals. By contrast, PDV seems not to have become established in European harbor seals following the 1988 epidemic and a second event of similar magnitude and extent occurred in 2002. PDV is a distinct species within the Morbillivirus genus with minor sequence variation between outbreaks over time. There is now mounting evidence of PDV-like viruses in the North Pacific/Western Arctic with serological and molecular evidence of infection in pinnipeds and sea otters. However, despite the absence of associated mortality in the region, there is concern that the virus may infect the large Pacific harbor seal and northern elephant seal populations or the endangered Hawaiian monk seals. Here, we review the current state of knowledge on PDV with particular focus on developments in diagnostics, pathogenesis, immune response, vaccine development, phylogenetics and modeling over the past 20 years. PMID:25533658

  18. Phocine distemper virus: current knowledge and future directions.

    PubMed

    Duignan, Pádraig J; Van Bressem, Marie-Françoise; Baker, Jason D; Barbieri, Michelle; Colegrove, Kathleen M; De Guise, Sylvain; de Swart, Rik L; Di Guardo, Giovanni; Dobson, Andrew; Duprex, W Paul; Early, Greg; Fauquier, Deborah; Goldstein, Tracey; Goodman, Simon J; Grenfell, Bryan; Groch, Kátia R; Gulland, Frances; Hall, Ailsa; Jensen, Brenda A; Lamy, Karina; Matassa, Keith; Mazzariol, Sandro; Morris, Sinead E; Nielsen, Ole; Rotstein, David; Rowles, Teresa K; Saliki, Jeremy T; Siebert, Ursula; Waltzek, Thomas; Wellehan, James F X

    2014-12-01

    Phocine distemper virus (PDV) was first recognized in 1988 following a massive epidemic in harbor and grey seals in north-western Europe. Since then, the epidemiology of infection in North Atlantic and Arctic pinnipeds has been investigated. In the western North Atlantic endemic infection in harp and grey seals predates the European epidemic, with relatively small, localized mortality events occurring primarily in harbor seals. By contrast, PDV seems not to have become established in European harbor seals following the 1988 epidemic and a second event of similar magnitude and extent occurred in 2002. PDV is a distinct species within the Morbillivirus genus with minor sequence variation between outbreaks over time. There is now mounting evidence of PDV-like viruses in the North Pacific/Western Arctic with serological and molecular evidence of infection in pinnipeds and sea otters. However, despite the absence of associated mortality in the region, there is concern that the virus may infect the large Pacific harbor seal and northern elephant seal populations or the endangered Hawaiian monk seals. Here, we review the current state of knowledge on PDV with particular focus on developments in diagnostics, pathogenesis, immune response, vaccine development, phylogenetics and modeling over the past 20 years.

  19. Fatal canine distemper virus infection of giant pandas in China.

    PubMed

    Feng, Na; Yu, Yicong; Wang, Tiecheng; Wilker, Peter; Wang, Jianzhong; Li, Yuanguo; Sun, Zhe; Gao, Yuwei; Xia, Xianzhu

    2016-06-16

    We report an outbreak of canine distemper virus (CDV) infection among endangered giant pandas (Ailuropoda melanoleuca). Five of six CDV infected giant pandas died. The surviving giant panda was previously vaccinated against CDV. Genomic sequencing of CDV isolated from one of the infected pandas (giant panda/SX/2014) suggests it belongs to the Asia-1 cluster. The hemagglutinin protein of the isolated virus and virus sequenced from lung samples originating from deceased giant pandas all possessed the substitutions V26M, T213A, K281R, S300N, P340Q, and Y549H. The presence of the Y549H substitution is notable as it is found at the signaling lymphocytic activation molecule (SLAM) receptor-binding site and has been implicated in the emergence of highly pathogenic CDV and host switching. These findings demonstrate that giant pandas are susceptible to CDV and suggest that surveillance and vaccination among all captive giant pandas are warranted to support conservation efforts for this endangered species.

  20. The Hemagglutinin of Canine Distemper Virus Determines Tropism and Cytopathogenicity

    PubMed Central

    von Messling, Veronika; Zimmer, Gert; Herrler, Georg; Haas, Ludwig; Cattaneo, Roberto

    2001-01-01

    Canine distemper virus (CDV) and measles virus (MV) cause severe illnesses in their respective hosts. The viruses display a characteristic cytopathic effect by forming syncytia in susceptible cells. For CDV, the proficiency of syncytium formation varies among different strains and correlates with the degree of viral attenuation. In this study, we examined the determinants for the differential fusogenicity of the wild-type CDV isolate 5804Han89 (CDV5804), the small- and large-plaque-forming variants of the CDV vaccine strain Onderstepoort (CDVOS and CDVOL, respectively), and the MV vaccine strain Edmonston B (MVEdm). The cotransfection of different combinations of fusion (F) and hemagglutinin (H) genes in Vero cells indicated that the H protein is the main determinant of fusion efficiency. To verify the significance of this observation in the viral context, a reverse genetic system to generate recombinant CDVs was established. This system is based on a plasmid containing the full-length antigenomic sequence of CDVOS. The coding regions of the H proteins of all CDV strains and MVEdm were introduced into the CDV and MV genetic backgrounds, and recombinant viruses rCDV-H5804, rCDV-HOL, rCDV-HEdm, rMV-H5804, rMV-HOL, and rMV-HOS were recovered. Thus, the H proteins of the two morbilliviruses are interchangeable and fully functional in a heterologous complex. This is in contrast with the glycoproteins of other members of the family Paramyxoviridae, which do not function efficiently with heterologous partners. The fusogenicity, growth characteristics, and tropism of the recombinant viruses were examined and compared with those of the parental strains. All these characteristics were found to be predominantly mediated by the H protein regardless of the viral backbone used. PMID:11413309

  1. A recombinant canine distemper virus expressing a modified rabies virus glycoprotein induces immune responses in mice.

    PubMed

    Li, Zhili; Wang, Jigui; Yuan, Daoli; Wang, Shuang; Sun, Jiazeng; Yi, Bao; Hou, Qiang; Mao, Yaping; Liu, Weiquan

    2015-06-01

    Canine distemper virus (CDV) and rabies virus (RV) are two important pathogens of the dog. CDV, a member of the morbillivirus genus, has shown promise as an expression vector. The glycoprotein from RV is a main contributor to protective immunity and capable of eliciting the production of virus-neutralizing antibodies. In this study, we recovered an attenuated strain of canine distemper virus and constructed a recombinant virus, rCDV-RV-G, expressing a modified (R333Q) rabies virus glycoprotein (RV-G) of RV Flury strain LEP. RV-G expression by the recombinant viruses was confirmed. Furthermore, G was proved to be incorporated into the surface of CDV particles. While replication of the recombinant virus was slightly reduced compared with the parental CDV, it stably expressed the RV-G over ten serial passages. Inoculation of mice induced specific neutralizing antibodies against both RV-G and CDV. Therefore, the rCDV-RV-G has the potential as a vaccine that may be used to control rabies virus infection in dogs and other animals. PMID:25764477

  2. Neutralizing antibodies to phocine distemper virus in Atlantic walruses (Odobenus rosmarus rosmarus) from Arctic Canada.

    PubMed

    Duignan, P J; Saliki, J T; St Aubin, D J; House, J A; Geraci, J R

    1994-01-01

    The first evidence of phocine distemper virus (PDV) infection in Atlantic walruses (Odobenus rosmarus rosmarus) from Nottingham Island, Northwest Territories, Canada, is reported. Blood samples were collected from three male walruses killed by Inuit hunters in the fall of 1990. Differential virus neutralization test for each animal yielded higher titers against PDV than against other members of the Morbillivirus genus including canine distemper, peste des petits ruminants, rinderpest and measles viruses. Thus, PDV infection may be enzootic in walruses of the eastern Canadian Arctic.

  3. Fatal vaccine-induced canine distemper virus infection in black-footed ferrets

    USGS Publications Warehouse

    Carpenter, J.W.; Appel, M.J.G.; Erickson, R.C.; Novilla, M.N.

    1976-01-01

    Four black-footed ferrets that were live-trapped in South Dakota and transported to the Patuxent Wildlife Research Center died within 21 days after vaccination with modified live canine distemper virus. Immunofluorescence, European ferret inoculation, virus isolation attempts, and serum-neutralization tests indicated insufficient attenuation of the vaccine for this species.

  4. A canine distemper virus epidemic in Serengeti lions (Panthera leo).

    PubMed

    Roelke-Parker, M E; Munson, L; Packer, C; Kock, R; Cleaveland, S; Carpenter, M; O'Brien, S J; Pospischil, A; Hofmann-Lehmann, R; Lutz, H; Mwamengele, G L; Mgasa, M N; Machange, G A; Summers, B A; Appel, M J

    1996-02-01

    Canine distemper virus (CDV) is thought to have caused several fatal epidemics in canids within the Serengeti-Mara ecosystem of East Africa, affecting silver-backed jackals (Canis mesomelas) and bat-eared foxes (Otocyon megalotis) in 1978 (ref. 1), and African wild dogs (Lycaon pictus) in 1991 (refs 2, 3). The large, closely monitored Serengeti lion population was not affected in these epidemics. However, an epidemic caused by a morbillivirus closely related to CDV emerged abruptly in the lion population of the Serengeti National Park, Tanzania, in early 1994, resulting in fatal neurological disease characterized by grand mal seizures and myoclonus; the lions that died had encephalitis and pneumonia. Here we report the identification of CDV from these lions, and the close phylogenetic relationship between CDV isolates from lions and domestic dogs. By August 1994, 85% of the Serengeti lion population had anti-CDV antibodies, and the epidemic spread north to lions in the Maasai Mara National reserve, Kenya, and uncounted hyaenas, bat-eared foxes, and leopards were also affected.

  5. Canine Distemper Virus in Wild Felids of Costa Rica.

    PubMed

    Avendaño, Roberto; Barrueta, Flor; Soto-Fournier, Sofía; Chavarría, Max; Monge, Otto; Gutiérrez-Espeleta, Gustavo A; Chaves, Andrea

    2016-04-28

    Several highly infectious diseases can be transmitted through feces and cause elevated mortality among carnivore species. One such infectious agent, canine distemper virus (CDV; Paramyxoviridae: Morbillivirus), has been reported to affect wild carnivores, among them several felid species. We screened free-ranging and captive wild carnivores in Costa Rica for CDV. Between 2006 and 2012, we collected 306 fecal samples from 70 jaguars (Panther onca), 71 ocelots ( Leopardus pardalis ), five jaguarundis (Puma yaguaroundi), 105 pumas ( Puma concolor ), five margays ( Leopardus wiedii ), 23 coyotes ( Canis latrans ), and 27 undetermined Leopardus spp. We found CDV in six individuals: one captive jaguarundi (rescued in 2009), three free-ranging ocelots (samples collected in 2012), and two free-ranging pumas (samples collected in 2007). Phylogenetic analyses were performed using sequences of the phosphoprotein (P) gene. We provide evidence of CDV in wild carnivores in Costa Rica and sequence data from a Costa Rican CDV isolate, adding to the very few sequence data available for CDV isolates from wild Central American carnivores. PMID:26967127

  6. Measles Vaccination of Nonhuman Primates Provides Partial Protection against Infection with Canine Distemper Virus

    PubMed Central

    de Vries, Rory D.; Ludlow, Martin; Verburgh, R. Joyce; van Amerongen, Geert; Yüksel, Selma; Nguyen, D. Tien; McQuaid, Stephen; Osterhaus, Albert D. M. E.; Duprex, W. Paul

    2014-01-01

    ABSTRACT Measles virus (MV) is being considered for global eradication, which would likely reduce compliance with MV vaccination. As a result, children will grow up without MV-specific immunity, creating a potential niche for closely related animal morbilliviruses such as canine distemper virus (CDV). Natural CDV infection causing clinical signs has never been reported in humans, but recent outbreaks in captive macaques have shown that CDV can cause disease in primates. We studied the virulence and tropism of recombinant CDV expressing enhanced green fluorescent protein in naive and measles-vaccinated cynomolgus macaques. In naive animals CDV caused viremia and fever and predominantly infected CD150+ lymphocytes and dendritic cells. Virus was reisolated from the upper and lower respiratory tracts, but infection of epithelial or neuronal cells was not detectable at the time points examined, and the infections were self-limiting. This demonstrates that CDV readily infects nonhuman primates but suggests that additional mutations are necessary to achieve full virulence in nonnatural hosts. Partial protection against CDV was observed in measles-vaccinated macaques, as demonstrated by accelerated control of virus replication and limited shedding from the upper respiratory tract. While neither CDV infection nor MV vaccination induced detectable cross-reactive neutralizing antibodies, MV-specific neutralizing antibody levels of MV-vaccinated macaques were boosted by CDV challenge infection, suggesting that cross-reactive VN epitopes exist. Rapid increases in white blood cell counts in MV-vaccinated macaques following CDV challenge suggested that cross-reactive cellular immune responses were also present. This study demonstrates that zoonotic morbillivirus infections can be controlled by measles vaccination. IMPORTANCE Throughout history viral zoonoses have had a substantial impact on human health. Given the drive toward global eradication of measles, it is essential to

  7. Vaccinia virus recombinants expressing either the measles virus fusion or hemagglutinin glycoprotein protect dogs against canine distemper virus challenge.

    PubMed

    Taylor, J; Pincus, S; Tartaglia, J; Richardson, C; Alkhatib, G; Briedis, D; Appel, M; Norton, E; Paoletti, E

    1991-08-01

    cDNA clones of the genes encoding either the hemagglutinin (HA) or fusion (F) proteins of the Edmonston strain of measles virus (MV) were expressed in vaccinia virus recombinants. Immunofluorescence analysis detected both proteins on the plasma membranes of unfixed cells as well as internally in fixed cells. Immunoprecipitation of metabolically radiolabeled infected-cell extracts by using specific sera demonstrated a 76-kDa HA polypeptide and gene products of 60, 44, and 23 kDa which correspond to a MV F precursor and cleavage products F0, F1, and F2, respectively. Neither recombinant induced cell fusion of Vero cells when inoculated individually, but efficient cell fusion was readily observed upon coinfection of cells with both recombinants. Inoculation of dogs with the vaccinia virus-MV F recombinant (VV-MVF) did not give rise to detectable MV-neutralizing antibody. Inoculation of dogs with the vaccinia virus-MV HA recombinant (VV-MVHA) or coinoculation with both recombinants (VV-MVF and VV-MVHA) induced significant MV-neutralizing titers that were increased following a booster inoculation. Inoculation of dogs with the vaccinia virus recombinants or with MV failed to induce canine distemper virus (CDV)-neutralizing antibodies. Upon challenge with a lethal dose of virulent CDV, signs of infection were observed in dogs inoculated with (VV-MVF). No symptoms of disease were observed in dogs that had been vaccinated with VV-MVHA or with VV-MVHA and VV-MVF and then challenged with CDV. All dogs vaccinated with the recombinant viruses as well as those inoculated with MV or a vaccine strain of CDV survived CDV challenge. PMID:1830113

  8. Vaccinia virus recombinants expressing either the measles virus fusion or hemagglutinin glycoprotein protect dogs against canine distemper virus challenge.

    PubMed Central

    Taylor, J; Pincus, S; Tartaglia, J; Richardson, C; Alkhatib, G; Briedis, D; Appel, M; Norton, E; Paoletti, E

    1991-01-01

    cDNA clones of the genes encoding either the hemagglutinin (HA) or fusion (F) proteins of the Edmonston strain of measles virus (MV) were expressed in vaccinia virus recombinants. Immunofluorescence analysis detected both proteins on the plasma membranes of unfixed cells as well as internally in fixed cells. Immunoprecipitation of metabolically radiolabeled infected-cell extracts by using specific sera demonstrated a 76-kDa HA polypeptide and gene products of 60, 44, and 23 kDa which correspond to a MV F precursor and cleavage products F0, F1, and F2, respectively. Neither recombinant induced cell fusion of Vero cells when inoculated individually, but efficient cell fusion was readily observed upon coinfection of cells with both recombinants. Inoculation of dogs with the vaccinia virus-MV F recombinant (VV-MVF) did not give rise to detectable MV-neutralizing antibody. Inoculation of dogs with the vaccinia virus-MV HA recombinant (VV-MVHA) or coinoculation with both recombinants (VV-MVF and VV-MVHA) induced significant MV-neutralizing titers that were increased following a booster inoculation. Inoculation of dogs with the vaccinia virus recombinants or with MV failed to induce canine distemper virus (CDV)-neutralizing antibodies. Upon challenge with a lethal dose of virulent CDV, signs of infection were observed in dogs inoculated with (VV-MVF). No symptoms of disease were observed in dogs that had been vaccinated with VV-MVHA or with VV-MVHA and VV-MVF and then challenged with CDV. All dogs vaccinated with the recombinant viruses as well as those inoculated with MV or a vaccine strain of CDV survived CDV challenge. Images PMID:1830113

  9. CANINE DISTEMPER VIRUS ANTIBODY TITERS IN DOMESTIC CATS AFTER DELIVERY OF A LIVE ATTENUATED VIRUS VACCINE.

    PubMed

    Ramsay, Edward; Sadler, Ryan; Rush, Robert; Seimon, Tracie; Tomaszewicz, Ania; Fleetwood, Ellen A; McAloose, Denise; Wilkes, Rebecca P

    2016-06-01

    Three methods for delivering a live attenuated canine distemper virus (CDV) vaccine to domestic cats ( Felis catus ) were investigated, as models for developing vaccination protocols for tigers (Panthera tigris). Twenty domestic cats were randomly divided into four treatment groups: saline injection (negative controls); and oral, intranasal, and subcutaneous vaccinates. Cats were injected with saline or a CDV vaccine (Nobivac DP, Merck) at wk 0 and 4. Blood and nasal swabs were collected at wk 0 (prior to the initial vaccination) and weekly thereafter for 9 wk. Urine samples were collected on wk 1 to 9 after initial vaccination. Forty-nine weeks following the initial vaccination series, three cats from the subcutaneous group and three cats from the intranasal group were revaccinated. Blood was collected immediately prior, and 7 and 21 days subsequent to revaccination. Nasal swabs and urine samples were collected from each cat prior to wk 49 revaccination and daily for 7 days thereafter. Nasal swabs and urine were analyzed by quantitative PCR for vaccine virus presence. Sera were tested for CDV antibodies by virus neutralization. All cats were sero-negative for CDV antibodies at the beginning of the study, and saline-injected cats remained sero-negative throughout the study. A dramatic anamnestic response was seen following wk 4 subcutaneous vaccinations, with titers peaking at wk 6 (geometric mean = 2,435.5). Following wk 49 revaccination, subcutaneous vaccinates again mounted impressive titers (wk 52 geometric mean = 2,048). Revaccination of the intranasal group cats at wk 49 produced a small increase in titers (wk 52 geometric mean = 203). CDV viral RNA was detected in six nasal swabs but no urine samples, demonstrating low viral shedding postvaccination. The strong antibody response to subcutaneous vaccination and the lack of adverse effects suggest this vaccine is safe and potentially protective against CDV infection in domestic cats.

  10. CANINE DISTEMPER VIRUS ANTIBODY TITERS IN DOMESTIC CATS AFTER DELIVERY OF A LIVE ATTENUATED VIRUS VACCINE.

    PubMed

    Ramsay, Edward; Sadler, Ryan; Rush, Robert; Seimon, Tracie; Tomaszewicz, Ania; Fleetwood, Ellen A; McAloose, Denise; Wilkes, Rebecca P

    2016-06-01

    Three methods for delivering a live attenuated canine distemper virus (CDV) vaccine to domestic cats ( Felis catus ) were investigated, as models for developing vaccination protocols for tigers (Panthera tigris). Twenty domestic cats were randomly divided into four treatment groups: saline injection (negative controls); and oral, intranasal, and subcutaneous vaccinates. Cats were injected with saline or a CDV vaccine (Nobivac DP, Merck) at wk 0 and 4. Blood and nasal swabs were collected at wk 0 (prior to the initial vaccination) and weekly thereafter for 9 wk. Urine samples were collected on wk 1 to 9 after initial vaccination. Forty-nine weeks following the initial vaccination series, three cats from the subcutaneous group and three cats from the intranasal group were revaccinated. Blood was collected immediately prior, and 7 and 21 days subsequent to revaccination. Nasal swabs and urine samples were collected from each cat prior to wk 49 revaccination and daily for 7 days thereafter. Nasal swabs and urine were analyzed by quantitative PCR for vaccine virus presence. Sera were tested for CDV antibodies by virus neutralization. All cats were sero-negative for CDV antibodies at the beginning of the study, and saline-injected cats remained sero-negative throughout the study. A dramatic anamnestic response was seen following wk 4 subcutaneous vaccinations, with titers peaking at wk 6 (geometric mean = 2,435.5). Following wk 49 revaccination, subcutaneous vaccinates again mounted impressive titers (wk 52 geometric mean = 2,048). Revaccination of the intranasal group cats at wk 49 produced a small increase in titers (wk 52 geometric mean = 203). CDV viral RNA was detected in six nasal swabs but no urine samples, demonstrating low viral shedding postvaccination. The strong antibody response to subcutaneous vaccination and the lack of adverse effects suggest this vaccine is safe and potentially protective against CDV infection in domestic cats. PMID:27468028

  11. SLAM- and Nectin-4-Independent Noncytolytic Spread of Canine Distemper Virus in Astrocytes

    PubMed Central

    Alves, Lisa; Khosravi, Mojtaba; Avila, Mislay; Ader-Ebert, Nadine; Bringolf, Fanny; Zurbriggen, Andreas; Vandevelde, Marc

    2015-01-01

    ABSTRACT Measles and canine distemper viruses (MeV and CDV, respectively) first replicate in lymphatic and epithelial tissues by using SLAM and nectin-4 as entry receptors, respectively. The viruses may also invade the brain to establish persistent infections, triggering fatal complications, such as subacute sclerosis pan-encephalitis (SSPE) in MeV infection or chronic, multiple sclerosis-like, multifocal demyelinating lesions in the case of CDV infection. In both diseases, persistence is mediated by viral nucleocapsids that do not require packaging into particles for infectivity but are directly transmitted from cell to cell (neurons in SSPE or astrocytes in distemper encephalitis), presumably by relying on restricted microfusion events. Indeed, although morphological evidence of fusion remained undetectable, viral fusion machineries and, thus, a putative cellular receptor, were shown to contribute to persistent infections. Here, we first showed that nectin-4-dependent cell-cell fusion in Vero cells, triggered by a demyelinating CDV strain, remained extremely limited, thereby supporting a potential role of nectin-4 in mediating persistent infections in astrocytes. However, nectin-4 could not be detected in either primary cultured astrocytes or the white matter of tissue sections. In addition, a bioengineered “nectin-4-blind” recombinant CDV retained full cell-to-cell transmission efficacy in primary astrocytes. Combined with our previous report demonstrating the absence of SLAM expression in astrocytes, these findings are suggestive for the existence of a hitherto unrecognized third CDV receptor expressed by glial cells that contributes to the induction of noncytolytic cell-to-cell viral transmission in astrocytes. IMPORTANCE While persistent measles virus (MeV) infection induces SSPE in humans, persistent canine distemper virus (CDV) infection causes chronic progressive or relapsing demyelination in carnivores. Common to both central nervous system (CNS

  12. Optic neuritis caused by canine distemper virus in a Jack Russell terrier.

    PubMed

    Richards, Tara R; Whelan, Nick C; Pinard, Chantale L; Alcala, Fernanda Castillo; Wolfe, Katheryn C

    2011-04-01

    An atypical case of canine distemper (CD) was diagnosed in a vaccinated healthy adult dog. The patient was presented circling, seizuring, and blind. Postmortem examination resulted in a diagnosis of CD. Optic neuritis was diagnosed, a finding not previously described in the context of CD virus infection presenting solely with neurological signs.

  13. Optic neuritis caused by canine distemper virus in a Jack Russell terrier

    PubMed Central

    Richards, Tara R.; Whelan, Nick C.; Pinard, Chantale L.; Alcala, Fernanda Castillo; Wolfe, Katheryn C.

    2011-01-01

    An atypical case of canine distemper (CD) was diagnosed in a vaccinated healthy adult dog. The patient was presented circling, seizuring, and blind. Postmortem examination resulted in a diagnosis of CD. Optic neuritis was diagnosed, a finding not previously described in the context of CD virus infection presenting solely with neurological signs. PMID:21731093

  14. Characteristics of a Canine Distemper Virus Outbreak in Dichato, Chile Following the February 2010 Earthquake

    PubMed Central

    Garde, Elena; Pérez, Guillermo; Acosta-Jamett, Gerardo; Bronsvoort, Barend Mark

    2013-01-01

    Simple Summary A disease outbreak in domestic dogs was reported by a coastal Chilean community following the February 2010 earthquake and tsunami. Using clinical exams and diagnostic testing, canine distemper virus was confirmed. Most dogs seen had never been vaccinated, and the majority of those with positive results were recorded in dogs less than two years of age. There were no facilities to contain or treat dogs locally, and no plan to shelter free-roaming dogs. This observational study demonstrates the great need for disaster preparedness planning in developing countries that includes companion animals. Abstract Following the earthquake and tsunami disaster in Chile in February 2010, residents of Dichato reported high morbidity and mortality in dogs, descriptions of which resembled canine distemper virus (CDV). To assess the situation, free vaccine clinics were offered in April and May. Owner information, dog history and signalment were gathered; dogs received physical examinations and vaccines protecting against CDV, and other common canine pathogens. Blood was collected to screen for IgM antibodies to CDV. In total, 208 dogs received physical exams and vaccines were given to 177. IgM antibody titres to CDV were obtained for 104 dogs. Fifty-four dogs (51.9%) tested positive for CDV at the cut off titre of >1:50, but a total of 91.4% of dogs had a detectable titre >1:10. Most of the positive test results were in dogs less than 2 years of age; 33.5% had been previously vaccinated against CDV, and owners of 84 dogs (42.2%) reported clinical signs characteristic of CDV in their dogs following the disaster. The presence of endemic diseases in dog populations together with poor pre-disaster free-roaming dog management results in a potential for widespread negative effects following disasters. Creation of preparedness plans that include animal welfare, disease prevention and mitigation should be developed. PMID:26479537

  15. Distinct cell tropism of canine distemper virus strains to adult olfactory ensheathing cells and Schwann cells in vitro.

    PubMed

    Techangamsuwan, Somporn; Haas, Ludwig; Rohn, Karl; Baumgärtner, Wolfgang; Wewetzer, Konstantin

    2009-09-01

    Canine distemper virus (CDV) can enter the brain via infection of olfactory neurons. Whether olfactory ensheathing cells (OECs) are also infected by CDV, and if yes, how they respond to the virus has remained enigmatic. Here, we exposed adult canine OECs in vitro to several attenuated (CDV-2544, CDV-R252, CDV-Ond, CDV-OndeGFP) and one virulent CDV strain (CDV-5804PeGFP) and studied their susceptibility compared to Schwann cells, a closely related cell type sharing the phagocytizing activity. We show that OECs and Schwann cells were infected by CDV strains albeit to different levels. Ten days post-infection (dpi), a mild to severe cytopathic effect ranging from single cell necrosis to layer detachment was noted. The percentage of infection increased during 10 dpi and viral progenies were detected in each culture using virus titration. Interestingly, CDV-2544, CDV-OndeGFP, and CDV-5804PeGFP predominantly infected OECs, while CDV-Ond targeted Schwann cells. No significant differences were found between the virulent and attenuated CDV strains. The observation of a CDV strain-specific cell tropism is evidence for significant molecular differences between OECs and Schwann cells. Whether these differences are either related to strain-specific distemper pathogenesis or support a role of OECs during CDV infection and virus spread needs to be addressed in future studies.

  16. Unusual Necrotizing Encephalitis in Raccoons and Skunks Concurrently Infected With Canine Distemper Virus and Sarcocystis sp.

    PubMed

    Kubiski, S V; Sisó, S; Church, M E; Cartoceti, A N; Barr, B; Pesavento, P A

    2016-05-01

    Canine distemper virus commonly infects free-ranging, terrestrial mesopredators throughout the United States. Due to the immunosuppressive effects of the virus, concurrent opportunistic infections are also common. Among these, secondary systemic protozoal infections have been described in a number of species. We report an unusual presentation of necrotizing encephalitis associated withSarcocystissp in four raccoons and one skunk concurrently infected with canine distemper virus. Lesions were characterized by variably sized necrotizing cavitations composed of abundant mineral admixed with inflammatory cells and protozoa.Sarcocystissp was confirmed via immunohistochemistry using a monoclonal antibody toSarcocystis neurona The pathologic changes are similar to lesions in human AIDS patients infected withToxoplasma gondii.

  17. Unusual Necrotizing Encephalitis in Raccoons and Skunks Concurrently Infected With Canine Distemper Virus and Sarcocystis sp.

    PubMed

    Kubiski, S V; Sisó, S; Church, M E; Cartoceti, A N; Barr, B; Pesavento, P A

    2016-05-01

    Canine distemper virus commonly infects free-ranging, terrestrial mesopredators throughout the United States. Due to the immunosuppressive effects of the virus, concurrent opportunistic infections are also common. Among these, secondary systemic protozoal infections have been described in a number of species. We report an unusual presentation of necrotizing encephalitis associated withSarcocystissp in four raccoons and one skunk concurrently infected with canine distemper virus. Lesions were characterized by variably sized necrotizing cavitations composed of abundant mineral admixed with inflammatory cells and protozoa.Sarcocystissp was confirmed via immunohistochemistry using a monoclonal antibody toSarcocystis neurona The pathologic changes are similar to lesions in human AIDS patients infected withToxoplasma gondii. PMID:26374278

  18. Morbillivirus Experimental Animal Models: Measles Virus Pathogenesis Insights from Canine Distemper Virus

    PubMed Central

    da Fontoura Budaszewski, Renata; von Messling, Veronika

    2016-01-01

    Morbilliviruses share considerable structural and functional similarities. Even though disease severity varies among the respective host species, the underlying pathogenesis and the clinical signs are comparable. Thus, insights gained with one morbillivirus often apply to the other members of the genus. Since the Canine distemper virus (CDV) causes severe and often lethal disease in dogs and ferrets, it is an attractive model to characterize morbillivirus pathogenesis mechanisms and to evaluate the efficacy of new prophylactic and therapeutic approaches. This review compares the cellular tropism, pathogenesis, mechanisms of persistence and immunosuppression of the Measles virus (MeV) and CDV. It then summarizes the contributions made by studies on the CDV in dogs and ferrets to our understanding of MeV pathogenesis and to vaccine and drugs development. PMID:27727184

  19. Canine distemper virus infection among wildlife before and after the epidemic

    PubMed Central

    SUZUKI, Junko; NISHIO, Yohei; KAMEO, Yuki; TERADA, Yutaka; KUWATA, Ryusei; SHIMODA, Hiroshi; SUZUKI, Kazuo; MAEDA, Ken

    2015-01-01

    In 2007–2008, a canine distemper virus (CDV) epidemic occurred among wild animals in Wakayama Prefecture, Japan, and many mammals, including the wild boar and deer, were infected. In this study, CDV prevalence among wild animals was surveyed before and after the epidemic. At first, an enzyme-linked immunosorbent assay (ELISA) with horseradish peroxidase-conjugated protein A/G was established to detect CDV antibodies in many mammalian species. This established ELISA was available for testing dogs, raccoons and raccoon dogs as well as virus-neutralization test. Next, a serological survey of wild mammalians was conducted, and it was indicated that many wild mammalians, particularly raccoons, were infected with CDV during the epidemic, but few were infected before and after the epidemic. On the other hand, many raccoon dogs died during the epidemic, but CDV remained prevalent in the remaining population, and a small epidemic occurred in raccoon dogs in 2012–2013. These results indicated that the epidemic of 2007–2008 may have been intensified by transmission to raccoons. PMID:26074342

  20. Epizootiological investigations of canine distemper virus in free-ranging carnivores from Germany.

    PubMed

    Frölich, K; Czupalla, O; Haas, L; Hentschke, J; Dedek, J; Fickel, J

    2000-06-12

    Canine distemper virus (CDV) infects a broad range of carnivores. To assess whether wild carnivores may play a role in the epidemiology of CDV in domestic dogs in Germany, the seroprevalence of CDV was determined. In sera from red foxes (30 of 591 (5%)) and stone martens (2 of 10 (20%)) antiviral antibodies were detected using a neutralization assay, whereas sera of raccoons, two mink, one pine marten and one raccoon dog were negative. In foxes, there was a significantly higher prevalence in urban and suburban compared to rural regions. When testing lung and spleen tissue samples (fox, badger, stone marten, polecat, raccoon dog) 13 of 253 (5.1%) foxes, 2 of 13 (15.4%) stone martens and 2 of 6 (33%) badgers were virus positive using RT-PCR. Phylogenetic analysis based on partial sequences of the F gene revealed a distinct relatedness to canine CDV isolates. Together, the data support the concept of transmission of CDV between domestic dogs and wild carnivores. PMID:10831852

  1. Canine distemper virus infection among wildlife before and after the epidemic.

    PubMed

    Suzuki, Junko; Nishio, Yohei; Kameo, Yuki; Terada, Yutaka; Kuwata, Ryusei; Shimoda, Hiroshi; Suzuki, Kazuo; Maeda, Ken

    2015-11-01

    In 2007-2008, a canine distemper virus (CDV) epidemic occurred among wild animals in Wakayama Prefecture, Japan, and many mammals, including the wild boar and deer, were infected. In this study, CDV prevalence among wild animals was surveyed before and after the epidemic. At first, an enzyme-linked immunosorbent assay (ELISA) with horseradish peroxidase-conjugated protein A/G was established to detect CDV antibodies in many mammalian species. This established ELISA was available for testing dogs, raccoons and raccoon dogs as well as virus-neutralization test. Next, a serological survey of wild mammalians was conducted, and it was indicated that many wild mammalians, particularly raccoons, were infected with CDV during the epidemic, but few were infected before and after the epidemic. On the other hand, many raccoon dogs died during the epidemic, but CDV remained prevalent in the remaining population, and a small epidemic occurred in raccoon dogs in 2012-2013. These results indicated that the epidemic of 2007-2008 may have been intensified by transmission to raccoons.

  2. Importance of canine distemper virus (CDV) infection in free-ranging Iberian lynxes (Lynx pardinus).

    PubMed

    Meli, Marina L; Simmler, Pascale; Cattori, Valentino; Martínez, Fernando; Vargas, Astrid; Palomares, Francisco; López-Bao, José V; Simón, Miguel A; López, Guillermo; León-Vizcaino, Luis; Hofmann-Lehmann, Regina; Lutz, Hans

    2010-11-20

    Canine distemper virus (CDV) is a morbillivirus that is the etiological agent of one of the most important viral diseases affecting canids and an expanding range of other carnivores. Using real-time RT-PCR, CDV RNA was detected in organs of an Iberian lynx (Lynx pardinus) found dead in the Doñana National Park, Southwestern Andalusia, Spain. This finding may be of great importance for the conservation of the species; at present the Iberian lynx is the most critically endangered wild felid. The aim of the present study was to elucidate the significance of CDV for the Iberian lynx population. High viral loads were evident in the dead lynx, suggesting an etiological involvement of CDV in its death. When carnivores from the same region were analyzed by CDV RT-PCR, a stone marten (Martes foina) was positive. Phylogenetic analyses demonstrated high identity of the two detected CDVs and a close relationship to the European dog lineage of CDV. Antibodies to CDV were detected in 14.8% of 88 tested free-ranging Iberian lynxes. The sample seroprevalence was significantly higher in lynxes from the Doñana Natural Space (22.9%) than Sierra Morena (5%). The stone marten and a red fox (Vulpes vulpes) also tested seropositive. In conclusion, CDV is present in the Iberian lynx population, especially in the Doñana region, with sporadic cases of disease. To reduce the infectious pressure of CDV on this endangered population, a mass dog vaccination should be considered.

  3. Antibody study in canine distemper virus nucleocapsid protein gene-immunized mice.

    PubMed

    Yuan, B; Li, X Y; Zhu, T; Yuan, L; Hu, J P; Chen, J; Gao, W; Ren, W Z

    2015-01-01

    The gene for the nucleocapsid (N) protein of canine distemper virus was cloned into the pMD-18T vector, and positive recombinant plasmids were obtained by enzyme digestion and sequencing. After digestion by both EcoRI and KpnI, the plasmid was directionally cloned into the eukaryotic expression vector pcDNA; the positive clone pcDNA-N was screened by electrophoresis and then transfected into COS-7 cells. Immunofluorescence analysis results showed that the canine distemper virus N protein was expressed in the cytoplasm of transfected COS-7 cells. After emulsification in Freund's adjuvant, the recombinant plasmid pcDNA-N was injected into the abdominal cavity of 8-week-old BABL/c mice, with the pcDNA original vector used as a negative control. Mice were immunized 3 times every 2 weeks. The blood of immunized mice was drawn 2 weeks after completing the immunizations to measure titer levels. The antibody titer in the pcDNA-N test was 10(1.62 ± 0.164), while in the control group this value was 10(0.52 ± 0.56), indicating that specific humoral immunity was induced in canine distemper virus nucleocapsid protein-immunized mice. PMID:25966074

  4. Duration of Maternal Antibodies against Canine Distemper Virus and Hendra Virus in Pteropid Bats.

    PubMed

    Epstein, Jonathan H; Baker, Michelle L; Zambrana-Torrelio, Carlos; Middleton, Deborah; Barr, Jennifer A; Dubovi, Edward; Boyd, Victoria; Pope, Brian; Todd, Shawn; Crameri, Gary; Walsh, Allyson; Pelican, Katey; Fielder, Mark D; Davies, Angela J; Wang, Lin-Fa; Daszak, Peter

    2013-01-01

    Old World frugivorous bats have been identified as natural hosts for emerging zoonotic viruses of significant public health concern, including henipaviruses (Nipah and Hendra virus), Ebola virus, and Marburg virus. Epidemiological studies of these viruses in bats often utilize serology to describe viral dynamics, with particular attention paid to juveniles, whose birth increases the overall susceptibility of the population to a viral outbreak once maternal immunity wanes. However, little is understood about bat immunology, including the duration of maternal antibodies in neonates. Understanding duration of maternally derived immunity is critical for characterizing viral dynamics in bat populations, which may help assess the risk of spillover to humans. We conducted two separate studies of pregnant Pteropus bat species and their offspring to measure the half-life and duration of antibodies to 1) canine distemper virus antigen in vaccinated captive Pteropus hypomelanus; and 2) Hendra virus in wild-caught, naturally infected Pteropus alecto. Both of these pteropid bat species are known reservoirs for henipaviruses. We found that in both species, antibodies were transferred from dam to pup. In P. hypomelanus pups, titers against CDV waned over a mean period of 228.6 days (95% CI: 185.4-271.8) and had a mean terminal phase half-life of 96.0 days (CI 95%: 30.7-299.7). In P. alecto pups, antibodies waned over 255.13 days (95% CI: 221.0-289.3) and had a mean terminal phase half-life of 52.24 days (CI 95%: 33.76-80.83). Each species showed a duration of transferred maternal immunity of between 7.5 and 8.5 months, which was longer than has been previously estimated. These data will allow for more accurate interpretation of age-related Henipavirus serological data collected from wild pteropid bats.

  5. Duration of Maternal Antibodies against Canine Distemper Virus and Hendra Virus in Pteropid Bats

    PubMed Central

    Zambrana-Torrelio, Carlos; Middleton, Deborah; Barr, Jennifer A.; DuBovi, Edward; Boyd, Victoria; Pope, Brian; Todd, Shawn; Crameri, Gary; Walsh, Allyson; Pelican, Katey; Fielder, Mark D.; Davies, Angela J.; Wang, Lin-Fa; Daszak, Peter

    2013-01-01

    Old World frugivorous bats have been identified as natural hosts for emerging zoonotic viruses of significant public health concern, including henipaviruses (Nipah and Hendra virus), Ebola virus, and Marburg virus. Epidemiological studies of these viruses in bats often utilize serology to describe viral dynamics, with particular attention paid to juveniles, whose birth increases the overall susceptibility of the population to a viral outbreak once maternal immunity wanes. However, little is understood about bat immunology, including the duration of maternal antibodies in neonates. Understanding duration of maternally derived immunity is critical for characterizing viral dynamics in bat populations, which may help assess the risk of spillover to humans. We conducted two separate studies of pregnant Pteropus bat species and their offspring to measure the half-life and duration of antibodies to 1) canine distemper virus antigen in vaccinated captive Pteropus hypomelanus; and 2) Hendra virus in wild-caught, naturally infected Pteropus alecto. Both of these pteropid bat species are known reservoirs for henipaviruses. We found that in both species, antibodies were transferred from dam to pup. In P. hypomelanus pups, titers against CDV waned over a mean period of 228.6 days (95% CI: 185.4–271.8) and had a mean terminal phase half-life of 96.0 days (CI 95%: 30.7–299.7). In P. alecto pups, antibodies waned over 255.13 days (95% CI: 221.0–289.3) and had a mean terminal phase half-life of 52.24 days (CI 95%: 33.76–80.83). Each species showed a duration of transferred maternal immunity of between 7.5 and 8.5 months, which was longer than has been previously estimated. These data will allow for more accurate interpretation of age-related Henipavirus serological data collected from wild pteropid bats. PMID:23826322

  6. Prevalence of antibodies to canine parvovirus and distemper virus in wolves in the Canadian Rocky Mountains.

    PubMed

    Nelson, Brynn; Hebblewhite, Mark; Ezenwa, Vanessa; Shury, Todd; Merrill, Evelyn H; Paquet, Paul C; Schmiegelow, Fiona; Seip, Dale; Skinner, Geoff; Webb, Nathan

    2012-01-01

    Wild carnivores are often exposed to diseases via contact with peridomestic host species that travel through the wildland-urban interfaces. To determine the antibody prevalences and relationships to human activity for two common canid pathogens, we sampled 99 wolves (Canis lupus) from 2000 to 2008 for antibodies to canine parvovirus (CPV) and canine distemper virus (CDV) in Banff and Jasper National Parks and surrounding areas of the Canadian Rockies. This population was the source for wolves reintroduced into the Northern Rockies of the US. Of 99 wolves sampled, 94 had detectable antibody to CPV (95%), 24 were antibody-positive for CDV (24%), and 24 had antibodies to both pathogens (24%). We tested whether antibody prevalences for CPV and CDV were higher closer to human activity (roads, town sites, First Nation reserves) and as a function of sex and age class. Wolves ≥2 yr old were more likely to be have antibodies to CPV. For CDV, male wolves, wolves ≥2 yr, and those closer to First Nation reserves were more likely to have antibodies. Overall, however, we found minimal support for human influence on antibody prevalence for CDV and CPV. The similarity between our antibody prevalence results and results from recent studies in Yellowstone National Park suggests that at least in the case of CDV, and perhaps CPV, these could be important pathogens with potential effects on wolf populations.

  7. Characteristics of a Canine Distemper Virus Outbreak in Dichato, Chile Following the February 2010 Earthquake.

    PubMed

    Garde, Elena; Pérez, Guillermo; Acosta-Jamett, Gerardo; Bronsvoort, Barend Mark

    2013-01-01

    Following the earthquake and tsunami disaster in Chile in February 2010, residents of Dichato reported high morbidity and mortality in dogs, descriptions of which resembled canine distemper virus (CDV). To assess the situation, free vaccine clinics were offered in April and May. Owner information, dog history and signalment were gathered; dogs received physical examinations and vaccines protecting against CDV, and other common canine pathogens. Blood was collected to screen for IgM antibodies to CDV. In total, 208 dogs received physical exams and vaccines were given to 177. IgM antibody titres to CDV were obtained for 104 dogs. Fifty-four dogs (51.9%) tested positive for CDV at the cut off titre of >1:50, but a total of 91.4% of dogs had a detectable titre >1:10. Most of the positive test results were in dogs less than 2 years of age; 33.5% had been previously vaccinated against CDV, and owners of 84 dogs (42.2%) reported clinical signs characteristic of CDV in their dogs following the disaster. The presence of endemic diseases in dog populations together with poor pre-disaster free-roaming dog management results in a potential for widespread negative effects following disasters. Creation of preparedness plans that include animal welfare, disease prevention and mitigation should be developed. PMID:26479537

  8. Prevalence of antibodies to canine parvovirus and distemper virus in wolves in the Canadian Rocky Mountains.

    PubMed

    Nelson, Brynn; Hebblewhite, Mark; Ezenwa, Vanessa; Shury, Todd; Merrill, Evelyn H; Paquet, Paul C; Schmiegelow, Fiona; Seip, Dale; Skinner, Geoff; Webb, Nathan

    2012-01-01

    Wild carnivores are often exposed to diseases via contact with peridomestic host species that travel through the wildland-urban interfaces. To determine the antibody prevalences and relationships to human activity for two common canid pathogens, we sampled 99 wolves (Canis lupus) from 2000 to 2008 for antibodies to canine parvovirus (CPV) and canine distemper virus (CDV) in Banff and Jasper National Parks and surrounding areas of the Canadian Rockies. This population was the source for wolves reintroduced into the Northern Rockies of the US. Of 99 wolves sampled, 94 had detectable antibody to CPV (95%), 24 were antibody-positive for CDV (24%), and 24 had antibodies to both pathogens (24%). We tested whether antibody prevalences for CPV and CDV were higher closer to human activity (roads, town sites, First Nation reserves) and as a function of sex and age class. Wolves ≥2 yr old were more likely to be have antibodies to CPV. For CDV, male wolves, wolves ≥2 yr, and those closer to First Nation reserves were more likely to have antibodies. Overall, however, we found minimal support for human influence on antibody prevalence for CDV and CPV. The similarity between our antibody prevalence results and results from recent studies in Yellowstone National Park suggests that at least in the case of CDV, and perhaps CPV, these could be important pathogens with potential effects on wolf populations. PMID:22247375

  9. Low Titers of Canine Distemper Virus Antibody in Wild Fishers (Martes pennanti) in the Eastern USA.

    PubMed

    Peper, Steven T; Peper, Randall L; Mitcheltree, Denise H; Kollias, George V; Brooks, Robert P; Stevens, Sadie S; Serfass, Thomas L

    2016-01-01

    Canine distemper virus (CDV) infects species in the order Carnivora. Members of the family Mustelidae are among the species most susceptible to CDV and have a high mortality rate after infection. Assessing an animal's pathogen or disease load prior to any reintroduction project is important to help protect the animal being reintroduced, as well as the wildlife and livestock in the area of relocation. We screened 58 fishers for CDV antibody prior to their release into Pennsylvania, US, as part of a reintroduction program. Five of the 58 (9%) fishers had a weak-positive reaction for CDV antibody at a dilution of 1:16. None of the fishers exhibited any clinical sign of canine distemper while being held prior to release.

  10. Low Titers of Canine Distemper Virus Antibody in Wild Fishers (Martes pennanti) in the Eastern USA.

    PubMed

    Peper, Steven T; Peper, Randall L; Mitcheltree, Denise H; Kollias, George V; Brooks, Robert P; Stevens, Sadie S; Serfass, Thomas L

    2016-01-01

    Canine distemper virus (CDV) infects species in the order Carnivora. Members of the family Mustelidae are among the species most susceptible to CDV and have a high mortality rate after infection. Assessing an animal's pathogen or disease load prior to any reintroduction project is important to help protect the animal being reintroduced, as well as the wildlife and livestock in the area of relocation. We screened 58 fishers for CDV antibody prior to their release into Pennsylvania, US, as part of a reintroduction program. Five of the 58 (9%) fishers had a weak-positive reaction for CDV antibody at a dilution of 1:16. None of the fishers exhibited any clinical sign of canine distemper while being held prior to release. PMID:26555109

  11. Broadly reactive pan-paramyxovirus reverse transcription polymerase chain reaction and sequence analysis for the detection of Canine distemper virus in a case of canine meningoencephalitis of unknown etiology

    PubMed Central

    Schatzberg, Scott J.; Li, Qiang; Porter, Brian F.; Barber, Renee M.; Claiborne, Mary Kate; Levine, Jonathan M.; Levine, Gwendolyn J.; Israel, Sarah K.; Young, Benjamin D.; Kiupel, Matti; Greene, Craig; Ruone, Susan; Anderson, Larry; Tong, Suxiang

    2016-01-01

    Despite the immunologic protection associated with routine vaccination protocols, Canine distemper virus (CDV) remains an important pathogen of dogs. Antemortem diagnosis of systemic CDV infection may be made by reverse transcription polymerase chain reaction (RT-PCR) and/or immunohistochemical testing for CDV antigen; central nervous system infection often requires postmortem confirmation via histopathology and immunohistochemistry. An 8-month-old intact male French Bulldog previously vaccinated for CDV presented with multifocal neurologic signs. Based on clinical and postmortem findings, the dog’s disease was categorized as a meningoencephalitis of unknown etiology. Broadly reactive, pan-paramyxovirus RT-PCR using consensus-degenerate hybrid oligonucleotide primers, combined with sequence analysis, identified CDV amplicons in the dog’s brain. Immunohistochemistry confirmed the presence of CDV antigens, and a specific CDV RT-PCR based on the phosphoprotein gene identified a wild-type versus vaccinal virus strain. This case illustrates the utility of broadly reactive PCR and sequence analysis for the identification of pathogens in diseases with unknown etiology. PMID:19901287

  12. EVALUATION OF TWO CANINE DISTEMPER VIRUS VACCINES IN CAPTIVE TIGERS (PANTHERA TIGRIS).

    PubMed

    Sadler, Ryan A; Ramsay, Edward; McAloose, Denise; Rush, Robert; Wilkes, Rebecca P

    2016-06-01

    Canine distemper virus (CDV) has caused clinical disease and death in nondomestic felids in both captive settings and in the wild. Outbreaks resulting in high mortality rates in tigers (Panthera tigris) have prompted some zoos to vaccinate tigers for CDV. In this study, six tigers received a recombinant canarypox-vectored CDV vaccine (1 ml s.c.) and were revaccinated with 3 ml s.c. (mean) 39 days later. Blood collection for CDV antibody detection via serum neutralization was performed on (mean) days 0, 26, and 66 post-initial vaccination. No tigers had detectable antibodies at days 0 or 26, and only two tigers had low (16 and 32) antibody titers at day 66. Eight additional tigers received a live, attenuated CDV vaccine (1 ml s.c.) on day 0 and were revaccinated with 1 ml s.c. (mean) 171 days later. Blood collection for CDV antibody detection via serum neutralization was performed on (mean) days 0, 26, 171, and 196. Seven of eight tigers receiving the live, attenuated vaccine had no detectable titers prior to vaccination, but all animals had titers of >128 (range 128-1,024) at day 26. At 171 days, all tigers still had detectable titers (geometric mean 69.8, range 16-256), and at 196 days (2 wk post-revaccination) all but two showed an increase to >128 (range 32-512). To determine safety, an additional 41 tigers were vaccinated with 2 ml of a recombinant vaccine containing only CDV components, and an additional 38 tigers received 1 ml of the live, attenuated vaccine, administered either subcutaneously or intramuscularly; no serious adverse effects were noted. Although both vaccines appear safe, the live, attenuated vaccine produced a stronger and more consistent serologic response in tigers.

  13. EVALUATION OF TWO CANINE DISTEMPER VIRUS VACCINES IN CAPTIVE TIGERS (PANTHERA TIGRIS).

    PubMed

    Sadler, Ryan A; Ramsay, Edward; McAloose, Denise; Rush, Robert; Wilkes, Rebecca P

    2016-06-01

    Canine distemper virus (CDV) has caused clinical disease and death in nondomestic felids in both captive settings and in the wild. Outbreaks resulting in high mortality rates in tigers (Panthera tigris) have prompted some zoos to vaccinate tigers for CDV. In this study, six tigers received a recombinant canarypox-vectored CDV vaccine (1 ml s.c.) and were revaccinated with 3 ml s.c. (mean) 39 days later. Blood collection for CDV antibody detection via serum neutralization was performed on (mean) days 0, 26, and 66 post-initial vaccination. No tigers had detectable antibodies at days 0 or 26, and only two tigers had low (16 and 32) antibody titers at day 66. Eight additional tigers received a live, attenuated CDV vaccine (1 ml s.c.) on day 0 and were revaccinated with 1 ml s.c. (mean) 171 days later. Blood collection for CDV antibody detection via serum neutralization was performed on (mean) days 0, 26, 171, and 196. Seven of eight tigers receiving the live, attenuated vaccine had no detectable titers prior to vaccination, but all animals had titers of >128 (range 128-1,024) at day 26. At 171 days, all tigers still had detectable titers (geometric mean 69.8, range 16-256), and at 196 days (2 wk post-revaccination) all but two showed an increase to >128 (range 32-512). To determine safety, an additional 41 tigers were vaccinated with 2 ml of a recombinant vaccine containing only CDV components, and an additional 38 tigers received 1 ml of the live, attenuated vaccine, administered either subcutaneously or intramuscularly; no serious adverse effects were noted. Although both vaccines appear safe, the live, attenuated vaccine produced a stronger and more consistent serologic response in tigers. PMID:27468029

  14. Identification of a new genotype of canine distemper virus circulating in America.

    PubMed

    Gámiz, César; Martella, Vito; Ulloa, Raúl; Fajardo, Raúl; Quijano-Hernandéz, Israel; Martínez, Simón

    2011-08-01

    Canine Distemper is a highly contagious viral systemic disease that affects a wide variety of terrestrial carnivores. Canine Distemper virus (CDV) appears genetically heterogeneous, markedly in the hemagglutinin protein (H), showing geographic patterns of diversification that are useful to monitor CDV molecular epidemiology. In Mexico the activity of canine distemper remains high in dogs, likely because vaccine prophylaxis coverage in canine population is under the levels required to control effectively the disease. By phylogenetic analysis based on the nucleoprotein (N) and on the H genes, Mexican CDV strains collected between 2007 and 2010 were distinguished into several genovariants, all which constituted a unique group, clearly distinct from field and vaccine strains circulating worldwide, but resembling a CDV strain, 19876, identified in Missouri, USA, 2004, that was genetically unrelated to other North-American CDV strains. Gathering information on the genetic heterogeneity of CDV on a global scale appears pivotal in order to investigate the origin and modalities of introduction of unusual/novel CDV strains, as well as to understand if vaccine breakthroughs or disease epidemics may be somewhat related to genetic/antigenic or biological differences between field and vaccine strains.

  15. Identification of a new genotype of canine distemper virus circulating in America.

    PubMed

    Gámiz, César; Martella, Vito; Ulloa, Raúl; Fajardo, Raúl; Quijano-Hernandéz, Israel; Martínez, Simón

    2011-08-01

    Canine Distemper is a highly contagious viral systemic disease that affects a wide variety of terrestrial carnivores. Canine Distemper virus (CDV) appears genetically heterogeneous, markedly in the hemagglutinin protein (H), showing geographic patterns of diversification that are useful to monitor CDV molecular epidemiology. In Mexico the activity of canine distemper remains high in dogs, likely because vaccine prophylaxis coverage in canine population is under the levels required to control effectively the disease. By phylogenetic analysis based on the nucleoprotein (N) and on the H genes, Mexican CDV strains collected between 2007 and 2010 were distinguished into several genovariants, all which constituted a unique group, clearly distinct from field and vaccine strains circulating worldwide, but resembling a CDV strain, 19876, identified in Missouri, USA, 2004, that was genetically unrelated to other North-American CDV strains. Gathering information on the genetic heterogeneity of CDV on a global scale appears pivotal in order to investigate the origin and modalities of introduction of unusual/novel CDV strains, as well as to understand if vaccine breakthroughs or disease epidemics may be somewhat related to genetic/antigenic or biological differences between field and vaccine strains. PMID:21713437

  16. Genotypic lineages and restriction fragment length polymorphism of canine distemper virus isolates in Thailand.

    PubMed

    Radtanakatikanon, Araya; Keawcharoen, Juthatip; Charoenvisal, Na Taya; Poovorawan, Yong; Prompetchara, Eakachai; Yamaguchi, Ryoji; Techangamsuwan, Somporn

    2013-09-27

    Canine distemper virus (CDV) is known to cause multisystemic disease in all families of terrestrial carnivores. Attenuated live vaccines have been used to control CDV in a variety of species for many decades, yet a number of CDV infections in vaccinated dogs are still observed. The aims of this study were to investigate the genetic diversity of CDV lineages based on phosphoprotein (P), hemagglutinin (H) and fusion protein (F) genes and to develop the restriction fragment length polymorphism (RFLP) technique for effective differentiation among individual wild-type and vaccine lineages in Thailand. Four commercial vaccine products, thirteen conjunctival swabs and various tissues from 9 necropsied dogs suspected of having CDV infections were included. Virus isolation was performed using Vero cell expressing canine signaling lymphocyte activation molecules (Vero-DST cells). Reverse-transcription polymerase chain reaction (RT-PCR) on 3 gene regions from the dog derived specimens and the vaccines were carried out, then RFLP analysis upon F-gene amplified fragments was developed. Nucleotide sequence and phylogenetic analysis were compared with other CDV lineages in Genbank. Phylogenetic relationships revealed that CDV field isolates were separated from the vaccine lineage and could be divided into two clusters; one of which belonged to the Asia-1 lineage and another, not related to any previous recognized lineages was proposed as 'Asia-4'. RFLP patterns demonstrating concordance with phylogenetic trees of the distemper virus allowed for differentiation between the Asia-1, Asia-4 and vaccine lineages. Thus, RFLP technique is able to effectively distinguish individual wild-type canine distemper virus from vaccine lineages in Thailand.

  17. Pathogenesis of canine distemper virus in experimentally infected raccoon dogs, foxes, and minks.

    PubMed

    Zhao, Jianjun; Shi, Ning; Sun, Yangang; Martella, Vito; Nikolin, Veljko; Zhu, Chunsheng; Zhang, Hailing; Hu, Bo; Bai, Xue; Yan, Xijun

    2015-10-01

    Canine distemper virus (CDV) infects a broad range of carnivores and causes a highly contagious disease with severe immunosuppression. The disease severity markedly varies in different species. To investigate the pathogenesis of CDV in raccoon dog (Nyctereutes procyonoides), fox (Vulpes vulpes) and mink (Neovison vison) species, three groups of CDV sero-negative animals were infected with CDV strain LN(10)1. This CDV strain belongs to the Asia-1 genotype, which is epidemiologically predominant in carnivores in China. CDV infection provoked marked differences in virulence in the three species that were studied. Raccoon dogs developed fever, severe conjunctivitis, and pathological lesions, with 100% (5/5) mortality and with high viral RNA loads in organs within 15 days post infection (dpi). In infected foxes, the onset of the disease was delayed, with 40% (2/5) mortality by 21 dpi. Infected minks developed only mild clinical signs and pathological lesions, and mortality was not observed. Raccoon dogs and foxes showed more severe immune suppression (lymphopenia, decreased lymphocyte proliferation, viremia and low-level virus neutralizing antibodies) than minks. We also observed a distinct pattern of cytokine mRNA transcripts at different times after infection. Decreased IFN-γ and IL-4 mRNA responses were evident in the animals with fatal disease, while up-regulation of these cytokines was observed in the animals surviving the infection. Increased TNF-α response was detected in animals with mild or severe clinical signs. Based on the results, we could distinguish three different patterns of disease after experimental CDV infection, e.g. a mild form in minks, a moderate form in foxes and a severe disease in raccoon dogs. The observed differences in susceptibility to CDV could be related to distinct host cytokine profiles. Comparative evaluation of CDV pathogenesis in various animal species is pivotal to generate models suitable for the evaluation of CDV

  18. Characterization of a novel Canine distemper virus causing disease in wildlife.

    PubMed

    Pope, Jenny P; Miller, Debra L; Riley, Matthew C; Anis, Eman; Wilkes, Rebecca P

    2016-09-01

    Canine distemper virus (CDV) is a common cause of a multisystemic disease in both domestic dogs and wildlife species, including raccoons and foxes. Outbreaks of CDV in domestic dogs in eastern Tennessee have occurred since 2012, and it was determined that these outbreaks resulted from a novel genotype of CDV. We hypothesized that this virus is also infecting area wildlife and may be a source of the virus for these outbreaks in dogs. From 2013 to 2014, autopsies were performed and tissues collected from raccoons (Procyon lotor; n = 50) and gray foxes (Urocyon cinereoargenteus; n = 8) for CDV testing. A real-time reverse transcription PCR was used to document the presence of CDV in tissue samples, and a portion of the virus was subsequently sequenced for phylogenetic analysis. A high percentage of wildlife, both with (86%) and without (55%) clinical signs, tested positive for CDV, with the majority (77%) testing positive for the novel genotype. Microscopic findings, including syncytia in the lungs and viral inclusion bodies in urothelium, astrocytes, neurons, and bronchiolar epithelium, were also consistent with canine distemper. Minimal inflammation in the central nervous system of affected animals was indicative of the acute neurologic form of the disease. Pneumonia and parasitism were also commonly found in CDV-infected animals. Based on these results, CDV appears to be prevalent in eastern Tennessee wildlife. Subclinical or clinically recovered shedders are a potential source of this novel genotype for domestic dogs, and this genotype is genetically distinct from vaccine strains. PMID:27400957

  19. Fatal combined infection with canine distemper virus and orthopoxvirus in a group of Asian marmots (Marmota caudata).

    PubMed

    Origgi, F C; Sattler, U; Pilo, P; Waldvogel, A S

    2013-09-01

    A fatal combined infection with canine distemper virus (CDV) and orthopoxvirus (OPXV) in Asian marmots (Marmota caudata) is reported in this article. A total of 7 Asian marmots from a small zoological garden in Switzerland were found dead in hibernation during a routine check in the winter of 2011. The marmots died in February 2011. No clinical signs of disease were observed at any time. The viruses were detected in all individuals for which the tissues were available (n = 3). Detection of the viruses was performed by reverse transcription polymerase chain reaction. The most consistent gross lesion was a neck and thorax edema. A necrotizing pharyngitis and a multifocal necrotizing pneumonia were observed histologically. Numerous large intracytoplasmic eosinophilic inclusions were seen in the epithelial cells of the pharynx, of the airways, and in the skin keratinocytes. Brain lesions were limited to mild multifocal gliosis. Phylogenetic analysis revealed that the marmot CDV strain was closely related to the clusters of CDVs detected in Switzerland in wild carnivores during a local outbreak in 2002 and the 2009-2010 nationwide epidemic, suggesting a spillover of this virus from wildlife. The OPXV was most closely related to a strain of cowpoxvirus, a poxvirus species considered endemic in Europe. This is the first reported instance of CDV infection in a rodent species and of a combined CDV and OPXV infection.

  20. Rabies virus and canine distemper virus in wild and domestic carnivores in Northern Kenya: are domestic dogs the reservoir?

    PubMed

    Prager, K C; Mazet, Jonna A K; Dubovi, Edward J; Frank, Laurence G; Munson, Linda; Wagner, Aaron P; Woodroffe, Rosie

    2012-12-01

    Rabies virus (RV) and canine distemper virus (CDV) can cause significant mortality in wild carnivore populations, and RV threatens human lives. We investigated serological patterns of exposure to CDV and RV in domestic dogs (Canis familiaris), African wild dogs (Lycaon pictus), black-backed jackals (Canis mesomelas), spotted hyenas (Crocuta crocuta), striped hyenas (Hyaena hyaena) and African lions (Panthera leo), over a 10-year period, in a Kenyan rangeland to assess the role domestic dogs may play in the transmission dynamics of these two important canid pathogens. Observed patterns of RV exposure suggested that repeated introduction, rather than maintenance, occurred in the wild carnivore species studied. However, RV appeared to have been maintained in domestic dogs: exposure was more likely in domestic dogs than in the wild carnivores; was detected consistently over time without variation among years; and was detected in juveniles (≤1-year-old) as well as adults (>1-year-old). We conclude that this domestic dog population could be a RV reservoir. By contrast, the absence of evidence of CDV exposure for each carnivore species examined in the study area, for specific years, suggested repeated introduction, rather than maintenance, and that CDV may require a larger reservoir population than RV. This reservoir could be a larger domestic dog population; another wildlife species; or a "metareservoir" consisting of multiple interconnected carnivore populations. Our findings suggest that RV risks to people and wild carnivores might be controlled by domestic dog vaccination, but that CDV control, if required, would need to target the species of concern. PMID:23459924

  1. Rabies virus and canine distemper virus in wild and domestic carnivores in Northern Kenya: are domestic dogs the reservoir?

    PubMed

    Prager, K C; Mazet, Jonna A K; Dubovi, Edward J; Frank, Laurence G; Munson, Linda; Wagner, Aaron P; Woodroffe, Rosie

    2012-12-01

    Rabies virus (RV) and canine distemper virus (CDV) can cause significant mortality in wild carnivore populations, and RV threatens human lives. We investigated serological patterns of exposure to CDV and RV in domestic dogs (Canis familiaris), African wild dogs (Lycaon pictus), black-backed jackals (Canis mesomelas), spotted hyenas (Crocuta crocuta), striped hyenas (Hyaena hyaena) and African lions (Panthera leo), over a 10-year period, in a Kenyan rangeland to assess the role domestic dogs may play in the transmission dynamics of these two important canid pathogens. Observed patterns of RV exposure suggested that repeated introduction, rather than maintenance, occurred in the wild carnivore species studied. However, RV appeared to have been maintained in domestic dogs: exposure was more likely in domestic dogs than in the wild carnivores; was detected consistently over time without variation among years; and was detected in juveniles (≤1-year-old) as well as adults (>1-year-old). We conclude that this domestic dog population could be a RV reservoir. By contrast, the absence of evidence of CDV exposure for each carnivore species examined in the study area, for specific years, suggested repeated introduction, rather than maintenance, and that CDV may require a larger reservoir population than RV. This reservoir could be a larger domestic dog population; another wildlife species; or a "metareservoir" consisting of multiple interconnected carnivore populations. Our findings suggest that RV risks to people and wild carnivores might be controlled by domestic dog vaccination, but that CDV control, if required, would need to target the species of concern.

  2. Canine distemper virus neutralization activity is low in human serum and it is sensitive to an amino acid substitution in the hemagglutinin protein

    SciTech Connect

    Zhang, Xinsheng; Wallace, Olivia L.; Domi, Arban; Wright, Kevin J.; Driscoll, Jonathan; Anzala, Omu; Sanders, Eduard J.; Kamali, Anatoli; Allen, Susan; Fast, Pat; Gilmour, Jill; Price, Matt A.; Parks, Christopher L.

    2015-08-15

    Serum was analyzed from 146 healthy adult volunteers in eastern Africa to evaluate measles virus (MV) and canine distemper virus (CDV) neutralizing antibody (nAb) prevalence and potency. MV plaque reduction neutralization test (PRNT) results indicated that all sera were positive for MV nAbs. Furthermore, the 50% neutralizing dose (ND50) for the majority of sera corresponded to antibody titers induced by MV vaccination. CDV nAbs titers were low and generally were detected in sera with high MV nAb titers. A mutant CDV was generated that was less sensitive to neutralization by human serum. The mutant virus genome had 10 nucleotide substitutions, which coded for single amino acid substitutions in the fusion (F) and hemagglutinin (H) glycoproteins and two substitutions in the large polymerase (L) protein. The H substitution occurred in a conserved region involved in receptor interactions among morbilliviruses, implying that this region is a target for cross-reactive neutralizing antibodies. - Highlights: • Screened 146 serum samples for measles virus (MV) and canine distemper virus (CDV) neutralizing antibody (nAb). • MV nAb is prevalent in the sera. • CDV neutralizing activity is generally low or absent and when detected it is present in sera with high MV nAb titers. • A neutralization-resistant CDV mutant was isolated using human serum selection. • A mutation was identified in the receptor-binding region of CDV hemagglutinin protein that confers the neutralization resistance.

  3. Canine Distemper Virus: an Emerging Disease in Wild Endangered Amur Tigers (Panthera tigris altaica)

    PubMed Central

    Seimon, Tracie A.; Miquelle, Dale G.; Chang, Tylis Y.; Newton, Alisa L.; Korotkova, Irina; Ivanchuk, Galina; Lyubchenko, Elena; Tupikov, Andre; Slabe, Evgeny; McAloose, Denise

    2013-01-01

    ABSTRACT Fewer than 500 Amur tigers (Panthera tigris altaica) remain in the wild. Due to low numbers and their solitary and reclusive nature, tiger sightings across their range in the Russian Far East and China are rare; sightings of sick tigers are rarer still. Serious neurologic disease observed in several wild tigers since 2001 suggested disease emergence in this endangered species. To investigate this possibility, histology, immunohistochemistry (IHC), in situ hybridization (ISH), and reverse transcription-PCR (RT-PCR) were performed on tissues from 5 affected tigers that died or were destroyed in 2001, 2004, or 2010. Our results reveal canine distemper virus (CDV) infection as the cause of neurologic disease in two tigers and definitively establish infection in a third. Nonsuppurative encephalitis with demyelination, eosinophilic nuclear viral inclusions, and positive immunolabeling for CDV by IHC and ISH were present in the two tigers with available brain tissue. CDV phosphoprotein (P) and hemagglutinin (H) gene products were obtained from brains of these two tigers by RT-PCR, and a short fragment of CDV P gene sequence was detected in lymph node tissue of a third tiger. Phylogenetically, Amur tiger CDV groups with an Arctic-like strain in Baikal seals (Phoca siberica). Our results, which include mapping the location of positive tigers and recognition of a cluster of cases in 2010, coupled with a lack of reported CDV antibodies in Amur tigers prior to 2000 suggest wide geographic distribution of CDV across the tiger range and recent emergence of CDV as a significant infectious disease threat to endangered Amur tigers in the Russian Far East. PMID:23943758

  4. A comparison of the immune responses of dogs exposed to canine distemper virus (CDV) — Differences between vaccinated and wild-type virus exposed dogs

    PubMed Central

    Perrone, Danielle; Bender, Scott; Niewiesk, Stefan

    2010-01-01

    Canine distemper virus (CDV)-specific immune response was measured in different dog populations. Three groups of vaccinated or wild-type virus exposed dogs were tested: dogs with a known vaccination history, dogs without a known vaccination history (shelter dogs), and dogs with potential exposure to wild-type CDV. The use of a T-cell proliferation assay demonstrated a detectable CDV-specific T-cell response from both spleen and blood lymphocytes of dogs. Qualitatively, antibody assays [enzyme-linked immunosorbent assay (ELISA) and neutralization assay] predicted the presence of a T-cell response well, although quantitatively neither antibody assays nor the T-cell assay correlated well with each other. An interesting finding from our study was that half of the dogs in shelters were not vaccinated (potentially posing a public veterinary health problem) and that antibody levels in dogs living in an environment with endemic CDV were lower than in vaccinated animals. PMID:20885846

  5. Coinfection with Hepatozoon sp. and Canine Distemper Virus in a Yellow-throated Marten ( Martes flavigula koreana) in Korea.

    PubMed

    Park, Surim; Choi, Ul Soo; Kim, Eun Ju; Lee, Jong Hyun; Lee, Hae Beom; Cho, Ho Seong; Kim, Wonil; Lim, Chae Woong; Kim, Bumseok

    2016-04-28

    We describe coinfection with Hepatozoon sp. and canine distemper virus (CDV) in a yellow-throated marten ( Martes flavigula koreana). We found Hepatozoon cysts in muscular tissue and viral inclusion bodies in the brain. Hepatozoon sp., and CDV was confirmed in blood and brain, respectively, by PCR.

  6. Canine Distemper Virus Epithelial Cell Infection Is Required for Clinical Disease but Not for Immunosuppression

    PubMed Central

    Sawatsky, Bevan; Wong, Xiao-Xiang; Hinkelmann, Sarah; Cattaneo, Roberto

    2012-01-01

    To characterize the importance of infection of epithelial cells for morbillivirus pathogenesis, we took advantage of the severe disease caused by canine distemper virus (CDV) in ferrets. To obtain a CDV that was unable to enter epithelial cells but retained the ability to enter immune cells, we transferred to its attachment (H) protein two mutations shown to interfere with the interaction of measles virus H with its epithelial receptor, human nectin-4. As expected for an epithelial receptor (EpR)-blind CDV, this virus infected dog and ferret epithelial cells inefficiently and did not cause cell fusion or syncytium formation. On the other hand, the EpR-blind CDV replicated in cells expressing canine signaling lymphocyte activation molecule (SLAM), the morbillivirus immune cell receptor, with similar kinetics to those of wild-type CDV. While ferrets infected with wild-type CDV died within 12 days after infection, after developing severe rash and fever, animals infected with the EpR-blind virus showed no clinical signs of disease. Nevertheless, both viruses spread rapidly and efficiently in immune cells, causing similar levels of leukopenia and inhibition of lymphocyte proliferation activity, two indicators of morbillivirus immunosuppression. Infection was documented for airway epithelia of ferrets infected with wild-type CDV but not for those of animals infected with the EpR-blind virus, and only animals infected with wild-type CDV shed virus. Thus, epithelial cell infection is necessary for clinical disease and efficient virus shedding but not for immunosuppression. PMID:22278252

  7. Plaque identification of strand-forming canine distemper virus by staphylococcal protein A-mediated reverse passive haemadsorption.

    PubMed

    Johnson, G C; Fulks, K; Krakowka, S

    1985-02-01

    The R252 neurotropic isolate of canine distemper virus (CDV) produces cytopathic effects (CPE) dominated by strand formation rather than by the formation of multinucleate giant cells. The lack of well-defined CPE and consequent rapid spread of infection throughout the cell monolayer has hindered plaque purification of this virus by conventional methods. However, the use of an immunological detection system which utilizes binding of hyperimmune dog serum to virus-infected cells, followed by the identification of those sites by staphylococcal Protein A-coupled sheep red blood cells (reverse passive haemadsorption) allowed infected foci in cell monolayers to be detected as early as 4 days after infection, coincident with the appearance of the first immunofluorescently identified viral foci. Foci of haemadsorption were specific to sites of CDV infection as demonstrated by blocking experiments. Material recovered from the plaques was successful in infecting Vero cells. Thus, immunologically mediated adsorption of Protein A coupled red blood cells can be used to identify and isolate foci of viral infection which exhibit minimal or no viral CPE without destroying viral replicative ability.

  8. Expression of a foreign gene by recombinant canine distemper virus recovered from cloned DNAs.

    PubMed

    Parks, Christopher L; Wang, Hai-Ping; Kovacs, Gerald R; Vasilakis, Nikos; Kowalski, Jacek; Nowak, Rebecca M; Lerch, Robert A; Walpita, Pramila; Sidhu, Mohinderjit S; Udem, Stephen A

    2002-02-26

    A canine distemper virus (CDV) genomic cDNA clone and expression plasmids required to establish a CDV rescue system were generated from a laboratory-adapted strain of the Onderstepoort vaccine virus. In addition, a CDV minireplicon was prepared and used in transient expression studies performed to identify optimal virus rescue conditions. Results from the transient expression experiments indicated that minireplicon-encoded reporter gene activity was increased when transfected cell cultures were maintained at 32 rather than 37 degrees C, and when the cellular stress response was induced by heat shock. Applying these findings to rescue of recombinant CDV (rCDV) resulted in efficient recovery of virus after transfected HEp2 or A549 cells were co-cultured with Vero cell monolayers. Nucleotide sequence determination and analysis of restriction site polymorphisms confirmed that rescued virus was rCDV. A rCDV strain also was engineered that contained the luciferase gene inserted between the P and M genes; this virus directed high levels of luciferase expression in infected cells. PMID:11864746

  9. Viral Oncolysis — Can Insights from Measles Be Transferred to Canine Distemper Virus?

    PubMed Central

    Lapp, Stefanie; Pfankuche, Vanessa M.; Baumgärtner, Wolfgang; Puff, Christina

    2014-01-01

    Neoplastic diseases represent one of the most common causes of death among humans and animals. Currently available and applied therapeutic options often remain insufficient and unsatisfactory, therefore new and innovative strategies and approaches are highly needed. Periodically, oncolytic viruses have been in the center of interest since the first anecdotal description of their potential usefulness as an anti-tumor treatment concept. Though first reports referred to an incidental measles virus infection causing tumor regression in a patient suffering from lymphoma several decades ago, no final treatment concept has been developed since then. However, numerous viruses, such as herpes-, adeno- and paramyxoviruses, have been investigated, characterized, and modified with the aim to generate a new anti-cancer treatment option. Among the different viruses, measles virus still represents a highly interesting candidate for such an approach. Numerous different tumors of humans including malignant lymphoma, lung and colorectal adenocarcinoma, mesothelioma, and ovarian cancer, have been studied in vitro and in vivo as potential targets. Moreover, several concepts using different virus preparations are now in clinical trials in humans and may proceed to a new treatment option. Surprisingly, only few studies have investigated viral oncolysis in veterinary medicine. The close relationship between measles virus (MV) and canine distemper virus (CDV), both are morbilliviruses, and the fact that numerous tumors in dogs exhibit similarities to their human counterpart, indicates that both the virus and species dog represent a highly interesting translational model for future research in viral oncolysis. Several recent studies support such an assumption. It is therefore the aim of the present communication to outline the mechanisms of morbillivirus-mediated oncolysis and to stimulate further research in this potentially expanding field of viral oncolysis in a highly suitable

  10. Cytotoxic T-lymphocyte activity specific for hemagglutinin (H) protein of canine distemper virus in dogs.

    PubMed

    Hirama, Kyoko; Togashi, Ken-ichi; Wakasa, Chiaki; Yoneda, Misako; Nishi, Toshiya; Endo, Yasuyuki; Miura, Ryuichi; Tsukiyama-Kohara, Kyoko; Kai, Chieko

    2003-01-01

    Cytotoxic T-lymphocyte (CTL) responses to hemagglutinin (H) protein of canine distemper virus (CDV) were evaluated in dogs using the replication-deficient adenovirus protein expression system. Skin fibroblasts were isolated from two dogs and were infected with recombinant adenovirus bearing the CDV-H gene (Ade-CDVH). CTL assay was performed using fibroblasts expressing CDV-H protein as target cells and peripheral blood lymphocytes (PBL) collected from the same dogs one week after immunization of CDV as effector cells. Specific cytotoxic activity was observed against autologous but not heterologous fibroblasts expressing CDV-H protein. These results indicate that the CTL epitope(s) were localized in the H protein. PMID:12576714

  11. Identification of canine helper T-cell epitopes from the fusion protein of canine distemper virus

    PubMed Central

    Ghosh, Souravi; Walker, John; Jackson, David C

    2001-01-01

    The fusion protein of canine distemper virus (CDV-F), a 662 amino-acid envelope protein, was used as the target molecule for identification of canine T helper (Th) epitopes. A library of 94 peptides, each 17 residues in length overlapping by 10 residues and covering the entire sequence of CDV-F, was screened using a lymphocyte proliferation assay with peripheral blood mononuclear cells (PBMC) obtained from dogs inoculated with canine distemper virus (CDV) vaccine. Initially we observed low and inconsistent proliferation of PBMC in response to these peptides, even when using cells obtained from dogs that had received multiple doses of CDV. Subsequently, the use of expanded cell populations derived by in vitro stimulation of canine PBMC with pools of peptides allowed the identification of a number of putative canine Th-epitopes within the protein sequence of CDV-F. There were two major clusters of Th-epitopes identified close to the cleavage site of the F0 fusion protein, while some others were scattered in both the F1 and F2 fragments of the protein. Some of these peptides, in particular peptide 35 (p35), were stimulatory in dogs of different breeds and ages. The identification of such promiscuous canine Th-epitopes encouraged us to assemble p35 in tandem with luteinising hormone releasing hormone (LHRH) a 10 amino-acid residue synthetic peptide representing a B-cell epitope which alone induces no antibody in dogs. The totally synthetic immunogen was able to induce the production of very high titres of antibodies against LHRH in all dogs tested. These results indicate that p35 could be an ideal candidate for use as a Th-epitope for use in outbred dogs. PMID:11576221

  12. Molecular phylogeography of canine distemper virus: Geographic origin and global spreading.

    PubMed

    Panzera, Yanina; Sarute, Nicolás; Iraola, Gregorio; Hernández, Martín; Pérez, Ruben

    2015-11-01

    Canine distemper virus (CDV) (Paramyxoviridae-Morbillivirus) is a worldwide spread virus causing a fatal systemic disease in a broad range of carnivore hosts. In this study we performed Bayesian inferences using 208 full-length hemagglutinin gene nucleotide sequences isolated in 16 countries during 37 years (1975-2011). The estimated time to the most recent common ancestor suggested that current CDV strains emerged in the United States in the 1880s. This ancestor diversified through time into two ancestral clades, the current America 1 lineage that recently spread to Asia, and other ancestral clade that diversified and spread worldwide to originate the remaining eight lineages characterized to date. The spreading of CDV was characterized by several migratory events with posterior local differentiation, and expansion of the virus host range. A significant genetic flow between domestic and wildlife hosts is displayed; being domestic hosts the main viral reservoirs worldwide. This study is an extensive and integrative description of spatio/temporal population dynamics of CDV lineages that provides a novel evolutionary paradigm about the origin and dissemination of the current strains of the virus.

  13. Canine distemper virus neutralization activity is low in human serum and it is sensitive to an amino acid substitution in the hemagglutinin protein.

    PubMed

    Zhang, Xinsheng; Wallace, Olivia L; Domi, Arban; Wright, Kevin J; Driscoll, Jonathan; Anzala, Omu; Sanders, Eduard J; Kamali, Anatoli; Karita, Etienne; Allen, Susan; Fast, Pat; Gilmour, Jill; Price, Matt A; Parks, Christopher L

    2015-08-01

    Serum was analyzed from 146 healthy adult volunteers in eastern Africa to evaluate measles virus (MV) and canine distemper virus (CDV) neutralizing antibody (nAb) prevalence and potency. MV plaque reduction neutralization test (PRNT) results indicated that all sera were positive for MV nAbs. Furthermore, the 50% neutralizing dose (ND50) for the majority of sera corresponded to antibody titers induced by MV vaccination. CDV nAbs titers were low and generally were detected in sera with high MV nAb titers. A mutant CDV was generated that was less sensitive to neutralization by human serum. The mutant virus genome had 10 nucleotide substitutions, which coded for single amino acid substitutions in the fusion (F) and hemagglutinin (H) glycoproteins and two substitutions in the large polymerase (L) protein. The H substitution occurred in a conserved region involved in receptor interactions among morbilliviruses, implying that this region is a target for cross-reactive neutralizing antibodies. PMID:25880113

  14. Canine distemper virus neutralization activity is low in human serum and it is sensitive to an amino acid substitution in the hemagglutinin protein.

    PubMed

    Zhang, Xinsheng; Wallace, Olivia L; Domi, Arban; Wright, Kevin J; Driscoll, Jonathan; Anzala, Omu; Sanders, Eduard J; Kamali, Anatoli; Karita, Etienne; Allen, Susan; Fast, Pat; Gilmour, Jill; Price, Matt A; Parks, Christopher L

    2015-08-01

    Serum was analyzed from 146 healthy adult volunteers in eastern Africa to evaluate measles virus (MV) and canine distemper virus (CDV) neutralizing antibody (nAb) prevalence and potency. MV plaque reduction neutralization test (PRNT) results indicated that all sera were positive for MV nAbs. Furthermore, the 50% neutralizing dose (ND50) for the majority of sera corresponded to antibody titers induced by MV vaccination. CDV nAbs titers were low and generally were detected in sera with high MV nAb titers. A mutant CDV was generated that was less sensitive to neutralization by human serum. The mutant virus genome had 10 nucleotide substitutions, which coded for single amino acid substitutions in the fusion (F) and hemagglutinin (H) glycoproteins and two substitutions in the large polymerase (L) protein. The H substitution occurred in a conserved region involved in receptor interactions among morbilliviruses, implying that this region is a target for cross-reactive neutralizing antibodies.

  15. Arctic lineage-canine distemper virus as a cause of death in Apennine wolves (Canis lupus) in Italy.

    PubMed

    Di Sabatino, Daria; Lorusso, Alessio; Di Francesco, Cristina E; Gentile, Leonardo; Di Pirro, Vincenza; Bellacicco, Anna Lucia; Giovannini, Armando; Di Francesco, Gabriella; Marruchella, Giuseppe; Marsilio, Fulvio; Savini, Giovanni

    2014-01-01

    Canine distemper virus (CDV) infection is a primary threat affecting a wide number of carnivore species, including wild animals. In January 2013, two carcasses of Apennine wolves (Canis lupus) were collected in Ortona dei Marsi (L'Aquila province, Italy) by the local Veterinary Services. CDV was immediately identified either by RT-PCR or immunohistochemistry in lung and central nervous tissue samples. At the same time, severe clinical signs consistent with CDV infection were identified and taped (Videos S1-S3) from three wolves rescued in the areas surrounding the National Parks of the Abruzzi region by the Veterinary Services. The samples collected from these symptomatic animals also turned out CDV positive by RT-PCR. So far, 30 carcasses of wolves were screened and CDV was detected in 20 of them. The sequencing of the haemagglutinin gene and subsequent phylogenetic analysis demonstrated that the identified virus belonged to the CDV Arctic lineage. Strains belonging to this lineage are known to circulate in Italy and in Eastern Europe amongst domestic dogs. To the best of our knowledge this is the first report of CDV Arctic lineage epidemics in the wild population in Europe.

  16. Arctic Lineage-Canine Distemper Virus as a Cause of Death in Apennine Wolves (Canis lupus) in Italy

    PubMed Central

    Di Sabatino, Daria; Lorusso, Alessio; Di Francesco, Cristina E.; Gentile, Leonardo; Di Pirro, Vincenza; Bellacicco, Anna Lucia; Giovannini, Armando; Di Francesco, Gabriella; Marruchella, Giuseppe; Marsilio, Fulvio; Savini, Giovanni

    2014-01-01

    Canine distemper virus (CDV) infection is a primary threat affecting a wide number of carnivore species, including wild animals. In January 2013, two carcasses of Apennine wolves (Canis lupus) were collected in Ortona dei Marsi (L'Aquila province, Italy) by the local Veterinary Services. CDV was immediately identified either by RT-PCR or immunohistochemistry in lung and central nervous tissue samples. At the same time, severe clinical signs consistent with CDV infection were identified and taped (Videos S1–S3) from three wolves rescued in the areas surrounding the National Parks of the Abruzzi region by the Veterinary Services. The samples collected from these symptomatic animals also turned out CDV positive by RT-PCR. So far, 30 carcasses of wolves were screened and CDV was detected in 20 of them. The sequencing of the haemagglutinin gene and subsequent phylogenetic analysis demonstrated that the identified virus belonged to the CDV Arctic lineage. Strains belonging to this lineage are known to circulate in Italy and in Eastern Europe amongst domestic dogs. To the best of our knowledge this is the first report of CDV Arctic lineage epidemics in the wild population in Europe. PMID:24465373

  17. Use of hydrophilic extra-viral domain of canine distemper virus H protein for enzyme-linked immunosorbent assay development

    PubMed Central

    Cho, Ki-hyun; Kim, Jeongmi; Yoo, Hyun-ah; Kim, Dae-hee; Park, Seung-yong; Song, Chang-seon; Choi, In-soo

    2014-01-01

    Simple methods for measuring the levels of serum antibody against canine distemper virus (CDV) would assist in the effective vaccination of dogs. To develop an enzyme-linked immunosorbent assay (ELISA) specific for CDV, we expressed hydrophilic extra-viral domain (HEVD) protein of the A75/17-CDV H gene in a pET 28a plasmid-based Escherichia (E.) coli vector system. Expression was confirmed by dot and Western blotting. We proposed that detection of E. coli-expressed H protein might be conformation-dependent because intensities of the reactions observed with these two methods varied. The H gene HEVD protein was further purified and used as an antigen for an ELISA. Samples from dogs with undetectable to high anti-CDV antibody titers were analyzed using this HEVD-specific ELISA and a commercial CDV antibody detection kit (ImmunoComb). Levels of HEVD antigenicity measured with the assays and immunochromatography correlated. These data indicated that the HEDV protein may be used as antigen to develop techniques for detecting antibodies against CDV. PMID:25234325

  18. Phylogenetic evidence of a new canine distemper virus lineage among domestic dogs in Colombia, South America.

    PubMed

    Espinal, Maria A; Díaz, Francisco J; Ruiz-Saenz, Julian

    2014-08-01

    Canine distemper virus (CDV) is a highly contagious viral disease of carnivores affecting both wild and domestic populations. The hemagglutinin gene, encoding for the attachment protein that determines viral tropism, shows high heterogeneity among strains, allowing for the distinction of ten different lineages distributed worldwide according to a geographic pattern. We obtained the sequences of the full-length H gene of 15 wild-type CDV strains circulating in domestic dog populations from the Aburrá Valley, Colombia. A phylogenetic analysis of H gene nucleotide sequences from Colombian CDV viruses along with field isolates from different geographic regions and vaccine strains was performed. Colombian wild-type viruses formed a distinct monophyletic cluster clearly separated from the previously identified wild-type and vaccine lineages, suggesting that a novel genetic variant, quite different from vaccines and other lineages, is circulating among dog populations in the Aburrá Valley. We propose naming this new lineage as "South America 3". This information indicates that there are at least three different CDV lineages circulating in domestic and wild carnivore populations in South America. The first one, renamed Europe/South America 1, circulates in Brazil and Uruguay; the second, South America 2, appears to be restricted to Argentina; and the third, South America 3, which comprises all the strains characterized in this study, may also be circulating in other northern countries of South America. PMID:24950886

  19. Recombinant Canine Distemper Virus Strain Snyder Hill Expressing Green or Red Fluorescent Proteins Causes Meningoencephalitis in the Ferret

    PubMed Central

    Ludlow, M.; Nguyen, D. T.; Silin, D.; Lyubomska, O.; de Vries, R. D.; von Messling, V.; McQuaid, S.; De Swart, R. L.

    2012-01-01

    The propensity of canine distemper virus (CDV) to spread to the central nervous system is one of the primary features of distemper. Therefore, we developed a reverse genetics system based on the neurovirulent Snyder Hill (SH) strain of CDV (CDVSH) and show that this virus rapidly circumvents the blood-brain and blood-cerebrospinal fluid (CSF) barriers to spread into the subarachnoid space to induce dramatic viral meningoencephalitis. The use of recombinant CDVSH (rCDVSH) expressing enhanced green fluorescent protein (EGFP) or red fluorescent protein (dTomato) facilitated the sensitive pathological assessment of routes of virus spread in vivo. Infection of ferrets with these viruses led to the full spectrum of clinical signs typically associated with distemper in dogs during a rapid, fatal disease course of approximately 2 weeks. Comparison with the ferret-adapted CDV5804P and the prototypic wild-type CDVR252 showed that hematogenous infection of the choroid plexus is not a significant route of virus spread into the CSF. Instead, viral spread into the subarachnoid space in rCDVSH-infected animals was triggered by infection of vascular endothelial cells and the hematogenous spread of virus-infected leukocytes from meningeal blood vessels into the subarachnoid space. This resulted in widespread infection of cells of the pia and arachnoid mater of the leptomeninges over large areas of the cerebral hemispheres. The ability to sensitively assess the in vivo spread of a neurovirulent strain of CDV provides a novel model system to study the mechanisms of virus spread into the CSF and the pathogenesis of acute viral meningitis. PMID:22553334

  20. Identification of Amino Acid Substitutions with Compensational Effects in the Attachment Protein of Canine Distemper Virus

    PubMed Central

    Sattler, Ursula; Khosravi, Mojtaba; Avila, Mislay; Pilo, Paola; Langedijk, Johannes P.; Ader-Ebert, Nadine; Alves, Lisa A.; Plattet, Philippe

    2014-01-01

    ABSTRACT The hemagglutinin (H) gene of canine distemper virus (CDV) encodes the receptor-binding protein. This protein, together with the fusion (F) protein, is pivotal for infectivity since it contributes to the fusion of the viral envelope with the host cell membrane. Of the two receptors currently known for CDV (nectin-4 and the signaling lymphocyte activation molecule [SLAM]), SLAM is considered the most relevant for host susceptibility. To investigate how evolution might have impacted the host-CDV interaction, we examined the functional properties of a series of missense single nucleotide polymorphisms (SNPs) naturally accumulating within the H-gene sequences during the transition between two distinct but related strains. The two strains, a wild-type strain and a consensus strain, were part of a single continental outbreak in European wildlife and occurred in distinct geographical areas 2 years apart. The deduced amino acid sequence of the two H genes differed at 5 residues. A panel of mutants carrying all the combinations of the SNPs was obtained by site-directed mutagenesis. The selected mutant, wild type, and consensus H proteins were functionally evaluated according to their surface expression, SLAM binding, fusion protein interaction, and cell fusion efficiencies. The results highlight that the most detrimental functional effects are associated with specific sets of SNPs. Strikingly, an efficient compensational system driven by additional SNPs appears to come into play, virtually neutralizing the negative functional effects. This system seems to contribute to the maintenance of the tightly regulated function of the H-gene-encoded attachment protein. IMPORTANCE To investigate how evolution might have impacted the host-canine distemper virus (CDV) interaction, we examined the functional properties of naturally occurring single nucleotide polymorphisms (SNPs) in the hemagglutinin gene of two related but distinct strains of CDV. The hemagglutinin gene encodes

  1. Immunogenicity of an inactivated oil-emulsion canine distemper vaccine in African wild dogs.

    PubMed

    Cirone, Francesco; Elia, Gabriella; Campolo, Marco; Friedrich, Klaus; Martella, Vito; Pratelli, Annamaria; Buonavoglia, Canio

    2004-04-01

    The immunogenicity of an inactivated oil-emulsion vaccine against canine distemper virus was evaluated in nine captive African wild dogs (Lycaon pictus). Antibody levels were determined by neutralization test in Vero cells. No significant local or systemic adverse reactions were observed in the animals. Virus neutralizing antibody levels >1:20 were detected, especially in animals that were vaccinated twice. The use of oil adjuvants is suggested as a good way to enhance the immune response to inactivated canine distemper vaccine.

  2. Canine Distemper

    MedlinePlus

    Although this brochure provides basic information about canine distemper, your veterinarian is always your best source of health information. Consult your veterinarian for more information about canine distemper and its prevention. ...

  3. The V domain of dog PVRL4 (nectin-4) mediates canine distemper virus entry and virus cell-to-cell spread

    SciTech Connect

    Delpeut, Sebastien; Noyce, Ryan S.; Richardson, Christopher D.

    2014-04-15

    The entry of canine distemper virus (CDV) is a multistep process that involves the attachment of CDV hemagglutinin (H) to its cellular receptor, followed by fusion between virus and cell membranes. Our laboratory recently identified PVRL4 (nectin-4) to be the epithelial receptor for measles and canine distemper viruses. In this study, we demonstrate that the V domain of PVRL4 is critical for CDV entry and virus cell-to-cell spread. Furthermore, four key amino acid residues within the V domain of dog PVRL4 and two within the CDV hemagglutinin were shown to be essential for receptor-mediated virus entry. - Highlights: • PVRL4 (nectin-4) is the epithelial cell receptor for measles and canine distemper viruses. • V domain of PVRL4 is critical for CDV entry, cell-to-cell spread, and syncytia formation. • Chimeric PVRL1 backbone substituted with the V domain of PVRL4 can function as a receptor. • Amino acids (F132/P133/A134/G135) within the V domain are essential for PVRL4 receptor activity. • Amino acids (P493/Y539) within CDV H protein are essential for PVRL4 receptor interaction.

  4. An outbreak of canine distemper virus in tigers (Panthera tigris): possible transmission from wild animals to zoo animals.

    PubMed

    Nagao, Yumiko; Nishio, Yohei; Shiomoda, Hiroshi; Tamaru, Seiji; Shimojima, Masayuki; Goto, Megumi; Une, Yumi; Sato, Azusa; Ikebe, Yusuke; Maeda, Ken

    2012-06-01

    Canine distemper virus (CDV), a morbillivirus that causes one of the most contagious and lethal viral diseases known in canids, has an expanding host range, including wild animals. Since December 2009, several dead or dying wild raccoon dogs (Nyctereutes procyonoides) were found in and around one safari-style zoo in Japan, and CDV was isolated from four of these animals. In the subsequent months (January to February 2010), 12 tigers (Panthera tigris) in the zoo developed respiratory and gastrointestinal diseases, and CDV RNA was detected in fecal samples of the examined tigers. In March 2010, one of the tigers developed a neurological disorder and died; CDV was isolated from the lung of this animal. Sequence analysis of the complete hemagglutinin (H) gene and the signal peptide region of the fusion (F) gene showed high homology among these isolates (99.8-100%), indicating that CDV might have been transmitted from raccoon dog to tiger. In addition, these isolates belonged to genotype Asia-1 and had lower homology (<90%) to the vaccine strain (Onderstepoort). Seropositivity of lions (Panthera leo) in the zoo and wild bears (Ursus thibetanus) captured around this area supported the theory that a CDV epidemic had occurred in many mammal species in and around the zoo. These results indicate a risk of CDV transmission among many animal species, including large felids and endangered species.

  5. First report of clinical disease associated with canine distemper virus infection in a wild black bear (Ursus americana).

    PubMed

    Cottrell, W O; Keel, M Kevin; Brooks, J W; Mead, D G; Phillips, J E

    2013-10-01

    An approximately 1-yr-old black bear was discovered on the porch of a rural residence in southwestern Pennsylvania on October 26, 2011, where it remained during the day in spite of efforts to frighten it away. The bear exhibited periods of somnolence and sporadic tremors and seizures. It was euthanized by gunshot that evening. Immediately after euthanasia it was observed to have footpads that exuded fluid when compressed. It was submitted for necropsy the next day where roughened footpads were noted. Histologic examination of the brain demonstrated nonsuppurative encephalitis with eosinophilic intranuclear and intracytoplasmic inclusion bodies in neurons. The footpads were thickened and hyperkeratotic. Canine distemper virus (CDV) was detected by immunohistochemistry (IHC) in the brain and footpads, and by reverse transcription polymerase chain reaction (RT-PCR) from the brain tissue. Phylogenetic analysis indicated that the CDV cDNA from the bear had 98.2% nucleotide identity to the Rockborn-Candur vaccine and a canine isolate from 2004 in Missouri, USA, and 97.3% nucleotide identity to a raccoon CDV isolated in 2011 from Tennessee, USA. This represents a first report of CDV as a cause of encephalitis or footpad hyperkeratosis in a wild black bear. PMID:24502734

  6. 9 CFR 113.302 - Distemper Vaccine-Mink.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 9 Animals and Animal Products 1 2014-01-01 2014-01-01 false Distemper Vaccine-Mink. 113.302... Virus Vaccines § 113.302 Distemper Vaccine—Mink. Distemper Vaccine—Mink shall be prepared from virus... immunogenic shall be used for preparing the production seed virus for vaccine production. All serials...

  7. 9 CFR 113.302 - Distemper Vaccine-Mink.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 9 Animals and Animal Products 1 2013-01-01 2013-01-01 false Distemper Vaccine-Mink. 113.302... Virus Vaccines § 113.302 Distemper Vaccine—Mink. Distemper Vaccine—Mink shall be prepared from virus... immunogenic shall be used for preparing the production seed virus for vaccine production. All serials...

  8. 9 CFR 113.302 - Distemper Vaccine-Mink.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 9 Animals and Animal Products 1 2012-01-01 2012-01-01 false Distemper Vaccine-Mink. 113.302... Virus Vaccines § 113.302 Distemper Vaccine—Mink. Distemper Vaccine—Mink shall be prepared from virus... immunogenic shall be used for preparing the production seed virus for vaccine production. All serials...

  9. 9 CFR 113.302 - Distemper Vaccine-Mink.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 9 Animals and Animal Products 1 2011-01-01 2011-01-01 false Distemper Vaccine-Mink. 113.302... Virus Vaccines § 113.302 Distemper Vaccine—Mink. Distemper Vaccine—Mink shall be prepared from virus... immunogenic shall be used for preparing the production seed virus for vaccine production. All serials...

  10. Efficacy of Recombinant Canine Distemper Virus Expressing Leishmania Antigen against Leishmania Challenge in Dogs

    PubMed Central

    Yoneda, Misako; Takenaka, Akiko; Doki, Miho; Goto, Yasuyuki; Sanjoba, Chizu; Endo, Yasuyuki; Fujiyuki, Tomoko; Sugai, Akihiro; Tsukiyama-Kohara, Kyoko; Matsumoto, Yoshitsugu; Sato, Hiroki; Kai, Chieko

    2015-01-01

    Canine distemper virus (CDV) vaccination confers long-term protection against CDV reinfection. To investigate the utility of CDV as a polyvalent vaccine vector for Leishmania, we generated recombinant CDVs, based on an avirulent Yanaka strain, that expressed Leishmania antigens: LACK, TSA, or LmSTI1 (rCDV–LACK, rCDV–TSA, and rCDV–LmSTI1, respectively). Dogs immunized with rCDV-LACK were protected against challenge with lethal doses of virulent CDV, in the same way as the parental Yanaka strain. To evaluate the protective effects of the recombinant CDVs against cutaneous leishmaniasis in dogs, dogs were immunized with one recombinant CDV or a cocktail of three recombinant CDVs, before intradermal challenge (in the ears) with infective-stage promastigotes of Leishmania major. Unvaccinated dogs showed increased nodules with ulcer formation after 3 weeks, whereas dogs immunized with rCDV–LACK showed markedly smaller nodules without ulceration. Although the rCDV–TSA- and rCDV–LmSTI1-immunized dogs showed little protection against L. major, the cocktail of three recombinant CDVs more effectively suppressed the progression of nodule formation than immunization with rCDV–LACK alone. These results indicate that recombinant CDV is suitable for use as a polyvalent live attenuated vaccine for protection against both CDV and L. major infections in dogs. PMID:26162094

  11. Efficacy of Recombinant Canine Distemper Virus Expressing Leishmania Antigen against Leishmania Challenge in Dogs.

    PubMed

    Miura, Ryuichi; Kooriyama, Takanori; Yoneda, Misako; Takenaka, Akiko; Doki, Miho; Goto, Yasuyuki; Sanjoba, Chizu; Endo, Yasuyuki; Fujiyuki, Tomoko; Sugai, Akihiro; Tsukiyama-Kohara, Kyoko; Matsumoto, Yoshitsugu; Sato, Hiroki; Kai, Chieko

    2015-01-01

    Canine distemper virus (CDV) vaccination confers long-term protection against CDV reinfection. To investigate the utility of CDV as a polyvalent vaccine vector for Leishmania, we generated recombinant CDVs, based on an avirulent Yanaka strain, that expressed Leishmania antigens: LACK, TSA, or LmSTI1 (rCDV-LACK, rCDV-TSA, and rCDV-LmSTI1, respectively). Dogs immunized with rCDV-LACK were protected against challenge with lethal doses of virulent CDV, in the same way as the parental Yanaka strain. To evaluate the protective effects of the recombinant CDVs against cutaneous leishmaniasis in dogs, dogs were immunized with one recombinant CDV or a cocktail of three recombinant CDVs, before intradermal challenge (in the ears) with infective-stage promastigotes of Leishmania major. Unvaccinated dogs showed increased nodules with ulcer formation after 3 weeks, whereas dogs immunized with rCDV-LACK showed markedly smaller nodules without ulceration. Although the rCDV-TSA- and rCDV-LmSTI1-immunized dogs showed little protection against L. major, the cocktail of three recombinant CDVs more effectively suppressed the progression of nodule formation than immunization with rCDV-LACK alone. These results indicate that recombinant CDV is suitable for use as a polyvalent live attenuated vaccine for protection against both CDV and L. major infections in dogs. PMID:26162094

  12. Canine distemper virus as a threat to wild tigers in Russia and across their range.

    PubMed

    Gilbert, Martin; Soutyrina, Svetlana V; Seryodkin, Ivan V; Sulikhan, Nadezhda; Uphyrkina, Olga V; Goncharuk, Mikhail; Matthews, Louise; Cleaveland, Sarah; Miquelle, Dale G

    2015-07-01

    Canine distemper virus (CDV) has recently been identified in populations of wild tigers in Russia and India. Tiger populations are generally too small to maintain CDV for long periods, but are at risk of infections arising from more abundant susceptible hosts that constitute a reservoir of infection. Because CDV is an additive mortality factor, it could represent a significant threat to small, isolated tiger populations. In Russia, CDV was associated with the deaths of tigers in 2004 and 2010, and was coincident with a localized decline of tigers in Sikhote-Alin Biosphere Zapovednik (from 25 tigers in 2008 to 9 in 2012). Habitat continuity with surrounding areas likely played an important role in promoting an ongoing recovery. We recommend steps be taken to assess the presence and the impact of CDV in all tiger range states, but should not detract focus away from the primary threats to tigers, which include habitat loss and fragmentation, poaching and retaliatory killing. Research priorities include: (i) recognition and diagnosis of clinical cases of CDV in tigers when they occur; and (ii) collection of baseline data on the health of wild tigers. CDV infection of individual tigers need not imply a conservation threat, and modeling should complement disease surveillance and targeted research to assess the potential impact to tiger populations across the range of ecosystems, population densities and climate extremes occupied by tigers. Describing the role of domestic and wild carnivores as contributors to a local CDV reservoir is an important precursor to considering control measures.

  13. Phylogenetic characterization of canine distemper virus isolates from naturally infected dogs and a marten in Korea.

    PubMed

    An, Dong-Jun; Yoon, Sook-Hee; Park, Jee-Yong; No, In-Sun; Park, Bong-Kyun

    2008-12-10

    We sequenced the hemagglutinin (H) genes from four canine distemper virus (CDV) isolates obtained from three dogs and a marten in Korea. These sequences were included in subsequent H gene-focused phylogenetic tree analysis of 89 CDV strains. This analysis revealed eight clades designated as EU1, EU2, EU3, NA1, NA2, Asia 1, Asia 2 and Vaccine. Three of the Korean isolates (97Jindo, 98Marten and 07D111) occurred in the Asia 2 group that also contains many Japanese CDV strains isolated in 1998. The remaining Korean strain (07Q72) fell into the Asia 1 group. The 21 H protein sequences of 25 Asia 1 strains are generally predicted to bear nine potential N-linked glycosylation sites. In contrast, the 9 H protein sequences of 12 Asia 2 strains had eight potential N-linked glycosylation sites. The remaining strains had six (98Marten and 07D111) and seven (97Jindo) potential N-linked glycosylation sites.

  14. Canine distemper virus as a threat to wild tigers in Russia and across their range.

    PubMed

    Gilbert, Martin; Soutyrina, Svetlana V; Seryodkin, Ivan V; Sulikhan, Nadezhda; Uphyrkina, Olga V; Goncharuk, Mikhail; Matthews, Louise; Cleaveland, Sarah; Miquelle, Dale G

    2015-07-01

    Canine distemper virus (CDV) has recently been identified in populations of wild tigers in Russia and India. Tiger populations are generally too small to maintain CDV for long periods, but are at risk of infections arising from more abundant susceptible hosts that constitute a reservoir of infection. Because CDV is an additive mortality factor, it could represent a significant threat to small, isolated tiger populations. In Russia, CDV was associated with the deaths of tigers in 2004 and 2010, and was coincident with a localized decline of tigers in Sikhote-Alin Biosphere Zapovednik (from 25 tigers in 2008 to 9 in 2012). Habitat continuity with surrounding areas likely played an important role in promoting an ongoing recovery. We recommend steps be taken to assess the presence and the impact of CDV in all tiger range states, but should not detract focus away from the primary threats to tigers, which include habitat loss and fragmentation, poaching and retaliatory killing. Research priorities include: (i) recognition and diagnosis of clinical cases of CDV in tigers when they occur; and (ii) collection of baseline data on the health of wild tigers. CDV infection of individual tigers need not imply a conservation threat, and modeling should complement disease surveillance and targeted research to assess the potential impact to tiger populations across the range of ecosystems, population densities and climate extremes occupied by tigers. Describing the role of domestic and wild carnivores as contributors to a local CDV reservoir is an important precursor to considering control measures. PMID:25939829

  15. Canine Distemper Virus Fusion Activation: Critical Role of Residue E123 of CD150/SLAM

    PubMed Central

    Khosravi, Mojtaba; Bringolf, Fanny; Röthlisberger, Silvan; Bieringer, Maria; Schneider-Schaulies, Jürgen; Zurbriggen, Andreas; Origgi, Francesco

    2015-01-01

    ABSTRACT Measles virus (MeV) and canine distemper virus (CDV) possess tetrameric attachment proteins (H) and trimeric fusion proteins, which cooperate with either SLAM or nectin 4 receptors to trigger membrane fusion for cell entry. While the MeV H-SLAM cocrystal structure revealed the binding interface, two distinct oligomeric H assemblies were also determined. In one of the conformations, two SLAM units were sandwiched between two discrete H head domains, thus spotlighting two binding interfaces (“front” and “back”). Here, we investigated the functional relevance of both interfaces in activating the CDV membrane fusion machinery. While alanine-scanning mutagenesis identified five critical regulatory residues in the front H-binding site of SLAM, the replacement of a conserved glutamate residue (E at position 123, replaced with A [E123A]) led to the most pronounced impact on fusion promotion. Intriguingly, while determination of the interaction of H with the receptor using soluble constructs revealed reduced binding for the identified SLAM mutants, no effect was recorded when physical interaction was investigated with the full-length counterparts of both molecules. Conversely, although mutagenesis of three strategically selected residues within the back H-binding site of SLAM did not substantially affect fusion triggering, nevertheless, the mutants weakened the H-SLAM interaction recorded with the membrane-anchored protein constructs. Collectively, our findings support a mode of binding between the attachment protein and the V domain of SLAM that is common to all morbilliviruses and suggest a major role of the SLAM residue E123, located at the front H-binding site, in triggering the fusion machinery. However, our data additionally support the hypothesis that other microdomain(s) of both glycoproteins (including the back H-binding site) might be required to achieve fully productive H-SLAM interactions. IMPORTANCE A complete understanding of the measles virus

  16. Evaluation of Trapper-Collected Nobuto Filter-Paper Blood Samples for Distemper and Parvovirus Antibody Detection in Coyotes (Canis latrans) and Raccoons (Procyon lotor).

    PubMed

    Kamps, Amanda J; Dubay, Shelli A; Langenberg, Julie; Maes, Roger K

    2015-07-01

    Blood samples are often collected from free-ranging wildlife for antibody detection. However, filter-paper (FP) strips are more cost efficient and easy to collect and store. We evaluated trapper-collected FP strips and body-cavity blood for canine distemper (CDV) and parvovirus (CPV-2) antibody detection in raccoons (Procyon lotor) and coyotes (Canis latrans). From 2008 to 2010, licensed trappers near Madison and Milwaukee, Wisconsin, US collected paired samples from harvested animals. Canine distemper antibodies were detected using virus neutralization and parvovirus antibodies were detected using hemagglutination inhibition. Titers ≥ 1:32 for CDV and ≥ 1:25 for CPV-2 were considered evidence of exposure. Using Cohen's kappa test of agreement, FP strip titers agreed with sera for CDV in coyotes (n = 28, K = 0.772) and raccoons (n = 29, K = 0.858) and for CPV-2 in coyotes (n = 40, K = 0.775) and raccoons (n = 70, K = 0.646). However, raccoons determined to be exposed to CPV-2 from sera were unexposed by FP strips in 35% of the samples. Titer results may be affected by quality and volume of blood samples, interval between collection and processing, small sample sizes, and diagnostic testing procedures. Filter-paper strips can be useful for detecting CDV and CPV-2 exposure in coyotes and raccoons with correct field sample collection and appropriate diagnostic testing procedures.

  17. Evaluation of Trapper-Collected Nobuto Filter-Paper Blood Samples for Distemper and Parvovirus Antibody Detection in Coyotes (Canis latrans) and Raccoons (Procyon lotor).

    PubMed

    Kamps, Amanda J; Dubay, Shelli A; Langenberg, Julie; Maes, Roger K

    2015-07-01

    Blood samples are often collected from free-ranging wildlife for antibody detection. However, filter-paper (FP) strips are more cost efficient and easy to collect and store. We evaluated trapper-collected FP strips and body-cavity blood for canine distemper (CDV) and parvovirus (CPV-2) antibody detection in raccoons (Procyon lotor) and coyotes (Canis latrans). From 2008 to 2010, licensed trappers near Madison and Milwaukee, Wisconsin, US collected paired samples from harvested animals. Canine distemper antibodies were detected using virus neutralization and parvovirus antibodies were detected using hemagglutination inhibition. Titers ≥ 1:32 for CDV and ≥ 1:25 for CPV-2 were considered evidence of exposure. Using Cohen's kappa test of agreement, FP strip titers agreed with sera for CDV in coyotes (n = 28, K = 0.772) and raccoons (n = 29, K = 0.858) and for CPV-2 in coyotes (n = 40, K = 0.775) and raccoons (n = 70, K = 0.646). However, raccoons determined to be exposed to CPV-2 from sera were unexposed by FP strips in 35% of the samples. Titer results may be affected by quality and volume of blood samples, interval between collection and processing, small sample sizes, and diagnostic testing procedures. Filter-paper strips can be useful for detecting CDV and CPV-2 exposure in coyotes and raccoons with correct field sample collection and appropriate diagnostic testing procedures. PMID:25973631

  18. Molecular Determinants Defining the Triggering Range of Prefusion F Complexes of Canine Distemper Virus

    PubMed Central

    Avila, Mislay; Alves, Lisa; Khosravi, Mojtaba; Ader-Ebert, Nadine; Origgi, Francesco; Schneider-Schaulies, Jürgen; Zurbriggen, Andreas; Plemper, Richard K.

    2014-01-01

    ABSTRACT The morbillivirus cell entry machinery consists of a fusion (F) protein trimer that refolds to mediate membrane fusion following receptor-induced conformational changes in its binding partner, the tetrameric attachment (H) protein. To identify molecular determinants that control F refolding, we generated F chimeras between measles virus (MeV) and canine distemper virus (CDV). We located a central pocket in the globular head domain of CDV F that regulates the stability of the metastable, prefusion conformational state of the F trimer. Most mutations introduced into this “pocket'” appeared to mediate a destabilizing effect, a phenotype associated with enhanced membrane fusion activity. Strikingly, under specific triggering conditions (i.e., variation of receptor type and H protein origin), some F mutants also exhibited resistance to a potent morbillivirus entry inhibitor, which is known to block F triggering by enhancing the stability of prefusion F trimers. Our data reveal that the molecular nature of the F stimulus and the intrinsic stability of metastable prefusion F both regulate the efficiency of F refolding and escape from small-molecule refolding blockers. IMPORTANCE With the aim to better characterize the thermodynamic basis of morbillivirus membrane fusion for cell entry and spread, we report here that the activation energy barrier of prefusion F trimers together with the molecular nature of the triggering “stimulus” (attachment protein and receptor types) define a “triggering range,” which governs the initiation of the membrane fusion process. A central “pocket” microdomain in the globular F head contributes substantially to the regulation of the conformational stability of the prefusion complexes. The triggering range also defines the mechanism of viral escape from entry inhibitors and describes how the cellular environment can affect membrane fusion efficiency. PMID:24371057

  19. Isolation and sequence analysis of a canine distemper virus from a raccoon dog in Jilin Province, China.

    PubMed

    Cheng, Yuening; Wang, Jianke; Zhang, Miao; Zhao, Jianjun; Shao, Xiqun; Ma, Zengjun; Zhao, Hang; Lin, Peng; Wu, Hua

    2015-10-01

    Canine distemper virus (CDV) is a major pathogen not only in raccoon dogs but also in a variety of carnivorous animals, including domesticated animals, particularly if they have not been vaccinated. In this study, a wild-type strain of CDV was isolated from lung tissue from a raccoon dog kept at a fur farm in Jilin Province, China. Cytopathic effects typical of CDV infection were observed after three blind passages in Vero cells, yielding a virus titer of 10(4.6) TCID50/mL. Virus identification was carried out by RT-PCR, immunofluorescence, electron microscopy, and genome sequencing. The results showed that the isolated virus, termed the SY strain, corresponded to the Asia-1 genotype of CDV and has a genome of 15,690 nucleotides. This represents the first complete nucleotide sequence of a CDV strain circulating in raccoon dogs in China.

  20. Isolation and sequence analysis of a canine distemper virus from a raccoon dog in Jilin Province, China.

    PubMed

    Cheng, Yuening; Wang, Jianke; Zhang, Miao; Zhao, Jianjun; Shao, Xiqun; Ma, Zengjun; Zhao, Hang; Lin, Peng; Wu, Hua

    2015-10-01

    Canine distemper virus (CDV) is a major pathogen not only in raccoon dogs but also in a variety of carnivorous animals, including domesticated animals, particularly if they have not been vaccinated. In this study, a wild-type strain of CDV was isolated from lung tissue from a raccoon dog kept at a fur farm in Jilin Province, China. Cytopathic effects typical of CDV infection were observed after three blind passages in Vero cells, yielding a virus titer of 10(4.6) TCID50/mL. Virus identification was carried out by RT-PCR, immunofluorescence, electron microscopy, and genome sequencing. The results showed that the isolated virus, termed the SY strain, corresponded to the Asia-1 genotype of CDV and has a genome of 15,690 nucleotides. This represents the first complete nucleotide sequence of a CDV strain circulating in raccoon dogs in China. PMID:26265248

  1. Canine distemper virus (CDV) infection of ferrets as a model for testing Morbillivirus vaccine strategies: NYVAC- and ALVAC-based CDV recombinants protect against symptomatic infection.

    PubMed

    Stephensen, C B; Welter, J; Thaker, S R; Taylor, J; Tartaglia, J; Paoletti, E

    1997-02-01

    Canine distemper virus (CDV) infection of ferrets causes an acute systemic disease involving multiple organ systems, including the respiratory tract, lymphoid system, and central nervous system (CNS). We have tested candidate CDV vaccines incorporating the fusion (F) and hemagglutinin (HA) proteins in the highly attenuated NYVAC strain of vaccinia virus and in the ALVAC strain of canarypox virus, which does not productively replicate in mammalian hosts. Juvenile ferrets were vaccinated twice with these constructs, or with an attenuated live-virus vaccine, while controls received saline or the NYVAC and ALVAC vectors expressing rabies virus glycoprotein. Control animals did not develop neutralizing antibody and succumbed to distemper after developing fever, weight loss, leukocytopenia, decreased activity, conjunctivitis, an erythematous rash typical of distemper, CNS signs, and viremia in peripheral blood mononuclear cells (as measured by reverse transcription-PCR). All three CDV vaccines elicited neutralizing titers of at least 1:96. All vaccinated ferrets survived, and none developed viremia. Both recombinant vaccines also protected against the development of symptomatic distemper. However, ferrets receiving the live-virus vaccine lost weight, became lymphocytopenic, and developed the erythematous rash typical of CDV. These data show that ferrets are an excellent model for evaluating the ability of CDV vaccines to protect against symptomatic infection. Because the pathogenesis and clinical course of CDV infection of ferrets is quite similar to that of other Morbillivirus infections, including measles, this model will be useful in testing new candidate Morbillivirus vaccines. PMID:8995676

  2. Canine distemper virus (CDV) infection of ferrets as a model for testing Morbillivirus vaccine strategies: NYVAC- and ALVAC-based CDV recombinants protect against symptomatic infection.

    PubMed Central

    Stephensen, C B; Welter, J; Thaker, S R; Taylor, J; Tartaglia, J; Paoletti, E

    1997-01-01

    Canine distemper virus (CDV) infection of ferrets causes an acute systemic disease involving multiple organ systems, including the respiratory tract, lymphoid system, and central nervous system (CNS). We have tested candidate CDV vaccines incorporating the fusion (F) and hemagglutinin (HA) proteins in the highly attenuated NYVAC strain of vaccinia virus and in the ALVAC strain of canarypox virus, which does not productively replicate in mammalian hosts. Juvenile ferrets were vaccinated twice with these constructs, or with an attenuated live-virus vaccine, while controls received saline or the NYVAC and ALVAC vectors expressing rabies virus glycoprotein. Control animals did not develop neutralizing antibody and succumbed to distemper after developing fever, weight loss, leukocytopenia, decreased activity, conjunctivitis, an erythematous rash typical of distemper, CNS signs, and viremia in peripheral blood mononuclear cells (as measured by reverse transcription-PCR). All three CDV vaccines elicited neutralizing titers of at least 1:96. All vaccinated ferrets survived, and none developed viremia. Both recombinant vaccines also protected against the development of symptomatic distemper. However, ferrets receiving the live-virus vaccine lost weight, became lymphocytopenic, and developed the erythematous rash typical of CDV. These data show that ferrets are an excellent model for evaluating the ability of CDV vaccines to protect against symptomatic infection. Because the pathogenesis and clinical course of CDV infection of ferrets is quite similar to that of other Morbillivirus infections, including measles, this model will be useful in testing new candidate Morbillivirus vaccines. PMID:8995676

  3. Black-backed jackal exposure to rabies virus, canine distemper virus, and Bacillus anthracis in Etosha National Park, Namibia.

    PubMed

    Bellan, Steve E; Cizauskas, Carrie A; Miyen, Jacobeth; Ebersohn, Karen; Küsters, Martina; Prager, K C; Van Vuuren, Moritz; Sabeta, Claude; Getz, Wayne M

    2012-04-01

    Canine distemper virus (CDV) and rabies virus (RABV) occur worldwide in wild carnivore and domestic dog populations and pose threats to wildlife conservation and public health. In Etosha National Park (ENP), Namibia, anthrax is endemic and generates carcasses frequently fed on by an unusually dense population of black-backed jackals (Canis mesomelas). Using serology, phylogenetic analyses (on samples obtained from February 2009-July 2010), and historical mortality records (1975-2011), we assessed jackal exposure to Bacillus anthracis (BA; the causal bacterial agent of anthrax), CDV, and RABV. Prevalence of antibodies against BA (95%, n = 86) and CDV (71%, n = 80) was relatively high, while that of antibodies against RABV was low (9%, n = 81). Exposure to BA increased significantly with age, and all animals >6 mo old were antibody-positive. As with BA, prevalence of antibodies against CDV increased significantly with age, with similar age-specific trends during both years of the study. No significant effect of age was found on the prevalence of antibodies against RABV. Three of the seven animals with antibodies against RABV were monitored for more than 1 yr after sampling and showed no signs of active infection. Mortality records revealed that rabid animals are destroyed nearly every year inside the ENP tourist camps. Phylogenetic analyses demonstrated that jackal RABV in ENP is part of the same transmission cycle as other dog-jackal RABV cycles in Namibia. PMID:22493112

  4. Black-backed jackal exposure to rabies virus, canine distemper virus, and Bacillus anthracis in Etosha National Park, Namibia.

    PubMed

    Bellan, Steve E; Cizauskas, Carrie A; Miyen, Jacobeth; Ebersohn, Karen; Küsters, Martina; Prager, K C; Van Vuuren, Moritz; Sabeta, Claude; Getz, Wayne M

    2012-04-01

    Canine distemper virus (CDV) and rabies virus (RABV) occur worldwide in wild carnivore and domestic dog populations and pose threats to wildlife conservation and public health. In Etosha National Park (ENP), Namibia, anthrax is endemic and generates carcasses frequently fed on by an unusually dense population of black-backed jackals (Canis mesomelas). Using serology, phylogenetic analyses (on samples obtained from February 2009-July 2010), and historical mortality records (1975-2011), we assessed jackal exposure to Bacillus anthracis (BA; the causal bacterial agent of anthrax), CDV, and RABV. Prevalence of antibodies against BA (95%, n = 86) and CDV (71%, n = 80) was relatively high, while that of antibodies against RABV was low (9%, n = 81). Exposure to BA increased significantly with age, and all animals >6 mo old were antibody-positive. As with BA, prevalence of antibodies against CDV increased significantly with age, with similar age-specific trends during both years of the study. No significant effect of age was found on the prevalence of antibodies against RABV. Three of the seven animals with antibodies against RABV were monitored for more than 1 yr after sampling and showed no signs of active infection. Mortality records revealed that rabid animals are destroyed nearly every year inside the ENP tourist camps. Phylogenetic analyses demonstrated that jackal RABV in ENP is part of the same transmission cycle as other dog-jackal RABV cycles in Namibia.

  5. Antibody responses of red wolves to canine distemper virus and canine parvovirus vaccination.

    PubMed

    Harrenstien, L A; Munson, L; Ramsay, E C; Lucash, C F; Kania, S A; Potgieter, L N

    1997-07-01

    Twenty captive red wolves (Canis rufus), including 16 intended for release into Great Smoky Mountains National Park, Cades Cove, Tennessee (USA), and four housed at Knoxville Zoological Gardens, Inc., Knoxville, Tennessee, were evaluated for immunologic response to vaccination between June 1994 and April 1995. Wolves were vaccinated with modified-live (MLV) canine distemper virus (CDV) and canine parvovirus type-2 (CPV2). Sera were collected, and immunofluorescent staining was performed for determination of immunoglobulin titers (CDV IgM, CDV IgG, and CPV2 IgG). A capture enzyme-linked immunosorbent assay was performed for validation purposes, to confirm the reactivity of our standard diagnostic reagents with red wolf serum. All wolves produced a measurable antibody response to CDV and CPV2 vaccination. Titers against CDV and CPV2 varied widely among individual wolves, but between-litter differences in mean titers were not significant. No consistent response between the degree of response to CDV versus CPV2 vaccination was observed in individual wolves. No differences were seen between IgG responses of pups vaccinated with univalent vaccines given concurrently or during alternating weeks. Pups had an IgG response to CDV and CPV2 vaccination as early as 9 wk of age. Mean post-vaccination IgG titers against CDV were at or above the level normally measured in vaccinated domestic dogs. Mean post-vaccination IgG titers against CPV2 were below the level normally measured in domestic dogs. Adult previously-vaccinated wolves had measurable CDV and CPV2 IgG titers more than 1 yr after vaccination, but did not have significant IgG titer increases after revaccination. We conclude that red wolves are capable of producing an antibody response after vaccination with commercial canine products but that their response to CPV2 vaccination was minimal. This response can be assayed using tests developed for domestic dogs.

  6. Urban domestic dog populations as a source of canine distemper virus for wild carnivores in the Coquimbo region of Chile.

    PubMed

    Acosta-Jamett, G; Chalmers, W S K; Cunningham, A A; Cleaveland, S; Handel, I G; Bronsvoort, B M deC

    2011-09-28

    Urban areas can support dog populations dense enough to maintain canine distemper virus (CDV) and can be a source of infection for rural dogs and free-ranging carnivores. The aim of this study was to investigate the relationships between urban and rural domestic dog and wild carnivore populations and their effects on the epidemiology of CDV to explain retrospectively a CD outbreak in wild foxes in 2003. From 2005 to 2007 a cross-sectional household questionnaire survey was conducted in Coquimbo and Ovalle cities, in three towns and in rural sites along two transects from these cities to the Fray Jorge National Park (FJNP) in the Coquimbo region, Chile. Blood samples were collected from unvaccinated dogs at surveyed households and from free-ranging foxes in rural areas along the transects. The seroprevalence of CDV in domestic dogs was higher in urban than in rural areas and in the later was highest in dogs born before 2001-2002. The seroprevalence of CDV in foxes was higher in areas closer to human settlements. A high seroprevalence in dogs born before 2001-2002 further supports a link between CDV patterns in rural dog and fox populations. In our study area, urban dogs are proposed to be the source of CDV infection to wild carnivores. The large dog population size and density detected in Coquimbo and Ovalle provides optimal conditions for maintaining a large and dense susceptible population of dogs, which can act as a reservoir for highly infectious diseases and could have been the source of infection in the CD outbreak in wild foxes. PMID:21641130

  7. Urban domestic dog populations as a source of canine distemper virus for wild carnivores in the Coquimbo region of Chile.

    PubMed

    Acosta-Jamett, G; Chalmers, W S K; Cunningham, A A; Cleaveland, S; Handel, I G; Bronsvoort, B M deC

    2011-09-28

    Urban areas can support dog populations dense enough to maintain canine distemper virus (CDV) and can be a source of infection for rural dogs and free-ranging carnivores. The aim of this study was to investigate the relationships between urban and rural domestic dog and wild carnivore populations and their effects on the epidemiology of CDV to explain retrospectively a CD outbreak in wild foxes in 2003. From 2005 to 2007 a cross-sectional household questionnaire survey was conducted in Coquimbo and Ovalle cities, in three towns and in rural sites along two transects from these cities to the Fray Jorge National Park (FJNP) in the Coquimbo region, Chile. Blood samples were collected from unvaccinated dogs at surveyed households and from free-ranging foxes in rural areas along the transects. The seroprevalence of CDV in domestic dogs was higher in urban than in rural areas and in the later was highest in dogs born before 2001-2002. The seroprevalence of CDV in foxes was higher in areas closer to human settlements. A high seroprevalence in dogs born before 2001-2002 further supports a link between CDV patterns in rural dog and fox populations. In our study area, urban dogs are proposed to be the source of CDV infection to wild carnivores. The large dog population size and density detected in Coquimbo and Ovalle provides optimal conditions for maintaining a large and dense susceptible population of dogs, which can act as a reservoir for highly infectious diseases and could have been the source of infection in the CD outbreak in wild foxes.

  8. Genetically distant American Canine distemper virus lineages have recently caused epizootics with somewhat different characteristics in raccoons living around a large suburban zoo in the USA

    PubMed Central

    Lednicky, John A; Dubach, Jean; Kinsel, Michael J; Meehan, Thomas P; Bocchetta, Maurizio; Hungerford, Laura L; Sarich, Nicolene A; Witecki, Kelley E; Braid, Michael D; Pedrak, Casandra; Houde, Christiane M

    2004-01-01

    Background Mortality rates have differed during distemper outbreaks among free-ranging raccoons (Procyon lotor) living around a large Chicago-area zoo, and appeared higher in year 2001 than in 1998 and 2000. We hypothesized that a more lethal variant of the local Canine distemper virus (CDV) lineage had emerged in 2001, and sought the genetic basis that led to increased virulence. However, a more complex model surfaced during preliminary analyses of CDV genomic sequences in infected tissues and of virus isolated in vitro from the raccoons. Results Phylogenetic analyses of subgenomic CDV fusion (F) -, phosphoprotein (P) -, and complete hemagglutinin (H) – gene sequences indicated that distinct American CDV lineages caused the distemper epizootics. The 1998 outbreak was caused by viruses that are likely from an old CDV lineage that includes CDV Snyder Hill and Lederle, which are CDV strains from the early 1950's. The 2000 and 2001 viruses appear to stem from the lineage of CDV A75/17, which was isolated in the mid 1970's. Only the 2001 viruses formed large syncytia in brain and/or lung tissue, and during primary isolation in-vitro in Vero cells, demonstrating at least one phenotypic property by which they differed from the other viruses. Conclusions Two different American CDV lineages caused the raccoon distemper outbreaks. The 1998 viruses are genetically distant to the 2000/2001 viruses. Since CDV does not cause persistent infections, the cycling of different CDV lineages within the same locale suggests multiple reintroductions of the virus to area raccoons. Our findings establish a precedent for determining whether the perceived differences in mortality rates are actual and attributable in part to inherent differences between CDV strains arising from different CDV lineages. PMID:15507154

  9. Canine distemper virus-associated encephalitis in free-living lynx (Lynx canadensis) and bobcats (Lynx rufus) of eastern Canada.

    PubMed

    Daoust, Pierre-Yves; McBurney, Scott R; Godson, Dale L; van de Bildt, Marco W G; Osterhaus, Albert D M E

    2009-07-01

    Between 1993 and 1999, encephalitis caused by morbillivirus was diagnosed by immunohistochemistry and histology in six lynx (Lynx canadensis) and one bobcat (Lynx rufus) in the eastern Canadian provinces of New Brunswick and Nova Scotia. Five of the six cases in lynx occurred within an 11-mo period in 1996-97. A second bobcat with encephalitis caused by unidentified protozoa and a nematode larva also had immunohistochemical evidence of neurologic infection by morbillivirus. The virus was identified as canine distemper virus (CDV) by reverse transcriptase polymerase chain reaction and nucleotide sequencing in four of five animals from which frozen tissue samples were available, and it was isolated in cell culture from one of them. To our knowledge, this is the first report of disease caused by CDV in free-living felids in North America.

  10. The humoral immune response to recombinant nucleocapsid antigen of canine distemper virus in dogs vaccinated with attenuated distemper virus or DNA encoding the nucleocapsid of wild-type virus.

    PubMed

    Griot-Wenk, M E; Cherpillod, P; Koch, A; Zurbriggen, R; Bruckner, L; Wittek, R; Zurbriggen, A

    2001-06-01

    This study compared the humoral immune response against the nucleocapsid-(N) protein of canine distemper virus (CDV) of dogs vaccinated with a multivalent vaccine against parvo-, adeno-, and parainfluenza virus and leptospira combined with either the attenuated CDV Onderstepoort strain (n = 15) or an expression plasmid containing the N-gene of CDV (n = 30). The vaccinations were applied intramuscularly three times at 2-week intervals beginning at the age of 6 weeks. None of the pre-immune sera recognized the recombinant N-protein, confirming the lack of maternal antibodies at this age. Immunization with DNA vaccine for CDV resulted in positive serum N-specific IgG response. However, their IgG (and IgA) titres were lower than those of CDV-vaccinated dogs. Likewise, DNA-vaccinated dogs did not show an IgM peak. There was no increase in N-specific serum IgE titres in either group. Serum titres to the other multivalent vaccine components were similar in both groups. PMID:11475904

  11. Persistence of canine distemper virus in the Greater Yellowstone Ecosystem's carnivore community

    USGS Publications Warehouse

    Almberg, E.S.; Cross, P.C.; Smith, D.W.

    2010-01-01

    Canine distemper virus (CDV) is an acute, highly immunizing pathogen that should require high densities and large populations of hosts for long-term persistence, yet CDV persists among terrestrial carnivores with small, patchily distributed groups. We used CDV in the Greater Yellowstone ecosystem's (GYE) wolves (Canis lupus) and coyotes (Canis latrans) as a case study for exploring how metapopulation structure, host demographics, and multi-host transmission affect the critical community size and spatial scale required for CDV persistence. We illustrate how host spatial connectivity and demographic turnover interact to affect both local epidemic dynamics, such as the length and variation in inter-epidemic periods, and pathogen persistence using stochastic, spatially explicit susceptible-exposed-infectious-recovered simulation models. Given the apparent absence of other known persistence mechanisms (e.g., a carrier or environmental state, densely populated host, chronic infection, or a vector), we suggest that CDV requires either large spatial scales or multi-host transmission for persistence. Current GYE wolf populations are probably too small to support endemic CDV. Coyotes are a plausible reservoir host, but CDV would still require 50 000-100 000 individuals for moderate persistence (>50% over 10 years), which would equate to an area of 1-3 times the size of the GYE (60000-200000 km2). Coyotes, and carnivores in general, are not uniformly distributed; therefore, this is probably a gross underestimate of the spatial scale of CDV persistence. However, the presence of a second competent host species can greatly increase the probability of long-term CDV persistence at much smaller spatial scales. Although no management of CDV is currently recommended for the GYE, wolf managers in the region should expect periodic but unpredictable CDV-related population declines as often as every 2-5 years. Awareness and monitoring of such outbreaks will allow corresponding

  12. Persistence of canine distemper virus in the Greater Yellowstone ecosystem's carnivore community.

    PubMed

    Almberg, Emily S; Cross, Paul C; Smith, Douglas W

    2010-10-01

    Canine distemper virus (CDV) is an acute, highly immunizing pathogen that should require high densities and large populations of hosts for long-term persistence, yet CDV persists among terrestrial carnivores with small, patchily distributed groups. We used CDV in the Greater Yellowstone ecosystem's (GYE) wolves (Canis lupus) and coyotes (Canis latrans) as a case study for exploring how metapopulation structure, host demographics, and multi-host transmission affect the critical community size and spatial scale required for CDV persistence. We illustrate how host spatial connectivity and demographic turnover interact to affect both local epidemic dynamics, such as the length and variation in inter-epidemic periods, and pathogen persistence using stochastic, spatially explicit susceptible-exposed-infectious-recovered simulation models. Given the apparent absence of other known persistence mechanisms (e.g., a carrier or environmental state, densely populated host, chronic infection, or a vector), we suggest that CDV requires either large spatial scales or multi-host transmission for persistence. Current GYE wolf populations are probably too small to support endemic CDV. Coyotes are a plausible reservoir host, but CDV would still require 50000-100000 individuals for moderate persistence (> 50% over 10 years), which would equate to an area of 1-3 times the size of the GYE (60000-200000 km2). Coyotes, and carnivores in general, are not uniformly distributed; therefore, this is probably a gross underestimate of the spatial scale of CDV persistence. However, the presence of a second competent host species can greatly increase the probability of long-term CDV persistence at much smaller spatial scales. Although no management of CDV is currently recommended for the GYE, wolf managers in the region should expect periodic but unpredictable CDV-related population declines as often as every 2-5 years. Awareness and monitoring of such outbreaks will allow corresponding adjustments

  13. Genotypes of Canine Distemper Virus Determined by Analysis of the Hemagglutinin Genes of Recent Isolates from Dogs in Japan

    PubMed Central

    Mochizuki, Masami; Hashimoto, Michiru; Hagiwara, Shigeyuki; Yoshida, Yoshio; Ishiguro, Shinryo

    1999-01-01

    Canine distemper of domestic dogs is caused by canine distemper virus (CDV), a member of the morbilliviruses. It has been a highly contagious disease of great veterinary importance for centuries, but for the last several decades it has been controlled satisfactorily by modified live vaccines. In the 1990s, however, it was described that CDV strains genetically different from vaccine strains may have caused the disease in vaccinated dogs. The highest antigenic variation is found in the H protein. Therefore, in the present study, hemagglutinin (H) genes obtained from current vaccines and field isolates and amplified directly from clinical specimens were genetically analyzed by restriction fragment length polymorphism assay and sequencing. Phylogenetic analysis of the H-gene amino acid sequences revealed that at least two CDV genotypes are circulating among dogs in Japan; one is a genotype to which almost all Japanese CDV isolates belong and the other has not been previously described. Both are separate and independent from the other lineages or genotypes of vaccine strains, as well as European and U.S. CDV isolates. The results suggest that CDV has also evolved in Japan, and further studies will be needed for an evaluation and possible improvement of the efficacies of current CDV vaccines. PMID:10449479

  14. Three-year duration of immunity in dogs vaccinated with a canarypox-vectored recombinant canine distemper virus vaccine.

    PubMed

    Larson, L J; Schultz, R D

    2007-01-01

    Two studies evaluated the duration of serologic response to the recombinant, canarypox-vectored canine distemper virus vaccine (Recombitek, Merial). Serologic duration of immunity was shown to be at least 36 months. Thus, Recombitek provides protection when administered less frequently than the manufacturer's label. After the initial vaccination protocol of two or more doses administered approximately 4 weeks apart, with the last dose given at 12 to 16 weeks of age or older, and re-vaccination at 1 year of age, Recombitek can confidently be readministered every 3 years with assurance of protection in immunocompetent dogs. This allows the vaccine to be administered in accordance with the recommendations of the American Animal Hospital Association Canine Vaccine Task Force and others. PMID:17616944

  15. Serologic responses after vaccination of fennec foxes (Vulpes zerda) and meerkats (Suricata suricatta) with a live, canarypox-vectored canine distemper virus vaccine.

    PubMed

    Coke, Rob L; Backues, Kay A; Hoover, John P; Saliki, Jeremiah T; Ritchey, Jerry W; West, Gary D

    2005-06-01

    Fennec foxes (Vulpes zerda) and meerkats (Suricata suricatta) are considered to be susceptible to canine distemper virus (CDV) infection. Although no definitive clinical cases of natural CDV infections have been reported, mortalities due to CDV have been suspected and are reported in other closely related species. A commercially available monovalent, live, canarypox-vectored CDV vaccine induced neutralizing antibody titers that were maintained for at least a year in both fennec foxes and meerkats.

  16. Serologic responses after vaccination of fennec foxes (Vulpes zerda) and meerkats (Suricata suricatta) with a live, canarypox-vectored canine distemper virus vaccine.

    PubMed

    Coke, Rob L; Backues, Kay A; Hoover, John P; Saliki, Jeremiah T; Ritchey, Jerry W; West, Gary D

    2005-06-01

    Fennec foxes (Vulpes zerda) and meerkats (Suricata suricatta) are considered to be susceptible to canine distemper virus (CDV) infection. Although no definitive clinical cases of natural CDV infections have been reported, mortalities due to CDV have been suspected and are reported in other closely related species. A commercially available monovalent, live, canarypox-vectored CDV vaccine induced neutralizing antibody titers that were maintained for at least a year in both fennec foxes and meerkats. PMID:17323579

  17. Construction of an Expression System for Bioactive IL-18 and Generation of Recombinant Canine Distemper Virus Expressing IL-18

    PubMed Central

    LIU, Yuxiu; SATO, Hiroki; HAMANA, Masahiro; MOONAN, Navita Anisia; YONEDA, Misako; XIA, Xianzhu; KAI, Chieko

    2014-01-01

    ABSTRACT Interleukin 18 (IL-18) plays an important role in the T-helper-cell type 1 immune response against intracellular parasites, bacteria and viral infections. It has been widely used as an adjuvant for vaccines and as an anticancer agent. However, IL-18 protein lacks a typical signal sequence and requires cleavage into its mature active form by caspase 1. In this study, we constructed mammalian expression vectors carrying cDNA encoding mature canine IL-18 (cIL-18) or mouse IL-18 (mIL-18) fused to the human IL-2 (hIL-2) signal sequence. The expressed proIL-18 proteins were processed to their mature forms in the cells. The supernatants of cells transfected with these plasmids induced high interferon-γ production in canine peripheral blood mononuclear cells or mouse splenocytes, respectively, indicating the secretion of bioactive IL-18. Using reverse genetics, we also generated a recombinant canine distemper virus that expresses cIL-18 or mIL-18 fused to the hIL-2 signal sequence. As expected, both recombinant viruses produced mature IL-18 in the infected cells, which secreted bioactive IL-18. These results indicate that the signal sequence from hIL-2 is suitable for the secretion of mature IL-18. These recombinant viruses can also potentially be used as immunoadjuvants and agents for anticancer therapies in vivo. PMID:24898077

  18. Construction of an expression system for bioactive IL-18 and generation of recombinant canine distemper virus expressing IL-18.

    PubMed

    Liu, Yuxiu; Sato, Hiroki; Hamana, Masahiro; Moonan, Navita Anisia; Yoneda, Misako; Xia, Xianzhu; Kai, Chieko

    2014-09-01

    Interleukin 18 (IL-18) plays an important role in the T-helper-cell type 1 immune response against intracellular parasites, bacteria and viral infections. It has been widely used as an adjuvant for vaccines and as an anticancer agent. However, IL-18 protein lacks a typical signal sequence and requires cleavage into its mature active form by caspase 1. In this study, we constructed mammalian expression vectors carrying cDNA encoding mature canine IL-18 (cIL-18) or mouse IL-18 (mIL-18) fused to the human IL-2 (hIL-2) signal sequence. The expressed proIL-18 proteins were processed to their mature forms in the cells. The supernatants of cells transfected with these plasmids induced high interferon-γ production in canine peripheral blood mononuclear cells or mouse splenocytes, respectively, indicating the secretion of bioactive IL-18. Using reverse genetics, we also generated a recombinant canine distemper virus that expresses cIL-18 or mIL-18 fused to the hIL-2 signal sequence. As expected, both recombinant viruses produced mature IL-18 in the infected cells, which secreted bioactive IL-18. These results indicate that the signal sequence from hIL-2 is suitable for the secretion of mature IL-18. These recombinant viruses can also potentially be used as immunoadjuvants and agents for anticancer therapies in vivo. PMID:24898077

  19. Canine distemper virus persistence in demyelinating encephalitis by swift intracellular cell-to-cell spread in astrocytes is controlled by the viral attachment protein.

    PubMed

    Wyss-Fluehmann, Gaby; Zurbriggen, Andreas; Vandevelde, Marc; Plattet, Philippe

    2010-05-01

    The mechanism of viral persistence, the driving force behind the chronic progression of inflammatory demyelination in canine distemper virus (CDV) infection, is associated with non-cytolytic viral cell-to-cell spread. Here, we studied the molecular mechanisms of viral spread of a recombinant fluorescent protein-expressing virulent CDV in primary canine astrocyte cultures. Time-lapse video microscopy documented that CDV spread was very efficient using cell processes contacting remote target cells. Strikingly, CDV transmission to remote cells could occur in less than 6 h, suggesting that a complete viral cycle with production of extracellular free particles was not essential in enabling CDV to spread in glial cells. Titration experiments and electron microscopy confirmed a very low CDV particle production despite higher titers of membrane-associated viruses. Interestingly, confocal laser microscopy and lentivirus transduction indicated expression and functionality of the viral fusion machinery, consisting of the viral fusion (F) and attachment (H) glycoproteins, at the cell surface. Importantly, using a single-cycle infectious recombinant H-knockout, H-complemented virus, we demonstrated that H, and thus potentially the viral fusion complex, was necessary to enable CDV spread. Furthermore, since we could not detect CD150/SLAM expression in brain cells, the presence of a yet non-identified glial receptor for CDV was suggested. Altogether, our findings indicate that persistence in CDV infection results from intracellular cell-to-cell transmission requiring the CDV-H protein. Viral transfer, happening selectively at the tip of astrocytic processes, may help the virus to cover long distances in the astroglial network, "outrunning" the host's immune response in demyelinating plaques, thus continuously eliciting new lesions.

  20. Phylogenetic analysis of canine distemper virus in South America clade 1 reveals unique molecular signatures of the local epidemic.

    PubMed

    Fischer, Cristine D B; Gräf, Tiago; Ikuta, Nilo; Lehmann, Fernanda K M; Passos, Daniel T; Makiejczuk, Aline; Silveira, Marcos A T; Fonseca, André S K; Canal, Cláudio W; Lunge, Vagner R

    2016-07-01

    Canine distemper virus (CDV) is a highly contagious pathogen for domestic dogs and several wild carnivore species. In Brazil, natural infection of CDV in dogs is very high due to the large non-vaccinated dog population, a scenario that calls for new studies on the molecular epidemiology. This study investigates the phylodynamics and amino-acid signatures of CDV epidemic in South America by analyzing a large dataset compiled from publicly available sequences and also by collecting new samples from Brazil. A population of 175 dogs with canine distemper (CD) signs was sampled, from which 89 were positive for CDV, generating 42 new CDV sequences. Phylogenetic analysis of the new and publicly available sequences revealed that Brazilian sequences mainly clustered in South America 1 (SA1) clade, which has its origin estimated to the late 1980's. The reconstruction of the demographic history in SA1 clade showed an epidemic expanding until the recent years, doubling in size every nine years. SA1 clade epidemic distinguished from the world CDV epidemic by the emergence of the R580Q strain, a very rare and potentially detrimental substitution in the viral genome. The R580Q substitution was estimated to have happened in one single evolutionary step in the epidemic history in SA1 clade, emerging shortly after introduction to the continent. Moreover, a high prevalence (11.9%) of the Y549H mutation was observed among the domestic dogs sampled here. This finding was associated (p<0.05) with outcome-death and higher frequency in mixed-breed dogs, the later being an indicator of a continuous exchange of CDV strains circulating among wild carnivores and domestic dogs. The results reported here highlight the diversity of the worldwide CDV epidemic and reveal local features that can be valuable for combating the disease.

  1. In vitro inhibition of canine distemper virus by flavonoids and phenolic acids: implications of structural differences for antiviral design.

    PubMed

    Carvalho, O V; Botelho, C V; Ferreira, C G T; Ferreira, H C C; Santos, M R; Diaz, M A N; Oliveira, T T; Soares-Martins, J A P; Almeida, M R; Silva, A

    2013-10-01

    Infection caused by canine distemper virus (CDV) is a highly contagious disease with high incidence and lethality in the canine population. Antiviral activity of flavonoids quercetin, morin, rutin and hesperidin, and phenolic cinnamic, trans-cinnamic and ferulic acids were evaluated in vitro against the CDV using the time of addition assay to determine which step of the viral replicative cycle was affected. All flavonoids displayed great viral inhibition when they were added at the times 0 (adsorption) and 1h (penetration) of the viral replicative cycle. Both quercetin and hesperidin presented antiviral activity at the time 2h (intracellular). In the other hand, cinnamic acid showed antiviral activity at the times 0 and 2h while trans-cinnamic acid showed antiviral effect at the times -1h (pre-treatment) and 0 h. Ferulic acid inhibited CDV replicative cycle at the times 0 and 1h. Our study revealed promising candidates to be considered in the treatment of CDV. Structural differences among compounds and correlation to their antiviral activity were also explored. Our analysis suggest that these compounds could be useful in order to design new antiviral drugs against CDV as well as other viruses of great meaning in veterinary medicine.

  2. Effect of vaccination with recombinant canine distemper virus vaccine immediately before exposure under shelter-like conditions.

    PubMed

    Larson, L J; Schultz, R D

    2006-01-01

    Vaccination with modified-live virus (MLV) canine distemper virus (CDV) vaccine has historically been recommended for animals in high-risk environments because of the rapid onset of immunity following vaccination. Recombinant CDV (rCDV) vaccine was deemed a suitable alternative to MLV-CDV vaccination in pet dogs, but insufficient data precluded its use where CDV was a serious threat to puppies, such as in shelters, kennels, and pet stores. In this study, dogs experimentally challenged hours after a single dose of rCDV or MLV vaccine became sick but recovered, whereas unvaccinated dogs became sick and died. Dogs vaccinated with a single dose of rCDV or MLV vaccine 1 week before being experimentally challenged remained healthy and showed no clinical signs. Dogs given one dose of rCDV vaccine hours before being placed in a CDV-contaminated environment did not become sick. These findings support the hypothesis that rCDV vaccine has a similar time-to-immunity as MLV-CDV vaccines and can likewise protect dogs in high-risk environments after one dose. PMID:16871493

  3. The fusion protein of wild-type canine distemper virus is a major determinant of persistent infection

    SciTech Connect

    Plattet, Philippe; Rivals, Jean-Paul; Zuber, BenoIt; Brunner, Jean-Marc; Zurbriggen, Andreas; Wittek, Riccardo . E-mail: Riccardo.Wittek@unil.ch

    2005-07-05

    The wild-type A75/17 canine distemper virus (CDV) strain induces a persistent infection in the central nervous system but infects cell lines very inefficiently. In contrast, the genetically more distant Onderstepoort CDV vaccine strain (OP-CDV) induces extensive syncytia formation. Here, we investigated the roles of wild-type fusion (F{sub WT}) and attachment (H{sub WT}) proteins in Vero cells expressing, or not, the canine SLAM receptor by transfection experiments and by studying recombinants viruses expressing different combinations of wild-type and OP-CDV glycoproteins. We show that low fusogenicity is not due to a defect of the envelope proteins to reach the cell surface and that H{sub WT} determines persistent infection in a receptor-dependent manner, emphasizing the role of SLAM as a potent enhancer of fusogenicity. However, importantly, F{sub WT} reduced cell-to-cell fusion independently of the cell surface receptor, thus demonstrating that the fusion protein of the neurovirulent A75/17-CDV strain plays a key role in determining persistent infection.

  4. Accuracy of a point-of-care ELISA test kit for predicting the presence of protective canine parvovirus and canine distemper virus antibody concentrations in dogs.

    PubMed

    Litster, A L; Pressler, B; Volpe, A; Dubovi, E

    2012-08-01

    Canine parvovirus (CPV) and canine distemper virus (CDV) are highly infectious and often fatal diseases with worldwide distributions, and are important population management considerations in animal shelters. A point-of-care ELISA test kit is available to detect serum antibodies to CPV and CDV, and presumptively to predict protective status. The aim of this study was to determine the diagnostic accuracy of the test compared to CPV hemagglutination inhibition titers and CDV serum neutralization titers determined by a reference laboratory, using sera collected from dogs housed at animal shelters. The ELISA test was used under both field and laboratory conditions and duplicate specimens were processed using an extra wash step. The test kit yielded accurate results (CPV: sensitivity 92.3%, specificity 93.5%; CDV: sensitivity 75.7%, specificity 91.8%) under field conditions. CDV sensitivity was improved by performing the test under laboratory conditions and using an optical density (OD) meter (laboratory performed 94.0%; OD 88.1%). Point-of-care ELISA testing for serum CPV and CDV antibody titers was demonstrated to be a useful tool for determining antibody status when making decisions regarding the need for CPV and/or CDV vaccination and also in animal shelters for population management.

  5. Experimental Adaptation of Wild-Type Canine Distemper Virus (CDV) to the Human Entry Receptor CD150

    PubMed Central

    Bieringer, Maria; Han, Jung Woo; Kendl, Sabine; Khosravi, Mojtaba; Plattet, Philippe; Schneider-Schaulies, Jürgen

    2013-01-01

    Canine distemper virus (CDV), a close relative of measles virus (MV), is widespread and well known for its broad host range. When the goal of measles eradication may be achieved, and when measles vaccination will be stopped, CDV might eventually cross the species barrier to humans and emerge as a new human pathogen. In order to get an impression how fast such alterations may occur, we characterized required adaptive mutations to the human entry receptors CD150 (SLAM) and nectin-4 as first step to infect human target cells. Recombinant wild-type CDV-A75/17red adapted quickly to growth in human H358 epithelial cells expressing human nectin-4. Sequencing of the viral attachment proteins (hemagglutinin, H, and fusion protein, F) genes revealed that no adaptive alteration was required to utilize human nectin-4. In contrast, the virus replicated only to low titres (102 pfu/ml) in Vero cells expressing human CD150 (Vero-hSLAM). After three passages using these cells virus was adapted to human CD150 and replicated to high titres (105 pfu/ml). Sequence analyses revealed that only one amino acid exchange in the H-protein at position 540 Asp→Gly (D540G) was required for functional adaptation to human CD150. Structural modelling suggests that the adaptive mutation D540G in H reflects the sequence alteration from canine to human CD150 at position 70 and 71 from Pro to Leu (P70L) and Gly to Glu (G71E), and compensates for the gain of a negative charge in the human CD150 molecule. Using this model system our data indicate that only a minimal alteration, in this case one adaptive mutation, is required for adaptation of CDV to the human entry receptors, and help to understand the molecular basis why this adaptive mutation occurs. PMID:23554862

  6. Parasites in harbour seals ( Phoca vitulina) from the German Wadden Sea between two Phocine Distemper Virus epidemics

    NASA Astrophysics Data System (ADS)

    Lehnert, K.; Raga, J. A.; Siebert, U.

    2007-12-01

    Parasites were collected from 107 harbour seals ( Phoca vitulina) found on the coasts of Schleswig-Holstein, Germany, between 1997 and 2000. The prevalence of the parasites and their associated pathology were investigated. Eight species of parasites, primarily nematodes, were identified from the examined organs: two anisakid nematodes ( Pseudoterranova decipiens (sensu lato) , Contracaecum osculatum (sensu lato)) from the stomach, Otostrongylus circumlitus (Crenosomatidae) and Parafilaroides gymnurus (Filaroididae) from the respiratory tract, one filarioid nematode ( Acanthocheilonema spirocauda) from the heart, two acanthocephalans, Corynosoma strumosum and C. semerme (Polymorphidae), from the intestine and an ectoparasite, Echinophthirius horridus (Anoplura, Insecta). Lungworm infection was the most prominent parasitological finding and secondary bacterial bronchopneumonia the most pathogenic lesion correlated with the parasites. Heavy nematode burdens in the respiratory tract were highly age-related and more frequent in young seals. A positive correlation was observed between high levels of pulmonary infection and severity of bronchopneumonia. The prevalence of lungworms in this study was higher than in seals that died during the 1988/1989 Phocine Distemper Virus epidemic, and the prevalence of acanthocephalans and heartworms had decreased compared to findings from the first die-off.

  7. RT-PCR and sequence analysis of the full-length fusion protein of Canine Distemper Virus from domestic dogs.

    PubMed

    Romanutti, Carina; Gallo Calderón, Marina; Keller, Leticia; Mattion, Nora; La Torre, José

    2016-02-01

    During 2007-2014, 84 out of 236 (35.6%) samples from domestic dogs submitted to our laboratory for diagnostic purposes were positive for Canine Distemper Virus (CDV), as analyzed by RT-PCR amplification of a fragment of the nucleoprotein gene. Fifty-nine of them (70.2%) were from dogs that had been vaccinated against CDV. The full-length gene encoding the Fusion (F) protein of fifteen isolates was sequenced and compared with that of those of other CDVs, including wild-type and vaccine strains. Phylogenetic analysis using the F gene full-length sequences grouped all the Argentinean CDV strains in the SA2 clade. Sequence identity with the Onderstepoort vaccine strain was 89.0-90.6%, and the highest divergence was found in the 135 amino acids corresponding to the F protein signal-peptide, Fsp (64.4-66.7% identity). In contrast, this region was highly conserved among the local strains (94.1-100% identity). One extra putative N-glycosylation site was identified in the F gene of CDV Argentinean strains with respect to the vaccine strain. The present report is the first to analyze full-length F protein sequences of CDV strains circulating in Argentina, and contributes to the knowledge of molecular epidemiology of CDV, which may help in understanding future disease outbreaks. PMID:26611227

  8. RT-PCR and sequence analysis of the full-length fusion protein of Canine Distemper Virus from domestic dogs.

    PubMed

    Romanutti, Carina; Gallo Calderón, Marina; Keller, Leticia; Mattion, Nora; La Torre, José

    2016-02-01

    During 2007-2014, 84 out of 236 (35.6%) samples from domestic dogs submitted to our laboratory for diagnostic purposes were positive for Canine Distemper Virus (CDV), as analyzed by RT-PCR amplification of a fragment of the nucleoprotein gene. Fifty-nine of them (70.2%) were from dogs that had been vaccinated against CDV. The full-length gene encoding the Fusion (F) protein of fifteen isolates was sequenced and compared with that of those of other CDVs, including wild-type and vaccine strains. Phylogenetic analysis using the F gene full-length sequences grouped all the Argentinean CDV strains in the SA2 clade. Sequence identity with the Onderstepoort vaccine strain was 89.0-90.6%, and the highest divergence was found in the 135 amino acids corresponding to the F protein signal-peptide, Fsp (64.4-66.7% identity). In contrast, this region was highly conserved among the local strains (94.1-100% identity). One extra putative N-glycosylation site was identified in the F gene of CDV Argentinean strains with respect to the vaccine strain. The present report is the first to analyze full-length F protein sequences of CDV strains circulating in Argentina, and contributes to the knowledge of molecular epidemiology of CDV, which may help in understanding future disease outbreaks.

  9. Contact rates between wild and domestic canids: no evidence of parvovirus or canine distemper virus in crab-eating foxes.

    PubMed

    Courtenay, O; Quinnell, R J; Chalmers, W S

    2001-07-01

    Evaluating the risk of disease spill-over from domestic dogs to wildlife depends on knowledge of inter-specific contact rates and/or exposure to aetiological agents in dog environments. Here, contact rates of crab-eating foxes (Cerdocyon thous) with sympatric domestic dog populations were measured over 25months in Amazon Brazil. Foxes and dogs were serologically and clinically monitored for exposure to canine parvovirus (CPV-2) and canine distemper virus (CDV), pathogens known to have caused wildlife population declines elsewhere. Twenty-two of 24 (92%) tagged foxes visited one or more houses in a median 2 (range 1-3) villages per night where dog densities ranged from 7.2 to 15.4 per km(2) (mean 9.5 per km(2)). Foxes spent an average 6.4% (0-40.3%) of their 10h nocturnal activity period in villages, the equivalent of 38m (range 0-242) per night. The rate of potential exposure to disease agents was thus high, though varied by 3 orders of magnitude for individual foxes. Overall, 46% of the fox population was responsible for 80% of all contacts. None of the 37 monitored foxes however showed serological or clinical evidence of infection with CPV-2 or CDV. Seroprevalences for CPV-2 and CDV antibodies in the local domestic dog population were 13% (3/23) and 9% (2/23), respectively, and 89% of 97 monitored pups born during the study presented clinical signs consistent with active CPV-2 infection (haemorrhagic diarrhoea, vomiting, rapid morbidity and emaciation). Although there was no evidence for infection with either virus in foxes, the high level of contact of foxes with peridomestic habitats suggests that the probability of potential spill-over infections from dogs to foxes is high.

  10. Estimating the Potential Impact of Canine Distemper Virus on the Amur Tiger Population (Panthera tigris altaica) in Russia

    PubMed Central

    Gilbert, Martin; Miquelle, Dale G.; Goodrich, John M.; Reeve, Richard; Cleaveland, Sarah; Matthews, Louise; Joly, Damien O.

    2014-01-01

    Lethal infections with canine distemper virus (CDV) have recently been diagnosed in Amur tigers (Panthera tigris altaica), but long-term implications for the population are unknown. This study evaluates the potential impact of CDV on a key tiger population in Sikhote-Alin Biosphere Zapovednik (SABZ), and assesses how CDV might influence the extinction potential of other tiger populations of varying sizes. An individual-based stochastic, SIRD (susceptible-infected-recovered/dead) model was used to simulate infection through predation of infected domestic dogs, and/or wild carnivores, and direct tiger-to-tiger transmission. CDV prevalence and effective contact based on published and observed data was used to define plausible low- and high-risk infection scenarios. CDV infection increased the 50-year extinction probability of tigers in SABZ by 6.3% to 55.8% compared to a control population, depending on risk scenario. The most significant factors influencing model outcome were virus prevalence in the reservoir population(s) and its effective contact rate with tigers. Adjustment of the mortality rate had a proportional impact, while inclusion of epizootic infection waves had negligible additional impact. Small populations were found to be disproportionately vulnerable to extinction through CDV infection. The 50-year extinction risk in populations consisting of 25 individuals was 1.65 times greater when CDV was present than that of control populations. The effects of density dependence do not protect an endangered population from the impacts of a multi-host pathogen, such as CDV, where they coexist with an abundant reservoir presenting a persistent threat. Awareness of CDV is a critical component of a successful tiger conservation management policy. PMID:25354196

  11. Two Domains of the V Protein of Virulent Canine Distemper Virus Selectively Inhibit STAT1 and STAT2 Nuclear Import▿

    PubMed Central

    Röthlisberger, Anne; Wiener, Dominique; Schweizer, Matthias; Peterhans, Ernst; Zurbriggen, Andreas; Plattet, Philippe

    2010-01-01

    Canine distemper virus (CDV) causes in dogs a severe systemic infection, with a high frequency of demyelinating encephalitis. Among the six genes transcribed by CDV, the P gene encodes the polymerase cofactor protein (P) as well as two additional nonstructural proteins, C and V; of these V was shown to act as a virulence factor. We investigated the molecular mechanisms by which the P gene products of the neurovirulent CDV A75/17 strain disrupt type I interferon (IFN-α/β)-induced signaling that results in the establishment of the antiviral state. Using recombinant knockout A75/17 viruses, the V protein was identified as the main antagonist of IFN-α/β-mediated signaling. Importantly, immunofluorescence analysis illustrated that the inhibition of IFN-α/β-mediated signaling correlated with impaired STAT1/STAT2 nuclear import, whereas the phosphorylation state of these proteins was not affected. Coimmunoprecipitation assays identified the N-terminal region of V (VNT) responsible for STAT1 targeting, which correlated with its ability to inhibit the activity of the IFN-α/β-mediated antiviral state. Conversely, while the C-terminal domain of V (VCT) could not function autonomously, when fused to VNT it optimally interacted with STAT2 and subsequently efficiently suppressed the IFN-α/β-mediated signaling pathway. The latter result was further supported by a single mutation at position 110 within the VNT domain of CDV V protein, resulting in a mutant that lost STAT1 binding while retaining a partial STAT2 association. Taken together, our results identified the CDV VNT and VCT as two essential modules that complement each other to interfere with the antiviral state induced by IFN-α/β-mediated signaling. Hence, our experiments reveal a novel mechanism of IFN-α/β evasion among the morbilliviruses. PMID:20427537

  12. Estimating the potential impact of canine distemper virus on the Amur tiger population (Panthera tigris altaica) in Russia.

    PubMed

    Gilbert, Martin; Miquelle, Dale G; Goodrich, John M; Reeve, Richard; Cleaveland, Sarah; Matthews, Louise; Joly, Damien O

    2014-01-01

    Lethal infections with canine distemper virus (CDV) have recently been diagnosed in Amur tigers (Panthera tigris altaica), but long-term implications for the population are unknown. This study evaluates the potential impact of CDV on a key tiger population in Sikhote-Alin Biosphere Zapovednik (SABZ), and assesses how CDV might influence the extinction potential of other tiger populations of varying sizes. An individual-based stochastic, SIRD (susceptible-infected-recovered/dead) model was used to simulate infection through predation of infected domestic dogs, and/or wild carnivores, and direct tiger-to-tiger transmission. CDV prevalence and effective contact based on published and observed data was used to define plausible low- and high-risk infection scenarios. CDV infection increased the 50-year extinction probability of tigers in SABZ by 6.3% to 55.8% compared to a control population, depending on risk scenario. The most significant factors influencing model outcome were virus prevalence in the reservoir population(s) and its effective contact rate with tigers. Adjustment of the mortality rate had a proportional impact, while inclusion of epizootic infection waves had negligible additional impact. Small populations were found to be disproportionately vulnerable to extinction through CDV infection. The 50-year extinction risk in populations consisting of 25 individuals was 1.65 times greater when CDV was present than that of control populations. The effects of density dependence do not protect an endangered population from the impacts of a multi-host pathogen, such as CDV, where they coexist with an abundant reservoir presenting a persistent threat. Awareness of CDV is a critical component of a successful tiger conservation management policy.

  13. Recent host range expansion of canine distemper virus and variation in its receptor, the signaling lymphocyte activation molecule, in carnivores.

    PubMed

    Ohishi, Kazue; Suzuki, Rintaro; Maeda, Taro; Tsuda, Miwako; Abe, Erika; Yoshida, Takao; Endo, Yasuyuki; Okamura, Maki; Nagamine, Takashi; Yamamoto, Hanae; Ueda, Miya; Maruyama, Tadashi

    2014-07-01

    The signaling lymphocyte activation molecule (SLAM) is a receptor for morbilliviruses. To understand the recent host range expansion of canine distemper virus (CDV) in carnivores, we determined the nucleotide sequences of SLAMs of various carnivores and generated three-dimensional homology SLAM models. Thirty-four amino acid residues were found for the candidates binding to CDV on the interface of the carnivore SLAMs. SLAM of the domestic dog (Canis lupus familiaris) were similar to those of other members of the suborder Caniformia, indicating that the animals in this group have similar sensitivity to dog CDV. However, they were different at nine positions from those of felids. Among the nine residues, four of domestic cat (Felis catus) SLAM (72, 76, 82, and 129) and three of lion (Panthera leo persica) SLAM (72, 82, and 129) were associated with charge alterations, suggesting that the felid interfaces have lower affinities to dog CDV. Only the residue at 76 was different between domestic cat and lion SLAM interfaces. The domestic cat SLAM had threonine at 76, whereas the lion SLAM had arginine, a positively charged residue like that of the dog SLAM. The cat SLAM with threonine is likely to have lower affinity to CDV-H and to confer higher resistance against dog CDV. Thus, the four residues (72, 76, 82, and 129) on carnivore SLAMs are important for the determination of affinity and sensitivity with CDV. Additionally, the CDV-H protein of felid strains had a substitution of histidine for tyrosine at 549 of dog CDV-H and may have higher affinity to lion SLAM. Three-dimensional model construction is a new risk assessment method of morbillivirus infectivity. Because the method is applicable to animals that have no information about virus infection, it is especially useful for morbillivirus risk assessment and wildlife conservation.

  14. Estimating the potential impact of canine distemper virus on the Amur tiger population (Panthera tigris altaica) in Russia.

    PubMed

    Gilbert, Martin; Miquelle, Dale G; Goodrich, John M; Reeve, Richard; Cleaveland, Sarah; Matthews, Louise; Joly, Damien O

    2014-01-01

    Lethal infections with canine distemper virus (CDV) have recently been diagnosed in Amur tigers (Panthera tigris altaica), but long-term implications for the population are unknown. This study evaluates the potential impact of CDV on a key tiger population in Sikhote-Alin Biosphere Zapovednik (SABZ), and assesses how CDV might influence the extinction potential of other tiger populations of varying sizes. An individual-based stochastic, SIRD (susceptible-infected-recovered/dead) model was used to simulate infection through predation of infected domestic dogs, and/or wild carnivores, and direct tiger-to-tiger transmission. CDV prevalence and effective contact based on published and observed data was used to define plausible low- and high-risk infection scenarios. CDV infection increased the 50-year extinction probability of tigers in SABZ by 6.3% to 55.8% compared to a control population, depending on risk scenario. The most significant factors influencing model outcome were virus prevalence in the reservoir population(s) and its effective contact rate with tigers. Adjustment of the mortality rate had a proportional impact, while inclusion of epizootic infection waves had negligible additional impact. Small populations were found to be disproportionately vulnerable to extinction through CDV infection. The 50-year extinction risk in populations consisting of 25 individuals was 1.65 times greater when CDV was present than that of control populations. The effects of density dependence do not protect an endangered population from the impacts of a multi-host pathogen, such as CDV, where they coexist with an abundant reservoir presenting a persistent threat. Awareness of CDV is a critical component of a successful tiger conservation management policy. PMID:25354196

  15. The hemagglutinin envelope protein of canine distemper virus (CDV) confers cell tropism as illustrated by CDV and measles virus complementation analysis.

    PubMed Central

    Stern, L B; Greenberg, M; Gershoni, J M; Rozenblatt, S

    1995-01-01

    Measles virus (MV) and canine distemper virus (CDV) are morbilliviruses that cause acute illnesses and several persistent central nervous system infections in humans and in dogs, respectively. Characteristically, the cytopathic effect of these viruses is the formation of syncytia in permissive cells. In this study, a vaccinia virus expression system was used to express MV and CDV hemagglutinin (HA) and fusion (F) envelope proteins. We found that cotransfecting F and HA genes of MV or F and HA genes of CDV resulted in extensive syncytium formation in permissive cells while transfecting either F or HA alone did not. Similar experiments with heterologous pairs of proteins, CDV-F with MV-HA or MV-F with CDV-HA, caused significant cell fusion in both cases. These results indicate that in this expression system, cell fusion requires both F and HA; however, the functions of these proteins are interchangeable between the two types of morbilliviruses. Human-mouse somatic hybrids were used to determine the human chromosome conferring susceptibility to either MV and CDV. Of the 12 hybrids screened, none were sensitive to MV. Two of the hybrids containing human chromosome 19 formed syncytia following CDV infection. In addition, these two hybrids underwent cell fusion when cotransfected with CDV-F and CDV-HA (but not MV-F and MV-HA) glycoproteins by using the vaccinia virus expression system. To discover the viral component responsible for cell specificity, complementation experiments coexpressing CDV-HA with MV-F or CDV-F with MV-HA in the CDV-sensitive hybrids were performed. We found that syncytia were formed only in the presence of CDV-HA. These results support the idea that the HA protein is responsible for cell tropism. Furthermore, while the F protein is necessary for the fusion process, it is interchangeable with the F protein from other morbilliviruses. PMID:7853502

  16. Water system virus detection

    NASA Technical Reports Server (NTRS)

    Fraser, A. S.; Wells, A. F.; Tenoso, H. J.

    1975-01-01

    A monitoring system developed to test the capability of a water recovery system to reject the passage of viruses into the recovered water is described. A nonpathogenic marker virus, bacteriophage F2, is fed into the process stream before the recovery unit and the reclaimed water is assayed for its presence. Detection of the marker virus consists of two major components, concentration and isolation of the marker virus, and detection of the marker virus. The concentration system involves adsorption of virus to cellulose acetate filters in the presence of trivalent cations and low pH with subsequent desorption of the virus using volumes of high pH buffer. The detection of the virus is performed by a passive immune agglutination test utilizing specially prepared polystyrene particles. An engineering preliminary design was performed as a parallel effort to the laboratory development of the marker virus test system. Engineering schematics and drawings of a fully functional laboratory prototype capable of zero-G operation are presented. The instrument consists of reagent pump/metering system, reagent storage containers, a filter concentrator, an incubation/detector system, and an electronic readout and control system.

  17. Multiplex Amplification Refractory Mutation System Polymerase Chain Reaction (ARMS-PCR) for diagnosis of natural infection with canine distemper virus

    PubMed Central

    2010-01-01

    Background Canine distemper virus (CDV) is present worldwide and produces a lethal systemic infection of wild and domestic Canidae. Pre-existing antibodies acquired from vaccination or previous CDV infection might interfere the interpretation of a serologic diagnosis method. In addition, due to the high similarity of nucleic acid sequences between wild-type CDV and the new vaccine strain, current PCR derived methods cannot be applied for the definite confirmation of CD infection. Hence, it is worthy of developing a simple and rapid nucleotide-based assay for differentiation of wild-type CDV which is a cause of disease from attenuated CDVs after vaccination. High frequency variations have been found in the region spanning from the 3'-untranslated region (UTR) of the matrix (M) gene to the fusion (F) gene (designated M-F UTR) in a few CDV strains. To establish a differential diagnosis assay, an amplification refractory mutation analysis was established based on the highly variable region on M-F UTR and F regions. Results Sequences of frequent polymorphisms were found scattered throughout the M-F UTR region; the identity of nucleic acid between local strains and vaccine strains ranged from 82.5% to 93.8%. A track of AAA residue located 35 nucleotides downstream from F gene start codon highly conserved in three vaccine strains were replaced with TGC in the local strains; that severed as target sequences for deign of discrimination primers. The method established in the present study successfully differentiated seven Taiwanese CDV field isolates, all belonging to the Asia-1 lineage, from vaccine strains. Conclusions The method described herein would be useful for several clinical applications, such as confirmation of nature CDV infection, evaluation of vaccination status and verification of the circulating viral genotypes. PMID:20534175

  18. Rabies and Canine Distemper Virus Epidemics in the Red Fox Population of Northern Italy (2006–2010)

    PubMed Central

    De Benedictis, Paola; Citterio, Carlo; Obber, Federica; Lorenzetto, Monica; Pozza, Manuela Dalla; Cauchemez, Simon; Cattoli, Giovanni

    2013-01-01

    Since 2006 the red fox (Vulpes vulpes) population in north-eastern Italy has experienced an epidemic of canine distemper virus (CDV). Additionally, in 2008, after a thirteen-year absence from Italy, fox rabies was re-introduced in the Udine province at the national border with Slovenia. Disease intervention strategies are being developed and implemented to control rabies in this area and minimise risk to human health. Here we present empirical data and the epidemiological picture relating to these epidemics in the period 2006–2010. Of important significance for epidemiological studies of wild animals, basic mathematical models are developed to exploit information collected from the surveillance program on dead and/or living animals in order to assess the incidence of infection. These models are also used to estimate the rate of transmission of both diseases and the rate of vaccination, while correcting for a bias in early collection of CDV samples. We found that the rate of rabies transmission was roughly twice that of CDV, with an estimated effective contact between infected and susceptible fox leading to a new infection occurring once every 3 days for rabies, and once a week for CDV. We also inferred that during the early stage of the CDV epidemic, a bias in the monitoring protocol resulted in a positive sample being almost 10 times more likely to be collected than a negative sample. We estimated the rate of intake of oral vaccine at 0.006 per day, allowing us to estimate that roughly 68% of the foxes would be immunised. This was confirmed by field observations. Finally we discuss the implications for the eco-epidemiological dynamics of both epidemics in relation to control measures. PMID:23630599

  19. Dog overpopulation and burden of exposure to canine distemper virus and other pathogens on Santa Cruz Island, Galapagos.

    PubMed

    Diaz, Nicole M; Mendez, Gabriella S; Grijalva, C Jaime; Walden, Heather S; Cruz, Marilyn; Aragon, Eduardo; Hernandez, Jorge A

    2016-01-01

    Dog overpopulation and diseases are hazards to native island species and humans on the Galapagos. Vaccination and importation of dogs are prohibited on the Galapagos. Risk management of these hazards requires the use of science-based risk assessment and risk communication. The objectives of the study reported here were (i) to estimate the human:dog ratio and (ii) the prevalence of and identify exposure factors associated with positive antibody titers to canine distemper virus (CDV) and other pathogens, as well as infection with intestinal parasites in owned dogs on Santa Cruz Island, Galapagos in September 2014. The observed human:dog ratio was 6.148:1 which extrapolates to 2503 dogs (two times more than a recent dog count conducted by Galapagos Biosecurity Agency in March 2014). The proportion of spayed female dogs (50%) was higher, compared to neutered male dogs (30%) (p=0.04). Prevalence of dogs with positive antibody titers to CDV was 36% (95% CI=26, 46%), to canine parvovirus was 89% (95% CI=82, 95%), and to canine adenovirus was 40% (95% CI=30, 51%). The frequency of seropositive dogs to CDV was lower in urban dogs (26%), compared to rural dogs (53%) (p<0.05). A positive interaction effect between rural residence and spay/neuter status on seropositivity to CDV was observed, which we discuss in this report. Because vaccination is prohibited, the dog population on Santa Cruz is susceptible to an outbreak of CDV (particularly among urban dogs) with potential spill over to marine mammals. Dog's age (1-2 or 3-14 years old, compared to younger dogs), and residence (rural, urban) were associated with positive antibody titers to parvovirus, adenovirus, Ehrlichia spp., or Anaplasma spp., as well as infection with Ancylostoma spp., an intestinal parasite in dogs that can be transmitted to humans, particularly children. These results provide the most comprehensive assessment of dog overpopulation and exposure to CDV and other pathogens on the Galapagos to date.

  20. Dynamics of a morbillivirus at the domestic–wildlife interface: Canine distemper virus in domestic dogs and lions

    PubMed Central

    Viana, Mafalda; Matthiopoulos, Jason; Halliday, Jo; Packer, Craig; Craft, Meggan E.; Hampson, Katie; Czupryna, Anna; Dobson, Andrew P.; Dubovi, Edward J.; Ernest, Eblate; Fyumagwa, Robert; Hoare, Richard; Hopcraft, J. Grant C.; Horton, Daniel L.; Kaare, Magai T.; Kanellos, Theo; Lankester, Felix; Mentzel, Christine; Mlengeya, Titus; Mzimbiri, Imam; Takahashi, Emi; Willett, Brian; Haydon, Daniel T.; Lembo, Tiziana

    2015-01-01

    Morbilliviruses cause many diseases of medical and veterinary importance, and although some (e.g., measles and rinderpest) have been controlled successfully, others, such as canine distemper virus (CDV), are a growing concern. A propensity for host-switching has resulted in CDV emergence in new species, including endangered wildlife, posing challenges for controlling disease in multispecies communities. CDV is typically associated with domestic dogs, but little is known about its maintenance and transmission in species-rich areas or about the potential role of domestic dog vaccination as a means of reducing disease threats to wildlife. We address these questions by analyzing a long-term serological dataset of CDV in lions and domestic dogs from Tanzania’s Serengeti ecosystem. Using a Bayesian state–space model, we show that dynamics of CDV have changed considerably over the past three decades. Initially, peaks of CDV infection in dogs preceded those in lions, suggesting that spill-over from dogs was the main driver of infection in wildlife. However, despite dog-to-lion transmission dominating cross-species transmission models, infection peaks in lions became more frequent and asynchronous from those in dogs, suggesting that other wildlife species may play a role in a potentially complex maintenance community. Widespread mass vaccination of domestic dogs reduced the probability of infection in dogs and the size of outbreaks but did not prevent transmission to or peaks of infection in lions. This study demonstrates the complexity of CDV dynamics in natural ecosystems and the value of long-term, large-scale datasets for investigating transmission patterns and evaluating disease control strategies. PMID:25605919

  1. Dynamics of a morbillivirus at the domestic-wildlife interface: Canine distemper virus in domestic dogs and lions.

    PubMed

    Viana, Mafalda; Cleaveland, Sarah; Matthiopoulos, Jason; Halliday, Jo; Packer, Craig; Craft, Meggan E; Hampson, Katie; Czupryna, Anna; Dobson, Andrew P; Dubovi, Edward J; Ernest, Eblate; Fyumagwa, Robert; Hoare, Richard; Hopcraft, J Grant C; Horton, Daniel L; Kaare, Magai T; Kanellos, Theo; Lankester, Felix; Mentzel, Christine; Mlengeya, Titus; Mzimbiri, Imam; Takahashi, Emi; Willett, Brian; Haydon, Daniel T; Lembo, Tiziana

    2015-02-01

    Morbilliviruses cause many diseases of medical and veterinary importance, and although some (e.g., measles and rinderpest) have been controlled successfully, others, such as canine distemper virus (CDV), are a growing concern. A propensity for host-switching has resulted in CDV emergence in new species, including endangered wildlife, posing challenges for controlling disease in multispecies communities. CDV is typically associated with domestic dogs, but little is known about its maintenance and transmission in species-rich areas or about the potential role of domestic dog vaccination as a means of reducing disease threats to wildlife. We address these questions by analyzing a long-term serological dataset of CDV in lions and domestic dogs from Tanzania's Serengeti ecosystem. Using a Bayesian state-space model, we show that dynamics of CDV have changed considerably over the past three decades. Initially, peaks of CDV infection in dogs preceded those in lions, suggesting that spill-over from dogs was the main driver of infection in wildlife. However, despite dog-to-lion transmission dominating cross-species transmission models, infection peaks in lions became more frequent and asynchronous from those in dogs, suggesting that other wildlife species may play a role in a potentially complex maintenance community. Widespread mass vaccination of domestic dogs reduced the probability of infection in dogs and the size of outbreaks but did not prevent transmission to or peaks of infection in lions. This study demonstrates the complexity of CDV dynamics in natural ecosystems and the value of long-term, large-scale datasets for investigating transmission patterns and evaluating disease control strategies.

  2. Dynamics of a morbillivirus at the domestic-wildlife interface: Canine distemper virus in domestic dogs and lions.

    PubMed

    Viana, Mafalda; Cleaveland, Sarah; Matthiopoulos, Jason; Halliday, Jo; Packer, Craig; Craft, Meggan E; Hampson, Katie; Czupryna, Anna; Dobson, Andrew P; Dubovi, Edward J; Ernest, Eblate; Fyumagwa, Robert; Hoare, Richard; Hopcraft, J Grant C; Horton, Daniel L; Kaare, Magai T; Kanellos, Theo; Lankester, Felix; Mentzel, Christine; Mlengeya, Titus; Mzimbiri, Imam; Takahashi, Emi; Willett, Brian; Haydon, Daniel T; Lembo, Tiziana

    2015-02-01

    Morbilliviruses cause many diseases of medical and veterinary importance, and although some (e.g., measles and rinderpest) have been controlled successfully, others, such as canine distemper virus (CDV), are a growing concern. A propensity for host-switching has resulted in CDV emergence in new species, including endangered wildlife, posing challenges for controlling disease in multispecies communities. CDV is typically associated with domestic dogs, but little is known about its maintenance and transmission in species-rich areas or about the potential role of domestic dog vaccination as a means of reducing disease threats to wildlife. We address these questions by analyzing a long-term serological dataset of CDV in lions and domestic dogs from Tanzania's Serengeti ecosystem. Using a Bayesian state-space model, we show that dynamics of CDV have changed considerably over the past three decades. Initially, peaks of CDV infection in dogs preceded those in lions, suggesting that spill-over from dogs was the main driver of infection in wildlife. However, despite dog-to-lion transmission dominating cross-species transmission models, infection peaks in lions became more frequent and asynchronous from those in dogs, suggesting that other wildlife species may play a role in a potentially complex maintenance community. Widespread mass vaccination of domestic dogs reduced the probability of infection in dogs and the size of outbreaks but did not prevent transmission to or peaks of infection in lions. This study demonstrates the complexity of CDV dynamics in natural ecosystems and the value of long-term, large-scale datasets for investigating transmission patterns and evaluating disease control strategies. PMID:25605919

  3. The role of canine distemper virus and persistent organic pollutants in mortality patterns of Caspian seals (Pusa caspica).

    PubMed

    Wilson, Susan C; Eybatov, Tariel M; Amano, Masao; Jepson, Paul D; Goodman, Simon J

    2014-01-01

    Persistent organic pollutants are a concern for species occupying high trophic levels since they can cause immunosuppression and impair reproduction. Mass mortalities due to canine distemper virus (CDV) occurred in Caspian seals (Pusa caspica), in spring of 1997, 2000 and 2001, but the potential role of organochlorine exposure in these epizootics remains undetermined. Here we integrate Caspian seal mortality data spanning 1971-2008, with data on age, body condition, pathology and blubber organochlorine concentration for carcases stranded between 1997 and 2002. We test the hypothesis that summed PCB and DDT concentrations contributed to CDV associated mortality during epizootics. We show that age is the primary factor explaining variation in blubber organochlorine concentrations, and that organochlorine burden, age, sex, and body condition do not account for CDV infection status (positive/negative) of animals dying in epizootics. Most animals (57%, n = 67) had PCB concentrations below proposed thresholds for toxic effects in marine mammals (17 µg/g lipid weight), and only 3 of 67 animals had predicted TEQ values exceeding levels seen to be associated with immune suppression in harbour seals (200 pg/g lipid weight). Mean organonchlorine levels were higher in CDV-negative animals indicating that organochlorines did not contribute significantly to CDV mortality in epizootics. Mortality monitoring in Azerbaijan 1971-2008 revealed bi-annual stranding peaks in late spring, following the annual moult and during autumn migrations northwards. Mortality peaks comparable to epizootic years were also recorded in the 1970s-1980s, consistent with previous undocumented CDV outbreaks. Gompertz growth curves show that Caspian seals achieve an asymptotic standard body length of 126-129 cm (n = 111). Males may continue to grow slowly throughout life. Mortality during epizootics may exceed the potential biological removal level (PBR) for the population, but the low frequency of

  4. The Role of Canine Distemper Virus and Persistent Organic Pollutants in Mortality Patterns of Caspian Seals (Pusa caspica)

    PubMed Central

    Wilson, Susan C.; Eybatov, Tariel M.; Amano, Masao; Jepson, Paul D.; Goodman, Simon J.

    2014-01-01

    Persistent organic pollutants are a concern for species occupying high trophic levels since they can cause immunosuppression and impair reproduction. Mass mortalities due to canine distemper virus (CDV) occurred in Caspian seals (Pusa caspica), in spring of 1997, 2000 and 2001, but the potential role of organochlorine exposure in these epizootics remains undetermined. Here we integrate Caspian seal mortality data spanning 1971–2008, with data on age, body condition, pathology and blubber organochlorine concentration for carcases stranded between 1997 and 2002. We test the hypothesis that summed PCB and DDT concentrations contributed to CDV associated mortality during epizootics. We show that age is the primary factor explaining variation in blubber organochlorine concentrations, and that organochlorine burden, age, sex, and body condition do not account for CDV infection status (positive/negative) of animals dying in epizootics. Most animals (57%, n = 67) had PCB concentrations below proposed thresholds for toxic effects in marine mammals (17 µg/g lipid weight), and only 3 of 67 animals had predicted TEQ values exceeding levels seen to be associated with immune suppression in harbour seals (200 pg/g lipid weight). Mean organonchlorine levels were higher in CDV-negative animals indicating that organochlorines did not contribute significantly to CDV mortality in epizootics. Mortality monitoring in Azerbaijan 1971–2008 revealed bi-annual stranding peaks in late spring, following the annual moult and during autumn migrations northwards. Mortality peaks comparable to epizootic years were also recorded in the 1970s–1980s, consistent with previous undocumented CDV outbreaks. Gompertz growth curves show that Caspian seals achieve an asymptotic standard body length of 126–129 cm (n = 111). Males may continue to grow slowly throughout life. Mortality during epizootics may exceed the potential biological removal level (PBR) for the population, but the low

  5. Presence of antibodies to canine distemper virus, canine parvovirus and canine adenovirus type 1 in free-ranging jackals (Canis adustus and Canis mesomelas) in Zimbabwe.

    PubMed

    Spencer, J A; Bingham, J; Heath, R; Richards, B

    1999-09-01

    A survey of free-ranging jackals (Canis adustus and Canis mesomelas) in Zimbabwe was conducted to determine the prevalence of serum antibodies to canine distemper virus (CDV), canine parvovirus (CPV) and canine adenovirus type 1 (CAV-1). Sera from 16 Canis adustus and 22 Canis mesomelas were collected from 1990 to 1993 from various regions of Zimbabwe and assayed by means of immunofluorescent techniques. Seroprevalence in C. adustus and C. mesomelas respectively were 50% and 63.6% for CDV, 12.5% and 18.2% for CPV and 37.5 and 9.1 for CAV-1. These results demonstrate that jackals are infected with these viruses and may act as reservoirs of them, although their susceptibility to the viruses is not known.

  6. Water system virus detection

    NASA Technical Reports Server (NTRS)

    Fraser, A. S.; Wells, A. F.; Tenoso, H. J. (Inventor)

    1978-01-01

    The performance of a waste water reclamation system is monitored by introducing a non-pathogenic marker virus, bacteriophage F2, into the waste-water prior to treatment and, thereafter, testing the reclaimed water for the presence of the marker virus. A test sample is first concentrated by absorbing any marker virus onto a cellulose acetate filter in the presence of a trivalent cation at low pH and then flushing the filter with a limited quantity of a glycine buffer solution to desorb any marker virus present on the filter. Photo-optical detection of indirect passive immune agglutination by polystyrene beads indicates the performance of the water reclamation system in removing the marker virus. A closed system provides for concentrating any marker virus, initiating and monitoring the passive immune agglutination reaction, and then flushing the system to prepare for another sample.

  7. Vaccination against canine distemper virus infection in infant ferrets with and without maternal antibody protection, using recombinant attenuated poxvirus vaccines.

    PubMed

    Welter, J; Taylor, J; Tartaglia, J; Paoletti, E; Stephensen, C B

    2000-07-01

    Canine distemper virus (CDV) infection of ferrets is clinically and immunologically similar to measles, making this a useful model for the human disease. The model was used to determine if parenteral or mucosal immunization of infant ferrets at 3 and 6 weeks of age with attenuated vaccinia virus (NYVAC) or canarypox virus (ALVAC) vaccine strains expressing the CDV hemagglutinin (H) and fusion (F) protein genes (NYVAC-HF and ALVAC-HF) would induce serum neutralizing antibody and protect against challenge infection at 12 weeks of age. Ferrets without maternal antibody that were vaccinated parenterally with NYVAC-HF (n = 5) or ALVAC-HF (n = 4) developed significant neutralizing titers (log(10) inverse mean titer +/- standard deviation of 2.30 +/- 0.12 and 2.20 +/- 0.34, respectively) by the day of challenge, and all survived with no clinical or virologic evidence of infection. Ferrets without maternal antibody that were vaccinated intranasally (i.n.) developed lower neutralizing titers, with NYVAC-HF producing higher titers at challenge (1.11 +/- 0.57 versus 0.40 +/- 0.37, P = 0.02) and a better survival rate (6/7 versus 0/5, P = 0.008) than ALVAC-HF. Ferrets with maternal antibody that were vaccinated parenterally with NYVAC-HF (n = 7) and ALVAC-HF (n = 7) developed significantly higher antibody titers (1.64 +/- 0. 54 and 1.28 +/- 0.40, respectively) than did ferrets immunized with an attenuated CDV vaccine (0.46 +/- 0.59; n = 7) or the recombinant vectors expressing rabies glycoprotein (RG) (0.19 +/- 0.32; n = 8, P = 7 x 10(-6)). The NYVAC vaccine also protected against weight loss, and both the NYVAC and attenuated CDV vaccines protected against the development of some clinical signs of infection, although survival in each of the three vaccine groups was low (one of seven) and not significantly different from the RG controls (none of eight). Combined i.n.-parenteral immunization of ferrets with maternal antibody using NYVAC-HF (n = 9) produced higher titers (1

  8. Vaccination against Canine Distemper Virus Infection in Infant Ferrets with and without Maternal Antibody Protection, Using Recombinant Attenuated Poxvirus Vaccines

    PubMed Central

    Welter, Janet; Taylor, Jill; Tartaglia, James; Paoletti, Enzo; Stephensen, Charles B.

    2000-01-01

    Canine distemper virus (CDV) infection of ferrets is clinically and immunologically similar to measles, making this a useful model for the human disease. The model was used to determine if parenteral or mucosal immunization of infant ferrets at 3 and 6 weeks of age with attenuated vaccinia virus (NYVAC) or canarypox virus (ALVAC) vaccine strains expressing the CDV hemagglutinin (H) and fusion (F) protein genes (NYVAC-HF and ALVAC-HF) would induce serum neutralizing antibody and protect against challenge infection at 12 weeks of age. Ferrets without maternal antibody that were vaccinated parenterally with NYVAC-HF (n = 5) or ALVAC-HF (n = 4) developed significant neutralizing titers (log10 inverse mean titer ± standard deviation of 2.30 ± 0.12 and 2.20 ± 0.34, respectively) by the day of challenge, and all survived with no clinical or virologic evidence of infection. Ferrets without maternal antibody that were vaccinated intranasally (i.n.) developed lower neutralizing titers, with NYVAC-HF producing higher titers at challenge (1.11 ± 0.57 versus 0.40 ± 0.37, P = 0.02) and a better survival rate (6/7 versus 0/5, P = 0.008) than ALVAC-HF. Ferrets with maternal antibody that were vaccinated parenterally with NYVAC-HF (n = 7) and ALVAC-HF (n = 7) developed significantly higher antibody titers (1.64 ± 0.54 and 1.28 ± 0.40, respectively) than did ferrets immunized with an attenuated CDV vaccine (0.46 ± 0.59; n = 7) or the recombinant vectors expressing rabies glycoprotein (RG) (0.19 ± 0.32; n = 8, P = 7 × 10−6). The NYVAC vaccine also protected against weight loss, and both the NYVAC and attenuated CDV vaccines protected against the development of some clinical signs of infection, although survival in each of the three vaccine groups was low (one of seven) and not significantly different from the RG controls (none of eight). Combined i.n.-parenteral immunization of ferrets with maternal antibody using NYVAC-HF (n = 9) produced higher titers (1.63 ± 0

  9. Development of a challenge-protective vaccine concept by modification of the viral RNA-dependent RNA polymerase of canine distemper virus.

    PubMed

    Silin, D; Lyubomska, O; Ludlow, M; Duprex, W P; Rima, B K

    2007-12-01

    We demonstrate that insertion of the open reading frame of enhanced green fluorescent protein (EGFP) into the coding sequence for the second hinge region of the viral L (large) protein (RNA-dependent RNA polymerase) attenuates a wild-type canine distemper virus. Moreover, we show that single intranasal immunization with this recombinant virus provides significant protection against challenge with the virulent parental virus. Protection against wild-type challenge was gained either after recovery of cellular immunity postimmunization or after development of neutralizing antibodies. Insertion of EGFP seems to result in overattenuation of the virus, while our previous experiments demonstrated that the insertion of an epitope tag into a similar position did not affect L protein function. Thus, a desirable level of attenuation could be reached by manipulating the length of the insert (in the second hinge region of the L protein), providing additional tools for optimization of controlled attenuation. This strategy for controlled attenuation may be useful for a "quick response" in vaccine development against well-known and "new" viral infections and could be combined efficiently with other strategies of vaccine development and delivery systems. PMID:17898047

  10. Computer Viruses: Pathology and Detection.

    ERIC Educational Resources Information Center

    Maxwell, John R.; Lamon, William E.

    1992-01-01

    Explains how computer viruses were originally created, how a computer can become infected by a virus, how viruses operate, symptoms that indicate a computer is infected, how to detect and remove viruses, and how to prevent a reinfection. A sidebar lists eight antivirus resources. (four references) (LRW)

  11. Detection of dengue virus.

    PubMed

    Tripathi, Nagesh K; Shrivastava, Ambuj; Dash, Paban K; Jana, Asha M

    2011-01-01

    Global incidence of dengue has increased considerably over the past decade. Dengue fever (DF) is a self-limiting disease; however, dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS) are fatal. Since there is no therapy and vaccine against dengue, timely diagnosis is therefore necessary for patient management. Laboratory diagnosis is carried out by virus isolation, demonstration of viral antigen, presence of viral nucleic acid, and antibodies. Further, recombinant dengue envelope protein can be used to detect specific antibodies, both IgG and IgM against all four serotypes of virus using an E. coli vector. The purified protein can then be used for detection of dengue specific IgG or IgM antibodies in patient serum with higher sensitivity and specificity, than that of traditional assays. Molecular detection can be accomplished by a one-step, single-tube, rapid, multiplex, RT-PCR for serotype determination. Despite many advantages of the modern techniques, isolation of virus is still considered as "gold-standard" in dengue diagnosis.

  12. Vaccination of puppies born to immune dams with a canine adenovirus-based vaccine protects against a canine distemper virus challenge.

    PubMed

    Fischer, Laurent; Tronel, Jean Phillipe; Pardo-David, Camilla; Tanner, Patrick; Colombet, Guy; Minke, Jules; Audonnet, Jean-Christophe

    2002-10-01

    None of the currently available distemper vaccines provides a satisfactory solution for the immunization of very young carnivores in the face of maternal-derived immunity. Since mucosal immunization with replication-competent adenovirus-based vaccines has been proven effective in the face of passive immunity against the vector, it has the potential to provide a solution for the vaccination of young puppies born to canine distemper virus (CDV)-immune dams. We report the engineering and the characterization of two replication-competent canine adenovirus type 2 (CAV2)-based vaccines expressing, respectively, the CDV hemagglutinin (HA) and fusion (F) antigens. We first demonstrated that the intranasal vaccination with a mixture of both recombinant CAV2s provides an excellent level of protection in seronegative puppies, confirming the value of replication-competent adenovirus-based vectors for mucosal vaccination. In contrast, intranasal immunization with the same vaccine of puppies born to CDV- and CAV2-immune dams, failed to activate specific and protective immune responses. We hypothesized that an active CAV2 infection occurred while puppies were in close contact with the vaccinated dams in the breeding units and that the resulting active mucosal immunity interfered with the intranasal administration of CAV2-based CDV vaccine. However, when puppies born to CDV- and CAV2-immune dams were vaccinated subcutaneously with the CAV2-based CDV vaccine, significant seroconversion and solid protective immunity were triggered despite pre-existing systemic immunity to the vector. This latter result is surprising and suggests that subcutaneous vaccination with a replication-competent recombinant CAV2 may be an efficient strategy to overcome both passive and active adenovirus specific immunity in the dog. From a practical point of view, this could pave the way for an original strategy to vaccinate young puppies in the face of maternal-derived immunity. PMID:12297394

  13. Use of SLAM and PVRL4 and identification of pro-HB-EGF as cell entry receptors for wild type phocine distemper virus.

    PubMed

    Melia, Mary M; Earle, John Philip; Abdullah, Haniah; Reaney, Katherine; Tangy, Frederic; Cosby, Sara Louise

    2014-01-01

    Signalling lymphocyte activation molecule (SLAM) has been identified as an immune cell receptor for the morbilliviruses, measles (MV), canine distemper (CDV), rinderpest and peste des petits ruminants (PPRV) viruses, while CD46 is a receptor for vaccine strains of MV. More recently poliovirus like receptor 4 (PVRL4), also known as nectin 4, has been identified as a receptor for MV, CDV and PPRV on the basolateral surface of polarised epithelial cells. PVRL4 is also up-regulated by MV in human brain endothelial cells. Utilisation of PVRL4 as a receptor by phocine distemper virus (PDV) remains to be demonstrated as well as confirmation of use of SLAM. We have observed that unlike wild type (wt) MV or wtCDV, wtPDV strains replicate in African green monkey kidney Vero cells without prior adaptation, suggesting the use of a further receptor. We therefore examined candidate molecules, glycosaminoglycans (GAG) and the tetraspan proteins, integrin β and the membrane bound form of heparin binding epithelial growth factor (proHB-EGF),for receptor usage by wtPDV in Vero cells. We show that wtPDV replicates in Chinese hamster ovary (CHO) cells expressing SLAM and PVRL4. Similar wtPDV titres are produced in Vero and VeroSLAM cells but more limited fusion occurs in the latter. Infection of Vero cells was not inhibited by anti-CD46 antibody. Removal/disruption of GAG decreased fusion but not the titre of virus. Treatment with anti-integrin β antibody increased rather than decreased infection of Vero cells by wtPDV. However, infection was inhibited by antibody to HB-EGF and the virus replicated in CHO-proHB-EGF cells, indicating use of this molecule as a receptor. Common use of SLAM and PVRL4 by morbilliviruses increases the possibility of cross-species infection. Lack of a requirement for wtPDV adaptation to Vero cells raises the possibility of usage of proHB-EGF as a receptor in vivo but requires further investigation. PMID:25171206

  14. Use of SLAM and PVRL4 and identification of pro-HB-EGF as cell entry receptors for wild type phocine distemper virus.

    PubMed

    Melia, Mary M; Earle, John Philip; Abdullah, Haniah; Reaney, Katherine; Tangy, Frederic; Cosby, Sara Louise

    2014-01-01

    Signalling lymphocyte activation molecule (SLAM) has been identified as an immune cell receptor for the morbilliviruses, measles (MV), canine distemper (CDV), rinderpest and peste des petits ruminants (PPRV) viruses, while CD46 is a receptor for vaccine strains of MV. More recently poliovirus like receptor 4 (PVRL4), also known as nectin 4, has been identified as a receptor for MV, CDV and PPRV on the basolateral surface of polarised epithelial cells. PVRL4 is also up-regulated by MV in human brain endothelial cells. Utilisation of PVRL4 as a receptor by phocine distemper virus (PDV) remains to be demonstrated as well as confirmation of use of SLAM. We have observed that unlike wild type (wt) MV or wtCDV, wtPDV strains replicate in African green monkey kidney Vero cells without prior adaptation, suggesting the use of a further receptor. We therefore examined candidate molecules, glycosaminoglycans (GAG) and the tetraspan proteins, integrin β and the membrane bound form of heparin binding epithelial growth factor (proHB-EGF),for receptor usage by wtPDV in Vero cells. We show that wtPDV replicates in Chinese hamster ovary (CHO) cells expressing SLAM and PVRL4. Similar wtPDV titres are produced in Vero and VeroSLAM cells but more limited fusion occurs in the latter. Infection of Vero cells was not inhibited by anti-CD46 antibody. Removal/disruption of GAG decreased fusion but not the titre of virus. Treatment with anti-integrin β antibody increased rather than decreased infection of Vero cells by wtPDV. However, infection was inhibited by antibody to HB-EGF and the virus replicated in CHO-proHB-EGF cells, indicating use of this molecule as a receptor. Common use of SLAM and PVRL4 by morbilliviruses increases the possibility of cross-species infection. Lack of a requirement for wtPDV adaptation to Vero cells raises the possibility of usage of proHB-EGF as a receptor in vivo but requires further investigation.

  15. Display of neutralizing epitopes of Canine parvovirus and a T-cell epitope of the fusion protein of Canine distemper virus on chimeric tymovirus-like particles and its use as a vaccine candidate both against Canine parvo and Canine distemper.

    PubMed

    Chandran, Dev; Shahana, Pallichera Vijayan; Rani, Gudavelli Sudha; Sugumar, Parthasarthy; Shankar, Chinchkar Ramchandra; Srinivasan, Villuppanoor Alwar

    2009-12-10

    Expression of Physalis mottle tymovirus coat protein in Escherichia coli was earlier shown to self-assemble into empty capsids that were nearly identical to the capsids formed in vivo. Amino acid substitutions were made at the N-terminus of wild-type Physalis mottle virus coat protein with neutralizing epitopes of Canine parvovirus containing the antigenic sites 1-2, 4 and 6-7 and T-cell epitope of the fusion protein of Canine distemper virus in various combinations to yield PhMV1, PhMV2, PhMV3, PhMV4 and PhMV5. These constructs were cloned and expressed in E. coli. The chimeric proteins self-assembled into chimeric tymovirus-like particles (TVLPs) as determined by electron microscopy. The TVLPs were purified by ultracentrifugation and injected into guinea pigs and dogs to determine their immunogenicity. Initial immunogenicity studies in guinea pigs indicated that PhMV3 gave a higher response in comparison to the other TVLPs for both CPV and CDV and hence all further experiments in dogs were done with PhMV3. HI was done against different isolates obtained from various parts of the country. Protective titres indicated the broad spectrum of the vaccine. In conclusion the study indicated that the above chimeric VLP based vaccine could be used in dogs to generate a protective immune response against diseases caused by both Canine parvo and Canine distemper virus. PMID:19818723

  16. Canine distemper virus infection leads to an inhibitory phenotype of monocyte-derived dendritic cells in vitro with reduced expression of co-stimulatory molecules and increased interleukin-10 transcription.

    PubMed

    Qeska, Visar; Barthel, Yvonne; Herder, Vanessa; Stein, Veronika M; Tipold, Andrea; Urhausen, Carola; Günzel-Apel, Anne-Rose; Rohn, Karl; Baumgärtner, Wolfgang; Beineke, Andreas

    2014-01-01

    Canine distemper virus (CDV) exhibits a profound lymphotropism that causes immunosuppression and increased susceptibility of affected dogs to opportunistic infections. Similar to human measles virus, CDV is supposed to inhibit terminal differentiation of dendritic cells (DCs), responsible for disturbed repopulation of lymphoid tissues and diminished antigen presenting function in dogs. In order to testify the hypothesis that CDV-infection leads to an impairment of professional antigen presenting cells, canine DCs have been generated from peripheral blood monocytes in vitro and infected with CDV. Virus infection was confirmed and quantified by transmission electron microscopy, CDV-specific immunofluorescence, and virus titration. Flow cytometric analyses revealed a significant down-regulation of the major histocompatibility complex class II and co-stimulatory molecules CD80 and CD86 in CDV-infected DCs, indicative of disturbed antigen presenting capacity. Molecular analyses revealed an increased expression of the immune inhibitory cytokine interleukin-10 in DCs following infection. Results of the present study demonstrate that CDV causes phenotypical changes and altered cytokine expression of DCs, which represent potential mechanisms to evade host immune responses and might contribute to immune dysfunction and virus persistence in canine distemper.

  17. Canine Distemper Virus Infection Leads to an Inhibitory Phenotype of Monocyte-Derived Dendritic Cells In Vitro with Reduced Expression of Co-Stimulatory Molecules and Increased Interleukin-10 Transcription

    PubMed Central

    Herder, Vanessa; Stein, Veronika M.; Tipold, Andrea; Urhausen, Carola; Günzel-Apel, Anne-Rose; Rohn, Karl; Baumgärtner, Wolfgang; Beineke, Andreas

    2014-01-01

    Canine distemper virus (CDV) exhibits a profound lymphotropism that causes immunosuppression and increased susceptibility of affected dogs to opportunistic infections. Similar to human measles virus, CDV is supposed to inhibit terminal differentiation of dendritic cells (DCs), responsible for disturbed repopulation of lymphoid tissues and diminished antigen presenting function in dogs. In order to testify the hypothesis that CDV-infection leads to an impairment of professional antigen presenting cells, canine DCs have been generated from peripheral blood monocytes in vitro and infected with CDV. Virus infection was confirmed and quantified by transmission electron microscopy, CDV-specific immunofluorescence, and virus titration. Flow cytometric analyses revealed a significant down-regulation of the major histocompatibility complex class II and co-stimulatory molecules CD80 and CD86 in CDV-infected DCs, indicative of disturbed antigen presenting capacity. Molecular analyses revealed an increased expression of the immune inhibitory cytokine interleukin-10 in DCs following infection. Results of the present study demonstrate that CDV causes phenotypical changes and altered cytokine expression of DCs, which represent potential mechanisms to evade host immune responses and might contribute to immune dysfunction and virus persistence in canine distemper. PMID:24769532

  18. Possible lin between elevated accumulation of trace elements and canine distemper virus infection in the Caspian seals (Phoca caspica) stranded in 2000 and 2001

    NASA Astrophysics Data System (ADS)

    Shinsuke, T.; Takashi, K.; Yasumi, A.; Tokutaka, I.; Reiji, K.

    2003-05-01

    In the Caspian Sea, a die-off of thousands of Caspian seals (Phoca caspica) occurred in 1997 and 2000. While a direct cause for these deaths seems to be canine distemper virus (CDV) infection, immunosuppression due to environmental pollutants is considered as one of the possible explanations for the development of the disease. The purpose of this work is to examine whether exposure to trace metals could be one of the factors involved in the mass mortality of Caspian seals. Concentrations of 13 trace elements weredetermined in liver, kidney and muscle of Caspian seals found stranded along the coasts of the Caspian Sea in 2000 and 2001. Concentrations of toxic elemen ts (Ag, Cd, Hg, Tl and Pb) in the Caspian seals collected in 2000 and 2001 were comparable to or lower than those in healthy Caspian seals collected in 1993 and 1998 and in seals from other regions, suggesting that these elements would not be the causative agent for the death of Caspian seals. In contrast, Zn and Fe concentrations in the stranded Caspian seals were apparently higher than those in seals from other locations. These results suggest the disturbance in homeostatic control and nutritional statu s of essential elements in the stranded Caspian seals.

  19. The Fusion Protein Signal-Peptide-Coding Region of Canine Distemper Virus: A Useful Tool for Phylogenetic Reconstruction and Lineage Identification

    PubMed Central

    Sarute, Nicolás; Calderón, Marina Gallo; Pérez, Ruben; La Torre, José; Hernández, Martín; Francia, Lourdes; Panzera, Yanina

    2013-01-01

    Canine distemper virus (CDV; Paramyxoviridae, Morbillivirus) is the etiologic agent of a multisystemic infectious disease affecting all terrestrial carnivore families with high incidence and mortality in domestic dogs. Sequence analysis of the hemagglutinin (H) gene has been widely employed to characterize field strains, permitting the identification of nine CDV lineages worldwide. Recently, it has been established that the sequences of the fusion protein signal-peptide (Fsp) coding region are extremely variable, suggesting that analysis of its sequence might be useful for strain characterization studies. However, the divergence of Fsp sequences among worldwide strains and its phylogenetic resolution has not yet been evaluated. We constructed datasets containing the Fsp-coding region and H gene sequences of the same strains belonging to eight CDV lineages. Both datasets were used to evaluate their phylogenetic resolution. The phylogenetic analysis revealed that both datasets clustered the same strains into eight different branches, corresponding to CDV lineages. The inter-lineage amino acid divergence was fourfold greater for the Fsp peptide than for the H protein. The likelihood mapping revealed that both datasets display strong phylogenetic signals in the region of well-resolved topologies. These features indicate that Fsp-coding region sequence analysis is suitable for evolutionary studies as it allows for straightforward identification of CDV lineages. PMID:23675493

  20. Effects of body weight on antibody titers against canine parvovirus type 2, canine distemper virus, and canine adenovirus type 1 in vaccinated domestic adult dogs.

    PubMed

    Taguchi, Masayuki; Namikawa, Kazuhiko; Maruo, Takuya; Saito, Miyoko; Lynch, Jonathan; Sahara, Hiroeki

    2012-10-01

    The objective of this study was to determine whether post-vaccination antibody titers vary according to body weight in adult dogs. Antibody titers against canine parvovirus type 2 (CPV-2), canine distemper virus (CDV), and canine adenovirus type 1 (CAdV-1) were measured for 978 domestic adult dogs from 2 to 6 y of age. The dogs had been vaccinated approximately 12 mo earlier with a commercial combination vaccine. The dogs were divided into groups according to their weight. It was found that mean antibody titers in all weight groups were sufficient to prevent infection. Intergroup comparison, however, revealed that CPV-2 antibody titers were significantly higher in the Super Light (< 5 kg) group than in the Medium (10 to 19.9 kg) and Heavy (> 20 kg) groups and were also significantly higher in the Light (5 to 9.9 kg) group than in the Heavy group. Antibody titers against CDV were significantly higher in the Super Light, Light, and Medium groups than in the Heavy group. There were no significant differences among the groups for the CAdV-1 antibody titers.

  1. Induction of castration by immunization of male dogs with recombinant gonadotropin-releasing hormone (GnRH)-canine distemper virus (CDV) T helper cell epitope p35.

    PubMed

    Jung, Mi-Jeong; Moon, Young-Chan; Cho, Ik-Hyun; Yeh, Jung-Yong; Kim, Sun-Eui; Chang, Wha-Seok; Park, Seung-Young; Song, Chang-Seon; Kim, Hwi-Yool; Park, Keun-Kyu; McOrist, Steven; Choi, In-Soo; Lee, Joong-Bok

    2005-03-01

    Immunocastration is a considerable alternative to a surgical castration method especially in male animal species for alleviating unwanted male behaviors and characteristics. Induction of high titer of antibody specific for gonadotropin-releasing hormone (GnRH) correlates with the regression of testes. Fusion proteins composed of canine GnRH and T helper (Th) cell epitope p35 originated from canine distemper virus (CDV) F protein and goat rotavirus VP6 protein were produced in E. coli. When these fusion proteins were injected to male dogs which were previously immunized with CDV vaccine, the fusion protein of GnRH-CDV Th cell epitope p35 induced much higher antibody than that of GnRH-rotavirus VP6 protein or GnRH alone. The degeneration of spermatogenesis was also verified in the male dogs immunized with the fusion protein of GnRH-CDV Th cell epitope p35. These results indicate that canine GnRH conjugated to CDV Th cell epitope p35 acted as a strong immunogen and the antibody to GnRH specifically neutralized GnRH in the testes. This study also implies a potential application of GnRH-based vaccines for immunocastration of male pets. PMID:15785119

  2. Canine Distemper Virus Infects Canine Keratinocytes and Immune Cells by Using Overlapping and Distinct Regions Located on One Side of the Attachment Protein▿

    PubMed Central

    Langedijk, Johannes P. M.; Janda, Jozef; Origgi, Francesco C.; Örvell, Claes; Vandevelde, Marc; Zurbriggen, Andreas; Plattet, Philippe

    2011-01-01

    The morbilliviruses measles virus (MeV) and canine distemper virus (CDV) both rely on two surface glycoproteins, the attachment (H) and fusion proteins, to promote fusion activity for viral cell entry. Growing evidence suggests that morbilliviruses infect multiple cell types by binding to distinct host cell surface receptors. Currently, the only known in vivo receptor used by morbilliviruses is CD150/SLAM, a molecule expressed in certain immune cells. Here we investigated the usage of multiple receptors by the highly virulent and demyelinating CDV strain A75/17. We based our study on the assumption that CDV-H may interact with receptors similar to those for MeV, and we conducted systematic alanine-scanning mutagenesis on CDV-H throughout one side of the β-propeller documented in MeV-H to contain multiple receptor-binding sites. Functional and biochemical assays performed with SLAM-expressing cells and primary canine epithelial keratinocytes identified 11 residues mutation of which selectively abrogated fusion in keratinocytes. Among these, four were identical to amino acids identified in MeV-H as residues contacting a putative receptor expressed in polarized epithelial cells. Strikingly, when mapped on a CDV-H structural model, all residues clustered in or around a recessed groove located on one side of CDV-H. In contrast, reported CDV-H mutants with SLAM-dependent fusion deficiencies were characterized by additional impairments to the promotion of fusion in keratinocytes. Furthermore, upon transfer of residues that selectively impaired fusion induction in keratinocytes into the CDV-H of the vaccine strain, fusion remained largely unaltered. Taken together, our results suggest that a restricted region on one side of CDV-H contains distinct and overlapping sites that control functional interaction with multiple receptors. PMID:21849439

  3. Antibody response to canine distemper vaccine in African wild dogs.

    PubMed

    Spencer, J; Burroughs, R

    1992-07-01

    Antibody levels against canine distemper virus were measured by means of an immunofluorescent antibody test prior to, and after, administration of a modified-live virus booster vaccine to seven African wild dogs (Lycaon pictus). Positive seroconversion with no harmful side-effects was seen in all the animals.

  4. Prevalence of positive antibody test results for canine parvovirus (CPV) and canine distemper virus (CDV) and response to modified live vaccination against CPV and CDV in dogs entering animal shelters.

    PubMed

    Litster, Annette; Nichols, Jamieson; Volpe, Allison

    2012-05-25

    Canine parvovirus (CPV) and canine distemper virus (CDV) infections are relatively common in animal shelters and are important population management issues since the immune status of incoming dogs is usually unknown. This study aimed to determine the prevalence of positive antibody test results for CPV and CDV in incoming dogs aged ≥ 4 months and to measure antibody response over 2 weeks following vaccination with a modified live vaccine (MLV). Dogs aged 4-24 months entering an adoption-guarantee shelter (Shelter 1, n=51) and aged ≥ 4 months entering a limited admission shelter (Shelter 2; n=51) were enrolled. Dogs from Shelter 1 had been vaccinated with MLV at a municipal shelter 5 days before enrollment, whereas dogs from Shelter 2 had no known history of vaccination at enrollment. Sera were obtained on day 1, immediately prior to CPV/CDV MLV, and tested using an in-clinic ELISA kit to detect CPV/CDV antibodies. Dogs negative for CPV and/or CDV were retested at day 6-8 and those dogs still negative at day 6-8 were retested at day 13-15. Prior to CPV/CDV MLV on day 1, more dogs tested positive for CPV (Shelter 1 - 68.6%; Shelter 2 - 84.3%) than for CDV (Shelter 1 - 37.3%; Shelter 2 - 41.2%). On day 1, prior to MLV, all spayed/neutered animals tested CPV antibody-positive (n=17/102) and CPV antibody-positive dogs were older than serologically negative dogs (Shelter 1, P=0.0029; Shelter 2, P=0.0042). By day 13-15, almost all dogs were CPV antibody-positive (Shelter 1 - 97.9%; Shelter 2 - 100.0%) and CDV antibody-positive (Shelter 1 - 93.8%; Shelter 2 - 97.8%). MLV induces protective antibody titers against CPV/CDV in almost all dogs after 13-15 days.

  5. Canine distemper spillover in domestic dogs from urban wildlife.

    PubMed

    Kapil, Sanjay; Yeary, Teresa J

    2011-11-01

    Canine distemper virus (CDV) causes a major disease of domestic dogs that develops as a serious systemic infection in unvaccinated or improperly vaccinated dogs. Domesticated dogs are the main reservoir of CDV, a multihost pathogen. This virus of the genus Morbillivirus in the family Paramyxoviridae occurs in other carnivorous species including all members of the Canidae and Mustelidae families and in some members of the Procyonidae, Hyaenidae, Ursidae, and Viverridae families. Canine distemper also has been reported in the Felidae family and marine mammals. The spread and incidences of CDV epidemics in dogs and wildlife here and worldwide are increasing. PMID:22041204

  6. Use of quantitative real-time RT-PCR to investigate the correlation between viremia and viral shedding of canine distemper virus, and infection outcomes in experimentally infected dogs

    PubMed Central

    SEHATA, Go; SATO, Hiroaki; ITO, Toshihiro; IMAIZUMI, Yoshitaka; NORO, Taichi; OISHI, Eiji

    2015-01-01

    We used real-time RT-PCR and virus titration to examine canine distemper virus (CDV) kinetics in peripheral blood and rectal and nasal secretions from 12 experimentally infected dogs. Real-time RT-PCR proved extremely sensitive, and the correlation between the two methods for rectal and nasal (r=0.78, 0.80) samples on the peak day of viral RNA was good. Although the dogs showed diverse symptoms, viral RNA kinetics were similar; the peak of viral RNA in the symptomatic dogs was consistent with the onset of symptoms. These results indicate that real-time RT-PCR is sufficiently sensitive to monitor CDV replication in experimentally infected dogs regardless of the degree of clinical manifestation and suggest that the peak of viral RNA reflects active CDV replication. PMID:25728411

  7. Use of quantitative real-time RT-PCR to investigate the correlation between viremia and viral shedding of canine distemper virus, and infection outcomes in experimentally infected dogs.

    PubMed

    Sehata, Go; Sato, Hiroaki; Ito, Toshihiro; Imaizumi, Yoshitaka; Noro, Taichi; Oishi, Eiji

    2015-07-01

    We used real-time RT-PCR and virus titration to examine canine distemper virus (CDV) kinetics in peripheral blood and rectal and nasal secretions from 12 experimentally infected dogs. Real-time RT-PCR proved extremely sensitive, and the correlation between the two methods for rectal and nasal (r=0.78, 0.80) samples on the peak day of viral RNA was good. Although the dogs showed diverse symptoms, viral RNA kinetics were similar; the peak of viral RNA in the symptomatic dogs was consistent with the onset of symptoms. These results indicate that real-time RT-PCR is sufficiently sensitive to monitor CDV replication in experimentally infected dogs regardless of the degree of clinical manifestation and suggest that the peak of viral RNA reflects active CDV replication.

  8. Canine distemper epizootic in Everglades mink.

    PubMed

    Cunningham, M W; Shindle, D B; Allison, A B; Terrell, S P; Mead, D G; Owen, M

    2009-10-01

    Four free-ranging mink, Neovison vison, collected between June and September 2004 in the Fakahatchee Strand Preserve State Park (FSPSP, Florida, USA), were examined for canine distemper virus (CDV) infection. Microscopic lesions and viral inclusions consistent with CDV infection were observed in three mink. Virus isolation and reverse transcription-polymerase chain reaction performed on all mink were positive for CDV. Anecdotal records of mink observations in FSPSP suggest a postepizootic decline in the mink population followed by an apparent recovery. We recommend further research to assess the status of the Everglades mink and the impact of CDV on this and other American mink populations in Florida. PMID:19901388

  9. 9 CFR 113.47 - Detection of extraneous viruses by the fluorescent antibody technique.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... tested for: (i) Bluetongue virus; (ii) Bovine adenoviruses; (iii) Bovine parvovirus; and (iv) Bovine...) Canine distemper virus; and (iii) Canine parvovirus. (4) Equine cells shall, in addition, be tested for...) Porcine cells shall, in addition, be tested for: (i) Porcine adenovirus; (ii) Porcine parvovirus;...

  10. 9 CFR 113.47 - Detection of extraneous viruses by the fluorescent antibody technique.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... tested for: (i) Bluetongue virus; (ii) Bovine adenoviruses; (iii) Bovine parvovirus; and (iv) Bovine...) Canine distemper virus; and (iii) Canine parvovirus. (4) Equine cells shall, in addition, be tested for...) Porcine cells shall, in addition, be tested for: (i) Porcine adenovirus; (ii) Porcine parvovirus;...

  11. Phylogenetic analysis of the haemagglutinin gene of current wild-type canine distemper viruses from South Africa: lineage Africa.

    PubMed

    Woma, Timothy Y; van Vuuren, Moritz; Bosman, Ana-Mari; Quan, Melvyn; Oosthuizen, Marinda

    2010-07-14

    There are no reports of CDV isolations in southern Africa, and although CDV is said to have geographically distinct lineages, molecular information of African strains has not yet been documented. Viruses isolated in cell cultures were subjected to reverse transcription-polymerase chain reaction (RT-PCR), and the complete H gene was sequenced and phylogenetically analysed with other strains from GenBank. Phylogenetic comparisons of the complete H gene of CDV isolates from different parts of the world (available in GenBank) with wild-type South African isolates revealed nine clades. All South African isolates form a separate African clade of their own and thus are clearly separated from the American, European, Asian, Arctic and vaccine virus clades. It is likely that only the 'African lineage' of CDV may be circulating in South Africa currently, and the viruses isolated from dogs vaccinated against CDV are not the result of reversion to virulence of vaccine strains, but infection with wild-type strains.

  12. Jaundice and bilirubinemia as manifestations of canine distemper in raccoons and ferrets

    USGS Publications Warehouse

    Kilham, L.; Habermann, R.T.; Herman, C.M.

    1956-01-01

    1) Two strains of distemper virus have been isolated from wild raccoons and one strain from ferrets. 2) All strains isolated have induced bilirubinemia in raccoons and ferrets. Many raccoons with bilirubinemia also had jaundice. 3) Identification of these strains as members of the canine distemper virus complex has been by clinical and pathological findings consistent with this diagnosis as well as by cross-immunity tests.

  13. Control of canine distemper.

    PubMed

    Chappuis, G

    1995-05-01

    Control of canine distemper can realistically only be achieved by the use of vaccination. The types of vaccine in current use are described, together with some of the problems encountered such as interference by maternal antibodies, and usage in species other than dogs. Modified live viral vaccines, as used for more than thirty years, have proved very effective. Nevertheless there is scope for some improvement in vaccine efficacy and recent developments in genetic recombinant methods are described. PMID:8588329

  14. Distemper outbreak and its effect on African wild dog conservation.

    PubMed

    van de Bildt, Marco W G; Kuiken, Thijs; Visee, Aart M; Lema, Sangito; Fitzjohn, Tony R; Osterhaus, Albert D M E

    2002-02-01

    In December 2000, an infectious disease spread through a captive breeding group of African wild dogs (Lycaon pictus) in Tanzania, killing 49 of 52 animals within 2 months. The causative agent was identified as Canine distemper virus (CDV) by means of histologic examination, virus isolation, reverse transcriptase-polymerase chain reaction analysis, and nucleotide sequencing. This report emphasizes the importance of adequate protection against infectious diseases for the successful outcome of captive breeding programs of endangered species.

  15. Clinical trials with canine distemper vaccines in exotic carnivores.

    PubMed

    Montali, R J; Bartz, C R; Teare, J A; Allen, J T; Appel, M J; Bush, M

    1983-12-01

    Two types of killed canine distemper virus (CDV) vaccine and a modified-live CDV vaccine were clinically evaluated in four species of exotic carnivores. In 16 trials in which 13 red pandas (Ailurus fulgens) were given the killed vaccine, only 1 animal had a virus-neutralization titer that exceeded 1:100. A red panda given modified-live CDV vaccine deemed safe for gray foxes and ferrets died of bacterial pneumonia 16 days later. There was no pathologic evidence of canine distemper in that panda. The same modified-live vaccine proved to be immunogenic and safe in 12 bush dogs (Speothos venaticus), 5 maned wolves (Chrysocyon brachyurus), and 3 fennec foxes (Fennecus zerda) in which virus-neutralization titers often exceeded 1:512 and persisted for several months after vaccination.

  16. Canine distemper in endangered Ethiopian wolves.

    PubMed

    Gordon, Christopher H; Banyard, Ashley C; Hussein, Alo; Laurenson, M Karen; Malcolm, James R; Marino, Jorgelina; Regassa, Fekede; Stewart, Anne-Marie E; Fooks, Anthony R; Sillero-Zubiri, Claudio

    2015-05-01

    The Ethiopian wolf (Canis simensis) is the world's rarest canid; ≈500 wolves remain. The largest population is found within the Bale Mountains National Park (BMNP) in southeastern Ethiopia, where conservation efforts have demonstrated the negative effect of rabies virus on wolf populations. We describe previously unreported infections with canine distemper virus (CDV) among these wolves during 2005-2006 and 2010. Death rates ranged from 43% to 68% in affected subpopulations and were higher for subadult than adult wolves (83%-87% vs. 34%-39%). The 2010 CDV outbreak started 20 months after a rabies outbreak, before the population had fully recovered, and led to the eradication of several focal packs in BMNP's Web Valley. The combined effect of rabies and CDV increases the chance of pack extinction, exacerbating the typically slow recovery of wolf populations, and represents a key extinction threat to populations of this highly endangered carnivore. PMID:25898177

  17. Canine Distemper in Endangered Ethiopian Wolves

    PubMed Central

    Gordon, Christopher H.; Hussein, Alo; Laurenson, M. Karen; Malcolm, James R.; Marino, Jorgelina; Regassa, Fekede; Stewart, Anne-Marie E.; Fooks, Anthony R.; Sillero-Zubiri, Claudio

    2015-01-01

    The Ethiopian wolf (Canis simensis) is the world’s rarest canid; ≈500 wolves remain. The largest population is found within the Bale Mountains National Park (BMNP) in southeastern Ethiopia, where conservation efforts have demonstrated the negative effect of rabies virus on wolf populations. We describe previously unreported infections with canine distemper virus (CDV) among these wolves during 2005–2006 and 2010. Death rates ranged from 43% to 68% in affected subpopulations and were higher for subadult than adult wolves (83%–87% vs. 34%–39%). The 2010 CDV outbreak started 20 months after a rabies outbreak, before the population had fully recovered, and led to the eradication of several focal packs in BMNP’s Web Valley. The combined effect of rabies and CDV increases the chance of pack extinction, exacerbating the typically slow recovery of wolf populations, and represents a key extinction threat to populations of this highly endangered carnivore. PMID:25898177

  18. Canine distemper in endangered Ethiopian wolves.

    PubMed

    Gordon, Christopher H; Banyard, Ashley C; Hussein, Alo; Laurenson, M Karen; Malcolm, James R; Marino, Jorgelina; Regassa, Fekede; Stewart, Anne-Marie E; Fooks, Anthony R; Sillero-Zubiri, Claudio

    2015-05-01

    The Ethiopian wolf (Canis simensis) is the world's rarest canid; ≈500 wolves remain. The largest population is found within the Bale Mountains National Park (BMNP) in southeastern Ethiopia, where conservation efforts have demonstrated the negative effect of rabies virus on wolf populations. We describe previously unreported infections with canine distemper virus (CDV) among these wolves during 2005-2006 and 2010. Death rates ranged from 43% to 68% in affected subpopulations and were higher for subadult than adult wolves (83%-87% vs. 34%-39%). The 2010 CDV outbreak started 20 months after a rabies outbreak, before the population had fully recovered, and led to the eradication of several focal packs in BMNP's Web Valley. The combined effect of rabies and CDV increases the chance of pack extinction, exacerbating the typically slow recovery of wolf populations, and represents a key extinction threat to populations of this highly endangered carnivore.

  19. Detection of mink enteritis virus by loop-mediated isothermal amplification (LAMP).

    PubMed

    Wang, Jianke; Cheng, Shipeng; Yi, Li; Cheng, Yuening; Yang, Shen; Xu, Hongli; Li, Zhenguang; Shi, Xinchuan; Wu, Hua; Yan, Xijun

    2013-02-01

    Loop-mediated isothermal amplification (LAMP) method was discovered in the last decade but only used for the first time in the diagnosis of mink enteritis virus (MEV) infection in this study. The amplification could be completed within 60 min, under isothermal condition at 65°C, by employing a set of four primers targeting the VP2 gene of MEV. The LAMP was more sensitive than the conventional PCR, with a detection limit of 10(-1) median tissue culture infective doses (TCID(50))/ml per reaction, compared with 10 TCID(50)/ml for PCR analysis. No cross reactivity was observed for other related viruses, including canine distemper virus (CDV) and Aleutian mink disease parvovirus (AMDV). Eighty four of 230 clinical samples were found to be positive for MEV, which is higher than that determined by using the conventional PCR method (68). The results indicate the LAMP can be potentially used to determine MEV as a simple, rapid procedure. This assay would be an available alternative to PCR analysis for the diagnosis of MEV infection in mink, particularly in less well-equipped laboratories and in rural settings where resources are limited. PMID:23183142

  20. Differentiation of canine distemper virus isolates in fur animals from various vaccine strains by reverse transcription-polymerase chain reaction-restriction fragment length polymorphism according to phylogenetic relations in china

    PubMed Central

    2011-01-01

    In order to effectively identify the vaccine and field strains of Canine distemper virus (CDV), a new differential diagnostic test has been developed based on reverse transcription-polymerase chain reaction (RT-PCR) and restriction fragment length polymorphism (RFLP). We selected an 829 bp fragment of the nucleoprotein (N) gene of CDV. By RFLP analysis using BamHI, field isolates were distinguishable from the vaccine strains. Two fragments were obtained from the vaccine strains by RT-PCR-RFLP analysis while three were observed in the field strains. An 829 nucleotide region of the CDV N gene was analyzed in 19 CDV field strains isolated from minks, raccoon dogs and foxes in China between 2005 and 2007. The results suggest this method is precise, accurate and efficient. It was also determined that three different genotypes exist in CDV field strains in fur animal herds of the north of China, most of which belong to Asian type. Mutated field strains, JSY06-R1, JSY06-R2 and JDH07-F1 also exist in Northern China, but are most closely related to the standard virulent strain A75/17, designated in Arctic and America-2 genetype in the present study, respectively. PMID:21352564

  1. Specific detection of peste des petits ruminants virus antibodies in sheep and goat sera by the luciferase immunoprecipitation system.

    PubMed

    Berguido, Francisco J; Bodjo, Sanne Charles; Loitsch, Angelika; Diallo, Adama

    2016-01-01

    Peste des petits ruminants (PPR) is a contagious and often fatal transboundary animal disease affecting mostly sheep, goats and wild small ruminants. This disease is endemic in most of Africa, the Middle, Near East, and large parts of Asia. The causal agent is peste des petits ruminants virus (PPRV), which belongs to the genus Morbillivirus in the family Paramyxoviridae. This genus also includes measles virus (MV), canine distemper virus (CDV) and rinderpest virus (RPV). All are closely related viruses with serological cross reactivity. In this study, we have developed a Luciferase Immunoprecipitation System (LIPS) for the rapid detection of antibodies against PPRV in serum samples and for specific differentiation from antibodies against RPV. PPR and rinderpest (RP) serum samples were assayed by PPR-LIPS and two commercially available PPR cELISA tests. The PPR-LIPS showed high sensitivity and specificity for the samples tested and showed no cross reactivity with RPV unlike the commercial PPR cELISA tests which did cross react with RPV. Based on the results shown in this study, PPR-LIPS is presented as a good candidate for the specific serosurveillance of PPR.

  2. Specific detection of peste des petits ruminants virus antibodies in sheep and goat sera by the luciferase immunoprecipitation system.

    PubMed

    Berguido, Francisco J; Bodjo, Sanne Charles; Loitsch, Angelika; Diallo, Adama

    2016-01-01

    Peste des petits ruminants (PPR) is a contagious and often fatal transboundary animal disease affecting mostly sheep, goats and wild small ruminants. This disease is endemic in most of Africa, the Middle, Near East, and large parts of Asia. The causal agent is peste des petits ruminants virus (PPRV), which belongs to the genus Morbillivirus in the family Paramyxoviridae. This genus also includes measles virus (MV), canine distemper virus (CDV) and rinderpest virus (RPV). All are closely related viruses with serological cross reactivity. In this study, we have developed a Luciferase Immunoprecipitation System (LIPS) for the rapid detection of antibodies against PPRV in serum samples and for specific differentiation from antibodies against RPV. PPR and rinderpest (RP) serum samples were assayed by PPR-LIPS and two commercially available PPR cELISA tests. The PPR-LIPS showed high sensitivity and specificity for the samples tested and showed no cross reactivity with RPV unlike the commercial PPR cELISA tests which did cross react with RPV. Based on the results shown in this study, PPR-LIPS is presented as a good candidate for the specific serosurveillance of PPR. PMID:26506137

  3. DNA Microarray for Detection of Gastrointestinal Viruses

    PubMed Central

    Martínez, Miguel A.; Soto-del Río, María de los Dolores; Gutiérrez, Rosa María; Chiu, Charles Y.; Greninger, Alexander L.; Contreras, Juan Francisco; López, Susana; Arias, Carlos F.

    2014-01-01

    Gastroenteritis is a clinical illness of humans and other animals that is characterized by vomiting and diarrhea and caused by a variety of pathogens, including viruses. An increasing number of viral species have been associated with gastroenteritis or have been found in stool samples as new molecular tools have been developed. In this work, a DNA microarray capable in theory of parallel detection of more than 100 viral species was developed and tested. Initial validation was done with 10 different virus species, and an additional 5 species were validated using clinical samples. Detection limits of 1 × 103 virus particles of Human adenovirus C (HAdV), Human astrovirus (HAstV), and group A Rotavirus (RV-A) were established. Furthermore, when exogenous RNA was added, the limit for RV-A detection decreased by one log. In a small group of clinical samples from children with gastroenteritis (n = 76), the microarray detected at least one viral species in 92% of the samples. Single infection was identified in 63 samples (83%), and coinfection with more than one virus was identified in 7 samples (9%). The most abundant virus species were RV-A (58%), followed by Anellovirus (15.8%), HAstV (6.6%), HAdV (5.3%), Norwalk virus (6.6%), Human enterovirus (HEV) (9.2%), Human parechovirus (1.3%), Sapporo virus (1.3%), and Human bocavirus (1.3%). To further test the specificity and sensitivity of the microarray, the results were verified by reverse transcription-PCR (RT-PCR) detection of 5 gastrointestinal viruses. The RT-PCR assay detected a virus in 59 samples (78%). The microarray showed good performance for detection of RV-A, HAstV, and calicivirus, while the sensitivity for HAdV and HEV was low. Furthermore, some discrepancies in detection of mixed infections were observed and were addressed by reverse transcription-quantitative PCR (RT-qPCR) of the viruses involved. It was observed that differences in the amount of genetic material favored the detection of the most abundant

  4. DNA microarray for detection of gastrointestinal viruses.

    PubMed

    Martínez, Miguel A; Soto-Del Río, María de Los Dolores; Gutiérrez, Rosa María; Chiu, Charles Y; Greninger, Alexander L; Contreras, Juan Francisco; López, Susana; Arias, Carlos F; Isa, Pavel

    2015-01-01

    Gastroenteritis is a clinical illness of humans and other animals that is characterized by vomiting and diarrhea and caused by a variety of pathogens, including viruses. An increasing number of viral species have been associated with gastroenteritis or have been found in stool samples as new molecular tools have been developed. In this work, a DNA microarray capable in theory of parallel detection of more than 100 viral species was developed and tested. Initial validation was done with 10 different virus species, and an additional 5 species were validated using clinical samples. Detection limits of 1 × 10(3) virus particles of Human adenovirus C (HAdV), Human astrovirus (HAstV), and group A Rotavirus (RV-A) were established. Furthermore, when exogenous RNA was added, the limit for RV-A detection decreased by one log. In a small group of clinical samples from children with gastroenteritis (n = 76), the microarray detected at least one viral species in 92% of the samples. Single infection was identified in 63 samples (83%), and coinfection with more than one virus was identified in 7 samples (9%). The most abundant virus species were RV-A (58%), followed by Anellovirus (15.8%), HAstV (6.6%), HAdV (5.3%), Norwalk virus (6.6%), Human enterovirus (HEV) (9.2%), Human parechovirus (1.3%), Sapporo virus (1.3%), and Human bocavirus (1.3%). To further test the specificity and sensitivity of the microarray, the results were verified by reverse transcription-PCR (RT-PCR) detection of 5 gastrointestinal viruses. The RT-PCR assay detected a virus in 59 samples (78%). The microarray showed good performance for detection of RV-A, HAstV, and calicivirus, while the sensitivity for HAdV and HEV was low. Furthermore, some discrepancies in detection of mixed infections were observed and were addressed by reverse transcription-quantitative PCR (RT-qPCR) of the viruses involved. It was observed that differences in the amount of genetic material favored the detection of the most abundant

  5. Detection of respiratory viruses and Bordetella bronchiseptica in dogs with acute respiratory tract infections.

    PubMed

    Schulz, B S; Kurz, S; Weber, K; Balzer, H-J; Hartmann, K

    2014-09-01

    Canine infectious respiratory disease (CIRD) is an acute, highly contagious disease complex caused by a variety of infectious agents. At present, the role of viral and bacterial components as primary or secondary pathogens in CIRD is not fully understood. The aim of this study was to investigate the prevalence of canine parainfluenza virus (CPIV), canine adenovirus type 2 (CAV-2), canine influenza virus (CIV), canine respiratory coronavirus (CRCoV), canine herpes virus-1 (CHV-1), canine distemper virus (CDV) and Bordetella bronchiseptica in dogs with CIRD and to compare the data with findings in healthy dogs. Sixty-one dogs with CIRD and 90 clinically healthy dogs from Southern Germany were prospectively enrolled in this study. Nasal and pharyngeal swabs were collected from all dogs and were analysed for CPIV, CAV-2, CIV, CRCoV, CHV-1, CDV, and B. bronchiseptica by real-time PCR. In dogs with acute respiratory signs, 37.7% tested positive for CPIV, 9.8% for CRCoV and 78.7% for B. bronchiseptica. Co-infections with more than one agent were detected in 47.9% of B. bronchiseptica-positive, 82.6% of CPIV-positive, and 100% of CRCoV-positive dogs. In clinically healthy dogs, 1.1% tested positive for CAV-2, 7.8% for CPIV and 45.6% for B. bronchiseptica. CPIV and B. bronchiseptica were detected significantly more often in dogs with CIRD than in clinically healthy dogs (P < 0.001 for each pathogen) and were the most common infectious agents in dogs with CIRD in Southern Germany. Mixed infections with several pathogens were common. In conclusion, clinically healthy dogs can carry respiratory pathogens and could act as sources of infection for susceptible dogs. PMID:24980809

  6. Detection of respiratory viruses and Bordetella bronchiseptica in dogs with acute respiratory tract infections.

    PubMed

    Schulz, B S; Kurz, S; Weber, K; Balzer, H-J; Hartmann, K

    2014-09-01

    Canine infectious respiratory disease (CIRD) is an acute, highly contagious disease complex caused by a variety of infectious agents. At present, the role of viral and bacterial components as primary or secondary pathogens in CIRD is not fully understood. The aim of this study was to investigate the prevalence of canine parainfluenza virus (CPIV), canine adenovirus type 2 (CAV-2), canine influenza virus (CIV), canine respiratory coronavirus (CRCoV), canine herpes virus-1 (CHV-1), canine distemper virus (CDV) and Bordetella bronchiseptica in dogs with CIRD and to compare the data with findings in healthy dogs. Sixty-one dogs with CIRD and 90 clinically healthy dogs from Southern Germany were prospectively enrolled in this study. Nasal and pharyngeal swabs were collected from all dogs and were analysed for CPIV, CAV-2, CIV, CRCoV, CHV-1, CDV, and B. bronchiseptica by real-time PCR. In dogs with acute respiratory signs, 37.7% tested positive for CPIV, 9.8% for CRCoV and 78.7% for B. bronchiseptica. Co-infections with more than one agent were detected in 47.9% of B. bronchiseptica-positive, 82.6% of CPIV-positive, and 100% of CRCoV-positive dogs. In clinically healthy dogs, 1.1% tested positive for CAV-2, 7.8% for CPIV and 45.6% for B. bronchiseptica. CPIV and B. bronchiseptica were detected significantly more often in dogs with CIRD than in clinically healthy dogs (P < 0.001 for each pathogen) and were the most common infectious agents in dogs with CIRD in Southern Germany. Mixed infections with several pathogens were common. In conclusion, clinically healthy dogs can carry respiratory pathogens and could act as sources of infection for susceptible dogs.

  7. Severe canine distemper outbreak in unvaccinated dogs in Mozambique.

    PubMed

    Zacarias, Julieta; Dimande, Alberto; Achá, Sara; Dias, Paula T; Leonel, Elisa M; Messa, Aurora; Macucule, Baltazar; Júnior, José L; Bila, Custódio G

    2016-01-01

    Although significant animal suffering caused by preventable diseases is frequently seen in developing countries, reports of this are scarce. This report describes avoidable animal suffering owing to a suspected canine distemper (CD) outbreak in unvaccinated dogs owned by low-income families in Mozambique that killed approximately 200 animals. Affected dogs exhibited clinical signs, and gross and microscopic lesions compatible with CD. Immunohistochemical staining confirmed the presence of canine distemper virus (CDV) in the kidney of one dog from the cohort. This brief communication again illustrates that large outbreaks of CDV in unvaccinated dogs occur and that large-scale avoidable suffering and threats to the health of dogs and wild canines continue. Mass vaccination supported by government and non-government organisations is recommended. PMID:27543040

  8. Distemper: not a new disease in lions and tigers.

    PubMed Central

    Myers, D L; Zurbriggen, A; Lutz, H; Pospischil, A

    1997-01-01

    In light of recent canine distemper virus (CDV) epidemics, we set out to determine the historical significance of CDV infection in captive lions and tigers in Switzerland. The retrospective case material consisted of 42 lion and tiger necropsy cases from 1972 to 1992. Necropsy reports for all lions and tigers were reviewed. All existing paraffin tissues were immunohistochemically examined with a polyclonal antibody raised against CDV. The results for 19 of the 42 lions and tigers were classified as positive by immunohistochemistry; 23 results were negative or questionable. The results for four animals (three positive and one negative ) were further tested by in situ hybridization, and the results concurred with the immunohistochemistry findings. CDV infection of large cats is older and more widespread than previously thought. All large cats in captivity should be immunized even if canine distemper is not believed to be a problem for large cats in the area. PMID:9067652

  9. Detection of Respiratory Viruses by Molecular Methods

    PubMed Central

    Mahony, James B.

    2008-01-01

    Summary: Clinical laboratories historically diagnose seven or eight respiratory virus infections using a combination of techniques including enzyme immunoassay, direct fluorescent antibody staining, cell culture, and nucleic acid amplification tests. With the discovery of six new respiratory viruses since 2000, laboratories are faced with the challenge of detecting up to 19 different viruses that cause acute respiratory disease of both the upper and lower respiratory tracts. The application of nucleic acid amplification technology, particularly multiplex PCR coupled with fluidic or fixed microarrays, provides an important new approach for the detection of multiple respiratory viruses in a single test. These multiplex amplification tests provide a sensitive and comprehensive approach for the diagnosis of respiratory tract infections in individual hospitalized patients and the identification of the etiological agent in outbreaks of respiratory tract infection in the community. This review describes the molecular methods used to detect respiratory viruses and discusses the contribution that molecular testing, especially multiplex PCR, has made to our ability to detect respiratory viruses and to increase our understanding of the roles of various viral agents in acute respiratory disease. PMID:18854489

  10. Detection of pseudorabies virus by DNA hybridization

    SciTech Connect

    McFarlane, R.G.

    1985-01-01

    A DNA hybridization technique was developed in order to detect the presence of pseudorabies virus (PRV) deoxyribonucleic acid (DNA) in swine tissue. Seven, /sup 32/P-nick translated probes of high specific activity were prepared from transformed Escherichia coli plasmids into which Bacillus amyloliquefaciens H (Bam H1) restriction fragments of PRV-DNA had been inserted. Under optimal hybridization conditions, the minimum detection level of PRV-DNA was 10/sup -11/ g, which is equivalent to 1 copy of the PRV genome/80 host cells. PRV-DNA was detected in the DNA extracted from the tissues of 10 out of 11 swine previously shown to harbor infective virus. Furthermore, PRV-DNA was present in all four seropositive swine that had recovered from pseudorabies, where no infective virus or viral products were detected at necropsy. The PRV-DNA was present in either the anterior cerebral cortex in 2 swine, or the medulla oblongata and trigeminal ganglion in 1 swine. This perhaps indicates the portal of entry of the virus into the central nervous system. This DNA hybridization assay, which utilizes restriction fragments, may be useful for studying the dynamics and molecular biologic properties of the latency of pseudorabies virus in swine.

  11. Detection of Zika Virus in Urine

    PubMed Central

    Gourinat, Ann-Claire; O’Connor, Olivia; Calvez, Elodie; Goarant, Cyrille

    2015-01-01

    We describe the kinetics of Zika virus (ZIKV) detection in serum and urine samples of 6 patients. Urine samples were positive for ZIKV >10 days after onset of disease, which was a notably longer period than for serum samples. This finding supports the conclusion that urine samples are useful for diagnosis of ZIKV infections. PMID:25530324

  12. Detection of Zika virus in urine.

    PubMed

    Gourinat, Ann-Claire; O'Connor, Olivia; Calvez, Elodie; Goarant, Cyrille; Dupont-Rouzeyrol, Myrielle

    2015-01-01

    We describe the kinetics of Zika virus (ZIKV) detection in serum and urine samples of 6 patients. Urine samples were positive for ZIKV >10 days after onset of disease, which was a notably longer period than for serum samples. This finding supports the conclusion that urine samples are useful for diagnosis of ZIKV infections.

  13. Integrated capture and spectroscopic detection of viruses.

    PubMed

    Vargas, Crystal A; Wilhelm, Allison A; Williams, Jeremy; Lucas, Pierre; Reynolds, Kelly A; Riley, Mark R

    2009-10-01

    The goal of this work is to develop an online monitoring scheme for detection of viruses in flowing drinking water. The approach applies an electrodeposition process that is similar to the use of charged membrane filters previously employed for collection of viruses from aqueous samples. In the present approach, charged materials are driven onto a robust optical sensing element which has high transparency to infrared light. A spectroscopic measurement is performed using the evanescent wave that penetrates no more than 1 mum from the surface of an infrared optical element in an attenuated total reflectance measurement scheme. The infrared measurement provides quantitative information on the amount and identity of material deposited from the water. Initial studies of this sensing scheme used proteins reversibly electrodeposited onto germanium chips. The results of those studies were applied to design a method for collection of viruses onto an attenuated total reflectance crystal. Spectral signatures can be discriminated between three types of protein and two viruses. There is the potential to remove deposited material by reversing the voltage polarity. This work demonstrates a novel and practical scheme for detection of viruses in water systems with potential application to near-continual, automated monitoring of municipal drinking water.

  14. Detection of DNA viruses in prostate cancer

    PubMed Central

    Smelov, Vitaly; Bzhalava, Davit; Arroyo Mühr, Laila Sara; Eklund, Carina; Komyakov, Boris; Gorelov, Andrey; Dillner, Joakim; Hultin, Emilie

    2016-01-01

    We tested prostatic secretions from men with and without prostate cancer (13 cases and 13 matched controls) or prostatitis (18 cases and 18 matched controls) with metagenomic sequencing. A large number (>200) of viral reads was only detected among four prostate cancer cases (1 patient each positive for Merkel cell polyomavirus, JC polyomavirus and Human Papillomavirus types 89 or 40, respectively). Lower numbers of reads from a large variety of viruses were detected in all patient groups. Our knowledge of the biology of the prostate may be furthered by the fact that DNA viruses are commonly shed from the prostate and can be readily detected by metagenomic sequencing of expressed prostate secretions. PMID:27121729

  15. Rapid molecular detection of Lujo virus RNA.

    PubMed

    Atkinson, Barry; Chamberlain, John; Dowall, Stuart D; Cook, Nicola; Bruce, Christine; Hewson, Roger

    2014-01-01

    Lujo virus is an emerging arenavirus circulating in Southern Africa. Although to date there has only been a single outbreak of the novel haemorrhagic disease resulting from human infection with this virus, the case-fatality rate of exposed individuals, including nosocomial transmission, was 80%. The ability to identify viral haemorrhagic fevers accurately, especially those capable of nosocomial transmission, is of critical importance. Timely identification of these diseases allow medical professionals to isolate patients and implement barrier nursing techniques in order to prevent onward transmission of the virus. While rapid diagnostic methods are published for most viral haemorrhagic fevers, at present there are no such virus specific protocols for Lujo haemorrhagic fever. This report details the first set of diagnostic molecular assays designed to identify Lujo viral RNA rapidly, and demonstrates the potential functionality of these assays for use in the clinical setting. Although these assays have been designed and validated against a solitary isolate of Lujo virus, this represents the entirety of strains detected to date, and offer quick, cheap and easy methods for use in diagnostic laboratories.

  16. Detection, isolation, and persistence of viruses within bivalve mollusks

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Norovirus (NV), hepatitis A virus (HAV), and other virus transmission by molluscan shellfish is a significant issue. Research at the ARS-Dover DE laboratory has led to the development of improved methods for detecting these viruses. To identify pathogenic viruses within mollusks, a rapid highly-se...

  17. Detection of virus in shrimp using digital color correlation

    NASA Astrophysics Data System (ADS)

    Alvarez-Borrego, Josue; Chavez-Sanchez, Cristina; Bueno-Ibarra, Mario A.

    1999-07-01

    Detection of virus in shrimp tissue using digital color correlation is presented. Phase filters in three channels (red, green and blue) were used in order to detect HPV virus like target. These first results obtained showed that is possible to detect virus in shrimp tissue. More research must be made with color correlation in order to consider natural morphology of the virus, color, scale and rotation and noise in the samples.

  18. The Unknown Computer Viruses Detection Based on Similarity

    NASA Astrophysics Data System (ADS)

    Liu, Zhongda; Nakaya, Naoshi; Koui, Yuuji

    New computer viruses are continually being generated and they cause damage all over the world. In general, current anti-virus software detects viruses by matching a pattern based on the signature; thus, unknown viruses without any signature cannot be detected. Although there are some static analysis technologies that do not depend on signatures, virus writers often use code obfuscation techniques, which make it difficult to execute a code analysis. As is generally known, unknown viruses and known viruses share a common feature. In this paper we propose a new static analysis technology that can circumvent code obfuscation to extract the common feature and detect unknown viruses based on similarity. The results of evaluation experiments demonstrated that this technique is able to detect unknown viruses without false positives.

  19. Detection of Schmallenberg virus serum neutralising antibodies.

    PubMed

    Mansfield, Karen L; La Rocca, S Anna; Khatri, Meenakshi; Johnson, Nicholas; Steinbach, Falko; Fooks, Anthony R

    2013-03-01

    Schmallenberg virus (SBV) emerged in continental Europe in late 2011, and further work is required to assess the prevalence of SBV throughout Europe. Since its detection in Germany, SBV has now been detected in other European countries, including the United Kingdom. Infection with SBV can cause mild clinical signs in ruminants, including diarrhoea and reduced milk yield. However, the virus can have a devastating effect on the developing foetus leading to malformation in newborn offspring. This is a feature shared by other members of the Simbu group of orthobunyaviruses. Since disease in adult animals can be inapparent, serology offers the best method for monitoring for the presence of SBV and assisting in livestock management. This protocol describes a method for initial titration of SBV on African Green Monkey kidney (Vero) cells, and a plaque reduction neutralisation test (PRNT) for the detection of neutralising antibodies against SBV in cattle and sheep sera. This assay can be used to screen ruminant sera in order to confirm exposure to the virus, and the results obtained are comparable to a recently developed commercial enzyme linked immunosorbent assay (ELISA). Thus, these two assays constitute an effective diagnostic tool-box for providing confirmation of exposure to SBV.

  20. Biosensors for hepatitis B virus detection.

    PubMed

    Yao, Chun-Yan; Fu, Wei-Ling

    2014-09-21

    A biosensor is an analytical device used for the detection of analytes, which combines a biological component with a physicochemical detector. Recently, an increasing number of biosensors have been used in clinical research, for example, the blood glucose biosensor. This review focuses on the current state of biosensor research with respect to efficient, specific and rapid detection of hepatitis B virus (HBV). The biosensors developed based on different techniques, including optical methods (e.g., surface plasmon resonance), acoustic wave technologies (e.g., quartz crystal microbalance), electrochemistry (amperometry, voltammetry and impedance) and novel nanotechnology, are also discussed. PMID:25253948

  1. Biosensors for hepatitis B virus detection

    PubMed Central

    Yao, Chun-Yan; Fu, Wei-Ling

    2014-01-01

    A biosensor is an analytical device used for the detection of analytes, which combines a biological component with a physicochemical detector. Recently, an increasing number of biosensors have been used in clinical research, for example, the blood glucose biosensor. This review focuses on the current state of biosensor research with respect to efficient, specific and rapid detection of hepatitis B virus (HBV). The biosensors developed based on different techniques, including optical methods (e.g., surface plasmon resonance), acoustic wave technologies (e.g., quartz crystal microbalance), electrochemistry (amperometry, voltammetry and impedance) and novel nanotechnology, are also discussed. PMID:25253948

  2. Microbiology--Detection, Occurrence, and Removal of Viruses.

    ERIC Educational Resources Information Center

    Berg, Gerald

    1978-01-01

    Presents a literature review of the detection, survival, and destruction of viruses in wastewater sludges, covering publications of 1976-77. This review includes these topics: (1) detection methodology; (2) viruses in sludge, shellfish, and aerosols; and (3) indicators of viruses. A list of 59 references is also presented. (HM)

  3. Development of a two-step SYBR Green I based real time RT-PCR assay for detecting and quantifying peste des petits ruminants virus in clinical samples.

    PubMed

    Abera, Tsegalem; Thangavelu, Ardhanary

    2014-12-01

    A two-step SYBR Green I based real time RT-PCR targeting the matrix (M) gene of Peste des petits ruminants virus (PPRV) was developed. The specificity of the assay was assessed against viral nucleic acid extracted from a range of animal viruses of clinical and structural similarities to PPRV including canine distemper virus, measles virus, bluetongue virus and Newcastle disease virus. But none of the viruses and no template control showed an amplification signal. Sensitivity of the same assay was assessed based on plasmid DNA copy number and with respect to infectivity titre. The lower detection limit achieved was 2.88 plasmid DNA copies/μl with corresponding Ct value of 35.93. Based on tissue culture infectivity titre the lower detection limits were 0.0001TCID50/ml and 1TCID50/ml for the SYBR green I based real time RT-PCR and conventional RT-PCR, respectively. The calculated coefficient of variations values for intra- and inter-assay variability were low, ranging from 0.21% to 1.83% and 0.44% to 1.97%, respectively. The performance of newly developed assay was evaluated on a total of 36 clinical samples suspected of PPR and compared with conventional RT-PCR. The SYBR Green I based real time RT-PCR assay detected PPRV in 32 (88.8%) of clinical samples compared to 19 (52.7%) by conventional RT-PCR. Thus, the two-step SYBR Green I based real time RT-PCR assay targeting the M gene of PPRV reported in this study was highly sensitive, specific and reproducible for detection and quantitation of PPRV nucleic acids.

  4. Detection of measles virus nucleocapsid transcripts in circulating blood cells from patients with Paget disease.

    PubMed

    Reddy, S V; Singer, F R; Mallette, L; Roodman, G D

    1996-11-01

    Paget disease of bone is characterized by abnormalities in all phases of bone remodeling, but the fundamental cellular abnormality resides in the osteoclast (OCL). Osteoclasts in bone involved by Paget disease contain viral-like nuclear and cytoplasmic inclusions that react with antibodies directed against paramyxovirus nucleocapsid proteins, such as measles virus, respiratory syncytial virus, or canine distemper virus. However, the identity of the virus or the mechanisms responsible for its persistence or pathologic role in Paget disease is unclear. Furthermore, although Paget disease persists for many years, it remains a highly localized process with new lesions rarely if ever developing in previously unaffected bones. Since osteoclasts are formed by fusion of mononuclear precursors derived from colony forming unit-granulocyte macrophage (CFU-GM), the granulocyte-macrophage progenitor, we used reverse transcriptase polymerase chain reaction (RT-PCR) analysis to determine if CFU-GM, more differentiated osteoclast precursors, and peripheral blood cells derived from CFU-GM express measles virus nucleocapsid (MV-N) transcripts. We found that osteoclast precursors, as well as peripheral blood mononuclear cells, express MV transcripts in 9 of 13 patients. Sequence analysis of the PCR amplified products confirmed nucleotide identity of MV-N transcripts expressed in peripheral blood and bone marrow-derived cells from the same patient. In contrast, MV-N transcripts were not detected in OCL precursors or the peripheral blood from 10 normal subjects. In situ hybridization studies using 35S-labeled antisense riboprobes to MV-N transcripts further confirmed the expression of MV transcripts in these cells. Sequence analysis of the PCR amplified product from one of these patients also identified a novel mutation that converted lysine441 to glutamic acid441 in the MV-N transcript. These data demonstrate that OCL precursors and circulating peripheral blood cells also express MV

  5. Rapid Detection of the Varicella Zoster Virus

    NASA Technical Reports Server (NTRS)

    Lewis, Michelle P.; Harding, Robert

    2011-01-01

    1.Technology Description-Researchers discovered that when the Varicella Zoster Virus (VZV) reactivates from latency in the body, the virus is consistently present in saliva before the appearance of skin lesions. A small saliva sample is mixed with a specialized reagent in a test kit. If the virus is present in the saliva sample, the mixture turns a red color. The sensitivity and specificity emanates from an antibody-antigen reaction. This technology is a rapid, non-invasive, point of-of-care testing kit for detecting the virus from a saliva sample. The device is easy to use and can be used in clinics and in remote locations to quickly detect VZV and begin treatment with antiviral drugs. 2.Market Opportunity- RST Bioscience will be the first and only company to market a rapid, same day test kit for the detection of VZV in saliva. The RST detection test kit will have several advantages over existing, competitive technology. The test kit is self contained and laboratory equipment is not required for analysis of the sample. Only a single saliva sample is required to be taken instead of blood or cerebral spinal fluid. The test kit is portable, sterile and disposable after use. RST detection test kits require no electrical power or expensive storage equipment and can be used in remote locations. 3.Market Analysis- According to the CDC, it is estimated that 1 million cases of shingles occur each year in the U.S. with more than half over the age of sixty. There is a high demand for rapid diagnostics by the public. The point-of-care testing (POCT) market is growing faster than other segments of in vitro diagnostics. According to a July 2007 InteLab Corporation industry report the overall market for POCT was forecast to increase from $10.3 billion in 2005 to $18.7 billion by 2011. The market value of this test kit has not been determined. 4.Competition- The VZV vaccine prevents 50% of cases and reduces neuralgia by 66%. The most popular test detects VZV-specific IgM antibody

  6. Real-Time Detection of a Virus Using Detection Dogs.

    PubMed

    Angle, T Craig; Passler, Thomas; Waggoner, Paul L; Fischer, Terrence D; Rogers, Bart; Galik, Patricia K; Maxwell, Herris S

    2015-01-01

    Viral infections are ubiquitous in humans, animals, and plants. Real-time methods to identify viral infections are limited and do not exist for use in harsh or resource-constrained environments. Previous research identified that tissues produce unique volatile organic compounds (VOC) and demonstrated that VOC concentrations change during pathologic states, including infection, neoplasia, or metabolic disease. Patterns of VOC expression may be pathogen specific and may be associated with an odor that could be used for disease detection. We investigated the ability of two trained dogs to detect cell cultures infected with bovine viral diarrhea virus (BVDV) and to discriminate BVDV-infected cell cultures from uninfected cell cultures and from cell cultures infected with bovine herpes virus 1 (BHV 1) and bovine parainfluenza virus 3 (BPIV 3). Dogs were trained to recognize cell cultures infected with two different biotypes of BVDV propagated in Madin-Darby bovine kidney cells using one of three culture media. For detection trials, one target and seven distractors were presented on a scent wheel by a dog handler unaware of the location of targets and distractors. Detection of BVDV-infected cell cultures by Dog 1 had a diagnostic sensitivity of 0.850 (95% CI: 0.701-0.942), which was lower than Dog 2 (0.967, 95% CI: 0.837-0.994). Both dogs exhibited very high diagnostic specificity (0.981, 95% CI: 0.960-0.993) and (0.993, 95% CI: 0.975-0.999), respectively. These findings demonstrate that trained dogs can differentiate between cultured cells infected with BVDV, BHV1, and BPIV3 and are a realistic real-time mobile pathogen sensing technology for viral pathogens. The ability to discriminate between target and distractor samples plausibly results from expression of unique VOC patterns in virus-infected and -uninfected cells. PMID:26779494

  7. Real-Time Detection of a Virus Using Detection Dogs

    PubMed Central

    Angle, T. Craig; Passler, Thomas; Waggoner, Paul L.; Fischer, Terrence D.; Rogers, Bart; Galik, Patricia K.; Maxwell, Herris S.

    2016-01-01

    Viral infections are ubiquitous in humans, animals, and plants. Real-time methods to identify viral infections are limited and do not exist for use in harsh or resource-constrained environments. Previous research identified that tissues produce unique volatile organic compounds (VOC) and demonstrated that VOC concentrations change during pathologic states, including infection, neoplasia, or metabolic disease. Patterns of VOC expression may be pathogen specific and may be associated with an odor that could be used for disease detection. We investigated the ability of two trained dogs to detect cell cultures infected with bovine viral diarrhea virus (BVDV) and to discriminate BVDV-infected cell cultures from uninfected cell cultures and from cell cultures infected with bovine herpes virus 1 (BHV 1) and bovine parainfluenza virus 3 (BPIV 3). Dogs were trained to recognize cell cultures infected with two different biotypes of BVDV propagated in Madin–Darby bovine kidney cells using one of three culture media. For detection trials, one target and seven distractors were presented on a scent wheel by a dog handler unaware of the location of targets and distractors. Detection of BVDV-infected cell cultures by Dog 1 had a diagnostic sensitivity of 0.850 (95% CI: 0.701–0.942), which was lower than Dog 2 (0.967, 95% CI: 0.837–0.994). Both dogs exhibited very high diagnostic specificity (0.981, 95% CI: 0.960–0.993) and (0.993, 95% CI: 0.975–0.999), respectively. These findings demonstrate that trained dogs can differentiate between cultured cells infected with BVDV, BHV1, and BPIV3 and are a realistic real-time mobile pathogen sensing technology for viral pathogens. The ability to discriminate between target and distractor samples plausibly results from expression of unique VOC patterns in virus-infected and -uninfected cells. PMID:26779494

  8. Protocol for Metagenomic Virus Detection in Clinical Specimens1

    PubMed Central

    Brinkmann, Annika; Dabrowski, Piotr W.; Radonić, Aleksandar; Nitsche, Andreas; Kurth, Andreas

    2015-01-01

    Sixty percent of emerging viruses have a zoonotic origin, making transmission from animals a major threat to public health. Prompt identification and analysis of these pathogens are indispensable to taking action toward prevention and protection of the affected population. We quantifiably compared classical and modern approaches of virus purification and enrichment in theory and experiments. Eventually, we established an unbiased protocol for detection of known and novel emerging viruses from organ tissues (tissue-based universal virus detection for viral metagenomics [TUViD-VM]). The final TUViD-VM protocol was extensively validated by using real-time PCR and next-generation sequencing. We could increase the amount of detectable virus nucleic acids and improved the detection of viruses <75,000-fold compared with other tested approaches. This TUViD-VM protocol can be used in metagenomic and virome studies to increase the likelihood of detecting viruses from any biological source. PMID:25532973

  9. Chikungunya Virus Growth and Fluorescent Labeling: Detection of Chikungunya Virus by Immunofluorescence Assay.

    PubMed

    Moi, Meng Ling; Takasaki, Tomohiko

    2016-01-01

    Immunofluorescence assay (IFA) is a highly versatile and sensitive assay for detection and titration of chikungunya virus (CHIKV). The IFA technique requires virus-infected cells (viral antigen) and antibodies specific to the viral antigens for detection. Suitable antibodies for detection include monoclonal antibodies specific to CHIKV structural and nonstructural proteins, polyclonal antibodies, and convalescent serum samples. Here, the details of virus antigen preparation, detection by IFA method, and applications are described. The described IFA method is potentially useful in a wide range of studies including virus growth kinetics and virus infection mechanism studies. Additionally, the described IFA method can be modified for applications in arbovirus diagnosis, including CHIKV.

  10. Simultaneous multiplex PCR detection of seven cucurbit-infecting viruses.

    PubMed

    Kwon, Ji Yeon; Hong, Jin Sung; Kim, Min Jea; Choi, Sun Hee; Min, Byeong Eun; Song, Eun Gyeong; Kim, Hyun Hee; Ryu, Ki Hyun

    2014-09-01

    Two multiplex polymerase chain reaction (PCR) systems using dual priming oligonucleotide (DPO) primers were developed for the simultaneous detection of seven cucurbit-infecting viruses. One system allows for the detection of papaya ringspot virus, watermelon mosaic virus, and zucchini yellow mosaic virus, whereas the other permits the detection of cucumber green mottle mosaic virus, cucumber fruit mottle mosaic virus, kyuri green mottle mosaic virus, and zucchini green mottle mosaic virus. Viral species-specific DPO primers developed in this study detected as little as 10 fg/μl of viral RNA under monoplex conditions and 10 pg/μl of viral RNA under multiplex conditions. Multiplex PCR using the DPO primer sets was capable of amplifying viral genes at annealing temperatures ranging from 53 °C to 63 °C. Whereas the use of conventional primers gave rise to non-specific bands, the DPO primers detected target viral genes in the absence of non-specific amplification. When these DPO multiplex primer sets were applied to virus-infected cucurbit samples obtained in the field, multiple infection as well as single infection was accurately identified. This novel approach could also detect multiple viruses in infected seeds. The reliability of multiplex PCR systems using DPO primers for plant virus detection is discussed.

  11. Simultaneous multiplex PCR detection of seven cucurbit-infecting viruses.

    PubMed

    Kwon, Ji Yeon; Hong, Jin Sung; Kim, Min Jea; Choi, Sun Hee; Min, Byeong Eun; Song, Eun Gyeong; Kim, Hyun Hee; Ryu, Ki Hyun

    2014-09-01

    Two multiplex polymerase chain reaction (PCR) systems using dual priming oligonucleotide (DPO) primers were developed for the simultaneous detection of seven cucurbit-infecting viruses. One system allows for the detection of papaya ringspot virus, watermelon mosaic virus, and zucchini yellow mosaic virus, whereas the other permits the detection of cucumber green mottle mosaic virus, cucumber fruit mottle mosaic virus, kyuri green mottle mosaic virus, and zucchini green mottle mosaic virus. Viral species-specific DPO primers developed in this study detected as little as 10 fg/μl of viral RNA under monoplex conditions and 10 pg/μl of viral RNA under multiplex conditions. Multiplex PCR using the DPO primer sets was capable of amplifying viral genes at annealing temperatures ranging from 53 °C to 63 °C. Whereas the use of conventional primers gave rise to non-specific bands, the DPO primers detected target viral genes in the absence of non-specific amplification. When these DPO multiplex primer sets were applied to virus-infected cucurbit samples obtained in the field, multiple infection as well as single infection was accurately identified. This novel approach could also detect multiple viruses in infected seeds. The reliability of multiplex PCR systems using DPO primers for plant virus detection is discussed. PMID:24937806

  12. Single-Reaction Multiplex Reverse Transcription PCR for Detection of Zika, Chikungunya, and Dengue Viruses.

    PubMed

    Waggoner, Jesse J; Gresh, Lionel; Mohamed-Hadley, Alisha; Ballesteros, Gabriela; Davila, Maria Jose Vargas; Tellez, Yolanda; Sahoo, Malaya K; Balmaseda, Angel; Harris, Eva; Pinsky, Benjamin A

    2016-07-01

    Clinical manifestations of Zika virus, chikungunya virus, and dengue virus infections can be similar. To improve virus detection, streamline molecular workflow, and decrease test costs, we developed and evaluated a multiplex real-time reverse transcription PCR for these viruses.

  13. Molecular and serological surveillance of canine enteric viruses in stray dogs from Vila do Maio, Cape Verde

    PubMed Central

    2014-01-01

    Background Infections caused by canine parvovirus, canine distemper virus and canine coronavirus are an important cause of mortality and morbidity in dogs worldwide. Prior to this study, no information was available concerning the incidence and prevalence of these viruses in Cape Verde archipelago. Results To provide information regarding the health status of the canine population in Vila do Maio, Maio Island, Cape Verde, 53 rectal swabs were collected from 53 stray dogs during 2010 and 93 rectal swabs and 88 blood samples were collected from 125 stray dogs in 2011. All rectal swabs (2010 n = 53; 2011 n = 93) were analysed for the presence of canine parvovirus, canine distemper virus and canine coronavirus nucleic acids by quantitative PCR methods. Specific antibodies against canine distemper virus and canine parvovirus were also assessed (2011 n = 88). From the 2010 sampling, 43.3% (23/53) were positive for canine parvovirus DNA, 11.3% (6/53) for canine distemper virus RNA and 1.9% (1/53) for canine coronavirus RNA. In 2011, the prevalence values for canine parvovirus and canine coronavirus were quite similar to those from the previous year, respectively 44.1% (41/93), and 1.1% (1/93), but canine distemper virus was not detected in any of the samples analysed (0%, 0/93). Antibodies against canine parvovirus were detected in 71.6% (63/88) blood samples and the seroprevalence found for canine distemper virus was 51.1% (45/88). Conclusions This study discloses the data obtained in a molecular and serological epidemiological surveillance carried out in urban populations of stray and domestic animals. Virus transmission and spreading occurs easily in large dog populations leading to high mortality rates particularly in unvaccinated susceptible animals. In addition, these animals can act as disease reservoirs for wild animal populations by occasional contact. Identification of susceptible wildlife of Maio Island is of upmost importance to evaluate the risk

  14. Porcupine quills in raccoons as an indicator of rabies, distemper, or both diseases: disease management implications.

    PubMed

    Rosatte, Rick; Wandeler, Alex; Muldoon, Frances; Campbell, Doug

    2007-03-01

    A relationship was detected between the presence of embedded porcupine quills and the diagnosis of rabies in raccoons in eastern Canada during 1999-2004. No relationship was found between the presence of quills in raccoons and the diagnosis of canine distemper. Raccoons with embedded quills should be submitted for rabies testing. PMID:17436909

  15. Detection of Pathogenic Viruses in Sewage Provided Early Warnings of Hepatitis A Virus and Norovirus Outbreaks

    PubMed Central

    Hellmér, Maria; Paxéus, Nicklas; Magnius, Lars; Enache, Lucica; Arnholm, Birgitta; Johansson, Annette; Bergström, Tomas

    2014-01-01

    Most persons infected with enterically transmitted viruses shed large amounts of virus in feces for days or weeks, both before and after onset of symptoms. Therefore, viruses causing gastroenteritis may be detected in wastewater, even if only a few persons are infected. In this study, the presence of eight pathogenic viruses (norovirus, astrovirus, rotavirus, adenovirus, Aichi virus, parechovirus, hepatitis A virus [HAV], and hepatitis E virus) was investigated in sewage to explore whether their identification could be used as an early warning of outbreaks. Samples of the untreated sewage were collected in proportion to flow at Ryaverket, Gothenburg, Sweden. Daily samples collected during every second week between January and May 2013 were pooled and analyzed for detection of viruses by concentration through adsorption to milk proteins and PCR. The largest amount of noroviruses was detected in sewage 2 to 3 weeks before most patients were diagnosed with this infection in Gothenburg. The other viruses were detected at lower levels. HAV was detected between weeks 5 and 13, and partial sequencing of the structural VP1protein identified three different strains. Two strains were involved in an ongoing outbreak in Scandinavia and were also identified in samples from patients with acute hepatitis A in Gothenburg during spring of 2013. The third strain was unique and was not detected in any patient sample. The method used may thus be a tool to detect incipient outbreaks of these viruses and provide early warning before the causative pathogens have been recognized in health care. PMID:25172863

  16. Detection of pathogenic viruses in sewage provided early warnings of hepatitis A virus and norovirus outbreaks.

    PubMed

    Hellmér, Maria; Paxéus, Nicklas; Magnius, Lars; Enache, Lucica; Arnholm, Birgitta; Johansson, Annette; Bergström, Tomas; Norder, Heléne

    2014-11-01

    Most persons infected with enterically transmitted viruses shed large amounts of virus in feces for days or weeks, both before and after onset of symptoms. Therefore, viruses causing gastroenteritis may be detected in wastewater, even if only a few persons are infected. In this study, the presence of eight pathogenic viruses (norovirus, astrovirus, rotavirus, adenovirus, Aichi virus, parechovirus, hepatitis A virus [HAV], and hepatitis E virus) was investigated in sewage to explore whether their identification could be used as an early warning of outbreaks. Samples of the untreated sewage were collected in proportion to flow at Ryaverket, Gothenburg, Sweden. Daily samples collected during every second week between January and May 2013 were pooled and analyzed for detection of viruses by concentration through adsorption to milk proteins and PCR. The largest amount of noroviruses was detected in sewage 2 to 3 weeks before most patients were diagnosed with this infection in Gothenburg. The other viruses were detected at lower levels. HAV was detected between weeks 5 and 13, and partial sequencing of the structural VP1protein identified three different strains. Two strains were involved in an ongoing outbreak in Scandinavia and were also identified in samples from patients with acute hepatitis A in Gothenburg during spring of 2013. The third strain was unique and was not detected in any patient sample. The method used may thus be a tool to detect incipient outbreaks of these viruses and provide early warning before the causative pathogens have been recognized in health care.

  17. VirusDetect: An automated pipeline for efficient virus discovery using deep sequencing of small RNAs

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Accurate detection of viruses in plants and animals is critical for agriculture production and human health. Deep sequencing and assembly of virus-derived siRNAs has proven to be a highly efficient approach for virus discovery. However, to date no computational tools specifically designed for both k...

  18. Virus detection and quantification using electrical parameters

    PubMed Central

    Ahmad, Mahmoud Al; Mustafa, Farah; Ali, Lizna M.; Rizvi, Tahir A.

    2014-01-01

    Here we identify and quantitate two similar viruses, human and feline immunodeficiency viruses (HIV and FIV), suspended in a liquid medium without labeling, using a semiconductor technique. The virus count was estimated by calculating the impurities inside a defined volume by observing the change in electrical parameters. Empirically, the virus count was similar to the absolute value of the ratio of the change of the virus suspension dopant concentration relative to the mock dopant over the change in virus suspension Debye volume relative to mock Debye volume. The virus type was identified by constructing a concentration-mobility relationship which is unique for each kind of virus, allowing for a fast (within minutes) and label-free virus quantification and identification. For validation, the HIV and FIV virus preparations were further quantified by a biochemical technique and the results obtained by both approaches corroborated well. We further demonstrate that the electrical technique could be applied to accurately measure and characterize silica nanoparticles that resemble the virus particles in size. Based on these results, we anticipate our present approach to be a starting point towards establishing the foundation for label-free electrical-based identification and quantification of an unlimited number of viruses and other nano-sized particles. PMID:25355078

  19. Virus detection and quantification using electrical parameters

    NASA Astrophysics Data System (ADS)

    Ahmad, Mahmoud Al; Mustafa, Farah; Ali, Lizna M.; Rizvi, Tahir A.

    2014-10-01

    Here we identify and quantitate two similar viruses, human and feline immunodeficiency viruses (HIV and FIV), suspended in a liquid medium without labeling, using a semiconductor technique. The virus count was estimated by calculating the impurities inside a defined volume by observing the change in electrical parameters. Empirically, the virus count was similar to the absolute value of the ratio of the change of the virus suspension dopant concentration relative to the mock dopant over the change in virus suspension Debye volume relative to mock Debye volume. The virus type was identified by constructing a concentration-mobility relationship which is unique for each kind of virus, allowing for a fast (within minutes) and label-free virus quantification and identification. For validation, the HIV and FIV virus preparations were further quantified by a biochemical technique and the results obtained by both approaches corroborated well. We further demonstrate that the electrical technique could be applied to accurately measure and characterize silica nanoparticles that resemble the virus particles in size. Based on these results, we anticipate our present approach to be a starting point towards establishing the foundation for label-free electrical-based identification and quantification of an unlimited number of viruses and other nano-sized particles.

  20. Tunable and label-free virus enrichment for ultrasensitive virus detection using carbon nanotube arrays

    PubMed Central

    Yeh, Yin-Ting; Tang, Yi; Sebastian, Aswathy; Dasgupta, Archi; Perea-Lopez, Nestor; Albert, Istvan; Lu, Huaguang; Terrones, Mauricio; Zheng, Si-Yang

    2016-01-01

    Viral infectious diseases can erupt unpredictably, spread rapidly, and ravage mass populations. Although established methods, such as polymerase chain reaction, virus isolation, and next-generation sequencing have been used to detect viruses, field samples with low virus count pose major challenges in virus surveillance and discovery. We report a unique carbon nanotube size-tunable enrichment microdevice (CNT-STEM) that efficiently enriches and concentrates viruses collected from field samples. The channel sidewall in the microdevice was made by growing arrays of vertically aligned nitrogen-doped multiwalled CNTs, where the intertubular distance between CNTs could be engineered in the range of 17 to 325 nm to accurately match the size of different viruses. The CNT-STEM significantly improves detection limits and virus isolation rates by at least 100 times. Using this device, we successfully identified an emerging avian influenza virus strain [A/duck/PA/02099/2012(H11N9)] and a novel virus strain (IBDV/turkey/PA/00924/14). Our unique method demonstrates the early detection of emerging viruses and the discovery of new viruses directly from field samples, thus creating a universal platform for effectively remediating viral infectious diseases. PMID:27730213

  1. Molecular beacon real-time PCR detection of swine viruses.

    PubMed

    McKillen, John; Hjertner, Bernt; Millar, Andrena; McNeilly, Francis; Belák, Sándor; Adair, Brian; Allan, Gordon

    2007-03-01

    Rapid and reliable detection of viral pathogens is critical for the management of the diseases threatening the economic competitiveness of the swine farming industry worldwide. Molecular beacon assays are one type of real-time polymerase chain reaction (PCR) technology capable of fast, specific, sensitive, and reliable viral detection. In this paper, the development of molecular beacon assays as novel tools for the rapid detection of Aujeszky's disease virus, African swine fever virus, porcine circovirus type 2 and porcine parvovirus is described. The assays are capable of rapidly detecting 2 x 10(1) copies of target and are linear between 2 x 10(9) and 2 x 10(2) copies. They can detect virus specifically in clinical samples such as whole blood, serum and tissue. In comparison to conventional PCR they are either as sensitive or more sensitive. As such these molecular beacon assays represent a powerful tool for the detection of these viruses in swine.

  2. Detection of Multiple Potato Viruses in the Field Suggests Synergistic Interactions among Potato Viruses in Pakistan.

    PubMed

    Hameed, Amir; Iqbal, Zafar; Asad, Shaheen; Mansoor, Shahid

    2014-12-01

    Viral diseases have been a major limiting factor threating sustainable potato (Solanum tuberosum L.) production in Pakistan. Surveys were conducted to serologically quantify the incidence of RNA viruses infecting potato; Potato virus X (PVX), Potato virus Y (PVY), Potato virus S (PVS), Potato virus A (PVA), Potato virus M (PVM) and Potato leaf roll virus (PLRV) in two major potato cultivars (Desiree and Cardinal). The results suggest the prevalence of multiple viruses in all surveyed areas with PVY, PVS and PVX dominantly widespread with infection levels of up to 50% in some regions. Co-infections were detected with the highest incidence (15.5%) for PVX and PVS. Additionally the data showed a positive correlation between co-infecting viruses with significant increase in absorbance value (virus titre) for at least one of the virus in an infected plant and suggested a synergistic interaction. To test this hypothesis, glasshouse grown potato plants were challenged with multiple viruses and analyzed for systemic infections and symptomology studies. The results obtained conclude that multiple viral infections dramatically increase disease epidemics as compared to single infection and an effective resistance strategy in targeting multiple RNA viruses is required to save potato crop. PMID:25506305

  3. Microcavity single virus detection and sizing with molecular sensitivity

    NASA Astrophysics Data System (ADS)

    Dantham, V. R.; Holler, S.; Kolchenko, V.; Wan, Z.; Arnold, S.

    2013-02-01

    We report the label-free detection and sizing of the smallest individual RNA virus, MS2 by a spherical microcavity. Mass of this virus is ~6 ag and produces a theoretical resonance shift ~0.25 fm upon adsorbing an individual virus at the equator of the bare microcavity, which is well below the r.m.s background noise of 2 fm. However, detection was accomplished with ease (S/N = 8, Q = 4x105) using a single dipole stimulated plasmonic-nanoshell as a microcavity wavelength shift enhancer. Analytical expressions based on the "reactive sensing principle" are developed to extract the radius of the virus from the measured signals. Estimated limit of detection for these experiments was ~0.4 ag or 240 kDa below the size of all known viruses, largest globular and elongated proteins [Phosphofructokinase (345 kDa) and Fibrinogen (390 kDa), respectively].

  4. Detection and diagnosis of rice-infecting viruses

    PubMed Central

    Uehara-Ichiki, Tamaki; Shiba, Takuya; Matsukura, Keiichiro; Ueno, Takanori; Hirae, Masahiro; Sasaya, Takahide

    2013-01-01

    Rice-infecting viruses have caused serious damage to rice production in Asian, American, and African countries, where about 30 rice viruses and diseases have been reported. To control these diseases, developing accurate, quick methods to detect and diagnose the viruses in the host plants and any insect vectors of the viruses is very important. Based on an antigen–antibody reaction, serological methods such as latex agglutination reaction and enzyme-linked immunosorbent assay have advanced to detect viral particles or major proteins derived from viruses. They aid in forecasting disease and surveying disease spread and are widely used for virus detection at plant protection stations and research laboratories. From the early 2000s, based on sequence information for the target virus, several other methods such as reverse transcription-polymerase chain reaction (RT-PCR) and reverse transcription-loop-mediated isothermal amplification have been developed that are sensitive, rapid, and able to differentiate closely related viruses. Recent techniques such as real-time RT-PCR can be used to quantify the pathogen in target samples and monitor population dynamics of a virus, and metagenomic analyses using next-generation sequencing and microarrays show potential for use in the diagnosis of rice diseases. PMID:24130554

  5. Label-free virus detection using silicon photonic microring resonators

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Viruses represent a continual threat to humans through a number of mechanisms, which include disease, bioterrorism, and destruction of both plant and animal food resources. Many contemporary techniques used for the detection of viruses and viral infections suffer from limitations such as the need fo...

  6. Detection of enteric viruses in shellfish

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Norovirus and hepatitis A virus contamination are significant threats to the safety of shellfish and other foods. Methods for the extraction and assay of these viruses from shellfish are complex, time consuming, and technically challenging. Here, we itemize some of the salient points in extracting...

  7. Detection of transient and persistent feline leukaemia virus infections.

    PubMed

    Jarrett, O; Golder, M C; Stewart, M F

    1982-03-01

    A study was made of cats persistently or transiently viraemic with feline leukaemia virus (FeLV) following experimental oronasal infection. Cats of two ages were exposed to the virus. One group was infected when eight weeks old in the expectation that most of the cats would become persistently viraemic, and the second group when 16 weeks old, so that some would show signs of a transient infection and then recover. The periods following infection when virus was detectable in the blood and in the oropharynx were determined for each group. Three methods for detecting viraemia were compared: virus isolation, immunofluorescence on blood smears and an enzyme-linked immunosorbent assay (ELISA). There was good overall agreement among the three tests in detecting virus-positive cats. Virus was found sooner after infection by virus isolation than by the other methods, and virus appeared in the blood slightly sooner in cats which developed persistent viraemia than in transiently viraemic cats. Infectious FeLV was isolated from the oropharynx of all of the persistently viraemic cats, in most cases simultaneously with virus in the plasma. Virus was also isolated from the mouth of most transiently viraemic cats. Under field conditions such transient excretion of virus lasting only a few days would rarely be detected in a single sampling. This might explain how FeLV is maintained in free range urban cats in the absence of a large number of cats with persistent active FeLV infection. For routine diagnosis, immunofluorescence would appear to offer the best chance of differentiating transient and persistent infections by FeLV.

  8. Immunological-based assays for specific detection of shrimp viruses.

    PubMed

    Chaivisuthangkura, Parin; Longyant, Siwaporn; Sithigorngul, Paisarn

    2014-02-12

    Among shrimp viral pathogens, white spot syndrome virus (WSSV) and yellow head virus (YHV) are the most lethal agents, causing serious problems for both the whiteleg shrimp, Penaeus (Litopenaeus) vannamei, and the black tiger shrimp, Penaeus (Penaeus) monodon. Another important virus that infects P. vannamei is infectious myonecrosis virus (IMNV), which induces the white discoloration of affected muscle. In the cases of taura syndrome virus and Penaeus stylirostris densovirus (PstDNV; formerly known as infectious hypodermal and hematopoietic necrosis virus), their impacts were greatly diminished after the introduction of tolerant stocks of P. vannamei. Less important viruses are Penaeus monodon densovirus (PmDNV; formerly called hepatopancreatic parvovirus), and Penaeus monodon nucleopolyhedrovirus (PemoNPV; previously called monodon baculovirus). For freshwater prawn, Macrobrachium rosenbergii nodavirus and extra small virus are considered important viral pathogens. Monoclonal antibodies (MAbs) specific to the shrimp viruses described above have been generated and used as an alternative tool in various immunoassays such as enzyme-linked immunosorbent assay, dot blotting, Western blotting and immunohistochemistry. Some of these MAbs were further developed into immunochromatographic strip tests for the detection of WSSV, YHV, IMNV and PemoNPV and into a dual strip test for the simultaneous detection of WSSV/YHV. The strip test has the advantages of speed, as the result can be obtained within 15 min, and simplicity, as laboratory equipment and specialized skills are not required. Therefore, strip tests can be used by shrimp farmers for the pond-side monitoring of viral infection. PMID:24567913

  9. Immunological-based assays for specific detection of shrimp viruses

    PubMed Central

    Chaivisuthangkura, Parin; Longyant, Siwaporn; Sithigorngul, Paisarn

    2014-01-01

    Among shrimp viral pathogens, white spot syndrome virus (WSSV) and yellow head virus (YHV) are the most lethal agents, causing serious problems for both the whiteleg shrimp, Penaeus (Litopenaeus) vannamei, and the black tiger shrimp, Penaeus (Penaeus) monodon. Another important virus that infects P. vannamei is infectious myonecrosis virus (IMNV), which induces the white discoloration of affected muscle. In the cases of taura syndrome virus and Penaeus stylirostris densovirus (PstDNV; formerly known as infectious hypodermal and hematopoietic necrosis virus), their impacts were greatly diminished after the introduction of tolerant stocks of P. vannamei. Less important viruses are Penaeus monodon densovirus (PmDNV; formerly called hepatopancreatic parvovirus), and Penaeus monodon nucleopolyhedrovirus (PemoNPV; previously called monodon baculovirus). For freshwater prawn, Macrobrachium rosenbergii nodavirus and extra small virus are considered important viral pathogens. Monoclonal antibodies (MAbs) specific to the shrimp viruses described above have been generated and used as an alternative tool in various immunoassays such as enzyme-linked immunosorbent assay, dot blotting, Western blotting and immunohistochemistry. Some of these MAbs were further developed into immunochromatographic strip tests for the detection of WSSV, YHV, IMNV and PemoNPV and into a dual strip test for the simultaneous detection of WSSV/YHV. The strip test has the advantages of speed, as the result can be obtained within 15 min, and simplicity, as laboratory equipment and specialized skills are not required. Therefore, strip tests can be used by shrimp farmers for the pond-side monitoring of viral infection. PMID:24567913

  10. Detection and molecular characterization of Egyptian isolates of grapevine viruses.

    PubMed

    Fattouh, F; Ratti, C; El-Ahwany, A M D; Aleem, E Abdel; Babini, A R; Autonell, C Rubies

    2014-01-01

    Selected commercial and/or local vineyards and nurseries in three different governorates of Egypt (Alexandria, El-Beheira and El-Menofia) were surveyed for symptoms indicative of infection by grapevine viruses. Leaf samples from red-fruited and white-fruited Vitis vinefera were tested for grapevine leafroll associated viruses (GLRaV-1, GLRaV-2, and GLRaV-3), grapevine viruses A and B (GVA, GVB), grapevine rupestris stem pitting virus (GRSPaV), grapevine fanleaf virus (GFLV), and grapevine fleck virus (GFKV) from early April to late October 2010. Incidence of these viruses was assessed by RT-PCR in 60 different samples. Selected amplicons were sequenced. While GVA was the most wide spread (30%), GLRaV-1, GVB, GFLV, and GFKV were not detected during the survey. However, GVA, GLRaV-2, GLRaV-3, and GRSPaV were detected in the form of single infection or in mixed infections of 2 to 4 viruses. Phylogenetic analysis was performed on all Egyptian isolates of GLRaV-2 (4), GLRaV-3 (7), GVA (3), and GRSPaV (6). GRSPaV was detected for the first time in Egypt. Phylogenetic analysis provided insights into the evolutionary relationship between the reported Egyptian isolates and other previously reported isolates. PMID:24957718

  11. Detection of dengue virus type 4 in Easter Island, Chile.

    PubMed

    Fernández, J; Vera, L; Tognarelli, J; Fasce, R; Araya, P; Villagra, E; Roos, O; Mora, J

    2011-10-01

    We report the detection of dengue virus type 4 (DENV-4) for the first time in Easter Island, Chile. The virus was detected in serum samples of two patients treated at the Hospital in Easter Island. The two samples were IgM positive, and the infection was confirmed by RT-PCR and genetic sequencing; viral isolation was possible with one of them. The Easter Island isolates were most closely related to genotype II of dengue type 4.

  12. Spirometry filters can be used to detect exhaled respiratory viruses.

    PubMed

    Mitchell, Alicia B; Mourad, Bassel; Tovey, Euan; Buddle, Lachlan; Peters, Matthew; Morgan, Lucy; Oliver, Brian G

    2016-09-26

    Respiratory viruses are very common in the community and contribute to the burden of illness for patients with chronic respiratory diseases, including acute exacerbations. Traditional sampling methods are invasive and problematic to repeat. Accordingly, we explored whether respiratory viruses could be isolated from disposable spirometry filters and whether detection of viruses in this context represented presence in the upper or lower respiratory tract. Discovery (n  =  53) and validation (n  =  49) cohorts were recruited from a hospital outpatient department during two different time periods. Spirometry mouthpiece filters were collected from all participants. Respiratory secretions were sampled from the upper and lower respiratory tract by nasal washing (NW), sputum, and bronchoalveolar lavage (BAL). All samples were examined using RT-PCR to identify a panel of respiratory viruses (rhinovirus, respiratory syncytial virus, influenza A, influenza B, parainfluenza virus 1, 2 & 3, and human metapneumovirus). Rhinovirus was quantified using qPCR. Paired filter-NW samples (n  =  29), filter-sputum samples (n  =  24), filter-BAL samples (n  =  39) and filter-NW-BAL samples (n  =  10) provided a range of comparisons. At least one virus was detected in any sample in 85% of participants in the discovery cohort versus 45% in the validation cohort. Overall, 72% of viruses identified in the paired comparator method matched those detected in spirometry filters. There was a high correlation between viruses identified in spirometry filters compared with viruses identified in both the upper and lower respiratory tract using traditional sampling methods. Our results suggest that examination of spirometry filters may be a novel and inexpensive sampling method for the presence of respiratory viruses in exhaled breath.

  13. Spirometry filters can be used to detect exhaled respiratory viruses.

    PubMed

    Mitchell, Alicia B; Mourad, Bassel; Tovey, Euan; Buddle, Lachlan; Peters, Matthew; Morgan, Lucy; Oliver, Brian G

    2016-01-01

    Respiratory viruses are very common in the community and contribute to the burden of illness for patients with chronic respiratory diseases, including acute exacerbations. Traditional sampling methods are invasive and problematic to repeat. Accordingly, we explored whether respiratory viruses could be isolated from disposable spirometry filters and whether detection of viruses in this context represented presence in the upper or lower respiratory tract. Discovery (n  =  53) and validation (n  =  49) cohorts were recruited from a hospital outpatient department during two different time periods. Spirometry mouthpiece filters were collected from all participants. Respiratory secretions were sampled from the upper and lower respiratory tract by nasal washing (NW), sputum, and bronchoalveolar lavage (BAL). All samples were examined using RT-PCR to identify a panel of respiratory viruses (rhinovirus, respiratory syncytial virus, influenza A, influenza B, parainfluenza virus 1, 2 & 3, and human metapneumovirus). Rhinovirus was quantified using qPCR. Paired filter-NW samples (n  =  29), filter-sputum samples (n  =  24), filter-BAL samples (n  =  39) and filter-NW-BAL samples (n  =  10) provided a range of comparisons. At least one virus was detected in any sample in 85% of participants in the discovery cohort versus 45% in the validation cohort. Overall, 72% of viruses identified in the paired comparator method matched those detected in spirometry filters. There was a high correlation between viruses identified in spirometry filters compared with viruses identified in both the upper and lower respiratory tract using traditional sampling methods. Our results suggest that examination of spirometry filters may be a novel and inexpensive sampling method for the presence of respiratory viruses in exhaled breath. PMID:27669334

  14. Using Small RNA Deep Sequencing Data to Detect Human Viruses.

    PubMed

    Wang, Fang; Sun, Yu; Ruan, Jishou; Chen, Rui; Chen, Xin; Chen, Chengjie; Kreuze, Jan F; Fei, ZhangJun; Zhu, Xiao; Gao, Shan

    2016-01-01

    Small RNA sequencing (sRNA-seq) can be used to detect viruses in infected hosts without the necessity to have any prior knowledge or specialized sample preparation. The sRNA-seq method was initially used for viral detection and identification in plants and then in invertebrates and fungi. However, it is still controversial to use sRNA-seq in the detection of mammalian or human viruses. In this study, we used 931 sRNA-seq runs of data from the NCBI SRA database to detect and identify viruses in human cells or tissues, particularly from some clinical samples. Six viruses including HPV-18, HBV, HCV, HIV-1, SMRV, and EBV were detected from 36 runs of data. Four viruses were consistent with the annotations from the previous studies. HIV-1 was found in clinical samples without the HIV-positive reports, and SMRV was found in Diffuse Large B-Cell Lymphoma cells for the first time. In conclusion, these results suggest the sRNA-seq can be used to detect viruses in mammals and humans. PMID:27066498

  15. Using Small RNA Deep Sequencing Data to Detect Human Viruses

    PubMed Central

    Wang, Fang; Sun, Yu; Ruan, Jishou; Chen, Rui; Chen, Xin; Chen, Chengjie; Kreuze, Jan F.; Fei, ZhangJun; Zhu, Xiao

    2016-01-01

    Small RNA sequencing (sRNA-seq) can be used to detect viruses in infected hosts without the necessity to have any prior knowledge or specialized sample preparation. The sRNA-seq method was initially used for viral detection and identification in plants and then in invertebrates and fungi. However, it is still controversial to use sRNA-seq in the detection of mammalian or human viruses. In this study, we used 931 sRNA-seq runs of data from the NCBI SRA database to detect and identify viruses in human cells or tissues, particularly from some clinical samples. Six viruses including HPV-18, HBV, HCV, HIV-1, SMRV, and EBV were detected from 36 runs of data. Four viruses were consistent with the annotations from the previous studies. HIV-1 was found in clinical samples without the HIV-positive reports, and SMRV was found in Diffuse Large B-Cell Lymphoma cells for the first time. In conclusion, these results suggest the sRNA-seq can be used to detect viruses in mammals and humans. PMID:27066498

  16. EPA METHODS FOR VIRUS DETECTION IN WATER

    EPA Science Inventory

    A number of different types of human enteric viruses cause waterborne outbreaks when individuals are exposed to contaminated drinking and recreational waters. Members of the enterovirus group cause numerous diseases, including gastroenteritis, encephalitis, meningitis, myocard...

  17. Molecular beacons: a new approach to plant virus detection.

    PubMed

    Eun, A J; Wong, S M

    2000-03-01

    ABSTRACT Molecular beacons are single-stranded nucleic acid molecules with a stem-loop conformation. The stem portion consists of complementary sequences at the 5' and 3' terminals of the molecule, while the loop portion consists of probe sequences that are complementary to the target sequences of choice. A fluorescent moiety is attached to one end, while a quenching moiety is attached to the opposite end. Reverse transcription-polymerase chain reactions are carried out with primers that amplify specific genome sequences of interest, yielding targets complementary to their respective molecular beacons for subsequent detection. Here, we have designed four molecular beacons specific to the RNA-dependent RNA polymerase and coat protein genes of two orchid viruses, namely Cymbidium mosaic virus (CymMV) and Odontoglossum ringspot virus (ORSV). This technology is successfully applied to detect as little as 0.5 ng of viral RNA of both orchid viruses simultaneously in 100 mg of coinfected Oncidium orchid leaves. This rapid and specific technique is applicable to the orchid industry, which routinely carries out virus indexing and screening for virus-resistant cultivars. We belief that use of this molecular beacon approach can be extended to the detection of multiple plant viruses in various crops.

  18. Controversial results of therapy with mesenchymal stem cells in the acute phase of canine distemper disease.

    PubMed

    Pinheiro, A O; Cardoso, M T; Vidane, A S; Casals, J B; Passarelli, D; Alencar, A L F; Sousa, R L M; Fantinato-Neto, P; Oliveira, V C; Lara, V M; Ambrósio, C E

    2016-05-23

    Distemper disease is an infectious disease reported in several species of domestic and wild carnivores. The high mortality rate of animals infected with canine distemper virus (CDV) treated with currently available therapies has driven the study of new efficacious treatments. Mesenchymal stem cell (MSC)-based therapy is a promising therapeutic option for many degenerative, hereditary, and inflammatory diseases. Therefore, the aim of this study was to characterize stem cells derived from the canine fetal olfactory epithelium and to assess the systemic response of animals infected with CDV to symptomatic therapy and treatment with MSCs. Eight domestic mongrel dogs (N = 8) were divided into two groups: support group (SG) (N = 5) and support group + cell therapy (SGCT) (N = 3), which were monitored over 15 days. Blood samples were collected on days 0, 6, 9, 12, and 15 to assess blood count and serum biochemistry (urea, creatinine, alanine transferase, alkaline phosphatase, gamma-glutamyl transferase, total protein, albumin, and globulin), and urine samples were obtained on days 0 and 15 for urinary evaluation (urine I). The results showed a high mortality rate (SG = 4 and SGCT = 2), providing inadequate data on the clinical course of CDV infection. MSC therapy resulted in no significant improvement when administered during the acute phase of canine distemper disease, and a prevalence of animals with high mortality rate was found in both groups due to the severity of symptoms.

  19. The 2000 canine distemper epidemic in Caspian seals (Phoca caspica): pathology and analysis of contributory factors.

    PubMed

    Kuiken, T; Kennedy, S; Barrett, T; Van de Bildt, M W G; Borgsteede, F H; Brew, S D; Codd, G A; Duck, C; Deaville, R; Eybatov, T; Forsyth, M A; Foster, G; Jepson, P D; Kydyrmanov, A; Mitrofanov, I; Ward, C J; Wilson, S; Osterhaus, A D M E

    2006-05-01

    More than 10,000 Caspian seals (Phoca caspica) were reported dead in the Caspian Sea during spring and summer 2000. We performed necropsies and extensive laboratory analyses on 18 seals, as well as examination of the pattern of strandings and variation in weather in recent years, to identify the cause of mortality and potential contributory factors. The monthly stranding rate in 2000 was up to 2.8 times the historic mean. It was preceded by an unusually mild winter, as observed before in mass mortality events of pinnipeds. The primary diagnosis in 11 of 13 seals was canine distemper, characterized by broncho-interstitial pneumonia, lymphocytic necrosis and depletion in lymphoid organs, and the presence of typical intracytoplasmic inclusion bodies in multiple epithelia. Canine distemper virus infection was confirmed by phylogenetic analysis of reverse transcriptase-polymerase chain reaction products. Organochlorine and zinc concentrations in tissues of seals with canine distemper were comparable to those of Caspian seals in previous years. Concurrent bacterial infections that may have contributed to the mortality of the seals included Bordetella bronchiseptica (4/8 seals), Streptococcus phocae (3/8), Salmonella dublin (1/8), and S. choleraesuis (1/8). A newly identified bacterium, Corynebacterium caspium, was associated with balanoposthitis in one seal. Several infectious and parasitic organisms, including poxvirus, Atopobacter phocae, Eimeria- and Sarcocystis-like organisms, and Halarachne sp. were identified in Caspian seals for the first time. PMID:16672579

  20. Controversial results of therapy with mesenchymal stem cells in the acute phase of canine distemper disease.

    PubMed

    Pinheiro, A O; Cardoso, M T; Vidane, A S; Casals, J B; Passarelli, D; Alencar, A L F; Sousa, R L M; Fantinato-Neto, P; Oliveira, V C; Lara, V M; Ambrósio, C E

    2016-01-01

    Distemper disease is an infectious disease reported in several species of domestic and wild carnivores. The high mortality rate of animals infected with canine distemper virus (CDV) treated with currently available therapies has driven the study of new efficacious treatments. Mesenchymal stem cell (MSC)-based therapy is a promising therapeutic option for many degenerative, hereditary, and inflammatory diseases. Therefore, the aim of this study was to characterize stem cells derived from the canine fetal olfactory epithelium and to assess the systemic response of animals infected with CDV to symptomatic therapy and treatment with MSCs. Eight domestic mongrel dogs (N = 8) were divided into two groups: support group (SG) (N = 5) and support group + cell therapy (SGCT) (N = 3), which were monitored over 15 days. Blood samples were collected on days 0, 6, 9, 12, and 15 to assess blood count and serum biochemistry (urea, creatinine, alanine transferase, alkaline phosphatase, gamma-glutamyl transferase, total protein, albumin, and globulin), and urine samples were obtained on days 0 and 15 for urinary evaluation (urine I). The results showed a high mortality rate (SG = 4 and SGCT = 2), providing inadequate data on the clinical course of CDV infection. MSC therapy resulted in no significant improvement when administered during the acute phase of canine distemper disease, and a prevalence of animals with high mortality rate was found in both groups due to the severity of symptoms. PMID:27323085

  1. Single virus particle mass detection using microresonators with nanoscale thickness

    NASA Astrophysics Data System (ADS)

    Gupta, A.; Akin, D.; Bashir, R.

    2004-03-01

    In this letter, we present the microfabrication and application of arrays of silicon cantilever beams as microresonator sensors with nanoscale thickness to detect the mass of individual virus particles. The dimensions of the fabricated cantilever beams were in the range of 4-5 μm in length, 1-2 μm in width and 20-30 nm in thickness. The virus particles we used in the study were vaccinia virus, which is a member of the Poxviridae family and forms the basis of the smallpox vaccine. The frequency spectra of the cantilever beams, due to thermal and ambient noise, were measured using a laser Doppler vibrometer under ambient conditions. The change in resonant frequency as a function of the virus particle mass binding on the cantilever beam surface forms the basis of the detection scheme. We have demonstrated the detection of a single vaccinia virus particle with an average mass of 9.5 fg. These devices can be very useful as components of biosensors for the detection of airborne virus particles.

  2. Label-free virus detection using silicon photonic microring resonators.

    PubMed

    McClellan, Melinda S; Domier, Leslie L; Bailey, Ryan C

    2012-01-15

    Viruses represent a continual threat to humans through a number of mechanisms, which include disease, bioterrorism, and destruction of both plant and animal food resources. Many contemporary techniques used for the detection of viruses and viral infections suffer from limitations such as the need for extensive sample preparation or the lengthy window between infection and measurable immune response, for serological methods. In order to develop a method that is fast, cost-effective, and features reduced sample preparation compared to many other virus detection methods, we report the application of silicon photonic microring resonators for the direct, label-free detection of intact viruses in both purified samples as well as in a complex, real-world analytical matrix. As a model system, we demonstrate the quantitative detection of Bean pod mottle virus, a pathogen of great agricultural importance, with a limit of detection of 10 ng/mL. By simply grinding a small amount of leaf sample in buffer with a mortar and pestle, infected leaves can be identified over a healthy control with a total analysis time of less than 45 min. Given the inherent scalability and multiplexing capability of the semiconductor-based technology, we feel that silicon photonic microring resonators are well-positioned as a promising analytical tool for a number of viral detection applications. PMID:22138465

  3. Integrated reverse transcription polymerase chain reaction systems for virus detection.

    PubMed

    Lien, Kang-Yi; Lee, Wan-Chi; Lei, Huan-Yao; Lee, Gwo-Bin

    2007-03-15

    The current study reports on an integrated microreverse transcription polymerase chain reaction (RT-PCR) system for molecular diagnosis of microorganisms automatically. By using antibodies-conjugated superparamagnetic beads, the developed system can detect viruses with higher sensitivity and specificity when compared with traditional biological diagnosis methods using a ribonucleic acid (RNA) extraction kit. The target viruses were first captured by the conjugated antibodies on the magnetic beads, and were enriched using a magnetic field generated by micro-electromagnets or permanent magnets. With this approach, the virus can be purified and concentrated first, then the virus RNA was extracted and transcripted to complementary deoxyribonucleic acid (cDNA), followed by a nucleic acid amplification process using a micro-RT-PCR module. The integrated microfluidic chip can perform the whole process automatically with the aid of integrated micropumps and microvalves. This study successfully performs the specific detection of two different types of viruses, Dengue virus serotype 2 and enterovirus (EV) 71 using this developed integrated system. Comparable to a large-scale apparatus, the integrated microsystem can perform mixing, incubation, purification, transportation, and nucleic acid amplification of virus, possibly making it a crucial platform for future diagnosis applications.

  4. Simultaneous detection of major pome fruit viruses and a viroid.

    PubMed

    Kumar, Surender; Singh, Lakhmir; Ram, Raja; Zaidi, Aijaz A; Hallan, Vipin

    2014-06-01

    A rapid and sensitive two-step RT-PCR protocol for simultaneous detection of major apple viruses, namely Apple mosaic virus (ApMV), Apple stem pitting virus (ASPV), Apple stem grooving virus (ASGV), Apple chlorotic leaf spot virus (ACLSV) and Apple scar skin viroid (ASSVd), was developed. Five specific primer pairs were tested and confirmed for these viruses and viroid together in a single tube, giving amplicons of ~198, ~330, ~370, ~547 and ~645 bp corresponding to ASGV, ASSVd, ASPV, ApMV and ACLSV, respectively. Using a guanidinium-based extraction buffer along with a commercial kit resulted in better quality RNA as compared to kit, suited for multiplex RT-PCR. A rapid CTAB method for RNA isolation from apple tissue was developed, which produce good yield and saves time. To the best of our knowledge, this is the first report on the simultaneous detection of five pathogens (four viruses and a viroid) from apple with NADH dehydrogenase subunit 5 (nad5) as an internal control.

  5. Laboratory Detection of Respiratory Viruses by Automated Techniques

    PubMed Central

    Pérez-Ruiz, Mercedes; Pedrosa-Corral, Irene; Sanbonmatsu-Gámez, Sara; Navarro-Marí, José-María

    2012-01-01

    Advances in clinical virology for detecting respiratory viruses have been focused on nucleic acids amplification techniques, which have converted in the reference method for the diagnosis of acute respiratory infections of viral aetiology. Improvements of current commercial molecular assays to reduce hands-on-time rely on two strategies, a stepwise automation (semi-automation) and the complete automation of the whole procedure. Contributions to the former strategy have been the use of automated nucleic acids extractors, multiplex PCR, real-time PCR and/or DNA arrays for detection of amplicons. Commercial fully-automated molecular systems are now available for the detection of respiratory viruses. Some of them could convert in point-of-care methods substituting antigen tests for detection of respiratory syncytial virus and influenza A and B viruses. This article describes laboratory methods for detection of respiratory viruses. A cost-effective and rational diagnostic algorithm is proposed, considering technical aspects of the available assays, infrastructure possibilities of each laboratory and clinic-epidemiologic factors of the infection PMID:23248735

  6. An integrated microfluidic system using magnetic beads for virus detection.

    PubMed

    Lee, Wan-Chi; Lien, Kang-Yi; Lee, Gwo-Bin; Lei, Huan-Yao

    2008-01-01

    An integrated system capable of sample pretreatment using antibody-conjugated magnetic beads and one-step reverse transcriptase-polymerase chain reaction (RT-PCR) on a microfluidic system was developed to accelerate the detection of RNA viruses such as dengue virus or enterovirus 71. The targeted virus in the sample was first captured by the specific antibody-conjugated magnetic beads, which were manipulated by micro-electromagnets made of micro-electro-mechanical systems technology. The RNA of the targeted virus then underwent thermal lysis and was reverse-transcripted to cDNA using a microRT-PCR module. The sensitivity to detect dengue virus is around 10-100 PFU, which is equivalent to the commercial RNA extraction kit and a large-scale RT-PCR machine. This microsystem can specifically detect 4 serotypes of dengue virus, as well as enterovirus 71. The specificity was warranted by both antibody and primer. The microfluidic system allows automatic process of sample including mixing, incubation, and reaction. The antibody-conjugated magnetic beads offer sample pretreatment of purification and concentration. The integration of antibody-conjugated magnetic beads into the microfluidic system is promising for fast molecular diagnosis of microorganisms.

  7. Rapid Detection and Identification of Respiratory Viruses by Direct Immunofluorescence

    PubMed Central

    D'Alessio, Donn; Williams, Stanley; Dick, Elliot C.

    1970-01-01

    The use of fluorescein-conjugated antiserum against respiratory syncytial (RS) and parainfluenza 1 and 3 viruses was compared with conventional techniques in the rapid detection of virus in tissue cultures inoculated with pharyngeal specimens known to contain these viruses. Twenty-three specimens were tested: 9 RS, 8 parainfluenza 1, and 6 parainfluenza 3. The fluorescent-antibody technique (FA) detected virus in 52% of the tissue cultures in 24 hr, and, by 72 hr, 22 of the 23 cultures were FA-positive whereas only 5 were positive by conventional techniques. Additionally, conjugated antisera were prepared against herpes simplex, influenza A2, and adenovirus type 5. All conjugates stained only the homologous virus and were 100- to 10,000-fold more sensitive than conventional techniques in detecting descending dilutions of virus inocula by 24 hr. With the procedures described, several antisera could be conjugated and ready for use within 24 hr. Serum fractionation was by ammonium sulfate precipitation, and with the procedure outlined virtually complete recovery of the globulin fraction and elimination of all of the albumin were accomplished. Images PMID:4098101

  8. Detection of bovine viral diarrhea virus (BVDV) in seropositive cattle.

    PubMed

    Gogorza, L M; Morán, P E; Larghi, J L; Seguí, R; Lissarrague, C; Saracco, M; Braun, M; Esteban, E N

    2005-11-15

    Detection of bovine virus diarrhoea virus (BVDV) in one vaccinated beef cattle and three non-vaccinated dairy herds was investigated on peripheral blood leukocytes (PBL) with or without previous treatment followed by a capture ELISA (cELISA). Using the combination of PHA and polycation treatment, PBL from 229 seropositive cattle were studied and could be classified in four different states of BVDV infection. Lysed PBL from four animals were directly positive in cELISA (Category I), PBL of 17 animals were positive after PHA stimulation (Category II), 15 animals were positive only after PHA stimulation plus polycation treatment (Category III), while virus could not be detected in 193 seropositive cattle. Wild-type BVDV strains were isolated by co-culture on polycation-treated MDBK cells from 11 of these seropositive animals. BVDV antibodies of these same animals were able to neutralize their own virus, indicating that virus persists in PBL in spite of strain-specific antibodies. No apparent change of leukocyte subpopulations could be detected in any category of virus-positive animals. Thus, BVDV may be present in the PBL of some cattle, even in the presence of a specific active immune response.

  9. New viruses in veterinary medicine, detected by metagenomic approaches.

    PubMed

    Belák, Sándor; Karlsson, Oskar E; Blomström, Anne-Lie; Berg, Mikael; Granberg, Fredrik

    2013-07-26

    In our world, which is faced today with exceptional environmental changes and dramatically intensifying globalisation, we are encountering challenges due to many new factors, including the emergence or re-emergence of novel, so far "unknown" infectious diseases. Although a broad arsenal of diagnostic methods is at our disposal, the majority of the conventional diagnostic tests is highly virus-specific or is targeted entirely towards a limited group of infectious agents. This specificity complicates or even hinders the detection of new or unexpected pathogens, such as new, emerging or re-emerging viruses or novel viral variants. The recently developed approaches of viral metagenomics provide an effective novel way to screen samples and detect viruses without previous knowledge of the infectious agent, thereby enabling a better diagnosis and disease control, in line with the "One World, One Health" principles (www.oneworldonehealth.org). Using metagenomic approaches, we have recently identified a broad variety of new viruses, such as novel bocaviruses, Torque Teno viruses, astroviruses, rotaviruses and kobuviruses in porcine disease syndromes, new virus variants in honeybee populations, as well as a range of other infectious agents in further host species. These findings indicate that the metagenomic detection of viral pathogens is becoming now a powerful, cultivation-independent, and useful novel diagnostic tool in veterinary diagnostic virology. PMID:23428379

  10. New viruses in veterinary medicine, detected by metagenomic approaches.

    PubMed

    Belák, Sándor; Karlsson, Oskar E; Blomström, Anne-Lie; Berg, Mikael; Granberg, Fredrik

    2013-07-26

    In our world, which is faced today with exceptional environmental changes and dramatically intensifying globalisation, we are encountering challenges due to many new factors, including the emergence or re-emergence of novel, so far "unknown" infectious diseases. Although a broad arsenal of diagnostic methods is at our disposal, the majority of the conventional diagnostic tests is highly virus-specific or is targeted entirely towards a limited group of infectious agents. This specificity complicates or even hinders the detection of new or unexpected pathogens, such as new, emerging or re-emerging viruses or novel viral variants. The recently developed approaches of viral metagenomics provide an effective novel way to screen samples and detect viruses without previous knowledge of the infectious agent, thereby enabling a better diagnosis and disease control, in line with the "One World, One Health" principles (www.oneworldonehealth.org). Using metagenomic approaches, we have recently identified a broad variety of new viruses, such as novel bocaviruses, Torque Teno viruses, astroviruses, rotaviruses and kobuviruses in porcine disease syndromes, new virus variants in honeybee populations, as well as a range of other infectious agents in further host species. These findings indicate that the metagenomic detection of viral pathogens is becoming now a powerful, cultivation-independent, and useful novel diagnostic tool in veterinary diagnostic virology.

  11. Dog distemper: imported into Europe from South America?.

    PubMed

    Blancou, Jean

    2004-01-01

    According to Charles Frédéric Heusinger (1853), dog distemper had been imported from Peru into Spain in the course of the 17th century. The disease was well described in 1746 by Ulloa in his work Relación histórica del viaje a la América meridional. During the course of the 1760s, the disease was reported in Spain, followed by England, Italy (1764) and Russia (1770). In 1763, 900 dogs died in a single day in Madrid. In 1844, Karle succeeded in the first experimental transmission of the disease by brushing the lips of young dogs with the discharge from sick animals. The causal agent of the disease was only discovered in 1905, when the virus was isolated by Henri Carré. In the meantime, Edward Jenner, who thought that the disease was a pox-like affection, claimed that it could be prevented by inoculation of the vaccinia virus. PMID:15376360

  12. A SYBR Green I based real time RT-PCR assay for specific detection and quantitation of Peste des petits ruminants virus

    PubMed Central

    2014-01-01

    Background Peste des petits ruminants (PPR) is an economically important disease of small ruminants such as sheep and goats. The disease is characterized by severe pyrexia, oculo-nasal discharge, pneumonia, necrosis and ulceration of the mucous membrane and inflammation of the gastro-intestinal tract leading to severe diarrhea. A SYBR Green I based real time RT-PCR targeting the N gene of PPRV has not been established for PPRV detection. Thus, the objective of present study was to develop highly sensitive N gene target SYBR Green I real time RT-PCR for specific detection and quantification of PPRV in clinical samples. A set of primers was designed to detect the nucleocapsid (N) gene of PPRV. Results The assay exhibited high specificity as all the viruses which have clinical and structural similarities to PPRV including Canine distemper virus (CDV), Measles virus (MV), Bluetongue virus (BTV) and Newcastle disease virus (NDV) failed to show an amplification signal. The lower detection limit of the assay was 5.11 copies/μl (Ct value of 33.67 ± 0.5) and 0.001 TCID50/ml (Ct value of 34.7 ± 0.5) based on plasmid copy number and tissue culture infectivity titre. The assay was 3-log more sensitive than the conventional RT-PCR. The coefficient of variation (CV) values for intra- and inter-assay variability were low, ranging from 0.32% - 2.31%, and 0.71% - 5.32%, respectively. To evaluate the performance of the newly developed assay, a total of 36 clinical samples suspected of PPR were screened for the presence of PPRV in parallel with conventional RT-PCR. The real time RT-PCR assay detected PPRV in 30 (83.3%) of clinical samples compared to 16 (44.4%) by conventional RT-PCR. Conclusions The two-step SYBR Green I based real time RT-PCR assay reported here is highly sensitive, specific, reproducible and rapid for detection and quantification of PPRV nucleic acids. PMID:24423231

  13. Rapid and selective detection of viruses using virus-imprinted polymer films.

    PubMed

    Karthik, A; Margulis, K; Ren, K; Zare, R N; Leung, L W

    2015-12-01

    We prepared a nanopatterned polymer film of polydimethylsiloxane (PDMS) via virus imprinting. The imprinted surface exhibited nanoscale cavities with the mean size of 120 ± 4 nm. These cavities demonstrated the ability to preferentially capture a target virus from an aqueous suspension of ultralow volume (5 μL) after only 1 minute of contact. Two inactivated viruses with similar shape, Influenza A (HK68) and Newcastle Disease Virus (NDV), were employed as model pathogens. The polymer film, which was first imprinted with HK68 and exposed sequentially to suspensions containing fluorescently labeled NDV and HK68, was able to preferentially bind HK68 at a capture ratio of 1 : 8.0. When we reversed the procedure and imprinted with NDV, the capture ratio was 1 : 7.6. These results were obtained within 20 minutes of static exposure. The suspensions contained viruses at concentrations close to those occurring physiologically in influenza infections. The limit of detection was approximately 8 fM. Production of virus-imprinted films can be readily scaled to large quantities and yields a disposable, simple-to-use device that allows for rapid detection of viruses.

  14. Rapid and selective detection of viruses using virus-imprinted polymer films

    NASA Astrophysics Data System (ADS)

    Karthik, A.; Margulis, K.; Ren, K.; Zare, R. N.; Leung, L. W.

    2015-11-01

    We prepared a nanopatterned polymer film of polydimethylsiloxane (PDMS) via virus imprinting. The imprinted surface exhibited nanoscale cavities with the mean size of 120 +/- 4 nm. These cavities demonstrated the ability to preferentially capture a target virus from an aqueous suspension of ultralow volume (5 μL) after only 1 minute of contact. Two inactivated viruses with similar shape, Influenza A (HK68) and Newcastle Disease Virus (NDV), were employed as model pathogens. The polymer film, which was first imprinted with HK68 and exposed sequentially to suspensions containing fluorescently labeled NDV and HK68, was able to preferentially bind HK68 at a capture ratio of 1 : 8.0. When we reversed the procedure and imprinted with NDV, the capture ratio was 1 : 7.6. These results were obtained within 20 minutes of static exposure. The suspensions contained viruses at concentrations close to those occurring physiologically in influenza infections. The limit of detection was approximately 8 fM. Production of virus-imprinted films can be readily scaled to large quantities and yields a disposable, simple-to-use device that allows for rapid detection of viruses.

  15. Concurrent detection of other respiratory viruses in children shedding viable human respiratory syncytial virus.

    PubMed

    Gagliardi, T B; Paula, F E; Iwamoto, M A; Proença-Modena, J L; Santos, A E; Camara, A A; Cervi, M C; Cintra, O A L; Arruda, E

    2013-10-01

    Human respiratory syncytial virus (HRSV) is an important cause of respiratory disease. The majority of studies addressing the importance of virus co-infections to the HRSV-disease have been based on the detection of HRSV by RT-PCR, which may not distinguish current replication from prolonged shedding of remnant RNA from previous HRSV infections. To assess whether co-detections of other common respiratory viruses are associated with increased severity of HRSV illnesses from patients who were shedding viable-HRSV, nasopharyngeal aspirates from children younger than 5 years who sought medical care for respiratory infections in Ribeirão Preto (Brazil) were tested for HRSV by immunofluorescence, RT-PCR and virus isolation in cell culture. All samples with viable-HRSV were tested further by PCR for other respiratory viruses. HRSV-disease severity was assessed by a clinical score scale. A total of 266 samples from 247 children were collected and 111 (42%) were HRSV-positive. HRSV was isolated from 70 (63%), and 52 (74%) of them were positive for at least one additional virus. HRSV-positive diseases were more severe than HRSV-negative ones, but there was no difference in disease severity between patients with viable-HRSV and those HRSV-positives by RT-PCR. Co-detection of other viruses did not correlate with increased disease severity. HRSV isolation in cell culture does not seem to be superior to RT-PCR to distinguish infections associated with HRSV replication in studies of clinical impact of HRSV. A high rate of co-detection of other respiratory viruses was found in samples with viable-HRSV, but this was not associated with more severe HRSV infection.

  16. Easy and Rapid Detection of Mumps Virus by Live Fluorescent Visualization of Virus-Infected Cells.

    PubMed

    Takahashi, Tadanobu; Agarikuchi, Takashi; Kurebayashi, Yuuki; Shibahara, Nona; Suzuki, Chihiro; Kishikawa, Akiko; Fukushima, Keijo; Takano, Maiko; Suzuki, Fumie; Wada, Hirohisa; Otsubo, Tadamune; Ikeda, Kiyoshi; Minami, Akira; Suzuki, Takashi

    2015-01-01

    Mumps viruses show diverse cytopathic effects (CPEs) of infected cells and viral plaque formation (no CPE or no plaque formation in some cases) depending on the viral strain, highlighting the difficulty in mumps laboratory studies. In our previous study, a new sialidase substrate, 2-(benzothiazol-2-yl)-4-bromophenyl 5-acetamido-3,5-dideoxy-α-D-glycero-D-galacto-2-nonulopyranosidonic acid (BTP3-Neu5Ac), was developed for visualization of sialidase activity. BTP3-Neu5Ac can easily and rapidly perform histochemical fluorescent visualization of influenza viruses and virus-infected cells without an antiviral antibody and cell fixation. In the present study, the potential utility of BTP3-Neu5Ac for rapid detection of mumps virus was demonstrated. BTP3-Neu5Ac could visualize dot-blotted mumps virus, virus-infected cells, and plaques (plaques should be called focuses due to staining of infected cells in this study), even if a CPE was not observed. Furthermore, virus cultivation was possible by direct pick-up from a fluorescent focus. In conventional methods, visible appearance of the CPE and focuses often requires more than 6 days after infection, but the new method with BTP3-Neu5Ac clearly visualized infected cells after 2 days and focuses after 4 days. The BTP3-Neu5Ac assay is a precise, easy, and rapid assay for confirmation and titration of mumps virus.

  17. Detection of Entebbe Bat Virus After 54 Years.

    PubMed

    Kading, Rebekah C; Kityo, Robert; Nakayiki, Teddie; Ledermann, Jeremy; Crabtree, Mary B; Lutwama, Julius; Miller, Barry R

    2015-09-01

    Entebbe bat virus (ENTV; Flaviviridae: Flavivirus), closely related to yellow fever virus, was first isolated from a little free-tailed bat (Chaerephon pumilus) in Uganda in 1957, but was not detected after that initial isolation. In 2011, we isolated ENTV from a little free-tailed bat captured from the attic of a house near where it had originally been found. Infectious virus was recovered from the spleen and lung, and the viral RNA was sequenced and compared with that of the original isolate. Across the polypeptide sequence, there were 76 amino acid substitutions, resulting in 97.8% identity at the amino acid level between the 1957 and 2011 isolates. Further study of this virus would provide valuable insights into the ecological and genetic factors governing the evolution and transmission of bat- and mosquito-borne flaviviruses. PMID:26101270

  18. Detection of Entebbe Bat Virus after 54 Years

    PubMed Central

    Kading, Rebekah C.; Kityo, Robert; Nakayiki, Teddie; Ledermann, Jeremy; Crabtree, Mary B.; Lutwama, Julius; Miller, Barry R.

    2015-01-01

    Entebbe bat virus (ENTV; Flaviviridae: Flavivirus), closely related to yellow fever virus, was first isolated from a little free-tailed bat (Chaerephon pumilus) in Uganda in 1957, but was not detected after that initial isolation. In 2011, we isolated ENTV from a little free-tailed bat captured from the attic of a house near where it had originally been found. Infectious virus was recovered from the spleen and lung, and the viral RNA was sequenced and compared with that of the original isolate. Across the polypeptide sequence, there were 76 amino acid substitutions, resulting in 97.8% identity at the amino acid level between the 1957 and 2011 isolates. Further study of this virus would provide valuable insights into the ecological and genetic factors governing the evolution and transmission of bat- and mosquito-borne flaviviruses. PMID:26101270

  19. Single-Reaction Multiplex Reverse Transcription PCR for Detection of Zika, Chikungunya, and Dengue Viruses

    PubMed Central

    Waggoner, Jesse J.; Gresh, Lionel; Mohamed-Hadley, Alisha; Ballesteros, Gabriela; Davila, Maria Jose Vargas; Tellez, Yolanda; Sahoo, Malaya K.; Balmaseda, Angel; Harris, Eva

    2016-01-01

    Clinical manifestations of Zika virus, chikungunya virus, and dengue virus infections can be similar. To improve virus detection, streamline molecular workflow, and decrease test costs, we developed and evaluated a multiplex real-time reverse transcription PCR for these viruses. PMID:27184629

  20. Nano-mechanical Resonantor Sensors for Virus Detection

    NASA Astrophysics Data System (ADS)

    Bashir, Rashid

    2005-03-01

    Micro and nanoscale cantilever beams can be used as highly sensitive mass detectors. Scaling down the area of the cantilever allows a decrease in minimum detectable mass limit while scaling down the thickness allows the resonant frequencies to be within measurable range. We have fabricated arrays of silicon cantilever beams as nanomechanical resonant sensors to detect the mass of individual virus particles. The dimensions of the fabricated cantilever beams were in the range of 4-5 μm in length, 1-2 μm in width and 20-30 nm in thickness. The virus particles we used in the study were vaccinia virus, which is a member of the Poxviridae family and forms the basis of the smallpox vaccine. The frequency spectra of the cantilever beams, due to thermal and ambient noise, were measured using a laser Doppler vibrometer under ambient conditions. The change in resonant frequency as a function of the virus particle mass binding on the cantilever beam surface forms the basis of the detection scheme. We have demonstrated the detection of a single vaccinia virus particle with an average mass of 9.5 fg. Specific capture of the antigens requires attachment of antibodies, which can be in the same range of thickness as these cantilever sensors, and can alter their mechanical properties. We have attached protein layers on both sides of 30nm thick cantilever beams and we show that the resonant frequencies can increase or decrease upon the attachment of protein layers to the cantilevers. In certain cases, the increase in spring constant out-weighs the increase in mass and the resonant frequencies can increase upon the attachment of the protein layers. These devices can be very useful as components of biosensors for the detection of air-borne virus particles.

  1. Optimization of virus detection in cells using massively parallel sequencing.

    PubMed

    McClenahan, Shasta D; Uhlenhaut, Christine; Krause, Philip R

    2014-01-01

    Massively parallel sequencing (MPS)-based virus detection has potential regulatory applications. We studied the ability of one of these approaches, based on degenerate oligonucleotide primer (DOP)-polymerase chain reaction (PCR), to detect viral sequences in cell lines known to express viral genes or particles. DOP-PCR was highly sensitive for the detection of small quantities of isolated viral sequences. Detected viral sequences included nodavirus, bracovirus, and endogenous retroviruses in High Five cells, porcine circovirus type 1 and porcine endogenous retrovirus in PK15 cells, human T-cell leukemia virus 1 in MJ cells, human papillomavirus 18 in HeLa cells, human herpesvirus 8 in BCBL-1 cells, and Epstein-Barr Virus in Raji cells. Illumina sequencing (for which primers were most efficiently added using PCR) provided greater sensitivity for virus detection than Roche 454 sequencing. Analyzing nucleic acids extracted both directly from samples and from capsid-enriched preparations provided useful information. Although there are limitations of these methods, these results indicate significant promise for the combination of nonspecific PCR and MPS in identifying contaminants in clinical and biological samples, including cell lines and reagents used to produce vaccines and therapeutic products. PMID:24309095

  2. Detection of viral hemorrhagic septicemia virus

    USGS Publications Warehouse

    Winton, James; Kurath, Gael; Batts, William

    2007-01-01

    Viral hemorrhagic septicemia virus (VHSV) is considered to be one of the most important viral pathogens of finfish and is listed as reportable by many nations and international organizations (Office International des Epizooties 2006). Prior to 1988, VHSV was thought to be limited to Europe (Wolf 1988; Smail 1999). Subsequently, it was shown that the virus is endemic among many marine and anadromous fish species in both the Pacific and Atlantic Oceans (Meyers and Winton 1995; Skall et al. 2005). Genetic analysis reveals that isolates of VHSV can be divided into four genotypes that generally correlate with geographic location with the North American isolates generally falling into VHSV Genotype IV (Snow et al. 2004). In 2005-2006, reports from the Great Lakes region indicated that wild fish had experienced disease or, in some cases, very large die-offs from VHSV (Elsayed et al. 2006, Lumsden et al. 2007). The new strain from the Great Lakes, now identified as VHSV Genotype IVb, appears most closely related to isolates of VHSV from mortalities that occurred during 2000-2004 in rivers and near-shore areas of New Brunswick and Nova Scotia, Canada (Gagne et al. 2007). The type IVb isolate found in the Great Lakes region is the only strain outside of Europe that has been associated with significant mortality in freshwater species.

  3. Detection of Chikungunya Virus in Nepal.

    PubMed

    Pandey, Basu Dev; Neupane, Biswas; Pandey, Kishor; Tun, Mya Myat Ngwe; Morita, Kouichi

    2015-10-01

    Chikungunya virus (CHIKV) is an emerging alphaviral disease and a public health problem in South Asia including Nepal in recent years. In this study, sera were collected from patients presenting with fever, headache, muscular pain, fatigue, and joint pain of both upper and lower extremities. A total of 169 serum samples were tested for CHIKV and dengue virus (DENV) by using Immunoglobulin M (IgM) and Immunoglobulin G (IgG) antibody using enzyme-linked immunosorbent assay (ELISA) method during August to November 2013. Results showed that 3.6% and 27.8% samples were positive for CHIKV and DENV IgM positive, respectively. Similarly, results of IgG showed 3.0% samples were positive for CHIKV IgG and 29.0% were for DENV IgG positive. Further, a 50% focal reduction neutralization test (FRNT50) was performed to confirm the presence of CHIKV, which demonstrated that 8.9% of CHIKV IgM and/or IgG ELISA positive possessed neutralizing anti-CHIK antibodies. To our knowledge, this is the first report in which the presence of CHIKV is confirmed in Nepalese patients by FRNT50. Basic scientists and clinicians need to consider CHIKV as a differential diagnosis in febrile Nepalese patients, and policy makers should consider appropriate surveillance and actions for control strategies.

  4. [Molecular detection of tomato yellow leaf curl virus (TYLCV)].

    PubMed

    Li, Chang-Bao; Cui, Yan-Ling; Zhang, Li-Ying; Li, Chuan-You

    2012-03-01

    Tomato yellow leaf curl virus (TYLCV) is currently considered as one of the most devastating viruses in cultivated tomatoes (Solanum lycopersicum) worldwide. We reported here the development of a PCR-based method to quickly detect TYLCV using the primer pairs (TYLCV-F: 5'-ACG CAT GCC TCT AAT CCA GTG TA-3' and TYLCV-R: 5'-CCA ATA AGG CGT AAG CGT GTA GAC-3'), which was designed based on the genome sequence of TYLCV. A TYLCV-specific band of 543 bp was amplified from infected tomato plants. This protocol provides a rapid, reliable, and sensitive tool for molecular detection and identification of TYLCV in the industrial seedling and virus resistance breeding to facilitate safe and sustainable production of tomato.

  5. [Molecular detection of tomato yellow leaf curl virus (TYLCV)].

    PubMed

    Li, Chang-Bao; Cui, Yan-Ling; Zhang, Li-Ying; Li, Chuan-You

    2012-03-01

    Tomato yellow leaf curl virus (TYLCV) is currently considered as one of the most devastating viruses in cultivated tomatoes (Solanum lycopersicum) worldwide. We reported here the development of a PCR-based method to quickly detect TYLCV using the primer pairs (TYLCV-F: 5'-ACG CAT GCC TCT AAT CCA GTG TA-3' and TYLCV-R: 5'-CCA ATA AGG CGT AAG CGT GTA GAC-3'), which was designed based on the genome sequence of TYLCV. A TYLCV-specific band of 543 bp was amplified from infected tomato plants. This protocol provides a rapid, reliable, and sensitive tool for molecular detection and identification of TYLCV in the industrial seedling and virus resistance breeding to facilitate safe and sustainable production of tomato. PMID:22425956

  6. Virus detection using Viro-Adembeads, a rapid capture system for viruses, and plaque assay in intentionally virus-contaminated beverages.

    PubMed

    Hatano, Ben; Kojima, Asato; Sata, Tetsutaro; Katano, Harutaka

    2010-01-01

    Intentional contamination of beverages with microbes is one type of bioterrorist threat. While bacteria and fungus can be easily collected by a centrifuge, viruses are difficult to collect from virus-contaminated beverages. In this study, we demonstrated that Viro-Adembeads, a rapid-capture system for viruses using anionic polymer-coated magnetic beads, collected viruses from beverages contaminated intentionally with vaccinia virus and human herpesvirus 8. Real-time PCR showed that the recovery rates of the contaminated viruses in green tea and orange juice were lower than those in milk and water. Plaque assay showed that green tea and orange juice cut the efficiency of vaccinia virus infection in CV-1 cells. These results suggest that the efficiency of virus detection depends on the kind of beverage being tested. Viro-Adembeads would be a useful tool for detecting viruses rapidly in virus-contaminated beverages used in a bioterrorist attack.

  7. Virus detection using Viro-Adembeads, a rapid capture system for viruses, and plaque assay in intentionally virus-contaminated beverages.

    PubMed

    Hatano, Ben; Kojima, Asato; Sata, Tetsutaro; Katano, Harutaka

    2010-01-01

    Intentional contamination of beverages with microbes is one type of bioterrorist threat. While bacteria and fungus can be easily collected by a centrifuge, viruses are difficult to collect from virus-contaminated beverages. In this study, we demonstrated that Viro-Adembeads, a rapid-capture system for viruses using anionic polymer-coated magnetic beads, collected viruses from beverages contaminated intentionally with vaccinia virus and human herpesvirus 8. Real-time PCR showed that the recovery rates of the contaminated viruses in green tea and orange juice were lower than those in milk and water. Plaque assay showed that green tea and orange juice cut the efficiency of vaccinia virus infection in CV-1 cells. These results suggest that the efficiency of virus detection depends on the kind of beverage being tested. Viro-Adembeads would be a useful tool for detecting viruses rapidly in virus-contaminated beverages used in a bioterrorist attack. PMID:20093763

  8. Nanofluidic devices for rapid detection of virus particles.

    SciTech Connect

    Gourley, Paul Lee; McDonald, Anthony Eugene; Hendricks, Judy K.

    2005-01-01

    Technologies that could quickly detect and identify virus particles would play a critical role in fighting bioterrorism and help to contain the rapid spread of disease. Of special interest is the ability to detect the presence and movement of virions without chemically modifying them by attaching molecular probes. This would be useful for rapid detection of pathogens in food or water supplies without the use of expensive chemical reagents. Such detection requires new devices to quickly screen for the presence of tiny pathogens. To develop such a device, we fabricated nanochannels to transport virus particles through ultrashort laser cavities and measured the lasing output as a sensor for virions. To understand this transduction mechanism, we also investigated light scattering from virions, both to determine the magnitude of the scattered signal and to use it to investigate the motion of virions.

  9. Assays to Detect West Nile Virus in Dead Birds

    PubMed Central

    Therrien, Joseph E.; Benson, Robert; Kramer, Laura; Kauffman, Elizabeth B.; Eidson, Millicent; Campbell, Scott

    2005-01-01

    Using oral swab samples to detect West Nile virus in dead birds, we compared the Rapid Analyte Measurement Platform (RAMP) assay with VecTest and real-time reverse-transcriptase–polymerase chain reaction. The sensitivities of RAMP and VecTest for testing corvid species were 91.0% and 82.1%, respectively. PMID:16318736

  10. The use of a handheld Raman system for virus detection

    NASA Astrophysics Data System (ADS)

    Song, Chunyuan; Driskell, Jeremy D.; Tripp, Ralph A.; Cui, Yiping; Zhao, Yiping

    2012-06-01

    The combination of surface enhanced Raman spectroscopy (SERS) with a handheld Raman system would lead to a powerful portable device for defense and security applications. The Thermo Scientific FirstDefender RM instrument is a 785-nm handheld Raman spectrometer intended for rapid field identification of unknown solid and liquid samples. Its sensitivity and effectiveness for SERS-based detection was initially confirmed by evaluating detection of 1,2-di(4- pyridyl)ethylene as a reporter molecule on a silver nanorod (AgNR) substrate, and the results are comparable to those from a confocal Bruker Raman system. As avian influenza A viruses (AIV) are recognized as an important emerging threat to public health, this portable handheld Raman spectrometer is used, for the first time, to detect and identify avian influenza A viruses using a multi-well AgNR SERS chip. The SERS spectra obtained had rich peaks which demonstrated that the instrument can be effectively used for SERS-based influenza virus detection. According to the SERS spectra, these different influenza viruses were distinguished from the negative control via the principal component analysis and by partial least squares-discriminate analysis. Together, these results show that the combination effective SERS substrates when combined with a portable Raman spectrometer provides a powerful field device for chemical and biological sensing.

  11. Multiplex detection of Solenopsis invicta viruses -1, -2, and -3

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A multiplex polymerase chain reaction (PCR) method was developed to detect simultaneously Solenopsis invicta viruses -1, -2, and -3 in their host, the red imported fire ant, Solenopsis invicta. cDNA synthesis was conducted in a single reaction containing three oligonucleotide primers specific for ...

  12. Detection of dengue virus in platelets isolated from dengue patients.

    PubMed

    Noisakran, Sansanee; Gibbons, Robert V; Songprakhon, Pucharee; Jairungsri, Aroonroong; Ajariyakhajorn, Chuanpis; Nisalak, Ananda; Jarman, Richard G; Malasit, Prida; Chokephaibulkit, Kulkanya; Perng, Guey Chuen

    2009-03-01

    Though thrombocytopenia or dysfunction of platelets is common in dengue virus infection, the role of platelets has not been established. We enrolled 33 hospitalized children with serologically confirmed dengue virus infection. Blood specimens were collected during hospitalization. Platelets and plasma were isolated from the whole blood. Detection of dengue virus in plasma and platelets was carried out by RT-PCR with primers that can differentiate different dengue serotypes simultaneously, and by electron transmission microscopy (EM). Dengue viral RNA was detected in the platelets and plasma by conventional RT-PCR. A significantly higher percentage of dengue viral RNA was detected in platelets than in plasma (p = 0.03). Platelets isolated 5 days after onset of fever were most likely positive for viral RNA. Concurrent infection or co-circulation with multiple dengue serotypes was observed in 12% of patients. Infrequently, negative-stranded dengue viral RNA was detected in platelets and in plasma. Importantly, EM confirmed the presence of dengue viral-like particles inside platelets prepared from dengue patients. Our findings suggest the presence of dengue virus in platelets may be associated with the dysfunction of platelets observed in dengue patients.

  13. Using Fluorescent Viruses for Detecting Bacteria in Water

    NASA Technical Reports Server (NTRS)

    Tabacco, Mary Beth; Qian, Xiaohua; Russo, Jaimie A.

    2009-01-01

    A method of detecting water-borne pathogenic bacteria is based partly on established molecular-recognition and fluorescent-labeling concepts, according to which bacteria of a species of interest are labeled with fluorescent reporter molecules and the bacteria can then be detected by fluorescence spectroscopy. The novelty of the present method lies in the use of bacteriophages (viruses that infect bacteria) to deliver the fluorescent reporter molecules to the bacteria of the species of interest.

  14. Rapid Fluorescent Detection Assay for Human Parainfluenza Viruses.

    PubMed

    Takahashi, Tadanobu; Takano, Maiko; Kurebayashi, Yuuki; Agarikuchi, Takashi; Suzuki, Chihiro; Fukushima, Keijo; Takahashi, Shunsaku; Otsubo, Tadamune; Ikeda, Kiyoshi; Minami, Akira; Suzuki, Takashi

    2015-01-01

    Human parainfluenza virus type 1 (hPIV1) does not form clear plaque by the conventional plaque formation assay because of slightly a cytopathic effects in many cell lines infected with hPIV1, thus making in virus titration, isolation and inhibitor evaluation difficult. We have succeeded in fluorescent histochemical visualization of sialidase activities of influenza A and B viruses, Newcastle disease virus and Sendai virus by using a novel fluorescent sialidase substrate, 2-(benzothiazol-2-yl)-4-bromophenyl 5-acetamido-3,5-dideoxy-α-D-glycero-D-galacto-2-nonulopyranosidonic acid (BTP3-Neu5Ac). In this study, we applied the BTP3-Neu5Ac assay for rapid detection of hPIV1 and hPIV type 3. The BTP3-Neu5Ac assay could histochemically visualize dot-blotted hPIVs on a membrane and hPIV-infected cells as local fluorescence under UV irradiation. We succeeded in distinct fluorescent visualization of hPIV1-infected cells in only 3 d using the BTP3-Neu5Ac assay. Due to there being no fixation, hPIV1 was isolated directly from fluorescent stained focus cells by the BTP3-Neu5Ac assay. Establishment of a sensitive, easy, and rapid fluorescent focus detection assay for hPIV, hPIV1 in particular will contribute greatly to progress in hPIV studies.

  15. Detection of Human Enteric Viruses in Freshwater from European Countries.

    PubMed

    D'Ugo, Emilio; Marcheggiani, Stefania; Fioramonti, Ilaria; Giuseppetti, Roberto; Spurio, Roberto; Helmi, Karim; Guillebault, Delphine; Medlin, Linda K; Simeonovski, Ivan; Boots, Bas; Breitenbach, Ulrich; Koker, Latife; Albay, Meric; Mancini, Laura

    2016-09-01

    The transmission of water-borne pathogens typically occurs by a faecal-oral route, through inhalation of aerosols, or by direct or indirect contact with contaminated water. Previous molecular-based studies have identified viral particles of zoonotic and human nature in surface waters. Contaminated water can lead to human health issues, and the development of rapid methods for the detection of pathogenic microorganisms is a valuable tool for the prevention of their spread. The aims of this work were to determine the presence and identity of representative human pathogenic enteric viruses in water samples from six European countries by quantitative polymerase chain reaction (q-PCR) and to develop two quantitative PCR methods for Adenovirus 41 and Mammalian Orthoreoviruses. A 2-year survey showed that Norovirus, Mammalian Orthoreovirus and Adenoviruses were the most frequently identified enteric viruses in the sampled surface waters. Although it was not possible to establish viability and infectivity of the viruses considered, the detectable presence of pathogenic viruses may represent a potential risk for human health. The methodology developed may aid in rapid detection of these pathogens for monitoring quality of surface waters. PMID:27117764

  16. [Simultaneous detection of respiratory viruses and influenza A virus subtypes using multiplex PCR].

    PubMed

    Ciçek, Candan; Bayram, Nuri; Anıl, Murat; Gülen, Figen; Pullukçu, Hüsnü; Saz, Eylem Ulaş; Telli, Canan; Cok, Gürsel

    2014-10-01

    This study was conducted to investigate the respiratory viruses and subtyping of influenza A virus when positive by multiplex PCR in patients with flu-like symptoms, after the pandemic caused by influenza A (H1N1)pdm09. Nasopharyngeal swab samples collected from 700 patients (313 female, 387 male; age range: 24 days-94 yrs, median age: 1 yr) between December 2010 - January 2013 with flu-like symptoms including fever, headache, sore throat, rhinitis, cough, myalgia as defined by the World Health Organization were included in the study. Nucleic acid extractions (Viral DNA/RNA Extraction Kit, iNtRON, South Korea) and cDNA synthesis (RevertAid First Strand cDNA Synthesis Kits, Fermentas, USA) were performed according to the manufacturer's protocol. Multiplex amplification of nucleic acids was performed using DPO (dual priming oligonucleotide) primers and RV5 ACE Screening Kit (Seegene, South Korea) in terms of the presence of influenza A (INF-A) virus, influenza B (INF-B) virus, respiratory syncytial virus (RSV), and the other respiratory viruses. PCR products were detected by automated polyacrylamide gel electrophoresis using Screen Tape multiple detection system. Specimens which were positive for viral nucleic acids have been further studied by using specific DPO primers, FluA ACE Subtyping and RV15 Screening (Seegene, South Korea) kits. Four INF-A virus subtypes [human H1 (hH1), human H3 (hH3), swine H1 (sH1), avian H5 (aH5)] and 11 other respiratory viruses [Adenovirus, parainfluenza virus (PIV) types 1-4, human bocavirus (HBoV), human metapneumovirus (HMPV), rhinovirus types A and B, human coronaviruses (HCoV) OC43, 229E/NL63] were investigated with those tests. In the study, 53.6% (375/700) of the patients were found to be infected with at least one virus and multiple respiratory virus infections were detected in 15.7% (59/375) of the positive cases, which were mostly (49/59, 83%) in pediatric patients. RSV and rhinovirus coinfections were the most prevalent (18

  17. Serum virus neutralization assay for detection and quantitation of serum neutralizing antibodies to influenza A virus in swine

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The serum virus neutralization (SVN) assay is a serological test to detect the presence and magnitude of functional systemic antibodies that prevent infectivity of a virus. The SVN assay is a highly sensitive and specific test that may be applied to influenza A viruses (IAV) in swine to measure the ...

  18. The production of peste des petits ruminants hyperimmune sera in rabbits and their application in virus diagnosis.

    PubMed

    Obi, T U; McCullough, K C; Taylor, W P

    1990-07-01

    Hyperimmune sera were produced by serial inoculation of rabbits with Vero cell-adapted, sucrose gradient-purified Nigerian peste des petits ruminants virus (PPRV) isolate. Two antisera produced, neutralized the homologous PPRV but not the heterologous rinderpest Kabette "O" virus. The antisera gave strong precipitin lines with purified PPRV antigens and were used to detect PPRV and rinderpest virus antigens from ante-mortem secretions and post-mortem tissue homogenates from PPR and rinderpest virus infected goats and cattle by the agar gel precipitation tests (AGPT). The hyperimmune sera gave good titration curves with both purified Nigerian goat and the United Arab Emirate wildlife PPRV isolates in the indirect enzyme linked immunosorbent assay (ELISA). Results of indirect ELISA showed that although there were some cross reactions with the rinderpest, canine-distemper and measles viruses, at 1:100 dilution, the antisera would give a positive signal with only the homologous PPR virus.

  19. Growth profiles of recent canine distemper isolates on Vero cells expressing canine signalling lymphocyte activation molecule (SLAM).

    PubMed

    Lan, N T; Yamaguchi, R; Uchida, K; Sugano, S; Tateyama, S

    2005-07-01

    Fresh samples of lymph node, lung and cerebrum taken post mortem from dogs no. 1, 2 and 3 yielded canine distemper virus (CDV) strains 007 Lm, 009 L and 011 C, respectively. These were titrated on Vero cells stably expressing canine signalling lymphocyte activation molecule (SLAM; Vero-DST cells). Growth curves of the three strains were produced by titration of the released virus and cell-associated virus at various timepoints. All three isolates, especially 007 Lm, grew well on Vero-DST cells. The titres of cell-associated virus of two strains (009 L and 011 C) were clearly lower than those of virus released into the culture supernate. The results indicate that Vero-DST cells are not only useful for primary isolation but also efficient for titrating virus from fresh tissues and for the study of growth profiles of recent CDV isolates.

  20. Gene detection, virus isolation, and sequence analysis of avian leukosis viruses in Taiwan country chickens.

    PubMed

    Chang, Shu-Wei; Hsu, Meng-Fang; Wang, Ching-Ho

    2013-06-01

    Avian leukosis virus (ALV) infection in Taiwan Country chickens (TCCs) was investigated by using gene detection, virus isolation, and sequence analysis. The blood samples of 61 TCC flocks at market ages from a slaughter house were screened for exogenous ALVs using polymerase chain reaction to investigate the ALV infection status. The buffy coats from three breeder and four commercial chicken flocks were cocultured with DF-1 cells to isolate the virus. The full proviral DNA genomes of two ALV isolates were sequenced, analyzed, and compared with reference ALV strains. The gene detection results showed that 60 and 43 of the 61 flocks were infected with subgroup A of ALV (ALV-A) and subgroup J of ALV (ALV-J), respectively. Virus isolation results showed that five ALV-As and two ALV-Js were isolated from those seven TCC flocks. The full sequences of the isolates showed that isolate TW-3577 possessed a myeloblastosis-associated virus 1 gp85 coding region and an ALV-J 3'-untranslated region (3'UTR) and was similar to ordinary ALV-A. However, TW-3593 was unique. The 3'UTR of this isolate displayed high identity to endogenous counterpart sequence and its gp85 was different from all subgroups. This unique ALV is common in Taiwan.

  1. Detection of Nonhemagglutinating Influenza A(H3) Viruses by Enzyme-Linked Immunosorbent Assay in Quantitative Influenza Virus Culture

    PubMed Central

    Els, C.; Sprong, L.; van Beek, R.; van der Vries, E.; Osterhaus, A. D. M. E.; Rimmelzwaan, G. F.

    2014-01-01

    To assess the efficacy of novel antiviral drugs against influenza virus in clinical trials, it is necessary to quantify infectious virus titers in respiratory tract samples from patients. Typically, this is achieved by inoculating virus-susceptible cells with serial dilutions of clinical specimens and detecting the production of progeny virus by hemagglutination, since influenza viruses generally have the capacity to bind and agglutinate erythrocytes of various species through their hemagglutinin (HA). This readout method is no longer adequate, since an increasing number of currently circulating influenza A virus H3 subtype (A[H3]) viruses display a reduced capacity to agglutinate erythrocytes. Here, we report the magnitude of this problem by analyzing the frequency of HA-deficient A(H3) viruses detected in The Netherlands from 1999 to 2012. Furthermore, we report the development and validation of an alternative method for monitoring the production of progeny influenza virus in quantitative virus cultures, which is independent of the capacity to agglutinate erythrocytes. This method is based on the detection of viral nucleoprotein (NP) in virus culture plates by enzyme-linked immunosorbent assay (ELISA), and it produced results similar to those of the hemagglutination assay using strains with good HA activity, including A/Brisbane/059/07 (H1N1), A/Victoria/210/09 (H3N2), other seasonal A(H1N1), A(H1N1)pdm09, and the majority of A(H3) virus strains isolated in 2009. In contrast, many A(H3) viruses that have circulated since 2010 failed to display HA activity, and infectious virus titers were determined only by detecting NP. The virus culture ELISA described here will enable efficacy testing of new antiviral compounds in clinical trials during seasons in which nonhemagglutinating influenza A viruses circulate. PMID:24622097

  2. Detection of nonhemagglutinating influenza a(h3) viruses by enzyme-linked immunosorbent assay in quantitative influenza virus culture.

    PubMed

    van Baalen, C A; Els, C; Sprong, L; van Beek, R; van der Vries, E; Osterhaus, A D M E; Rimmelzwaan, G F

    2014-05-01

    To assess the efficacy of novel antiviral drugs against influenza virus in clinical trials, it is necessary to quantify infectious virus titers in respiratory tract samples from patients. Typically, this is achieved by inoculating virus-susceptible cells with serial dilutions of clinical specimens and detecting the production of progeny virus by hemagglutination, since influenza viruses generally have the capacity to bind and agglutinate erythrocytes of various species through their hemagglutinin (HA). This readout method is no longer adequate, since an increasing number of currently circulating influenza A virus H3 subtype (A[H3]) viruses display a reduced capacity to agglutinate erythrocytes. Here, we report the magnitude of this problem by analyzing the frequency of HA-deficient A(H3) viruses detected in The Netherlands from 1999 to 2012. Furthermore, we report the development and validation of an alternative method for monitoring the production of progeny influenza virus in quantitative virus cultures, which is independent of the capacity to agglutinate erythrocytes. This method is based on the detection of viral nucleoprotein (NP) in virus culture plates by enzyme-linked immunosorbent assay (ELISA), and it produced results similar to those of the hemagglutination assay using strains with good HA activity, including A/Brisbane/059/07 (H1N1), A/Victoria/210/09 (H3N2), other seasonal A(H1N1), A(H1N1)pdm09, and the majority of A(H3) virus strains isolated in 2009. In contrast, many A(H3) viruses that have circulated since 2010 failed to display HA activity, and infectious virus titers were determined only by detecting NP. The virus culture ELISA described here will enable efficacy testing of new antiviral compounds in clinical trials during seasons in which nonhemagglutinating influenza A viruses circulate.

  3. Detection of novel viruses in porcine fecal samples from China

    PubMed Central

    2013-01-01

    Background Pigs are well known source of human infectious disease. To better understand the spectrum of viruses present in pigs, we utilized the 454 Life Sciences GS-FLX high-throughput sequencing platform to sequence stool samples from healthy pigs. Findings Total nucleic acid was extracted from stool samples of healthy piglets and randomly amplified. The amplified materials were pooled and processed using a high-throughput pyrosequencing technique. The raw sequences were deconvoluted on the basis of the barcode and then processed through a standardized bioinformatics pipeline. The unique reads (348, 70 and 13) had limited similarity to known astroviruses, bocaviruses and parechoviruses. Specific primers were synthesized to assess the prevalence of the viruses in healthy piglets. Our results indicate extremely high rates of positivity. Conclusions Several novel astroviruses, bocaviruses and Ljungan-like viruses were identified in stool samples from healthy pigs. The rates of isolation for the new viruses were high. The high detection rate, diverse sequences and categories indicate that pigs are well-established reservoirs for and likely sources of different enteric viruses. PMID:23363449

  4. Detection of enteric viruses in treated drinking water.

    PubMed Central

    Keswick, B H; Gerba, C P; DuPont, H L; Rose, J B

    1984-01-01

    The occurrence of viruses in conventionally treated drinking water derived from a heavily polluted source was evaluated by collecting and analyzing 38 large-volume (65- to 756-liter) samples of water from a 9 m3/s (205 X 10(6) gallons [776 X 10(6) liters] per day) water treatment plant. Samples of raw, clarified, filtered, and chlorinated finished water were concentrated by using the filter adsorption-elution technique. Of 23 samples of finished water, 19 (83%) contained viruses. None of the nine finished water samples collected during the dry season contained detectable total coliform bacteria. Seven of nine finished water samples collected during the dry season met turbidity, total coliform bacteria, and total residual chlorine standards. Of these, four contained virus. During the dry season the percent removals were 25 to 93% for enteric viruses, 89 to 100% for bacteria, and 81% for turbidity. During the rainy season the percent removals were 0 to 43% for enteric viruses, 80 to 96% for bacteria, and 63% for turbidity. None of the 14 finished water samples collected during the rainy season met turbidity standards, and all contained rotaviruses or enteroviruses. PMID:6331313

  5. Optofluidic wavelength division multiplexing for single-virus detection

    PubMed Central

    Ozcelik, Damla; Parks, Joshua W.; Wall, Thomas A.; Stott, Matthew A.; Cai, Hong; Parks, Joseph W.; Hawkins, Aaron R.; Schmidt, Holger

    2015-01-01

    Optical waveguides simultaneously transport light at different colors, forming the basis of fiber-optic telecommunication networks that shuttle data in dozens of spectrally separated channels. Here, we reimagine this wavelength division multiplexing (WDM) paradigm in a novel context––the differentiated detection and identification of single influenza viruses on a chip. We use a single multimode interference (MMI) waveguide to create wavelength-dependent spot patterns across the entire visible spectrum and enable multiplexed single biomolecule detection on an optofluidic chip. Each target is identified by its time-dependent fluorescence signal without the need for spectral demultiplexing upon detection. We demonstrate detection of individual fluorescently labeled virus particles of three influenza A subtypes in two implementations: labeling of each virus using three different colors and two-color combinatorial labeling. By extending combinatorial multiplexing to three or more colors, MMI-based WDM provides the multiplexing power required for differentiated clinical tests and the growing field of personalized medicine. PMID:26438840

  6. Detection method for avian influenza viruses in water.

    PubMed

    Rönnqvist, Maria; Ziegler, Thedi; von Bonsdorff, Carl-Henrik; Maunula, Leena

    2012-03-01

    Recent events have shown that humans may become infected with some pathogenic avian influenza A viruses (AIV). Since soil and water, including lakes, rivers, and seashores, may be contaminated by AIV excreted by birds, effective methods are needed for monitoring water for emerging viruses. Combining water filtration with molecular methods such as PCR is a fast and effective way for detecting viruses. The objective of this study was to apply a convenient method for the detection of AIV in natural water samples. Distilled water and lake, river, and seawater were artificially contaminated with AIV (H5N3) and passed through a filter system. AIV was detected from filter membrane by real-time RT-PCR. The performance of Zetapor, SMWP, and Sartobind D5F membranes in recovering influenza viruses was first evaluated using contaminated distilled water. SWMP, which gave the highest virus recoveries, was then compared with a pre-filter combined GF/F filter membrane in a trial using natural water samples. In this study, the cellulose membrane SMWP was found to be practical for recovery of AIVs in water. Viral yields varied between 62.1 and 65.9% in distilled water and between 1 and 16.7% in natural water samples. The borosilicate glass membrane GF/F combined with pre-filter was also feasible in filtering natural water samples with viral yields from 1.98 to 7.33%. The methods described can be used for monitoring fresh and seawater samples for the presence of AIV and to determine the source of AIV transmission in an outbreak situation. PMID:23412765

  7. Detection method for avian influenza viruses in water.

    PubMed

    Rönnqvist, Maria; Ziegler, Thedi; von Bonsdorff, Carl-Henrik; Maunula, Leena

    2012-03-01

    Recent events have shown that humans may become infected with some pathogenic avian influenza A viruses (AIV). Since soil and water, including lakes, rivers, and seashores, may be contaminated by AIV excreted by birds, effective methods are needed for monitoring water for emerging viruses. Combining water filtration with molecular methods such as PCR is a fast and effective way for detecting viruses. The objective of this study was to apply a convenient method for the detection of AIV in natural water samples. Distilled water and lake, river, and seawater were artificially contaminated with AIV (H5N3) and passed through a filter system. AIV was detected from filter membrane by real-time RT-PCR. The performance of Zetapor, SMWP, and Sartobind D5F membranes in recovering influenza viruses was first evaluated using contaminated distilled water. SWMP, which gave the highest virus recoveries, was then compared with a pre-filter combined GF/F filter membrane in a trial using natural water samples. In this study, the cellulose membrane SMWP was found to be practical for recovery of AIVs in water. Viral yields varied between 62.1 and 65.9% in distilled water and between 1 and 16.7% in natural water samples. The borosilicate glass membrane GF/F combined with pre-filter was also feasible in filtering natural water samples with viral yields from 1.98 to 7.33%. The methods described can be used for monitoring fresh and seawater samples for the presence of AIV and to determine the source of AIV transmission in an outbreak situation.

  8. Rapid detection of hendra virus using magnetic particles and quantum dots.

    PubMed

    Lisi, Fabio; Falcaro, Paolo; Buso, Dario; Hill, Anita J; Barr, Jennifer A; Crameri, Gary; Nguyen, Tich-Lam; Wang, Lin-Fa; Mulvaney, Paul

    2012-09-01

    A proof-of-concept for the development of a fast and portable Hendra virus biosensor is presented. Hendra virus, a deadly emerging pathogen in Australia, can be co-localized, concentrated and revealed using simultaneously magnetic and luminescent functional particles. This method should be applicable for the early detection of any other virus by targeting the specific virus with the corresponding antibody.

  9. Detection of evolutionarily distinct avian influenza a viruses in antarctica.

    PubMed

    Hurt, Aeron C; Vijaykrishna, Dhanasekaran; Butler, Jeffrey; Baas, Chantal; Maurer-Stroh, Sebastian; Silva-de-la-Fuente, M Carolina; Medina-Vogel, Gonzalo; Olsen, Bjorn; Kelso, Anne; Barr, Ian G; González-Acuña, Daniel

    2014-01-01

    ABSTRACT Distinct lineages of avian influenza viruses (AIVs) are harbored by spatially segregated birds, yet significant surveillance gaps exist around the globe. Virtually nothing is known from the Antarctic. Using virus culture, molecular analysis, full genome sequencing, and serology of samples from Adélie penguins in Antarctica, we confirmed infection by H11N2 subtype AIVs. Their genetic segments were distinct from all known contemporary influenza viruses, including South American AIVs, suggesting spatial separation from other lineages. Only in the matrix and polymerase acidic gene phylogenies did the Antarctic sequences form a sister relationship to South American AIVs, whereas distant phylogenetic relationships were evident in all other gene segments. Interestingly, their neuraminidase genes formed a distant relationship to all avian and human influenza lineages, and the polymerase basic 1 and polymerase acidic formed a sister relationship to the equine H3N8 influenza virus lineage that emerged during 1963 and whose avian origins were previously unknown. We also estimated that each gene segment had diverged for 49 to 80 years from its most closely related sequences, highlighting a significant gap in our AIV knowledge in the region. We also show that the receptor binding properties of the H11N2 viruses are predominantly avian and that they were unable to replicate efficiently in experimentally inoculated ferrets, suggesting their continuous evolution in avian hosts. These findings add substantially to our understanding of both the ecology and the intra- and intercontinental movement of Antarctic AIVs and highlight the potential risk of an incursion of highly pathogenic AIVs into this fragile environment. IMPORTANCE Avian influenza viruses (AIVs) are typically maintained and spread by migratory birds, resulting in the existence of distinctly different viruses around the world. However, AIVs have not previously been detected in Antarctica. In this study, we

  10. Detection of evolutionarily distinct avian influenza a viruses in antarctica.

    PubMed

    Hurt, Aeron C; Vijaykrishna, Dhanasekaran; Butler, Jeffrey; Baas, Chantal; Maurer-Stroh, Sebastian; Silva-de-la-Fuente, M Carolina; Medina-Vogel, Gonzalo; Olsen, Bjorn; Kelso, Anne; Barr, Ian G; González-Acuña, Daniel

    2014-01-01

    ABSTRACT Distinct lineages of avian influenza viruses (AIVs) are harbored by spatially segregated birds, yet significant surveillance gaps exist around the globe. Virtually nothing is known from the Antarctic. Using virus culture, molecular analysis, full genome sequencing, and serology of samples from Adélie penguins in Antarctica, we confirmed infection by H11N2 subtype AIVs. Their genetic segments were distinct from all known contemporary influenza viruses, including South American AIVs, suggesting spatial separation from other lineages. Only in the matrix and polymerase acidic gene phylogenies did the Antarctic sequences form a sister relationship to South American AIVs, whereas distant phylogenetic relationships were evident in all other gene segments. Interestingly, their neuraminidase genes formed a distant relationship to all avian and human influenza lineages, and the polymerase basic 1 and polymerase acidic formed a sister relationship to the equine H3N8 influenza virus lineage that emerged during 1963 and whose avian origins were previously unknown. We also estimated that each gene segment had diverged for 49 to 80 years from its most closely related sequences, highlighting a significant gap in our AIV knowledge in the region. We also show that the receptor binding properties of the H11N2 viruses are predominantly avian and that they were unable to replicate efficiently in experimentally inoculated ferrets, suggesting their continuous evolution in avian hosts. These findings add substantially to our understanding of both the ecology and the intra- and intercontinental movement of Antarctic AIVs and highlight the potential risk of an incursion of highly pathogenic AIVs into this fragile environment. IMPORTANCE Avian influenza viruses (AIVs) are typically maintained and spread by migratory birds, resulting in the existence of distinctly different viruses around the world. However, AIVs have not previously been detected in Antarctica. In this study, we

  11. Detection of avian nephritis virus in Australian chicken flocks.

    PubMed

    Hewson, Kylie A; O'Rourke, Denise; Noormohammadi, Amir H

    2010-09-01

    Avian nephritis virus (ANV) is thought to infect poultry flocks worldwide, but no confirmed case has been reported in Australia. The first such case is described in this study. Cases of young chickens with clinical signs of dehydration and diarrhea were submitted to our laboratory and histopathology detected interstitial nephritis. Vaccine strains of infectious bronchitis virus were detected in some of these cases but were not considered to be the causative agent. A total of seven fresh submissions from broiler chicken flocks were collected at 8-11 days of age. Degenerate PCR primers were designed based on published ANV polymerase gene sequences and used to analyze historic cases as well as the fresh submissions. Six of the seven fresh submissions, and one historic case, were positive for ANV with nucleotide sequencing confirming these results. These results establish ANV as an infectious pathogen circulating in Australian poultry.

  12. Comparison of sensitivities of virus isolation, antigen detection, and nucleic acid amplification for detection of equine influenza virus.

    PubMed

    Quinlivan, Michelle; Cullinane, Ann; Nelly, Maura; Van Maanen, Kees; Heldens, Jacco; Arkins, Sean

    2004-02-01

    Four seronegative foals aged 6 to 7 months were exposed to an aerosol of influenza strain A/Equi/2/Kildare/89 at 10(6) 50% egg infective doses (EID(50))/ml. Nasopharyngeal swabs were collected for 10 consecutive days after challenge. Virus isolation was performed in embryonated eggs, and the EID(50) was determined for all positive samples. The 50% tissue culture infective dose was determined using Madin-Darby canine kidney (MDCK) cells. Samples were also tested by an in vitro enzyme immunoassay test, Directigen Flu A, and by reverse transcription-PCR (RT-PCR) using nested primers from the nucleoprotein gene and a single set of primers from the matrix gene. RT-PCR using the matrix primers and virus isolation in embryonated eggs proved to be the most sensitive methods for the detection of virus. The Directigen Flu A test was the least sensitive method. The inclusion of 2% fetal calf serum in the viral transport medium inhibited the growth of virus from undiluted samples in MDCK cells but was essential for the maintenance of the virus titer in samples subjected to repeated freeze-thaw cycles. PMID:14766849

  13. First detection of Toscana virus in Corsica, France.

    PubMed

    Bichaud, L; Izri, A; de Lamballerie, X; Moureau, G; Charrel, R N

    2014-02-01

    Toscana virus (TOSV) was detected for the first time from Phlebotomus perniciosus sandflies in Corsica, a French Mediterranean island. Genetic analysis showed that Corsican TOSV belongs to lineage A, together with Italian, Tunisian, Turkish and other French strains. The demonstration of TOSV in Corsica indicates that autochthonous and tourist populations are at risk of infection. Hence, physicians must consider TOSV as a possible cause of aseptic meningitis and unidentified febrile illness during the warm season.

  14. A multiplex reverse transcription PCR assay for simultaneous detection of five tobacco viruses in tobacco plants.

    PubMed

    Dai, Jin; Cheng, Julong; Huang, Ting; Zheng, Xuan; Wu, Yunfeng

    2012-07-01

    Tobacco viruses including Tobacco mosaic virus (TMV), Cucumber mosaic virus (CMV), Tobacco etch virus (TEV), Potato virus Y (PVY) and Tobacco vein banding mosaic virus (TVBMV) are major viruses infecting tobacco and can cause serious crop losses. A multiplex reverse transcription polymerase chain reaction assay was developed to detect simultaneously and differentiate all five viruses. The system used specific primer sets for each virus producing five distinct fragments 237, 273, 347, 456 and 547 bp, representing TMV, CMV subgroup I, TEV, PVY(O) and TVBMV, respectively. These primers were used for detection of the different viruses by single PCR and multiplex PCR and the results were confirmed by DNA sequencing analysis. The protocol was used to detect viruses from different parts of China. The simultaneous and sensitive detection of different viruses using the multiplex PCR is more efficient and economical than other conventional methods for tobacco virus detection. This multiplex PCR provides a rapid and reliable method for the detection and identification of major tobacco viruses, and will be useful for epidemiological studies.

  15. Bioluminescent detection probe for tick-borne encephalitis virus immunoassay.

    PubMed

    Burakova, Ludmila P; Kudryavtsev, Alexander N; Stepanyuk, Galina A; Baykov, Ivan K; Morozova, Vera V; Tikunova, Nina V; Dubova, Maria A; Lyapustin, Victor N; Yakimenko, Valeri V; Frank, Ludmila A

    2015-07-01

    To facilitate the detection of the tick-borne encephalitis virus (TBEV), the causative agent of one of the most severe human neuroinfections, we have developed an immunoassay based on bioluminescent hybrid protein 14D5a-Rm7 as a detection probe. The protein containing Renilla luciferase as a reporter and a single-chain variable fragment (scFv) of murine immunoglobulin to TBEV as a recognition element was constructed, produced by bacterial expression, purified, and tested. Both domains were shown to reveal their specific biological properties-affinity to the target antigen and bioluminescent activity. Hybrid protein was applied as a label for solid-phase immunoassay of the antigens, associated with the tick-borne encephalitis virus (native glycoprotein E or extracts of the infected strain of lab ticks). The assay demonstrates high sensitivity (0.056 ng of glycoprotein E; 10(4)-10(5) virus particles or 0.1 pg virions) and simplicity and is competitive with conventional methods for detection of TBEV. PMID:25925861

  16. A strategy for detection of viruses in groundwater by PCR.

    PubMed

    Abbaszadegan, M; Stewart, P; LeChevallier, M

    1999-02-01

    We evaluated the use of the PCR for detection of enteric viruses in groundwater. To do this, we used an improved sample-processing technique and a large-volume amplification protocol. The objective of this study was to use advanced molecular techniques to develop a rapid and simple method which can be used by the water industry for detection of viral contamination in a variety of water samples. The strategy described here fulfills the water industry's need for a rapid, reliable, easily performed method for analyzing groundwater for virus contamination. Viruses were detected after concentration from at least 400 gallons (1,512 liters) of water by a filter adsorption and elution method, which resulted in a concentrate containing viruses. A total of 150 samples were analyzed by performing cell culture assays for enteroviruses and by performing reverse transcription PCR (RT-PCR) analyses for enteroviruses, hepatitis A virus, and rotavirus. Thirteen samples (8.7%) produced cellular cytopathic effects when the Buffalo green monkey cell line was used. When primers specific for enteroviruses were used in RT-PCR, 40 of 133 samples (30.1%) tested positive for the presence of enterovirus RNA. When hepatitis A virus-specific primers were used, 12 of 139 samples (8.6%) were considered positive for the presence of hepatitis A viral RNA. The RT-PCR analysis performed with rotavirus-specific primers identified 18 of 130 samples (13.8%) that were positive for rotavirus RNA sequences. Our sample-processing technique and large-volume PCR protocol (reaction volume, 300 microliter) resulted in sufficient removal or dilution of inhibitors so that more than 95% of the samples could be assayed by PCR. Because of its sensitivity for detecting viral nucleic acid sequences, PCR analysis should produce more positive results than cell culture analysis. Since either cell culture analysis or PCR can reveal only a "snapshot" of the quality of the groundwater being sampled, PCR seems to be a

  17. Effect of recombinant canine distemper vaccine on antibody titers in previously vaccinated dogs.

    PubMed

    Larson, L J; Hageny, T L; Haase, C J; Schultz, R D

    2006-01-01

    Two canine distemper virus (CDV) vaccine types are currently commercially available: modified-live virus (MLV) vaccines and a canarypox recombinant CDV (rCDV) vaccine (Recombitek, Merial). This study compared the ability of the rCDV vaccine and MLV vaccines to significantly enhance (boost) the antibody response of previously immunized adult and juvenile dogs. A significant (fourfold or greater) increase in titer occurred in significantly more dogs revaccinated with Recombitek C-4 or Recombitek C-6 than with the MLV-CDV vaccines. This study demonstrates that Recombitek, the only vaccine for dogs containing rCDV, is more likely to significantly boost the CDV antibody response in previously vaccinated dogs than are the MLV-CDV vaccines. Because rCDV vaccine can boost the antibody titer of dogs previously vaccinated with an MLV vaccine, it can and should be used when core vaccines are readministered. PMID:16871492

  18. Hendra virus detection using Loop-Mediated Isothermal Amplification.

    PubMed

    Foord, Adam J; Middleton, Deborah; Heine, Hans G

    2012-04-01

    Hendra virus (HeV) is a zoonotic paramyxovirus endemic in Australian Pteropus bats (fruit bats or flying foxes). Although bats appear to be unaffected by the virus, HeV can spread from fruit bats to horses, causing severe disease. Human infection results from close contact with the blood, body fluids and tissues of infected horses. HeV is a biosecurity level 4 (BSL-4) pathogen, with a high case-fatality rate in humans and horses. Current assays for HeV detection require complex instrumentation and are generally time consuming. The aim of this study was to develop a Loop-Mediated Isothermal Amplification (LAMP) assay to detect nucleic acid from all known HeV strains in horses without the requirement for complex laboratory equipment. A LAMP assay targeting a conserved region of the HeV P-gene was combined with a Lateral Flow Device (LFD) for detection of amplified product. All HeV isolates, the original HeV isolated in 1994 as well as the most recent isolates from 2011 were detected. Analytical sensitivity and specificity of the HeV-LAMP assay was equal to a TaqMan assay developed previously. Significantly, these assays detected HeV in horses before clinical signs were observed. The combined LAMP-LFD procedure is a sensitive method suitable for HeV diagnosis in a resource-limited situation or where rapid test results are critical.

  19. Hendra virus detection using Loop-Mediated Isothermal Amplification.

    PubMed

    Foord, Adam J; Middleton, Deborah; Heine, Hans G

    2012-04-01

    Hendra virus (HeV) is a zoonotic paramyxovirus endemic in Australian Pteropus bats (fruit bats or flying foxes). Although bats appear to be unaffected by the virus, HeV can spread from fruit bats to horses, causing severe disease. Human infection results from close contact with the blood, body fluids and tissues of infected horses. HeV is a biosecurity level 4 (BSL-4) pathogen, with a high case-fatality rate in humans and horses. Current assays for HeV detection require complex instrumentation and are generally time consuming. The aim of this study was to develop a Loop-Mediated Isothermal Amplification (LAMP) assay to detect nucleic acid from all known HeV strains in horses without the requirement for complex laboratory equipment. A LAMP assay targeting a conserved region of the HeV P-gene was combined with a Lateral Flow Device (LFD) for detection of amplified product. All HeV isolates, the original HeV isolated in 1994 as well as the most recent isolates from 2011 were detected. Analytical sensitivity and specificity of the HeV-LAMP assay was equal to a TaqMan assay developed previously. Significantly, these assays detected HeV in horses before clinical signs were observed. The combined LAMP-LFD procedure is a sensitive method suitable for HeV diagnosis in a resource-limited situation or where rapid test results are critical. PMID:22327143

  20. Detection of RNA viruses: current technologies and future perspectives.

    PubMed

    Cella, Lakshmi N; Blackstock, Daniel; Yates, Marylynn A; Mulchandani, Ashok; Chen, Wilfred

    2013-01-01

    RNA viruses constitute one of the major classes of pathogenic organisms causing human diseases, with varying degrees of severity. This review summarizes the conventional and emerging technologies that are available for the detection of these organisms. Cell culture-based techniques for viral detection have been popular since their inception and continue to be the gold standard against which all other techniques. Over many years, these techniques have undergone some radical changes, reducing the total time needed for detection and improving sensitivity, although even with their reliability and improved features they are being slowly replaced by nucleic acid-based technologies. These molecular detection techniques have revolutionized the area of viral detection by their high sensitivity, selectivity, and short detection time. The majority of nucleic acid-based techniques depend on amplifying viral RNA; however, there are some newer emerging techniques that detect viral RNA in live cells using various configurations of florescent probes. In addition, nucleic acid-based technology has made it possible for multiviral detection with either multiplex polymerase chain reaction assays or microarrays. Every technique described in this review has its own unique abilities, making them indispensable for viral detection. However, we believe that nucleic acid-based technologies will find widespread use after being standardized, limiting other technologies to very specific uses. PMID:23582035

  1. Quantitative PCR detection for abalone shriveling syndrome-associated virus.

    PubMed

    Jiang, Jing-Zhe; Zhu, Zhen-Ni; Zhang, Han; Liang, Ya-Yu; Guo, Zhi-Xun; Liu, Guang-Feng; Su, You-Lu; Wang, Jiang-Yong

    2012-09-01

    Haliotis diversicolor (small abalone) is an important seafood found along the southern coast of China. Since 1999, the yields of cultured abalone in China have been severely affected by an epidemic of continuous outbreaks of a fatal disease. A novel double-stranded DNA virus, abalone shriveling syndrome-associated virus (AbSV), was proven to be one of the main causative agent. Although the pathogenicity and genome of AbSV has been ascertained, the epidemiology of AbSV remains to be investigated. In this study, four pairs of AbSV-specific primers were designed on the basis of the AbSV genome, and were tested for their specificities and sensitivities in quantitative real-time PCRs (qPCRs) after optimization of the annealing temperature. The 3F3/3B3 primer pair was finally chosen with a good specificity and high efficiency of amplification, with a detection limit of about 10 copies of recombinant plasmid containing AbSV genes in a 20-μL reaction mixture. In the detection of AbSV in abalone samples along the southern coast of China, most of the diseased samples had more than 80 virus copies in 1ng host genome DNA. AbSV was also demonstrated in mature hybrid (LY) and juvenile (JH) abalones from assays of healthy animals collected in recent years.

  2. Biosensor for dengue virus detection: sensitive, rapid, and serotype specific.

    PubMed

    Baeumner, Antje J; Schlesinger, Nicole A; Slutzki, Naomi S; Romano, Joseph; Lee, Eun Mi; Montagna, Richard A

    2002-03-15

    A serotype-specific RNA biosensor was developed for the rapid detection of Dengue virus (serotypes 1-4) in blood samples. After RNA amplification, the biosensor allows the rapid detection of Dengue virus RNA in only 15 min. In addition, the biosensor is portable, inexpensive, and very easy to use, making it an ideal detection system for point-of-care and field applications. The biosensor is coupled to the isothermal nucleic acid sequence-based amplification (NASBA) technique with which small amounts of virus RNA are amplified using a simple water bath. During the NASBA reaction, a generic sequence is attached to all RNA molecules as described earlier (Wu, S. J.; Lee, E. M.; Putvatana, R.; Shurtliff, R. N.; Porter, K R.; Suharyono, W.; Watt, D. M.; King, C. C.; Murphy, G. S.; Hayes, C. G.; Romano, J. W. J. Clin. Microbiol. 2001, 39, 2794-2798.). It has been shown earlier that Dengue virus can be detected specifically using two DNA probes: a first probe hybridized with the attached generic sequence and, therefore, bound to every amplified RNA molecule; and a second probe either bound to all four Dengue virus serotypes or chosen to be specific for only one serotype. These probes were utilized in the biosensor described in this publication. For a generic Dengue virus biosensor, the second probe is complementary to a conserved region found in all Dengue serotypes. For identification of the individual Dengue virus serotypes, four serotype-specific probes were developed (Wu, S. J.; Lee, E. M.; Putvatana, R.; Shurtiff, R. N.; Porter, K. R.; Suharyono, W.; Watt, D. M.; King, C. C.; Murphy, G. S.; Hayes, C. G.; Romano, J. W. J. Clin. Microbiol. 2001, 39, 2794-2798.). The biosensor is a membrane-based DNA/RNA hybridization system using liposome amplification. The generic DNA probe (reporter probe) is coupled to the outside of dye-encapsulating liposomes. The conserved or Dengue serotype specific probes (capture probes) are immobilized on a polyethersulfone membrane strip

  3. Propagation, quantification, detection, and storage of West Nile virus.

    PubMed

    Brien, James D; Lazear, Helen M; Diamond, Michael S

    2013-01-01

    West Nile virus (WNV) is a member of the Flaviviridae family of enveloped, single-stranded, positive-sense RNA viruses. WNV, an emerging viral pathogen, is transmitted by mosquitoes to birds and mammals and is responsible for an increasing incidence of human disease in North America and Europe. Due to its ease of use in the laboratory and the availability of robust mouse models of disease, WNV provides an excellent experimental system for studying molecular virology and pathogenesis of infection by flaviviruses. Here, we describe common laboratory techniques used to propagate, quantify, detect, and store WNV. We also briefly describe appropriate safety precautions required for the laboratory use of WNV, which is classified as a Biosafety Level 3 pathogen by the United States Centers for Disease Control and Prevention.

  4. The microbial detection array for detection of emerging viruses in clinical samples--a useful panmicrobial diagnostic tool.

    PubMed

    Rosenstierne, Maiken W; McLoughlin, Kevin S; Olesen, Majken Lindholm; Papa, Anna; Gardner, Shea N; Engler, Olivier; Plumet, Sebastien; Mirazimi, Ali; Weidmann, Manfred; Niedrig, Matthias; Fomsgaard, Anders; Erlandsson, Lena

    2014-01-01

    Emerging viruses are usually endemic to tropical and sub-tropical regions of the world, but increased global travel, climate change and changes in lifestyle are believed to contribute to the spread of these viruses into new regions. Many of these viruses cause similar disease symptoms as other emerging viruses or common infections, making these unexpected pathogens difficult to diagnose. Broad-spectrum pathogen detection microarrays containing probes for all sequenced viruses and bacteria can provide rapid identification of viruses, guiding decisions about treatment and appropriate case management. We report a modified Whole Transcriptome Amplification (WTA) method that increases unbiased amplification, particular of RNA viruses. Using this modified WTA method, we tested the specificity and sensitivity of the Lawrence Livermore Microbial Detection Array (LLMDA) against a wide range of emerging viruses present in both non-clinical and clinical samples using two different microarray data analysis methods. PMID:24963710

  5. The microbial detection array for detection of emerging viruses in clinical samples--a useful panmicrobial diagnostic tool.

    PubMed

    Rosenstierne, Maiken W; McLoughlin, Kevin S; Olesen, Majken Lindholm; Papa, Anna; Gardner, Shea N; Engler, Olivier; Plumet, Sebastien; Mirazimi, Ali; Weidmann, Manfred; Niedrig, Matthias; Fomsgaard, Anders; Erlandsson, Lena

    2014-01-01

    Emerging viruses are usually endemic to tropical and sub-tropical regions of the world, but increased global travel, climate change and changes in lifestyle are believed to contribute to the spread of these viruses into new regions. Many of these viruses cause similar disease symptoms as other emerging viruses or common infections, making these unexpected pathogens difficult to diagnose. Broad-spectrum pathogen detection microarrays containing probes for all sequenced viruses and bacteria can provide rapid identification of viruses, guiding decisions about treatment and appropriate case management. We report a modified Whole Transcriptome Amplification (WTA) method that increases unbiased amplification, particular of RNA viruses. Using this modified WTA method, we tested the specificity and sensitivity of the Lawrence Livermore Microbial Detection Array (LLMDA) against a wide range of emerging viruses present in both non-clinical and clinical samples using two different microarray data analysis methods.

  6. Design of different strategies of multivalent DNA-based vaccination against rabies and canine distemper in mice and dogs

    PubMed Central

    2012-01-01

    Background During the vaccination campaigns, puppies younger than 3 months old are not targeted and remain unvaccinated for at least the first year of their lives. Almost half of the reported rabid dogs are 6 months or younger. Hence, we should recommend the vaccination against rabies of young puppies. Unfortunately, owing to the exposure of puppies to infections with either canine parvovirus (CPV) or distemper virus (CDV) after the intervention of the vaccinators, owners are reluctant to vaccinate puppies against rabies. Therefore, it is necessary to include the CPV and CDV valences in the vaccine against rabies. Multivalent DNA-based vaccination in dogs, including rabies and distemper valences, could help in raising vaccine coverage. Methods We have designed monovalent and multivalent DNA-based vaccine candidates for in vitro and in vivo assays. These plasmids encode to the rabies virus glycoprotein and/or the canine distemper virus hemagglutinin. The first strategy of multivalent DNA-based vaccination is by mixing plasmids encoding to a single antigen each. The second is by simply fusing the genes of the antigens together. The third is by adding the foot and mouth disease virus (FMDV) 2A oligopeptide gene into the antigen genes. The last strategy is by the design and use of a bicistronic plasmid with an “Internal Ribosome Entry Site” (IRES) domain. Results The monovalent construct against canine distemper was efficiently validated by inducing higher humoral immune responses compared to cell-culture-derived vaccine both in mice and dogs. All multivalent plasmids efficiently expressed both valences after in vitro transfection of BHK-21 cells. In BALB/c mice, the bicistronic IRES-dependant construct was the most efficient inducer of virus-neutralizing antibodies against both valences. It was able to induce better humoral immune responses compared to the administration of either cell-culture-derived vaccines or monovalent plasmids. The FMDV 2A was also efficient

  7. Detection of sweet potato viruses in Yunnan and genetic diversity analysis of the common viruses

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Two hundred seventy-nine samples with virus-like symptoms collected from 16 regions in Yunnan Province were tested by RT-PCR/PCR using virus-specific primers for 8 sweet potato viruses. Six viruses, Sweet potato chlorotic fleck virus (SPCFV), Sweet Potato feathery mottle virus (SPFMV), Sweet potato ...

  8. Detection of hepatitis B virus infection: A systematic review

    PubMed Central

    Ghosh, Mallika; Nandi, Srijita; Dutta, Shrinwanti; Saha, Malay Kumar

    2015-01-01

    AIM: To review published methods for detection of hepatitis B virus (HBV) infection. METHODS: A thorough search on Medline database was conducted to find original articles describing different methods or techniques of detection of HBV, which are published in English in last 10 years. Articles outlining methods of detection of mutants or drug resistance were excluded. Full texts and abstracts (if full text not available) were reviewed thoroughly. Manual search of references of retrieved articles were also done. We extracted data on different samples and techniques of detection of HBV, their sensitivity (Sn), specificity (Sp) and applicability. RESULTS: A total of 72 studies were reviewed. HBV was detected from dried blood/plasma spots, hepatocytes, ovarian tissue, cerumen, saliva, parotid tissue, renal tissue, oocytes and embryos, cholangiocarcinoma tissue, etc. Sensitivity of dried blood spot for detecting HBV was > 90% in all the studies. In case of seronegative patients, HBV DNA or serological markers have been detected from hepatocytes or renal tissue in many instances. Enzyme linked immunosorbent assay and Chemiluminescent immunoassay (CLIA) are most commonly used serological tests for detection. CLIA systems are also used for quantitation. Molecular techniques are used qualitatively as well as for quantitative detection. Among the molecular techniques version 2.0 of the CobasAmpliprep/CobasTaqMan assay and Abbott’s real time polymerase chain reaction kit were found to be most sensitive with a lower detection limit of only 6.25 IU/mL and 1.48 IU/mL respectively. CONCLUSION: Serological and molecular assays are predominant and reliable methods for HBV detection. Automated systems are highly sensitive and quantify HBV DNA and serological markers for monitoring. PMID:26483870

  9. Dengue virus detection in Aedes aegypti larvae from southeastern Brazil.

    PubMed

    Cecílio, Samyra Giarola; Júnior, Willer Ferreira Silva; Tótola, Antônio Helvécio; de Brito Magalhães, Cíntia Lopes; Ferreira, Jaqueline Maria Siqueira; de Magalhães, José Carlos

    2015-06-01

    The transmission of dengue, the most important arthropod-borne viral disease in Brazil, has been intensified over the past decades, along with the accompanying expansion and adaptation of its Aedes vectors. In the present study, we mapped dengue vectors in Ouro Preto and Ouro Branco, Minas Gerais, by installing ovitraps in 32 public schools. The traps were examined monthly between September, 2011 through July, 2012 and November, 2012 to April, 2013. The larvae were reared until the fourth stadium and identified according to species. The presence of dengue virus was detected by real time PCR and agarose gel electrophoresis. A total of 1,945 eggs was collected during the 17 months of the study. The Ovitrap Positivity Index (OPI) ranged from 0 to 28.13% and the Eggs Density Index (EDI) ranged from 0 to 59.9. The predominant species was Aedes aegypti, with 84.9% of the hatched larvae. Although the collection was low when compared to other ovitraps studies, vertical transmission could be detected. Of the 54 pools, dengue virus was detected in four Ae. aegypti pools. PMID:26047186

  10. ELISA for the detection of antibodies to Ebola viruses.

    PubMed

    Ksiazek, T G; West, C P; Rollin, P E; Jahrling, P B; Peters, C J

    1999-02-01

    EIAs for IgG and IgM antibodies directed against Ebola (EBO) viral antigens have been developed and evaluated using sera of animals and humans surviving infection with EBO viruses. The IgM capture assay detected anti-EBO (subtype Reston) antibodies in the sera of 5 of 5 experimentally infected animals at the time they succumbed to lethal infections. IgM antibodies were also detected in the serum of a human who was infected with EBO (subtype Reston) during a postmortem examination of an infected monkey. The antibody was detectable as early as day 6 after infection in experimentally infected animals and persisted for <90 days. The IgG response was less rapid; however, it persisted for >400 days in 3 animals who survived infection, and it persisted for approximately 10 years after infection in the sera of 2 humans. Although these data are limited by the number of sera available for verification, the IgM assay seems to have great promise as a diagnostic tool. Furthermore the long-term persistence of the IgG antibodies measured by this test strongly suggests that the ELISA will be useful in field investigations of EBO virus. PMID:9988184

  11. Rapid Detection of Herpes Viruses for Clinical Applications

    NASA Technical Reports Server (NTRS)

    Pierson, Duane; Mehta, Satish

    2013-01-01

    There are eight herpes viruses that infect humans, causing a wide range of diseases resulting in considerable morbidity and associated costs. Varicella zoster virus (VZV) is a human herpes virus that causes chickenpox in children and shingles in adults. Approximately 1,000,000 new cases of shingles occur each year; post-herpetic neuralgia (PHN) follows shingles in 100,000 to 200,000 people annually. PHN is characterized by debilitating, nearly unbearable pain for weeks, months, and even years. The onset of shingles is characterized by pain, followed by the zoster rash, leading to blisters and severe pain. The problem is that in the early stages, shingles can be difficult to diagnose; chickenpox in adults can be equally difficult to diagnose. As a result, both diseases can be misdiagnosed (false positive/negative). A molecular assay has been adapted for use in diagnosing VZV diseases. The polymerase chain reaction (PCR) assay is a non-invasive, rapid, sensitive, and highly specific method for VZV DNA detection. It provides unequivocal results and can effectively end misdiagnoses. This is an approximately two-hour assay that allows unequivocal diagnosis and rapid antiviral drug intervention. It has been demonstrated that rapid intervention can prevent full development of the disease, resulting in reduced likelihood of PHN. The technology was extended to shingles patients and demonstrated that VZV is shed in saliva and blood of all shingles patients. The amount of VZV in saliva parallels the medical outcome.

  12. Insect-specific flaviviruses, a worldwide widespread group of viruses only detected in insects.

    PubMed

    Calzolari, Mattia; Zé-Zé, Líbia; Vázquez, Ana; Sánchez Seco, Mari Paz; Amaro, Fátima; Dottori, Michele

    2016-06-01

    Several flaviviruses are important pathogens for humans and animals (Dengue viruses, Japanese encephalitis virus, Yellow-fever virus, Tick-borne encephalitis virus, West Nile virus). In recent years, numerous novel and related flaviviruses without known pathogenic capacity have been isolated worldwide in the natural mosquito population. However, phylogenetic studies have shown that genomic sequences of these viruses diverge from other flaviviruses. Moreover, these viruses seem to be exclusive of insects (they do not seem to grow on vertebrate cell lines), and were already defined as mosquito-only flaviviruses or insect-specific flaviviruses. At least eleven of these viruses were isolated worldwide, and sequences ascribable to other eleven putative viruses were detected in several mosquito species. A large part of the cycle of these viruses is not well known, and their persistence in the environment is poorly understood. These viruses are detected in a wide variety of distinct mosquito species and also in sandflies and chironomids worldwide; a single virus, or the genetic material ascribable to a virus, was detected in several mosquito species in different countries, often in different continents. Furthermore, some of these viruses are carried by invasive mosquitoes, and do not seem to have a depressive action on their fitness. The global distribution and the continuous detection of new viruses in this group point out the likely underestimation of their number, and raise interesting issues about their possible interactions with the pathogenic flaviviruses, and their influence on the bionomics of arthropod hosts. Some enigmatic features, as their integration in the mosquito genome, the recognition of their genetic material in DNA forms in field-collected mosquitoes, or the detection of the same virus in both mosquitoes and sandflies, indicate that the cycle of these viruses has unknown characteristics that could be of use to reach a deeper understanding of the cycle

  13. Mini review: current molecular methods for the detection and quantification of hepatitis B virus, hepatitis C virus, and human immunodeficiency virus type 1.

    PubMed

    Albertoni, Guilherme; Castelo Girão, Manoel João Batista; Schor, Nestor

    2014-08-01

    The detection of acute human immunodeficiency virus (HIV), hepatitis B virus (HBV), and hepatitis C virus (HCV) infection is vital for controlling the spread of HIV, HBV, and HCV to uninfected individuals. Considering that these viruses have high replication rates and are undetectable by serological markers, early detection upon transmission is crucial. Various nucleic acid assays have been developed for diagnostics and therapeutic monitoring of infections. In the past decade, rapid and sensitive molecular techniques such as PCR have revolutionized the detection of a variety of infectious viruses, including HIV, HCV, and HBV. Here, we describe two of the most commonly used licensed methods for the detection and quantification of HIV, HCV, and HBV: the cobas TaqScreen MPX (PCR) test and the Tigris System. We used transcription-mediated amplification to review and compare the development and efficiency of these technologies. PMID:24927665

  14. Rapid detection of pseudorabies virus by the shell vial technique.

    PubMed

    Tahir, R A; Goyal, S M

    1995-04-01

    Conventional 24-well microtiter plates and shell vials were seeded with pig kidney (PK-15) and bovine turbinate (BT) cells. The monolayers were inoculated with 244 clinical specimens from pigs suspected of having pseudorabies virus (PRV) infection. The results of a shell vial assay (SVA) were compared with those obtained in a 24-well plate cell culture assay in terms of sensitivity and speed of virus isolation. All samples were passaged only once in cell cultures in both assays. Samples producing cytopathic effects (cpe) in 1 or both assay systems and showing positive fluorescence in a direct fluorescent antibody assay were considered to be positive for PRV. Of the 244 samples examined, 118 (48.4%) and 121 (49.2%) were positive by the 24-well plate assay and SVA, respectively. Of the 118 samples positive in 24-well plates, 113 (95.8%) were positive in BT cells and 117 (99.2%) were positive in PK-15 cells. The SVA detected 121 positive samples of which 121 (100%) were positive in PK-15 cells and 113 (93.4%) were positive in BT cells. Virus-specific cpe appeared earlier in the SVA than in the 24-well assay. At 24 hours postinoculation, 91 (75.2%) samples were cpe positive by SVA, whereas only 15 (12.7%) were positive in 24-well plates. All but 2 of the 121 (98.3%) SVA-positive samples were positive within 48 hours postinoculation, whereas only 56 of 118 (47.5%) were positive in 24-well plates during the same time period. These results indicate that the SVA is comparable in sensitivity to 24-well plate assay but yields virus isolation results more quickly. Also, PK-15 cells appeared to be more sensitive than BT cells for PRV isolation. PMID:7619897

  15. Infectivity titration of hemorrhagic fever with renal syndrome virus: use of immune adherence hemagglutination for detection of virus growth.

    PubMed

    Matsuura, Y; Sugiyama, K; Morita, C; Morikawa, S; Shiga, S; Komatsu, T; Akao, Y; Kitamura, T

    1984-09-01

    Serial dilutions of hemorrhagic fever with renal syndrome viruses were inoculated into Vero-E6 cells in microplates. After 2 weeks of incubation, infected cells were disrupted by freezing and thawing, and virus antigens were detected by immune adherence hemagglutination. The infectivity titers of the virus as determined by this method were in close agreement with those obtained by the immunofluorescent antigen endpoint method. Then, a neutralization method was established. Japanese hemorrhagic fever with renal syndrome isolates, strains SR-11 and TR-352, were found to be distinct from Hantaan virus, strain 76-118, by the neutralization test.

  16. Distemper in raccoons and foxes suspected of having rabies

    USGS Publications Warehouse

    Habermann, R.T.; Herman, C.M.; Williams, F.P.

    1958-01-01

    1) Twenty-one raccoons and 3 red foxes were collected from areas where suspected rabies occurred. All were found to be nonrabid. 2) Distemper was diagnosed in 14 of the 21 raccoons by demonstrating intracytoplasmic and intranuclear inclusions in the brain and visceral tissues. Two of the 3 foxes were considered to have distemper; the clinical signs were typical and mouse inoculation tests were negative for rabies. 3) Deaths of the other 7 raccoons were attributed to: leishmaniasis 1, gastritis 1, bronchopneumonia 1, parasitism 2, car injury 1; 1 showed no significant lesions. The death of 1 fox was attributed to parasitism. 4) Distemper may be a frequent cause of death in raccoons and foxes, in epizootics which simulate rabies.

  17. Rapid and quantitative detection of hepatitis B virus

    PubMed Central

    Liu, Yue-Ping; Yao, Chun-Yan

    2015-01-01

    Despite availability of a universal vaccine, hepatitis B virus (HBV) infection has a huge impact on public health worldwide. Accurate and timely diagnosis of HBV infection is needed. Rapid developments have been made in the diagnostic and monitoring methods for HBV infection, including serological and molecular assays. In clinical practice, qualitative hepatitis B surface antigen (HBsAg) testing has long served as a diagnostic marker for individuals infected with HBV. More recently, HBsAg level has been used to predict treatment outcome when determined early during treatment or at baseline. However, identification of HBV DNA positive cases that do not have detectable HBsAg has encouraged the application of molecular tests. Hence, combination of quantitative detection of HBV DNA and HBsAg can be used to discriminate patients during the course of HBV infection and to monitor therapy. This article reviews the most commonly used quantitative methods for HBsAg and HBV DNA. PMID:26576084

  18. Biosensor based on nanocomposite material for pathogenic virus detection.

    PubMed

    Van Thu, Vu; Dung, Phuong Trung; Tam, Le Thi; Tam, Phuong Dinh

    2014-03-01

    This paper introduces a DNA biosensor based on a DNA/chitosan/multi-walled carbon nanotube nanocomposite for pathogenic virus detection. An easy, cost-effective approach to the immobilization of probe DNA sequences on the sensor surface was performed. Cyclic voltammograms were used to characterize the probe DNA sequence immobilization. Complementary sequence hybridization was examined by electrochemical impedance spectroscopy. Results revealed that the developed DNA sensor can detect a target DNA concentration as low as 0.01×10(-12) M. The sensitivity of the prepared sensor was 52.57 kΩ/fM. The reusability and storage stability of the DNA sensor were also investigated. Results showed that the electron-transfer resistance decreased to approximately 35% after 8 weeks and to approximately 80% after 12 weeks of storage.

  19. Accumulation of Extracellular Matrix in Advanced Lesions of Canine Distemper Demyelinating Encephalitis

    PubMed Central

    Seehusen, Frauke; Al-Azreg, Seham A.; Raddatz, Barbara B.; Haist, Verena; Puff, Christina; Spitzbarth, Ingo; Ulrich, Reiner; Baumgärtner, Wolfgang

    2016-01-01

    In demyelinating diseases, changes in the quality and quantity of the extracellular matrix (ECM) may contribute to demyelination and failure of myelin repair and axonal sprouting, especially in chronic lesions. To characterize changes in the ECM in canine distemper demyelinating leukoencephalitis (DL), histochemical and immunohistochemical investigations of formalin-fixed paraffin-embedded cerebella using azan, picrosirius red and Gomori`s silver stain as well as antibodies directed against aggrecan, type I and IV collagen, fibronectin, laminin and phosphacan showed alterations of the ECM in CDV-infected dogs. A significantly increased amount of aggrecan was detected in early and late white matter lesions. In addition, the positive signal for collagens I and IV as well as fibronectin was significantly increased in late lesions. Conversely, the expression of phosphacan was significantly decreased in early and more pronounced in late lesions compared to controls. Furthermore, a set of genes involved in ECM was extracted from a publically available microarray data set and was analyzed for differential gene expression. Gene expression of ECM molecules, their biosynthesis pathways, and pro-fibrotic factors was mildly up-regulated whereas expression of matrix remodeling enzymes was up-regulated to a relatively higher extent. Summarized, the observed findings indicate that changes in the quality and content of ECM molecules represent important, mainly post-transcriptional features in advanced canine distemper lesions. Considering the insufficiency of morphological regeneration in chronic distemper lesions, the accumulated ECM seems to play a crucial role upon regenerative processes and may explain the relatively small regenerative potential in late stages of this disease. PMID:27441688

  20. Antibody Derived Peptides for Detection of Ebola Virus Glycoprotein

    PubMed Central

    López-Pacheco, Felipe; Pérez-Chavarría, Roberto; González-Vázquez, Juan Carlos; González-González, Everardo; Trujillo-de Santiago, Grissel; Ponce-Ponce de León, César Alejandro; Zhang, Yu Shrike; Dokmeci, Mehmet Remzi; Khademhosseini, Ali; Alvarez, Mario Moisés

    2015-01-01

    Background Current Ebola virus (EBOV) detection methods are costly and impractical for epidemic scenarios. Different immune-based assays have been reported for the detection and quantification of Ebola virus (EBOV) proteins. In particular, several monoclonal antibodies (mAbs) have been described that bind the capsid glycoprotein (GP) of EBOV GP. However, the currently available platforms for the design and production of full-length mAbs are cumbersome and costly. The use of antibody fragments, rather than full-length antibodies, might represent a cost-effective alternative for the development of diagnostic and possibly even therapeutic alternatives for EBOV. Methods/Principal Findings We report the design and expression of three recombinant anti-GP mAb fragments in Escherichia coli cultures. These fragments contained the heavy and light variable portions of the three well-studied anti-GP full-length mAbs 13C6, 13F6, and KZ52, and are consequently named scFv-13C6, scFv-13F6, and Fab-KZ52, respectively. All three fragments exhibited specific anti-GP binding activity in ELISA experiments comparable to that of full-length anti-GP antibodies (i.e., the same order of magnitude) and they are easily and economically produced in bacterial cultures. Conclusion/Significance Antibody fragments might represent a useful, effective, and low cost alternative to full-length antibodies in Ebola related capture and diagnostics applications. PMID:26489048

  1. IMPROVED DETECTION OF HUMAN ENTERIC VIRUSES IN FOODS BY RT-PCR. (R826139)

    EPA Science Inventory

    Human enteric viruses (including hepatitis A virus (HAV) and Norwalk-like viruses (NLVs)) are now recognized as common causes of foodborne disease. While methods to detect these agents in clinical specimens have improved significantly over the last 10 years, applications to fo...

  2. Clinical Outcomes Associated With Respiratory Virus Detection Before Allogeneic Hematopoietic Stem Cell Transplant

    PubMed Central

    Campbell, Angela P.; Guthrie, Katherine A.; Englund, Janet A.; Farney, Robert M.; Minerich, Elisa L.; Kuypers, Jane; Corey, Lawrence; Boeckh, Michael

    2015-01-01

    Background. The management of respiratory virus infections prior to hematopoietic cell transplant (HCT) is difficult. We examined whether respiratory virus detection before HCT influenced the requirement for bronchoscopy, hospitalization, and overall survival following HCT. Methods. Pre-HCT and weekly post-HCT nasal washes were collected through day 100 from patients with and without symptoms. Samples were tested by multiplex polymerase chain reaction for respiratory syncytial virus, parainfluenza viruses 1–4, influenza A and B, human metapneumovirus, adenovirus, and human rhinoviruses, coronaviruses, and bocavirus. Results. Of 458 patients, 116 (25%) had respiratory viruses detected pre-HCT. Overall, patients with viruses detected pre-HCT had fewer days alive and out of the hospital and lower survival at day 100 (adjusted hazard ratio [aHR], 2.4; 95% confidence interval [CI], 1.3–4.5; P = .007) than patients with negative samples; this risk was also present with rhinovirus alone (aHR for mortality, 2.6; 95% CI, 1.2–5.5; P = .01). No difference in bronchoscopy incidence was seen in patients with and without respiratory viruses (aHR, 1.3; 95% CI, .8–2.0; P = .32). In symptomatic patients, those with respiratory viruses detected had increased overall mortality compared with patients without viruses detected (unadjusted HR, 3.5; 95% CI, 1.0–12.1; P = .05); among asymptomatic patients, detection of respiratory viruses was not associated with increased mortality. Conclusions. These data support routine testing for respiratory viruses among symptomatic patients before HCT, and delay of transplant with virus detection when feasible, even for detection of rhinovirus alone. Further study is needed to address whether asymptomatic patients should undergo screening for respiratory virus detection before HCT. PMID:25847977

  3. A novel multiplex isothermal amplification method for rapid detection and identification of viruses.

    PubMed

    Nyan, Dougbeh-Chris; Swinson, Kevin L

    2015-12-08

    A rapid multiplex isothermal amplification assay has been developed for detection and identification of multiple blood-borne viruses that infect millions of people world-wide. These infections may lead to chronic diseases or death if not diagnosed and treated in a timely manner. Sets of virus-specific oligonucleotides and oligofluorophores were designed and used in a reverse-transcription loop-mediated multiplexed isothermal amplification reaction for detection and gel electrophoretic identification of human Immunodeficiency virus (HIV), hepatitis-B virus (HBV), hepatitis-C virus (HCV), hepatitis-E virus (HEV), dengue virus (DENV), and West Nile (WNV) virus infection in blood plasma. Amplification was catalyzed with two thermostable enzymes for 30-60 minutes under isothermal condition, utilizing a simple digital heat source. Electrophoretic analysis of amplified products demonstrated simultaneous detection of 6 viruses that were distinctly identified by unique ladder-like banding patterns. Naked-eye fluorescent visualization of amplicons revealed intensely fluorescing products that indicated positive detection. The test demonstrated a 97% sensitivity and a 100% specificity, with no cross-reaction with other viruses observed. This portable detection tool may have clinical and field utility in the developing and developed world settings. This may enable rapid diagnosis and identification of viruses for targeted therapeutic intervention and prevention of disease transmission.

  4. A novel multiplex isothermal amplification method for rapid detection and identification of viruses.

    PubMed

    Nyan, Dougbeh-Chris; Swinson, Kevin L

    2015-01-01

    A rapid multiplex isothermal amplification assay has been developed for detection and identification of multiple blood-borne viruses that infect millions of people world-wide. These infections may lead to chronic diseases or death if not diagnosed and treated in a timely manner. Sets of virus-specific oligonucleotides and oligofluorophores were designed and used in a reverse-transcription loop-mediated multiplexed isothermal amplification reaction for detection and gel electrophoretic identification of human Immunodeficiency virus (HIV), hepatitis-B virus (HBV), hepatitis-C virus (HCV), hepatitis-E virus (HEV), dengue virus (DENV), and West Nile (WNV) virus infection in blood plasma. Amplification was catalyzed with two thermostable enzymes for 30-60 minutes under isothermal condition, utilizing a simple digital heat source. Electrophoretic analysis of amplified products demonstrated simultaneous detection of 6 viruses that were distinctly identified by unique ladder-like banding patterns. Naked-eye fluorescent visualization of amplicons revealed intensely fluorescing products that indicated positive detection. The test demonstrated a 97% sensitivity and a 100% specificity, with no cross-reaction with other viruses observed. This portable detection tool may have clinical and field utility in the developing and developed world settings. This may enable rapid diagnosis and identification of viruses for targeted therapeutic intervention and prevention of disease transmission. PMID:26643761

  5. A novel multiplex isothermal amplification method for rapid detection and identification of viruses

    PubMed Central

    Nyan, Dougbeh-Chris; Swinson, Kevin L.

    2015-01-01

    A rapid multiplex isothermal amplification assay has been developed for detection and identification of multiple blood-borne viruses that infect millions of people world-wide. These infections may lead to chronic diseases or death if not diagnosed and treated in a timely manner. Sets of virus-specific oligonucleotides and oligofluorophores were designed and used in a reverse-transcription loop-mediated multiplexed isothermal amplification reaction for detection and gel electrophoretic identification of human Immunodeficiency virus (HIV), hepatitis-B virus (HBV), hepatitis-C virus (HCV), hepatitis-E virus (HEV), dengue virus (DENV), and West Nile (WNV) virus infection in blood plasma. Amplification was catalyzed with two thermostable enzymes for 30–60 minutes under isothermal condition, utilizing a simple digital heat source. Electrophoretic analysis of amplified products demonstrated simultaneous detection of 6 viruses that were distinctly identified by unique ladder-like banding patterns. Naked-eye fluorescent visualization of amplicons revealed intensely fluorescing products that indicated positive detection. The test demonstrated a 97% sensitivity and a 100% specificity, with no cross-reaction with other viruses observed. This portable detection tool may have clinical and field utility in the developing and developed world settings. This may enable rapid diagnosis and identification of viruses for targeted therapeutic intervention and prevention of disease transmission. PMID:26643761

  6. F F{sub 1}-ATPase as biosensor to detect single virus

    SciTech Connect

    Liu, XiaoLong; Zhang, Yun; Yue, JiaChang . E-mail: yuejc@sun5.ibp.ac.cn; Jiang, PeiDong; Zhang, ZhenXi

    2006-04-21

    F F{sub 1}-ATPase within chromatophore was constructed as a biosensor (immuno-rotary biosensor) for the purpose of capturing single virus. Capture of virus was based on antibody-antigen reaction. The detection of virus based on proton flux change driven by ATP-synthesis of F F{sub 1}-ATPase, which was indicated by F1300, was directly observed by a fluorescence microscope. The results demonstrate that the biosensor loading of virus particles has remarkable signal-to-noise ratio (3.8:1) compared to its control at single molecular level, and will be convenient, quick, and even super-sensitive for detecting virus particles.

  7. Charge detection mass spectrometry: Instrumentation & applications to viruses

    NASA Astrophysics Data System (ADS)

    Pierson, Elizabeth E.

    For over three decades, electrospray ionization (ESI) has been used to ionize non-covalent complexes and subsequently transfer the intact ion into the gas phase for mass spectrometry (MS) analysis. ESI generates a distribution of multiple charged ions, resulting in an m/z spectrum comprised of a series of peaks, known as a charge state envelope. To obtain mass information, the number of charges for each peak must be deduced. For smaller biological analytes like peptides, the charge states are sufficiently resolved and this process is straightforward. For macromolecular complexes exceeding ~100 kDa, this process is complicated by the broadening and shifting of charge states due to incomplete desolvation, salt adduction, and inherent mass heterogeneity. As the analyte mass approaches the MDa regime, the m/z spectrum is often comprised of a broad distribution of unresolved charge states. In such cases, mass determination is precluded. Charge detection mass spectrometry (CDMS) is an emerging MS technique for determining the masses of heterogeneous, macromolecular complexes. In CDMS, the m/z and z of single ions are measured concurrently so that mass is easily calculated. With this approach, deconvolution of an m/z spectrum is unnecessary. This measurement is carried out by passing macroions through a conductive cylinder. The induced image charge on the cylindrical detector provides information about m/z and z: the m/z is related to its time-of-flight through the detector, and the z is related to the intensity of the image charge. We have applied CDMS to study the self-assembly of virus capsids. Late-stage intermediates in the assembly of hepatitis B virus, a devastating human pathogen, have been identified. This is the first time that such intermediates have been detected and represent a significant advancement towards understanding virus capsid assembly. CDMS has also been used to identify oversized, non-icosahedral polymorphs in the assembly of woodchuck hepatitis

  8. Vy-PER: eliminating false positive detection of virus integration events in next generation sequencing data.

    PubMed

    Forster, Michael; Szymczak, Silke; Ellinghaus, David; Hemmrich, Georg; Rühlemann, Malte; Kraemer, Lars; Mucha, Sören; Wienbrandt, Lars; Stanulla, Martin; Franke, Andre

    2015-01-01

    Several pathogenic viruses such as hepatitis B and human immunodeficiency viruses may integrate into the host genome. These virus/host integrations are detectable using paired-end next generation sequencing. However, the low number of expected true virus integrations may be difficult to distinguish from the noise of many false positive candidates. Here, we propose a novel filtering approach that increases specificity without compromising sensitivity for virus/host chimera detection. Our detection pipeline termed Vy-PER (Virus integration detection bY Paired End Reads) outperforms existing similar tools in speed and accuracy. We analysed whole genome data from childhood acute lymphoblastic leukemia (ALL), which is characterised by genomic rearrangements and usually associated with radiation exposure. This analysis was motivated by the recently reported virus integrations at genomic rearrangement sites and association with chromosomal instability in liver cancer. However, as expected, our analysis of 20 tumour and matched germline genomes from ALL patients finds no significant evidence for integrations by known viruses. Nevertheless, our method eliminates 12,800 false positives per genome (80× coverage) and only our method detects singleton human-phiX174-chimeras caused by optical errors of the Illumina HiSeq platform. This high accuracy is useful for detecting low virus integration levels as well as non-integrated viruses. PMID:26166306

  9. Vy-PER: eliminating false positive detection of virus integration events in next generation sequencing data.

    PubMed

    Forster, Michael; Szymczak, Silke; Ellinghaus, David; Hemmrich, Georg; Rühlemann, Malte; Kraemer, Lars; Mucha, Sören; Wienbrandt, Lars; Stanulla, Martin; Franke, Andre

    2015-07-13

    Several pathogenic viruses such as hepatitis B and human immunodeficiency viruses may integrate into the host genome. These virus/host integrations are detectable using paired-end next generation sequencing. However, the low number of expected true virus integrations may be difficult to distinguish from the noise of many false positive candidates. Here, we propose a novel filtering approach that increases specificity without compromising sensitivity for virus/host chimera detection. Our detection pipeline termed Vy-PER (Virus integration detection bY Paired End Reads) outperforms existing similar tools in speed and accuracy. We analysed whole genome data from childhood acute lymphoblastic leukemia (ALL), which is characterised by genomic rearrangements and usually associated with radiation exposure. This analysis was motivated by the recently reported virus integrations at genomic rearrangement sites and association with chromosomal instability in liver cancer. However, as expected, our analysis of 20 tumour and matched germline genomes from ALL patients finds no significant evidence for integrations by known viruses. Nevertheless, our method eliminates 12,800 false positives per genome (80× coverage) and only our method detects singleton human-phiX174-chimeras caused by optical errors of the Illumina HiSeq platform. This high accuracy is useful for detecting low virus integration levels as well as non-integrated viruses.

  10. Vy-PER: eliminating false positive detection of virus integration events in next generation sequencing data

    PubMed Central

    Forster, Michael; Szymczak, Silke; Ellinghaus, David; Hemmrich, Georg; Rühlemann, Malte; Kraemer, Lars; Mucha, Sören; Wienbrandt, Lars; Stanulla, Martin; Franke, Andre

    2015-01-01

    Several pathogenic viruses such as hepatitis B and human immunodeficiency viruses may integrate into the host genome. These virus/host integrations are detectable using paired-end next generation sequencing. However, the low number of expected true virus integrations may be difficult to distinguish from the noise of many false positive candidates. Here, we propose a novel filtering approach that increases specificity without compromising sensitivity for virus/host chimera detection. Our detection pipeline termed Vy-PER (Virus integration detection bY Paired End Reads) outperforms existing similar tools in speed and accuracy. We analysed whole genome data from childhood acute lymphoblastic leukemia (ALL), which is characterised by genomic rearrangements and usually associated with radiation exposure. This analysis was motivated by the recently reported virus integrations at genomic rearrangement sites and association with chromosomal instability in liver cancer. However, as expected, our analysis of 20 tumour and matched germline genomes from ALL patients finds no significant evidence for integrations by known viruses. Nevertheless, our method eliminates 12,800 false positives per genome (80× coverage) and only our method detects singleton human-phiX174-chimeras caused by optical errors of the Illumina HiSeq platform. This high accuracy is useful for detecting low virus integration levels as well as non-integrated viruses. PMID:26166306

  11. High-throughput detection method for influenza virus.

    PubMed

    Kumar, Pawan; Bartoszek, Allison E; Moran, Thomas M; Gorski, Jack; Bhattacharyya, Sanjib; Navidad, Jose F; Thakar, Monica S; Malarkannan, Subramaniam

    2012-01-01

    Influenza virus is a respiratory pathogen that causes a high degree of morbidity and mortality every year in multiple parts of the world. Therefore, precise diagnosis of the infecting strain and rapid high-throughput screening of vast numbers of clinical samples is paramount to control the spread of pandemic infections. Current clinical diagnoses of influenza infections are based on serologic testing, polymerase chain reaction, direct specimen immunofluorescence and cell culture (1,2). Here, we report the development of a novel diagnostic technique used to detect live influenza viruses. We used the mouse-adapted human A/PR/8/34 (PR8, H1N1) virus (3) to test the efficacy of this technique using MDCK cells (4). MDCK cells (10(4) or 5 x 10(3) per well) were cultured in 96- or 384-well plates, infected with PR8 and viral proteins were detected using anti-M2 followed by an IR dye-conjugated secondary antibody. M2 (5) and hemagglutinin (1) are two major marker proteins used in many different diagnostic assays. Employing IR-dye-conjugated secondary antibodies minimized the autofluorescence associated with other fluorescent dyes. The use of anti-M2 antibody allowed us to use the antigen-specific fluorescence intensity as a direct metric of viral quantity. To enumerate the fluorescence intensity, we used the LI-COR Odyssey-based IR scanner. This system uses two channel laser-based IR detections to identify fluorophores and differentiate them from background noise. The first channel excites at 680 nm and emits at 700 nm to help quantify the background. The second channel detects fluorophores that excite at 780 nm and emit at 800 nm. Scanning of PR8-infected MDCK cells in the IR scanner indicated a viral titer-dependent bright fluorescence. A positive correlation of fluorescence intensity to virus titer starting from 10(2)-10(5) PFU could be consistently observed. Minimal but detectable positivity consistently seen with 10(2)-10(3) PFU PR8 viral titers demonstrated the high

  12. Comparison of oral and intramuscular recombinant canine distemper vaccination in African wild dogs (Lycaon pictus).

    PubMed

    Connolly, Maren; Thomas, Patrick; Woodroffe, Rosie; Raphael, Bonnie L

    2013-12-01

    A series of three doses of recombinant canary-pox-vectored canine distemper virus vaccine was administered at 1-mo intervals, orally (n = 8) or intramuscularly (n = 13), to 21 previously unvaccinated juvenile African wild dogs (Lycaon pictus) at the Wildlife Conservation Society's Bronx Zoo. Titers were measured by serum neutralization at each vaccination and at intervals over a period of 3.5-21.5 mo after the initial vaccination. All postvaccination titers were negative for orally vaccinated animals at all sampling time points. Of the animals that received intramuscular vaccinations, 100% had presumed protective titers by the end of the course of vaccination, but only 50% of those sampled at 6.5 mo postvaccination had positive titers. None of the three animals sampled at 21.5 mo postvaccination had positive titers. PMID:24450046

  13. Comparison of oral and intramuscular recombinant canine distemper vaccination in African wild dogs (Lycaon pictus).

    PubMed

    Connolly, Maren; Thomas, Patrick; Woodroffe, Rosie; Raphael, Bonnie L

    2013-12-01

    A series of three doses of recombinant canary-pox-vectored canine distemper virus vaccine was administered at 1-mo intervals, orally (n = 8) or intramuscularly (n = 13), to 21 previously unvaccinated juvenile African wild dogs (Lycaon pictus) at the Wildlife Conservation Society's Bronx Zoo. Titers were measured by serum neutralization at each vaccination and at intervals over a period of 3.5-21.5 mo after the initial vaccination. All postvaccination titers were negative for orally vaccinated animals at all sampling time points. Of the animals that received intramuscular vaccinations, 100% had presumed protective titers by the end of the course of vaccination, but only 50% of those sampled at 6.5 mo postvaccination had positive titers. None of the three animals sampled at 21.5 mo postvaccination had positive titers.

  14. Novel optical sensors for detection of toxins, viruses and bacteria

    NASA Astrophysics Data System (ADS)

    Emmerson, Gregory D.; Sparrow, Ian J. G.; Bhatta, Devaki; SohnaSohna, Jean E.

    2008-10-01

    A novel optical sensor system for rapid, sensitive and robust biological detection is presented. Sensor elements based on integrated optical circuits confine all optical signals into a planar format, resulting in a small, low-cost and mechanically stable refractive index sensor, without any external bulk optics. Consequently, the sensor elements are able to operate in real-world environments, resilient to vibration and temperature changes, whilst maintaining refractive index resolution of 10-6. Oxide surfaces on the sensor are ideal for protein attachment and have a long lifetime in buffer solutions (>100hrs). Real-time, label-free detection of biological agents has been demonstrated using antibodies attached to the sensor surface. The sensor design results in a large penetration depth of the sensing light, up to 1μm into the sample liquid, conferring the ability to detect various classes of biological targets, spanning toxins, viruses and bacteria. Each sensing element utilizes parallel multiple wavelength data to provide additional information at the point of measurement, resulting in on-chip temperature and strain referencing, focused towards increased accuracy and reduction of false alarms. The large size range of biological detection, coupled with the long lifetime of the sensors makes the system ideally suited to applications ranging from medical diagnostics to confirmatory detectors for homeland security

  15. Rapid Detection of the Varicella Zoster Virus in Saliva

    NASA Technical Reports Server (NTRS)

    Pierson, Duane L.; Mehta, Satish K.; Cohrs, Randall J.; Gilden, Don H.; Harding, Robert E.

    2011-01-01

    Varicella zoster virus (VZV) causes chicken pox on first exposure (usually in children), and reactivates from latency causing shingles (usually in adults). Shingles can be extremely painful, causing nerve damage, organ damage, and blindness in some cases. The virus can be life-threatening in immune-compromised individuals. The virus is very difficult to culture for diagnosis, requiring a week or longer. This invention is a rapid test for VZV from a saliva sample and can be performed in a doctor s office. The kit is small, compact, and lightweight. Detec tion is sensitive, specific, and noninvasive (no needles); only a saliva sample is required. The test provides results in minutes. The entire test is performed in a closed system, with no exposure to infectious materials. The components are made mostly of inexpensive plastic injection molded parts, many of which can be purchased off the shelf and merely assembled. All biological waste is contained for fast, efficient disposal. This innovation was made possible because of discovery of a NASA scientists flight experiment showing the presence of VZV in saliva during high stress periods and disease. This finding enables clinicians to quickly screen patients for VZV and treat the ones that show positive results with antiviral medicines. This promotes a rapid recovery, easing of pain and symptoms, and reduces chances of complications from zoster. Screening of high-risk patients could be incorporated as part of a regular physical exam. These patients include the elderly, pregnant women, and immune-compromised individuals. In these patients, VZV can be a life-threatening disease. In both high- and low-risk patients, early detection and treatment with antiviral drugs can dramatically decrease or even eliminate the clinical manifestation of disease.

  16. Immunoassay for detection of antibodies to simian immunodeficiency virus and human immunodeficiency virus in serum.

    PubMed

    Otsyula, M G; Yee, J A; Suleman, M A; Marx, P A; Jennings, M B

    1996-04-01

    We developed a simple, inexpensive, rapid assay for the detection of antibodies to simian immunodeficiency virus (SIV) and human immunodeficiency virus type 1 (HIV-1) in serum. The immunoassay uses inactivated SIV and HIV-1 gp41 transmembrane recombinant protein as antigenic adsorbents on a nitrocellulose filter membrane. Diluted serum, with the addition of Protein-A-Gold, is gravity-filtered through the filter membrane, blocked, and buffer-washed. Antibodies to HIV or SIV or both in serum bind to the appropriate antigen, and the resulting antigen-antibody complex reacts with Protein-A-Gold to produce a readable pink color. Field evaluation of the test on 30 human and 70 nonhuman primate sera in Kenya and Zaire indicated that the test had at least 93 and 90% correlation with Western blot sensitivity and specificity respectively. Prior refrigeration of the test kit and incubation of sera during testing were not required. This result indicates that the test may be a rapid, economical, and simple test for detecting HIV, SIV, or both in serum. This immunoassay can be useful for carrying out HIV and SIV serosurveys in countries with limited or no laboratory facilities. PMID:8723237

  17. Detection of Bovine viral diarrhea virus-specific neutralizing antibodies in fresh colostrum: a modification of the virus neutralization test.

    PubMed

    Bedekovic, Tomislav; Mihaljevic, Zeljko; Jungic, Andreja; Lemo, Nina; Lojkic, Ivana; Cvetnic, Zeljko; Cac, Zeljko

    2013-03-01

    To eliminate cytotoxic effects of colostrum on cells, a modified virus neutralization test (VNT) for the detection of Bovine viral diarrhea virus-specific neutralizing antibodies in colostrum was developed. The new test was compared to the World Organization for Animal Health-recommended VNT and the results evaluated. The agreement of the new test compared to the standard VNT was determined to be 98%, whereas sensitivity and specificity of the modified VNT compared to the standard VNT were 100%. Bovine viral diarrhea virus-specific antibodies were detected in 42 sera samples and 38 colostrum samples. The antibody titers in serum and colostrum showed a high correlation (n = 56, r = 0.9719, P < 0.001). The modified virus neutralization technique described herein succeeds in eliminating cytotoxic effects and can be readily applied for the detection of specific antibodies against other infectious agents in colostrum. PMID:23417081

  18. Canine distemper infections, with special reference to South Africa, with a review of the literature.

    PubMed

    Leisewitz, A L; Carter, A; van Vuuren, M; van Blerk, L

    2001-09-01

    Canine distemper virus is a member of the genus Morbillivirus of the family Paramyxoviridae that causes severe disease in dogs and a range of wild mammals. The clinical signs relate essentially to the respiratory, gastrointestinal and central nervous systems. In South Africa, infection with Ehrlichia canis and canine parvovirus may present similarly Many dogs will initially present with a wide range of central nervous system signs without any history of systemic disease. A recent South African study evaluating ante mortem diagnosis highlighted the importance of recognising clinical signs, cerebrospinal fluid IgG titres, serum IgM titres and immunocytochemistry of epithelial tissue. A 2-year retrospective evaluation of cerebrospinal fluid samples collected from dogs presented to the Onderstepoort Veterinary Academic Hospital indicates that distemper infection is common, and this disease should routinely be suspected in cases of diverse neurological disease in dogs. The South African dog population is specifically at high risk for the disease because of the large pool of unvaccinated, reproductively-active dogs that expose the wildlife resources of the country to risk of fatal disease. Outbreaks of disease in dogs continue to occur in developed and developing communities in both vaccinated and unvaccinated dogs worldwide, and have also been described in a wide range of free-ranging wildlife, including seals, dolphins and lions, and in endangered zoo animals. Modified live virus vaccines have contributed markedly to disease control in the dog population but have caused mortality in some wild carnivores. New recombinant vaccines are being developed that will be safe in wild animals. The pathogenesis of CNS demyelination has been compared to various important demyelinating diseases in humans and, amongst other things, relates to down-regulation of the oligodendrocyte gene coding for myelin synthesis and non-immunocyte CNS cell expression of type II major

  19. Efficacy of two canine distemper vaccines in wild Nearctic river otters (Lontra canadensis).

    PubMed

    Peper, Steven T; Peper, Randall L; Kollias, George V; Brooks, Robert P; Stevens, Sadie S; Serfass, Thomas L

    2014-09-01

    Canine distemper virus (CDV), a contagious morbillivirus, infects families in the order Carnivora, including Nearctic river otters (Lontra canadensis). As a preventative measure, vaccinations against CDV are frequently given to mustelids in captive environments. The Pennsylvania River Otter Reintroduction Project (PRORP) used wild-caught river otters to evaluate the efficacy and need for vaccinations against CDV as part of any reintroduction project. The objectives of this study were to: 1) evaluate the prevalence of exposure to CDV in wild river otters, 2) determine the immunologic response of river otters (i.e., seroconversion) after vaccination with a single (primary) vaccine dose compared to a second (booster) dose of Galaxy-D, a modified live-virus canine distemper (CD) vaccine (MLV CDV), and 3) determine the immunologic response after being vaccinated with a primary vaccination compared to a booster dose of Fervac-D, an MLV CDV. River otters were injected subcutaneously in the nape of the neck with their designated vaccine. Timeframes for collection of blood samples and/or injection of booster vaccines varied depending on the parameters of PRORP. Ten of the 22 river otters had positive prevaccination titer levels to CD. Both vaccines, Galaxy-D and Fervac-D, produced sufficient seroconversion or rise of titer levels (86% and 57%, respectively) to recommend the use of vaccines in wild river otters. Future studies are recommended to evaluate currently produced CD vaccines. Future research should also focus on the number of days required between administration of primary and booster vaccines to achieve sufficient immune response. If only a primary dose is required, then hard-release reintroduction projects for river otters could be recommended. If primary and booster vaccines are required then soft-release reintroduction projects should be recommended. Soft-release projects should include captive management periods that allow for appropriate vaccination intervals

  20. Efficacy of two canine distemper vaccines in wild Nearctic river otters (Lontra canadensis).

    PubMed

    Peper, Steven T; Peper, Randall L; Kollias, George V; Brooks, Robert P; Stevens, Sadie S; Serfass, Thomas L

    2014-09-01

    Canine distemper virus (CDV), a contagious morbillivirus, infects families in the order Carnivora, including Nearctic river otters (Lontra canadensis). As a preventative measure, vaccinations against CDV are frequently given to mustelids in captive environments. The Pennsylvania River Otter Reintroduction Project (PRORP) used wild-caught river otters to evaluate the efficacy and need for vaccinations against CDV as part of any reintroduction project. The objectives of this study were to: 1) evaluate the prevalence of exposure to CDV in wild river otters, 2) determine the immunologic response of river otters (i.e., seroconversion) after vaccination with a single (primary) vaccine dose compared to a second (booster) dose of Galaxy-D, a modified live-virus canine distemper (CD) vaccine (MLV CDV), and 3) determine the immunologic response after being vaccinated with a primary vaccination compared to a booster dose of Fervac-D, an MLV CDV. River otters were injected subcutaneously in the nape of the neck with their designated vaccine. Timeframes for collection of blood samples and/or injection of booster vaccines varied depending on the parameters of PRORP. Ten of the 22 river otters had positive prevaccination titer levels to CD. Both vaccines, Galaxy-D and Fervac-D, produced sufficient seroconversion or rise of titer levels (86% and 57%, respectively) to recommend the use of vaccines in wild river otters. Future studies are recommended to evaluate currently produced CD vaccines. Future research should also focus on the number of days required between administration of primary and booster vaccines to achieve sufficient immune response. If only a primary dose is required, then hard-release reintroduction projects for river otters could be recommended. If primary and booster vaccines are required then soft-release reintroduction projects should be recommended. Soft-release projects should include captive management periods that allow for appropriate vaccination intervals

  1. Simultaneous detection of influenza A, influenza B, and respiratory syncytial viruses and subtyping of influenza A H3N2 virus and H1N1 (2009) virus by multiplex real-time PCR.

    PubMed

    Chen, Yu; Cui, Dawei; Zheng, Shufa; Yang, Shigui; Tong, Jia; Yang, Dagan; Fan, Jian; Zhang, Jie; Lou, Bin; Li, Xuefen; Zhuge, Xiaoling; Ye, Bo; Chen, Baode; Mao, Weilin; Tan, Yajun; Xu, Genyun; Chen, Zhenjin; Chen, Nan; Li, Lanjuan

    2011-04-01

    A multiplex real-time PCR assay was developed to simultaneously detect and discriminate influenza A virus subtypes, including novel H1N1 (2009) and seasonal H3N2 virus, influenza B virus, and respiratory syncytial virus (RSV) in a single test tube, with detection sensitivity and specificity of 99% and 100%, respectively, for the four pathogens. PMID:21270233

  2. Detection of influenza B virus in throat swabs using the polymerase chain reaction.

    PubMed

    Yamada, A; Imanishi, J

    1992-05-01

    An assay protocol based on exploiting the polymerase chain reaction (PCR) for the direct detection of influenza B virus in throat swabs is described. By the use of PCR with nested primers, it was possible to detect the virus in throat swabs. Dilution experiments showed that as little as 1 plaque forming unit of virus was sufficient for detecting the HA gene by the PCR. All throat swab samples from which influenza B virus had been isolated by conventional methods were also positive by the PCR method.

  3. Multi-Gene Detection and Identification of Mosquito-Borne RNA Viruses Using an Oligonucleotide Microarray

    PubMed Central

    Grubaugh, Nathan D.; McMenamy, Scott S.; Turell, Michael J.; Lee, John S.

    2013-01-01

    Background Arthropod-borne viruses are important emerging pathogens world-wide. Viruses transmitted by mosquitoes, such as dengue, yellow fever, and Japanese encephalitis viruses, infect hundreds of millions of people and animals each year. Global surveillance of these viruses in mosquito vectors using molecular based assays is critical for prevention and control of the associated diseases. Here, we report an oligonucleotide DNA microarray design, termed ArboChip5.1, for multi-gene detection and identification of mosquito-borne RNA viruses from the genera Flavivirus (family Flaviviridae), Alphavirus (Togaviridae), Orthobunyavirus (Bunyaviridae), and Phlebovirus (Bunyaviridae). Methodology/Principal Findings The assay utilizes targeted PCR amplification of three genes from each virus genus for electrochemical detection on a portable, field-tested microarray platform. Fifty-two viruses propagated in cell-culture were used to evaluate the specificity of the PCR primer sets and the ArboChip5.1 microarray capture probes. The microarray detected all of the tested viruses and differentiated between many closely related viruses such as members of the dengue, Japanese encephalitis, and Semliki Forest virus clades. Laboratory infected mosquitoes were used to simulate field samples and to determine the limits of detection. Additionally, we identified dengue virus type 3, Japanese encephalitis virus, Tembusu virus, Culex flavivirus, and a Quang Binh-like virus from mosquitoes collected in Thailand in 2011 and 2012. Conclusions/Significance We demonstrated that the described assay can be utilized in a comprehensive field surveillance program by the broad-range amplification and specific identification of arboviruses from infected mosquitoes. Furthermore, the microarray platform can be deployed in the field and viral RNA extraction to data analysis can occur in as little as 12 h. The information derived from the ArboChip5.1 microarray can help to establish public health

  4. African wild dogs (Lycaon pictus) endangered by a canine distemper epizootic among domestic dogs near the Masai Mara National Reserve, Kenya.

    PubMed

    Alexander, K A; Appel, M J

    1994-10-01

    A longitudinal study of canine distemper (CD) among domestic dogs on Malsai communal land to the north of the Masai Mara National Reserve in Kenya was conducted from 1989 to 1991. Prevalence of antibodies to CD was very low among domestic dogs in 1989 and 1990 (4%, n = 49; and 1%, n = 119, respectively) and no African wild dogs (Lycaon pictus; n = 16) collected simultaneously from the same area had detectable antibodies. Among 51 domestic dogs sampled in 1991, however, prevalence of CD antibodies rose significantly (P < 0.01) to 76%. Disease-related mortality rates among domestic dogs were estimated from 1990 to 1992; they rose significantly (P < 0.01) from 21% in 1990 to 50% in 1991 and then decreased significantly (P < 0.01) to 38% in 1992. The 1992 mortality rate remained significantly (P < 0.01) higher than that of 1990. Signs observed in clinically ill domestic dogs were consistent with CD and included listlessness, decreased appetite, bilateral serous to mucopurulent oculonasal discharge, and diarrhea. No carcasses could be retrieved for virus isolation and postmortem examination. Concurrent with this CD epizootic in domestic dogs, the known African wild dog packs in this region disappeared.

  5. New Approaches for Virus Detection through Multidisciplinary Partnerships.

    PubMed

    Fawcett, Helen; Ünlü, M Selim; Connor, John H

    2016-06-10

    A critical requirement for controlling outbreaks of viral infection is sensitive and accurate diagnostics, which can be expensive and are frequently located in resource-intensive clinical laboratories. Outbreaks of many viral infections occur in countries where healthcare resources are limited and clinical laboratories scarce. This creates a fulfillment gap, one that could be filled through the development of inexpensive, sensitive, easy to use, and portable diagnostics. Here we describe our efforts to develop a diagnostic technology that detects viruses without needing to label the particle directly. Our approach has the advantage of speed and assay simplicity while maintaining high sensitivity. Essential in this approach has been the assembly of an integrated, diverse, and interdisciplinary team that worked together to evaluate technologies, spin-out a company, and produce a product for infectious disease diagnostics. The synergy of different individuals with complementary skills has been critical for the development of our transformative technology. PMID:27627625

  6. Development of a HIV-1 Virus Detection System Based on Nanotechnology

    PubMed Central

    Lee, Jin-Ho; Oh, Byung-Keun; Choi, Jeong-Woo

    2015-01-01

    Development of a sensitive and selective detection system for pathogenic viral agents is essential for medical healthcare from diagnostics to therapeutics. However, conventional detection systems are time consuming, resource-intensive and tedious to perform. Hence, the demand for sensitive and selective detection system for virus are highly increasing. To attain this aim, different aspects and techniques have been applied to develop virus sensor with improved sensitivity and selectivity. Here, among those aspects and techniques, this article reviews HIV virus particle detection systems incorporated with nanotechnology to enhance the sensitivity. This review mainly focused on four different detection system including vertically configured electrical detection based on scanning tunneling microscopy (STM), electrochemical detection based on direct electron transfer in virus, optical detection system based on localized surface plasmon resonance (LSPR) and surface enhanced Raman spectroscopy (SERS) using plasmonic nanoparticle. PMID:25923937

  7. Dengue virus detection using impedance measured across nanoporous alumina membrane.

    PubMed

    Peh, Alister En Kai; Li, Sam Fong Yau

    2013-04-15

    The prevalence of dengue around the world makes it critical to develop a simple diagnostic device that can be easily handled by end users and provides fast results. In this paper, we described the use of a small and thin piece of alumina membrane, 60 μm thick and 13 mm in diameter as the sensing platform for the detection of dengue infection. The electrochemical setup is simplified by using the membrane as both the working and the counter electrode. This is achieved by coating both sides of the membrane with a submicron layer of platinum. Electrochemical impedance spectroscopy was utilized for the characterization of the immunosensor as well as the acquisition of data. The change in the pore resistance of the membrane displayed a good correlation with the concentration of the dengue 2 and dengue 3 viruses in plaque forming unit (PFU mL⁻¹), giving detection limit of 0.230 PFU mL⁻¹ and 0.710 PFU mL⁻¹ respectively. This thin piece of membrane sensor, coupled with the simple electrochemical setup, fast detection time of 40 min and high sensitivity, showed potential to be developed into a disposable point-of-care diagnostic tool for clinical uses.

  8. Evaluation of Commercially Available Chikungunya Virus Immunoglobulin M Detection Assays.

    PubMed

    Johnson, Barbara W; Goodman, Christin H; Holloway, Kimberly; de Salazar, P Martinez; Valadere, Anne M; Drebot, Michael A

    2016-07-01

    Commercial chikungunya virus (CHIKV)-specific IgM detection kits were evaluated at the Centers for Disease Control and Prevention (CDC), the Public Health Agency of Canada National Microbiology Laboratory, and the Caribbean Public Health Agency (CARPHA). The Euroimmun Anti-CHIKV IgM ELISA kit had ≥ 95% concordance with all three reference laboratory results. The limit of detection for low CHIK IgM+ samples, as measured by serial dilution of seven sera up to 1:12,800 ranged from 1:800 to 1:3,200. The Euroimmun IIFT kit evaluated at CDC and CARPHA performed well, but required more retesting of equivocal results. The InBios CHIKjj Detect MAC-ELISA had 100% and 98% concordance with CDC and CARPHA results, respectively, and had equal sensitivity to the CDC MAC-ELISA to 1:12,800 dilution in serially diluted samples. The Abcam Anti-CHIKV IgM ELISA had high performance at CARPHA, but at CDC, performance was inconsistent between lots. After replacement of the biotinylated IgM antibody controls with serum containing CHIKV-specific IgM and additional quality assurance/control measures, the Abcam kit was rereleased and reevaluated at CDC. The reformatted Abcam kit had 97% concordance with CDC results and limit of detection of 1:800 to 1:3,200. Two rapid tests and three other CHIKV MAC-ELISAs evaluated at CDC had low sensitivity, as the CDC CHIKV IgM in-house positive controls were below the level of detection. In conclusion, laboratories have options for CHIKV serological diagnosis using validated commercial kits. PMID:26976887

  9. Evaluation of Commercially Available Chikungunya Virus Immunoglobulin M Detection Assays

    PubMed Central

    Johnson, Barbara W.; Goodman, Christin H.; Holloway, Kimberly; de Salazar, P. Martinez; Valadere, Anne M.; Drebot, Michael A.

    2016-01-01

    Commercial chikungunya virus (CHIKV)–specific IgM detection kits were evaluated at the Centers for Disease Control and Prevention (CDC), the Public Health Agency of Canada National Microbiology Laboratory, and the Caribbean Public Health Agency (CARPHA). The Euroimmun Anti-CHIKV IgM ELISA kit had ≥ 95% concordance with all three reference laboratory results. The limit of detection for low CHIK IgM+ samples, as measured by serial dilution of seven sera up to 1:12,800 ranged from 1:800 to 1:3,200. The Euroimmun IIFT kit evaluated at CDC and CARPHA performed well, but required more retesting of equivocal results. The InBios CHIKjj Detect MAC-ELISA had 100% and 98% concordance with CDC and CARPHA results, respectively, and had equal sensitivity to the CDC MAC-ELISA to 1:12,800 dilution in serially diluted samples. The Abcam Anti-CHIKV IgM ELISA had high performance at CARPHA, but at CDC, performance was inconsistent between lots. After replacement of the biotinylated IgM antibody controls with serum containing CHIKV-specific IgM and additional quality assurance/control measures, the Abcam kit was rereleased and reevaluated at CDC. The reformatted Abcam kit had 97% concordance with CDC results and limit of detection of 1:800 to 1:3,200. Two rapid tests and three other CHIKV MAC-ELISAs evaluated at CDC had low sensitivity, as the CDC CHIKV IgM in-house positive controls were below the level of detection. In conclusion, laboratories have options for CHIKV serological diagnosis using validated commercial kits. PMID:26976887

  10. Multiplex RT-PCR method for the simultaneous detection of nine grapevine viruses.

    PubMed

    Gambino, Giorgio

    2015-01-01

    Viral diseases are a serious pathological problem for grapevines, and in recent years the need for increasingly specific and rapid diagnostic methods for the selection of propagation materials has grown. Arabis mosaic virus, Grapevine fanleaf virus, Grapevine virus A, Grapevine virus B, Grapevine rupestris stem pitting-associated virus, Grapevine fleck virus, and Grapevine leafroll-associated viruses 1, 2, and 3 are nine of the most widespread viruses that naturally infect grapevines. A multiplex RT-PCR was developed for simultaneous detection of these nine grapevine viruses, in combination with a plant RNA internal control used as an indicator of the effectiveness of the reaction. One to ten fragments specific for the viruses and an internal control were simultaneously amplified from infected samples and identified by their specific molecular sizes in agarose gel. The protocol reported is an update of previously published protocols for RNA extraction and multiplex diagnosis of viruses. After several years of use and hundreds of samples tested, and following validation in several laboratories, this multiplex RT-PCR provides a reliable and rapid method for detecting grapevine viruses from a large number of samples.

  11. The influenza virus nucleoprotein synthesized from cloned DNA in a simian virus 40 vector is detected in the nucleus.

    PubMed Central

    Lin, B C; Lai, C J

    1983-01-01

    We obtained DNA sequences coding for the nucleoprotein (NP) of an influenza A virus by reverse transcription of virion RNA with synthetic oligonucleotide primers. Terminal sequence analysis showed that the cloned gene contained a full-length copy of the virion RNA segment. The NP-specific DNA was inserted into the late region of a simian virus 40 vector, and the DNA recombinant was propagated in the presence of an early simian virus 40 temperature-sensitive mutant helper. Infection of African green monkey kidney cells with the recombinant produced a polypeptide immunoprecipitable with NP-specific antisera. The polypeptide product had a molecular weight of 56,000, identical to that of the nucleoprotein of influenza virus as estimated on polyacrylamide gels. The putative NP was detected in the nucleus of infected primate cells by an immunofluorescence assay. This nuclear localization of NP from recombinant DNA was similar to that seen during influenza virus infection. Images PMID:6296449

  12. THE APPLICATION OF EMERGING TECHNOLOGIES TO VIRUS DETECTION IN WATER

    EPA Science Inventory

    Human enteric viruses belonging to many different viral genera cause waterborne disease when susceptible individuals are exposed to contaminated drinking and recreational waters. Diseases resulting from infection with these viruses include gastroenteritis, hepatitis, encephali...

  13. DETECTION OF VIRUSES IN ENVIRONMENTAL WATERS, SEWAGE AND SEWAGE SLUDGES

    EPA Science Inventory

    There are many different groups of viruses that will be found in environmental waters. These include the many types of viruses whose hosts are natural aquatic organisms. There are also groups of viruses present in environmental waters which represent exogenous contaminants, whose...

  14. Cucurbit yellow stunting disorder virus detected in pigweed in Florida

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Samples were collected from pigweed, a species of Amaranth, that were in proximity to virus-infected watermelons in south Florida. No symptoms of virus infection were observed on the weed samples; however, testing by two independent methods found infection by Cucurbit yellow stunting disorder virus...

  15. A simplified method for simultaneous detection of Rice stripe virus and Rice black-streaked dwarf virus in insect vector.

    PubMed

    Li, Shuo; Wang, Xi; Xu, Jianxiang; Ji, Yinghua; Zhou, Yijun

    2015-01-01

    Rice stripe virus (RSV) and Rice black-streaked dwarf virus (RBSDV) are transmitted by their common vector small brown planthopper (SBPH) that cause serious crop losses in China. A simple reverse transcription-PCR method was developed for the simultaneous detection of RSV and RBSDV in single SBPH. Three primers targeted to RSV-RNA4 and RBSDV-S2 segments were designed to amplify respectively 1114-bp and 414-bp fragments in a reaction. The method is reliable, rapid and inexpensive for detecting the two viruses in vector, which could facilitate better forecasting and control of the virus diseases. Using this method, it was found that SBPH could carry RSV and RBSDV simultaneously. PMID:25455902

  16. A simplified method for simultaneous detection of Rice stripe virus and Rice black-streaked dwarf virus in insect vector.

    PubMed

    Li, Shuo; Wang, Xi; Xu, Jianxiang; Ji, Yinghua; Zhou, Yijun

    2015-01-01

    Rice stripe virus (RSV) and Rice black-streaked dwarf virus (RBSDV) are transmitted by their common vector small brown planthopper (SBPH) that cause serious crop losses in China. A simple reverse transcription-PCR method was developed for the simultaneous detection of RSV and RBSDV in single SBPH. Three primers targeted to RSV-RNA4 and RBSDV-S2 segments were designed to amplify respectively 1114-bp and 414-bp fragments in a reaction. The method is reliable, rapid and inexpensive for detecting the two viruses in vector, which could facilitate better forecasting and control of the virus diseases. Using this method, it was found that SBPH could carry RSV and RBSDV simultaneously.

  17. Detection of C-type virus by immunoferritin technique in bat lung cell line chronically infected with bovine leucosis virus.

    PubMed

    Mihailescu, D; Patrascu, I V; Apostol, I; Mazilu, M

    1980-01-01

    Reported in this paper are morphological studies and tests for the detection of Type-C particles from a line of bat lung cells chronically infected with bovine leucosis virus. The immunoferritin technique was used. Ferritin labelling of Type-C particles was regularly accompanied by black-spot arrangement of ferritin around the virus envelope, which provided evidence to the specificity of this immunochemical technique. PMID:6260052

  18. Target-size embracing dimension for sensitive detection of viruses with various sizes and influenza virus strains.

    PubMed

    Lin, Ying-Yi; Liao, Jiunn-Der; Yang, Mei-Lin; Wu, Chao-Liang

    2012-05-15

    The focused ion beam (FIB) technique was employed to precisely fabricate hexagon-like Au nano-rods (fibAu_h) arrays as a surface enhanced Raman scattering - active substrate. A "ring diameter" (D(R)) was created by the convergence of three fibAu_h with respect to the dimension of the target viruses (D(T)), such as adenovirus (Adeno), encephalomyocarditis virus (EMCV), influenza virus (H1N1) with different sizes. Three influenza A virus strains were also compared. The results indicate that as that with a D(R)/D(T) ratio of around 1, the discrimination ability for detecting the target viruses and SERS mechanism become obvious. The enhanced lightning rod effect surrounding the seized target virus is anticipated if its size and dimension is suitably embraced within three fibAu_h. Hence the as-designed fibAu_h sample with a target-size embracing dimension provides good discrimination ability for distinguishing virus of various sizes or virus strains.

  19. Detection and isolation of Bluetongue virus from commercial vaccine batches.

    PubMed

    Bumbarov, Velizar; Golender, Natalia; Erster, Oran; Khinich, Yevgeny

    2016-06-14

    In this report we describe the detection and identification of Bluetongue virus (BTV) contaminations in commercial vaccines. BTV RNA was detected in vaccine batches of Lumpy skin disease (LSD) and Sheep pox (SP) using quantitative PCR (qPCR) for VP1 and NS3 genes. Both batches were positive for VP1 and NS3 in qPCR. The LSD vaccine-derived sample was positive for VP1 and VP2 in conventional PCR. The SP vaccine-derived sample was examined by amplification of VP1, VP4, VP6, VP7, NS2 and NS3 gene segments in conventional PCR. The SP vaccine-derived sample was further propagated in embryonated chicken eggs (ECE) and Vero cells. Preliminary sequence analysis showed that the LSD vaccine-derived sequence was 98-99% similar to BTV9. Analysis of the six genomic segments from the SP vaccine-derived isolate showed the highest similarity to BTV26 (66.3-97.8%). These findings are particularly important due to the effect of BTV on cattle and sheep, for which the vaccines are intended. They also demonstrate the necessity of rigorous vaccine inspection and strict vaccine production control. PMID:27171751

  20. Viruses and human cancer: from detection to causality.

    PubMed

    Sarid, Ronit; Gao, Shou-Jiang

    2011-06-28

    The study of cancer is incomplete without taking into consideration of tumorigenic viruses. Initially, searches for human cancer viruses were fruitless despite an expansion of our knowledge in the same period concerning acute-transforming retroviruses in animals. However, over the last 40 years, we have witnessed rapid progress in the tumor virology field. Currently, acknowledged human cancer viruses include Epstein-Barr virus, hepatitis B virus, hepatitis C virus, high-risk human papilloma viruses, human T-cell lymphotropic virus type 1 and Kaposi's sarcoma-associated herpesvirus. Extensive epidemiological and mechanistic studies have led to the development of novel preventive and therapeutic approaches for managing some of these infections and associated cancers. In addition, recent advances in molecular technologies have enabled the discovery of a new potential human tumor virus, Merkel cell polyomavirus, but its association with cancer remains to be validated. It is anticipated that in the next few decades many additional human cancer viruses will be discovered and the mechanisms underlying viral oncogenesis delineated. Thus, it can be expected that better tools for preventing and treating virus-associated cancer will be available in the near future.

  1. Viruses and Human Cancer: From Detection to Causality

    PubMed Central

    Sarid, Ronit; Gao, Shou-Jiang

    2010-01-01

    The study of cancer is incomplete without taking into consideration of tumorigenic viruses. Initially, searches for human cancer viruses were fruitless despite an expansion of our knowledge in the same period concerning acute-transforming retroviruses in animals. However, over the last 40 years, we have witnessed rapid progress in the tumor virology field. Currently, acknowledged human cancer viruses include Epstein-Barr virus, Hepatitis B virus, Hepatitis C virus, High-risk human papilloma viruses, Human T-cell lymphotropic virus type 1 and Kaposi’s sarcoma-associated herpesvirus. Extensive epidemiological and mechanistic studies have led to the development of novel preventive and therapeutic approaches for managing some of these infections and associated cancers. In addition, recent advances in molecular technologies have enabled the discovery of a new potential human tumor virus, Merkel cell polyomavirus, but its association with cancer remains to be validated. It is anticipated that in the next few decades many additional human cancer viruses will be discovered and the mechanisms underlying viral oncogenesis delineated. Thus, it can be expected that better tools for preventing and treating virus-associated cancer will be available in the near future. PMID:20971551

  2. Detection of airborne viruses using electro-aerodynamic deposition and a field-effect transistor.

    PubMed

    Park, Kyu-Tae; Cho, Dong-Guk; Park, Ji-Woon; Hong, Seunghun; Hwang, Jungho

    2015-01-01

    We report a technique for the detection of aerosolized viruses. Conventional field-effect-transistor (FET)-based techniques use solution-based processes, thus require antibody binding to the detection region of the FET prior to the supply of the analyte. With the method described here, virus-antibody-bound particles are delivered to the FET during detection; therefore, neither a pre-treatment antibody binding step on the FET channel nor washing process for virus-antibody-binding are necessary. Our method is based on the concept that virus-antibody-bound particles are larger than the virus or antibody alone, and thus have larger charge numbers following aerosol charging. When these particles are charged by negative ions and electro-aerodynamically deposited on a substrate, there exists a location on the substrate where neither lone virus nor antibody particles land, and where only virus-antibody-bound particles are deposited. If this location coincides with the channel of the FET, the resulting variation in the current can be used to indicate the existence of a virus. By aerosolizing a mixed solution of the virus and the antibody, only the virus-antibody-bound particles were transported to the swCNT-FET, and the electric current in the swCNT-FET decreased to 30% of that measured with no deposited particles.

  3. Detection of airborne viruses using electro-aerodynamic deposition and a field-effect transistor.

    PubMed

    Park, Kyu-Tae; Cho, Dong-Guk; Park, Ji-Woon; Hong, Seunghun; Hwang, Jungho

    2015-01-01

    We report a technique for the detection of aerosolized viruses. Conventional field-effect-transistor (FET)-based techniques use solution-based processes, thus require antibody binding to the detection region of the FET prior to the supply of the analyte. With the method described here, virus-antibody-bound particles are delivered to the FET during detection; therefore, neither a pre-treatment antibody binding step on the FET channel nor washing process for virus-antibody-binding are necessary. Our method is based on the concept that virus-antibody-bound particles are larger than the virus or antibody alone, and thus have larger charge numbers following aerosol charging. When these particles are charged by negative ions and electro-aerodynamically deposited on a substrate, there exists a location on the substrate where neither lone virus nor antibody particles land, and where only virus-antibody-bound particles are deposited. If this location coincides with the channel of the FET, the resulting variation in the current can be used to indicate the existence of a virus. By aerosolizing a mixed solution of the virus and the antibody, only the virus-antibody-bound particles were transported to the swCNT-FET, and the electric current in the swCNT-FET decreased to 30% of that measured with no deposited particles. PMID:26642822

  4. Detection of airborne viruses using electro-aerodynamic deposition and a field-effect transistor

    NASA Astrophysics Data System (ADS)

    Park, Kyu-Tae; Cho, Dong-Guk; Park, Ji-Woon; Hong, Seunghun; Hwang, Jungho

    2015-12-01

    We report a technique for the detection of aerosolized viruses. Conventional field-effect-transistor (FET)-based techniques use solution-based processes, thus require antibody binding to the detection region of the FET prior to the supply of the analyte. With the method described here, virus-antibody-bound particles are delivered to the FET during detection; therefore, neither a pre-treatment antibody binding step on the FET channel nor washing process for virus-antibody-binding are necessary. Our method is based on the concept that virus-antibody-bound particles are larger than the virus or antibody alone, and thus have larger charge numbers following aerosol charging. When these particles are charged by negative ions and electro-aerodynamically deposited on a substrate, there exists a location on the substrate where neither lone virus nor antibody particles land, and where only virus-antibody-bound particles are deposited. If this location coincides with the channel of the FET, the resulting variation in the current can be used to indicate the existence of a virus. By aerosolizing a mixed solution of the virus and the antibody, only the virus-antibody-bound particles were transported to the swCNT-FET, and the electric current in the swCNT-FET decreased to 30% of that measured with no deposited particles.

  5. Application of a focus formation assay for detection and titration of porcine epidemic diarrhea virus.

    PubMed

    Cruz, Deu John M; Shin, Hyun-Jin

    2007-10-01

    A focus formation assay (FFA) for detection and titration of porcine epidemic diarrhea virus (PEDV) in a micro-culture system using Vero cells and PAP staining technique was evaluated. A linear correlation between the virus dilution and virus titer determined by FFA was observed between the range of 10 and 30 foci per well. Comparative analysis between FFA and plaque assay showed no significant difference in estimating the titer of cell adapted PEDV. However, the culture time required for detecting the virus was considerably shorter for FFA. In addition, FFA had higher sensitivity for detecting field isolates of PEDV as well as positive identification of the virus with the antibody specific reaction. A broader range of dilutions and number of replicates may be used for titration. A FFA may be applied as an alternative method for detection and titration of PEDV.

  6. Detection of yellowhead virus and Chinese baculovirus in penaeid shrimp by the Western blot technique.

    PubMed

    Nadala, E C; Tapay, L M; Cao, S; Loh, P C

    1997-12-01

    The continuing threat posed by viral diseases in cultured shrimp calls for the development of detection technologies for monitoring the animals, especially broodstock. Two of the most highly pathogenic viruses of penaeid shrimp are the yellow-head virus (YHV) and Chinese baculovirus (CBV, also called white spot baculovirus). A Western blot (WB) protocol capable of detecting YHV and CBV in the hemolymph of infected shrimp was developed. The use of the hemolymph as material for virus detection allowed for sample collection without sacrificing the animals. This protocol was highly specific, rapid, and sensitive enough to detect the presence of the viruses before the appearance of overt symptoms. It was also useful for demonstrating the growth of both viruses in primary shrimp lymphoid cell cultures.

  7. Detection of influenza viruses in throat swab by using polymerase chain reaction.

    PubMed

    Yamada, A; Imanishi, J; Nakajima, E; Nakajima, K; Nakajima, S

    1991-01-01

    An assay protocol based on exploiting the polymerase chain reaction (PCR) for the direct detection of influenza virus in throat swab is described. By use of the mixture of H1 and H3 primers, it was possible to determine the subtype of the influenza A viruses simultaneously. No visible band was detected after PCR of influenza B or A (H2N2) viruses with a pair of H1 or H3 primers. The dilution experiment showed that the influenza viruses, as few as 1.3-6 plaque-forming units, were sufficient for detecting the HA gene by PCR. All throat swab samples from which influenza viruses had been isolated by conventional method were also positively detected by PCR method.

  8. Finite Element Analysis on Nanomechanical Detection of Small Particles: Toward Virus Detection.

    PubMed

    Imamura, Gaku; Shiba, Kota; Yoshikawa, Genki

    2016-01-01

    Detection of small particles, including viruses and particulate matter (PM), has been attracting much attention in light of increasing need for environmental monitoring. Owing to their high versatility, a nanomechanical sensor is one of the most promising sensors which can be adapted to various monitoring systems. In this study, we present an optimization strategy to efficiently detect small particles with nanomechanical sensors. Adsorption of particles on the receptor layer of nanomechanical sensors and the resultant signal are analyzed using finite element analysis (FEA). We investigate the effect of structural parameters (e.g., adsorption position and embedded depth of a particle and thickness of the receptor layer) and elastic properties of the receptor layer (e.g., Young's modulus and Poisson's ratio) on the sensitivity. It is found that a membrane-type surface stress sensors (MSS) has the potential for robust detection of small particles.

  9. Finite Element Analysis on Nanomechanical Detection of Small Particles: Toward Virus Detection

    PubMed Central

    Imamura, Gaku; Shiba, Kota; Yoshikawa, Genki

    2016-01-01

    Detection of small particles, including viruses and particulate matter (PM), has been attracting much attention in light of increasing need for environmental monitoring. Owing to their high versatility, a nanomechanical sensor is one of the most promising sensors which can be adapted to various monitoring systems. In this study, we present an optimization strategy to efficiently detect small particles with nanomechanical sensors. Adsorption of particles on the receptor layer of nanomechanical sensors and the resultant signal are analyzed using finite element analysis (FEA). We investigate the effect of structural parameters (e.g., adsorption position and embedded depth of a particle and thickness of the receptor layer) and elastic properties of the receptor layer (e.g., Young's modulus and Poisson's ratio) on the sensitivity. It is found that a membrane-type surface stress sensors (MSS) has the potential for robust detection of small particles. PMID:27148181

  10. Detection of enteric viruses in activated sludge by feasible concentration methods

    PubMed Central

    Prado, Tatiana; Gaspar, Ana Maria Coimbra; Miagostovich, Marize Pereira

    2014-01-01

    Human enteric viruses are responsible to cause several diseases, including gastroenteritis and hepatitis, and can be present in high amounts in sewage sludge. This study compared virus recovery efficiency of two feasible concentration methods used for detecting human adenovirus (HAdV), rotavirus species A (RV-A), norovirus genogroup II (NoV GII) and hepatitis A virus (HAV) in sewage sludge from an activated sludge process. Twelve sewage sludge samples were collected bi-monthly from January to July, 2011. Ultracentrifugation was compared with a simplified protocol based on beef extract elution for recovering enteric viruses. Viruses were quantified by quantitative real-time PCR assays and virus recovery efficiency and limits of detection were determined. Methods showed mean recovery rates lower than 7.5%, presenting critical limits of detection (higher than 102 – 103 genome copies - GC L−1 for all viruses analyzed). Nevertheless, HAdV were detected in 90% of the analyzed sewage sludge samples (range: 1.8 × 104 to 1.1 × 105 GC L−1), followed by RV-A and NoV (both in 50%) and HAV (8%). Results suggesting that activated sludge is contaminated with high viral loads and HAdV are widely disseminated in these samples. The low virus recovery rates achieved, especially for HAV, indicate that other feasible concentration methods could be developed to improve virus recovery efficiency in these environmental matrices. PMID:24948954

  11. Reassessing the detection of B-virus-specific serum antibodies.

    PubMed

    Katz, David; Shi, Wei; Wildes, Martin J; Krug, Peter W; Hilliard, Julia K

    2012-12-01

    B virus, a natural pathogen of macaques, can cause a fatal zoonotic disease in humans. Serologic screening of macaques by titration ELISA (tELISA, screening test) and by Western blot analysis (WBA, confirmatory test) is one of the principle measures to prevent human infection. Here we slightly modified these 2 tests and reevaluated their correlation. We developed a high-throughput tELISA and used it to screen 278 sera simultaneously against the homologous BV antigen and the heterologous antigens of Papiine herpesvirus 2 and Human herpesvirus 1. More sera (35.6%) were positive by the BV-ELISA than by the HVP2-ELISA (21.6%) or HSV1-ELISA (19.8%). The superiority of the homologous tELISA over the heterologous tELISA was prominent in low-titer sera. WBA confirmed only 21% of the tELISA-positive sera with low or intermediate antibody titers. These sera might have contained antibodies to conformational epitopes that could not be detected by WBA, in which denatured antigens are used, but that could be detected by tELISA, which detects both linear and conformational epitopes. WBA confirmed 82% of the tELISA high-titer sera. However, WBA defined the remaining 18% of sera, which were negative by tELISA, as nonnegative. This finding can be attributed to the difficulties encountered with the subjective interpretation of results by WBA. Together, the current results indicate the inadequacy of WBA as a confirmatory assay for sera with low antibody titers.

  12. Spiranthes Mosaic Virus 3 and Bidens Mottle Virus,Two Potyviruses Detected in Phlox divaricata

    Technology Transfer Automated Retrieval System (TEKTRAN)

    This is the first report of Spiranthes mosaic virus in Florida and the first report of Bidens mottle virus in Phlox divaricata. This report provides an overview of this virus for growers, extension workers, crop consultants and research and regulatory scientists....

  13. Simultaneous detection of three lily-infecting viruses using a multiplex Luminex bead array.

    PubMed

    Lim, Mi Sang; Kim, Su Min; Choi, Sun Hee

    2016-05-01

    A Luminex bead array was applied to detect multiple-virus coinfection in lily plants exhibiting typical symptoms, and the efficiency of this detection system was assessed. Specific primer sets for the simultaneous detection of 4 targets in virus-infected lily plants were constructed and used for reverse transcription (RT)-polymerase chain reaction (PCR), and specific probes were used for Luminex-based assay. Each of the 4 targets was amplified, and the amplicons were used for Luminex bead array experiments. A Luminex bead array analysis of lily-infecting viruses was performed using the quadruplex RT-PCR products followed by hybridization between the biotinylated targets and anti-tagged microsphere beads. The hybridization products produced fluorescence signals that were detected by the Luminex system. Signal strengths were analyzed by their median fluorescence intensity (MFI) values. Detection of the different target elements was found to be very specific to the corresponding viruses in lilies, and coinfection with multiple viruses was specifically detected via the MFI signals. Therefore, the use of a Luminex bead array for the detection of co-infected multiple viruses in lily plants can be an improved system for screening and analyzing multiple-virus infection.

  14. Detection of plant viruses in natural environments by using RNA-Seq.

    PubMed

    Nagano, Atsushi J; Honjo, Mie N; Mihara, Motohiro; Sato, Masanao; Kudoh, Hiroshi

    2015-01-01

    Sequencing of RNA by next generation sequencers, RNA-Seq, is revolutionizing virus detection. In addition to the unbiased detection of various viruses from wild plants in natural environments, RNA-Seq also allows for the parallel collection of host plant transcriptome data. Host transcriptome data are highly valuable for studying the responses of hosts to viral infections, as well as viral host manipulation. When detecting viruses using RNA-Seq, it is critical to choose appropriate methods for the removal of rRNA from total RNA. Although viruses with polyadenylated genomes can be detected by RNA-Seq following mRNA purification using oligo-dT beads, viruses with non-polyadenylated genomes are not effectively detected. However, such viruses can be detected by RNA-Seq using the rRNA selective depression method. The high-throughput and cost-effective method of RNA-Seq library preparation which is described here allows us to detect a broad range of viruses in wild plants.

  15. A virus-MIPs fluorescent sensor based on FRET for highly sensitive detection of JEV.

    PubMed

    Liang, Caishuang; Wang, Huan; He, Kui; Chen, Chunyan; Chen, Xiaoming; Gong, Hang; Cai, Changqun

    2016-11-01

    Major stumbling blocks in the recognition and detection of virus are the unstable biological recognition element or the complex detection means. Here a fluorescent sensor based on virus-molecular imprinted polymers (virus-MIPs) was designed for specific recognition and highly sensitive detection of Japanese encephalitis virus (JEV). The virus-MIPs were anchored on the surface of silica microspheres modified by fluorescent dye, pyrene-1-carboxaldehyde (PC). The fluorescence intensity of PC can be enhanced by the principle of fluorescence resonance energy transfer (FRET), where virus acted as energy donor and PC acted as energy acceptor. The enhanced fluorescence intensity was proportional to the concentration of virus in the range of 24-960pM, with a limit of detection (LOD, 3σ) of 9.6pM, and the relative standard deviation was 1.99%. In additional, the specificity study confirmed the resultant MIPs has high-selectivity for JEV. This sensor would become a new key for the detection of virus because of its high sensitive, simple operation, high stability and low cost. PMID:27591625

  16. Simultaneous detection and differentiation of three viruses in pear plants by a multiplex RT-PCR.

    PubMed

    Yao, Bingyu; Wang, Guoping; Ma, Xiaofang; Liu, Wenbin; Tang, Huihui; Zhu, Hui; Hong, Ni

    2014-02-01

    A multiplex RT-PCR (mRT-PCR) assay was developed for detection and differentiation of the Apple stem pitting virus (ASPV), Apple stem grooving virus (ASGV) and Apple chlorotic leaf spot virus (ACLSV), which are viruses frequently occurring in pear trees. Different combinations of mixed primer pairs were tested for their specificity and sensitivity for the simultaneous detection of the three viruses. Three primer pairs were used to amplify their fragments of 247bp, 358bp and 500bp, respectively. The primer pair for ASPV was designed in this work, while the primer pairs for ACLSV and ASGV were from previous reports. The sensitivity and specificity of the mRT-PCR assay for the three viruses were comparable to that of each uniplex RT-PCR. The mRT-PCR was applied successfully for the detection of three viruses in leaves of pear and apple plants, but was unreliable in the detection of ASGV in dormant barks. In conclusion, this mRT-PCR provides a useful tool for the routine and rapid detection and the differentiation of three pear viruses.

  17. Detection of virus-specific RNA in simian sarcoma-leukemia virus-infected cells in in situ hybridization to viral complementary DNA.

    PubMed Central

    Kaufman, S L; Gallo, R C; Miller, N R

    1979-01-01

    An in situ molecular hybridization system which will detect retrovirus RNA in the cytoplasm of individual virus-infected cells has been developed. The technique was applied to cells infected with simian sarcoma-leukemia virus, where the virus-specific RNA was detected by hybridization to simian sarcoma-leukemia virus 3H-labeled complementary DNA. The system is useful for detecting viral RNA-containing cells in the presence of an excess of virus-negative cells and for determining which type of cell in a heterogenous population is expressing viral RNA. Images PMID:224220

  18. Detecting the emergence of novel, zoonotic viruses pathogenic to humans

    PubMed Central

    2015-01-01

    RNA viruses, with their high potential for mutation and epidemic spread, are the most common class of pathogens found as new causes of human illness. Despite great advances made in diagnostic technology since the 1950s, the annual rate at which novel virulent viruses have been found has remained at 2–3. Most emerging viruses are zoonoses; they have jumped from mammal or bird hosts to humans. An analysis of virus discovery indicates that the small number of novel viruses discovered annually is an artifact of inadequate surveillance in tropical and subtropical countries, where even established endemic pathogens are often misdiagnosed. Many of the emerging viruses of the future are already infecting humans but remain to be uncovered by a strategy of disease surveillance in selected populations. PMID:25416679

  19. 9 CFR 113.201 - Canine Distemper Vaccine, Killed Virus.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... final serum dilution. (ii) Vaccination. Each of the four vaccinates shall be injected as recommended on.... (iii) Serology. At the end of the post vaccination observation period, a second blood sample shall...

  20. 9 CFR 113.201 - Canine Distemper Vaccine, Killed Virus.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... final serum dilution. (ii) Vaccination. Each of the four vaccinates shall be injected as recommended on.... (iii) Serology. At the end of the post vaccination observation period, a second blood sample shall...

  1. 9 CFR 113.201 - Canine Distemper Vaccine, Killed Virus.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... final serum dilution. (ii) Vaccination. Each of the four vaccinates shall be injected as recommended on.... (iii) Serology. At the end of the post vaccination observation period, a second blood sample shall...

  2. 9 CFR 113.201 - Canine Distemper Vaccine, Killed Virus.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... final serum dilution. (ii) Vaccination. Each of the four vaccinates shall be injected as recommended on.... (iii) Serology. At the end of the post vaccination observation period, a second blood sample shall...

  3. 9 CFR 113.201 - Canine Distemper Vaccine, Killed Virus.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... final serum dilution. (ii) Vaccination. Each of the four vaccinates shall be injected as recommended on.... (iii) Serology. At the end of the post vaccination observation period, a second blood sample shall...

  4. Influenza virus titration, antigenic characterization, and serological methods for antibody detection.

    PubMed

    Klimov, Alexander; Balish, Amanda; Veguilla, Vic; Sun, Hong; Schiffer, Jarad; Lu, Xiuhua; Katz, Jacqueline M; Hancock, Kathy

    2012-01-01

    This chapter describes some commonly used methods of influenza virus titration, antigenic characterization, and serological methods by antibody detection. These methods are essential not only for virus characterization but also for identifying new antigenic variants, vaccine strain selection, and sero-epidemiologic studies of influenza virus transmission and prevalence. Virus titration methods such as the hemagglutination assay, 50% egg or tissue culture infectious dose, and plaque assay are employed to determine the amount of virus particles in a sample. The hemagglutination inhibition assay is a reliable, relatively simple and inexpensive technique to antigenically characterize isolates of influenza viruses. Serological methods such as virus neutralization and hemagglutination inhibition are the fundamental tools used in sero-epidemiologic studies of influenza virus transmission and prevalence and in the evaluation of vaccine immunogenicity. While serological methods rarely yield an early diagnosis of acute influenza virus infection, well-timed, paired acute, and convalescent serum samples may establish the diagnosis of a recent influenza infection even when attempts to detect the virus are negative.

  5. Functionalized magnetic microparticle-based colorimetric platform for influenza A virus detection.

    PubMed

    Chen, Chaohui; Zou, Zhong; Chen, Lu; Ji, Xinghu; He, Zhike

    2016-10-28

    A colorimetric platform for influenza A virus detection was developed by using the high efficiency of enzymatic catalysis and the reduction of gold ions with hydrogen peroxide. Aptamer-functionalized magnetic microparticles were synthesized to capture the influenza A virus. This was followed by the binding of ConA-GOx-AuNPs to the H3N2 virus through the ConA-glycan interaction. The sandwich complex was subsequently dispersed in glucose solution to trigger an enzymatic reaction to produce hydrogen peroxide, which controlled the growth of gold nanoparticles and produced colored solutions. The determination of H3N2 concentration was realized by comparing the two differently colored gold nanoparticles. This method could detect the target virus as low as 11.16 μg ml(-1). Furthermore, it opens new opportunities for sensitive and colorimetric detection of viruses and proteins. PMID:27655150

  6. Functionalized magnetic microparticle-based colorimetric platform for influenza A virus detection.

    PubMed

    Chen, Chaohui; Zou, Zhong; Chen, Lu; Ji, Xinghu; He, Zhike

    2016-10-28

    A colorimetric platform for influenza A virus detection was developed by using the high efficiency of enzymatic catalysis and the reduction of gold ions with hydrogen peroxide. Aptamer-functionalized magnetic microparticles were synthesized to capture the influenza A virus. This was followed by the binding of ConA-GOx-AuNPs to the H3N2 virus through the ConA-glycan interaction. The sandwich complex was subsequently dispersed in glucose solution to trigger an enzymatic reaction to produce hydrogen peroxide, which controlled the growth of gold nanoparticles and produced colored solutions. The determination of H3N2 concentration was realized by comparing the two differently colored gold nanoparticles. This method could detect the target virus as low as 11.16 μg ml(-1). Furthermore, it opens new opportunities for sensitive and colorimetric detection of viruses and proteins.

  7. Detection and transmission of infectious hematopoietic necrosis virus in rainbow trout

    USGS Publications Warehouse

    Amend, Donald F.

    1975-01-01

    Detection and transmission of Infectious Hematopoietic Necrosis Virus in rainbow trout (Salmo gairdneri) was studied at a commercial trout hatchery. Transmission of virus was demonstrated via water, feed and contaminated eggs. If eggs from carrier females were incubated several weeks in virus-free water, the resulting fry did not become infected. However, if fry subsequently became infected they were lifetime carriers. Infectious virus was readily detectable in most tissues of moribund fish; in carriers it was detected in sex products of spawning fish, and in samples from the intestine of post-spawning fish, but not in samples from blood, feces, kidney, or liver. The carrier rate was not significantly different between sexes. It was concluded that adult carriers are the reservoir of infection and that transmission occurs primarily when carriers shed virus and expose susceptible fish or eggs.

  8. Functionalized magnetic microparticle-based colorimetric platform for influenza A virus detection

    NASA Astrophysics Data System (ADS)

    Chen, Chaohui; Zou, Zhong; Chen, Lu; Ji, Xinghu; He, Zhike

    2016-10-01

    A colorimetric platform for influenza A virus detection was developed by using the high efficiency of enzymatic catalysis and the reduction of gold ions with hydrogen peroxide. Aptamer-functionalized magnetic microparticles were synthesized to capture the influenza A virus. This was followed by the binding of ConA-GOx-AuNPs to the H3N2 virus through the ConA-glycan interaction. The sandwich complex was subsequently dispersed in glucose solution to trigger an enzymatic reaction to produce hydrogen peroxide, which controlled the growth of gold nanoparticles and produced colored solutions. The determination of H3N2 concentration was realized by comparing the two differently colored gold nanoparticles. This method could detect the target virus as low as 11.16 μg ml-1. Furthermore, it opens new opportunities for sensitive and colorimetric detection of viruses and proteins.

  9. Detection of red-spotted grouper nervous necrosis virus by loop-mediated isothermal amplification.

    PubMed

    Xu, Hai-Dong; Feng, Juan; Guo, Zhi-Xun; Ou, You-Jun; Wang, Jiang-Yong

    2010-01-01

    Red-spotted grouper nervous necrosis virus (RGNNV) causes high mortality in marine fish larvae cultured in China. To control better an outbreak of this virus, a rapid, specific and sensitive detection method based on loop-mediated isothermal amplification (LAMP) was developed. A set of four primers, two outer and two inner, was designed from RGNNV genome RNA. The LAMP reaction mix was optimized. The method was specific as no cross-reaction was observed between white spot syndrome virus, koi herpesvirus, infectious spleen and kidney necrosis virus, mud crab reovirus, and grass carp hemorrhage virus. The sensitivity of LAMP was 100-fold higher than the nested PCR in detecting the presence of RGNNV. RGNNV was detected in the brain of Trachinotus ovatus that showed typical symptoms of NNV infection, with the standardized LAMP procedure.

  10. Micro flow cytometry utilizing a magnetic bead-based immunoassay for rapid virus detection.

    PubMed

    Yang, Sung-Yi; Lien, Kang-Yi; Huang, Kao-Jean; Lei, Huan-Yao; Lee, Gwo-Bin

    2008-12-01

    The current study presents a new miniature microfluidic flow cytometer integrated with several functional micro-devices capable of viral sample purification and detection by utilizing a magnetic bead-based immunoassay. The magnetic beads were conjugated with specific antibodies, which can recognize and capture target viruses. Another dye-labeled anti-virus antibody was then used to mark the bead-bound virus for the subsequent optical detection. Several essential components were integrated onto a single chip including a sample incubation module, a micro flow cytometry module and an optical detection module. The sample incubation module consisting of pneumatic micropumps and a membrane-type, active micromixer was used for purifying and enriching the target virus-bound magnetic beads with the aid of a permanent magnet. The micro flow cytometry module and the optical detection module were used to perform the functions of virus counting and collection. Experimental results showed that virus samples with a concentration of 10(3)PFU/ml can be automatically detected successfully by the developed system. In addition, the entire diagnosis procedure including sample incubation and virus detection took only about 40min. Consequently, the proposed micro flow cytometry may provide a powerful platform for rapid diagnosis and future biological applications.

  11. Ultrasensitive detection of influenza viruses with a glycan-based impedimetric biosensor

    PubMed Central

    Hushegyi, András; Pihíková, Dominika; Bertók, Tomáš; Adam, Vojtech; Kizek, René; Tkac, Jan

    2016-01-01

    An ultrasensitive impedimetric glycan-based biosensor for reliable and selective detection of inactivated, but intact influenza viruses H3N2 was developed. Such glycan-based approach has a distinct advantage over antibody-based detection of influenza viruses since glycans are natural viral receptors with a possibility to selectively distinguish between potentially pathogenic influenza subtypes by the glycan-based biosensors. Build-up of the biosensor was carefully optimized with atomic force microscopy applied for visualization of the biosensor surface after binding of viruses with the topology of an individual viral particle H3N2 analyzed. The glycan biosensor could detect a glycan binding lectin with a limit of detection (LOD) of 5 aM. The biosensor was finally applied for analysis of influenza viruses H3N2 with LOD of 13 viral particles in 1 μl, what is the lowest LOD for analysis of influenza viral particles by the glycan-based device achieved so far. The biosensor could detect H3N2 viruses selectively with a sensitivity ratio of 30 over influenza viruses H7N7. The impedimetric biosensor presented here is the most sensitive glycan-based device for detection of influenza viruses and among the most sensitive antibody or aptamer based biosensor devices. PMID:26765527

  12. Ultrasensitive detection of influenza viruses with a glycan-based impedimetric biosensor.

    PubMed

    Hushegyi, András; Pihíková, Dominika; Bertok, Tomas; Adam, Vojtech; Kizek, René; Tkac, Jan

    2016-05-15

    An ultrasensitive impedimetric glycan-based biosensor for reliable and selective detection of inactivated, but intact influenza viruses H3N2 was developed. Such glycan-based approach has a distinct advantage over antibody-based detection of influenza viruses since glycans are natural viral receptors with a possibility to selectively distinguish between potentially pathogenic influenza subtypes by the glycan-based biosensors. Build-up of the biosensor was carefully optimized with atomic force microscopy applied for visualization of the biosensor surface after binding of viruses with the topology of an individual viral particle H3N2 analyzed. The glycan biosensor could detect a glycan binding lectin with a limit of detection (LOD) of 5 aM. The biosensor was finally applied for analysis of influenza viruses H3N2 with LOD of 13 viral particles in 1 μl, what is the lowest LOD for analysis of influenza viral particles by the glycan-based device achieved so far. The biosensor could detect H3N2 viruses selectively with a sensitivity ratio of 30 over influenza viruses H7N7. The impedimetric biosensor presented here is the most sensitive glycan-based device for detection of influenza viruses and among the most sensitive antibody or aptamer based biosensor devices.

  13. Multiplex SYBR Green Real-Time PCR Assay for Detection of Respiratory Viruses

    PubMed Central

    Sultani, Mozhdeh; Mokhtari Azad, Talat; Eshragian, Mohammadreza; Shadab, Azadeh; Naseri, Maryam; Eilami, Owrang; Yavarian, Jila

    2015-01-01

    Background: It is often difficult for a physician to distinguish between viral and bacterial causes of respiratory infections and this may result in overuse of antibiotics. In many cases of community-acquired respiratory infections, clinicians treat patients empirically. The development of molecular methods for direct detection of viruses has been progressed recently. Objectives: The objective of this study was recognizing the panel of respiratory RNA viruses by multiplex SYBR Green real-time polymerase chain reaction (PCR). Materials and Methods: Randomized 172 influenza-negative respiratory specimens of all age groups of hospitalized patients were collected. After RNA extraction, cDNA was synthesized. Three SYBR Green multiplex real-time PCR assays were developed for simultaneous detection of 12 respiratory RNA viruses. Each set of multiplex methods detected four viruses, the first set: respiratory syncytial virus, human metapneumovirus, rhinovirus, enterovirus; the second set: parainfluenza viruses 1 - 4 (PIV1-4); the third set: coronaviruses NL63, 229E, severe acute respiratory syndrome (SARS), and OC43. Results: Application of the multiplex SYBR Green real-time PCR in clinical samples from 172 patients in a one-year study resulted in detection of 19 (11.04%) PIV3, 9 (5.23%) PIV4, and 1 (0.58%) coronavirus NL63. All the positive samples were detected during December to March (2011 - 2012). Conclusions: Multiplex SYBR Green real-time PCR is a rapid and relatively inexpensive method for detection of respiratory viruses. PMID:26468358

  14. 9 CFR 113.55 - Detection of extraneous agents in Master Seed Virus.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... Master Seed Virus. 113.55 Section 113.55 Animals and Animal Products ANIMAL AND PLANT HEALTH INSPECTION SERVICE, DEPARTMENT OF AGRICULTURE VIRUSES, SERUMS, TOXINS, AND ANALOGOUS PRODUCTS; ORGANISMS AND VECTORS STANDARD REQUIREMENTS Ingredient Requirements § 113.55 Detection of extraneous agents in Master Seed...

  15. Detecting the Presence of Nora Virus in "Drosophila" Utilizing Single Fly RT-PCR

    ERIC Educational Resources Information Center

    Munn, Bethany; Ericson, Brad; Carlson, Darby J.; Carlson, Kimberly A.

    2015-01-01

    A single fly RT-PCR protocol has recently been developed to detect the presence of the persistent, horizontally transmitted Nora virus in "Drosophila." Wild-caught flies from Ohio were tested for the presence of the virus, with nearly one-fifth testing positive. The investigation presented can serve as an ideal project for biology…

  16. Evidence of endemic Hendra virus infection in flying-foxes (Pteropus conspicillatus)--implications for disease risk management.

    PubMed

    Breed, Andrew C; Breed, Martin F; Meers, Joanne; Field, Hume E

    2011-01-01

    This study investigated the seroepidemiology of Hendra virus in a spectacled flying-fox (Pteropus conspicillatus) population in northern Australia, near the location of an equine and associated human Hendra virus infection in late 2004. The pattern of infection in the population was investigated using a serial cross-sectional serological study over a 25-month period, with blood sampled from 521 individuals over six sampling sessions. Antibody titres to the virus were determined by virus neutralisation test. In contrast to the expected episodic infection pattern, we observed that seroprevalence gradually increased over the two years suggesting infection was endemic in the population over the study period. Our results suggested age, pregnancy and lactation were significant risk factors for a detectable neutralizing antibody response. Antibody titres were significantly higher in females than males, with the highest titres occurring in pregnant animals. Temporal variation in antibody titres suggests that herd immunity to the virus may wax and wane on a seasonal basis. These findings support an endemic infection pattern of henipaviruses in bat populations suggesting their infection dynamics may differ significantly from the acute, self limiting episodic pattern observed with related viruses (e.g. measles virus, phocine distemper virus, rinderpest virus) hence requiring a much smaller critical host population size to sustain the virus. These findings help inform predictive modelling of henipavirus infection in bat populations, and indicate that the life cycle of the reservoir species should be taken into account when developing risk management strategies for henipaviruses.

  17. Evidence of endemic Hendra virus infection in flying-foxes (Pteropus conspicillatus)--implications for disease risk management.

    PubMed

    Breed, Andrew C; Breed, Martin F; Meers, Joanne; Field, Hume E

    2011-01-01

    This study investigated the seroepidemiology of Hendra virus in a spectacled flying-fox (Pteropus conspicillatus) population in northern Australia, near the location of an equine and associated human Hendra virus infection in late 2004. The pattern of infection in the population was investigated using a serial cross-sectional serological study over a 25-month period, with blood sampled from 521 individuals over six sampling sessions. Antibody titres to the virus were determined by virus neutralisation test. In contrast to the expected episodic infection pattern, we observed that seroprevalence gradually increased over the two years suggesting infection was endemic in the population over the study period. Our results suggested age, pregnancy and lactation were significant risk factors for a detectable neutralizing antibody response. Antibody titres were significantly higher in females than males, with the highest titres occurring in pregnant animals. Temporal variation in antibody titres suggests that herd immunity to the virus may wax and wane on a seasonal basis. These findings support an endemic infection pattern of henipaviruses in bat populations suggesting their infection dynamics may differ significantly from the acute, self limiting episodic pattern observed with related viruses (e.g. measles virus, phocine distemper virus, rinderpest virus) hence requiring a much smaller critical host population size to sustain the virus. These findings help inform predictive modelling of henipavirus infection in bat populations, and indicate that the life cycle of the reservoir species should be taken into account when developing risk management strategies for henipaviruses. PMID:22194920

  18. Evidence of Endemic Hendra Virus Infection in Flying-Foxes (Pteropus conspicillatus)—Implications for Disease Risk Management

    PubMed Central

    Breed, Andrew C.; Breed, Martin F.; Meers, Joanne; Field, Hume E.

    2011-01-01

    This study investigated the seroepidemiology of Hendra virus in a spectacled flying-fox (Pteropus conspicillatus) population in northern Australia, near the location of an equine and associated human Hendra virus infection in late 2004. The pattern of infection in the population was investigated using a serial cross-sectional serological study over a 25-month period, with blood sampled from 521 individuals over six sampling sessions. Antibody titres to the virus were determined by virus neutralisation test. In contrast to the expected episodic infection pattern, we observed that seroprevalence gradually increased over the two years suggesting infection was endemic in the population over the study period. Our results suggested age, pregnancy and lactation were significant risk factors for a detectable neutralizing antibody response. Antibody titres were significantly higher in females than males, with the highest titres occurring in pregnant animals. Temporal variation in antibody titres suggests that herd immunity to the virus may wax and wane on a seasonal basis. These findings support an endemic infection pattern of henipaviruses in bat populations suggesting their infection dynamics may differ significantly from the acute, self limiting episodic pattern observed with related viruses (e.g. measles virus, phocine distemper virus, rinderpest virus) hence requiring a much smaller critical host population size to sustain the virus. These findings help inform predictive modelling of henipavirus infection in bat populations, and indicate that the life cycle of the reservoir species should be taken into account when developing risk management strategies for henipaviruses. PMID:22194920

  19. Double-Stranded RNA Is Detected by Immunofluorescence Analysis in RNA and DNA Virus Infections, Including Those by Negative-Stranded RNA Viruses

    PubMed Central

    Son, Kyung-No; Liang, Zhiguo

    2015-01-01

    ABSTRACT Early biochemical studies of viral replication suggested that most viruses produce double-stranded RNA (dsRNA), which is essential for the induction of the host immune response. However, it was reported in 2006 that dsRNA could be detected by immunofluorescence antibody staining in double-stranded DNA and positive-strand RNA virus infections but not in negative-strand RNA virus infections. Other reports in the literature seemed to support these observations. This suggested that negative-strand RNA viruses produce little, if any, dsRNA or that more efficient viral countermeasures to mask dsRNA are mounted. Because of our interest in the use of dsRNA antibodies for virus discovery, particularly in pathological specimens, we wanted to determine how universal immunostaining for dsRNA might be in animal virus infections. We have detected the in situ formation of dsRNA in cells infected with vesicular stomatitis virus, measles virus, influenza A virus, and Nyamanini virus, which represent viruses from different negative-strand RNA virus families. dsRNA was also detected in cells infected with lymphocytic choriomeningitis virus, an ambisense RNA virus, and minute virus of mice (MVM), a single-stranded DNA (ssDNA) parvovirus, but not hepatitis B virus. Although dsRNA staining was primarily observed in the cytoplasm, it was also seen in the nucleus of cells infected with influenza A virus, Nyamanini virus, and MVM. Thus, it is likely that most animal virus infections produce dsRNA species that can be detected by immunofluorescence staining. The apoptosis induced in several uninfected cell lines failed to upregulate dsRNA formation. IMPORTANCE An effective antiviral host immune response depends on recognition of viral invasion and an intact innate immune system as a first line of defense. Double-stranded RNA (dsRNA) is a viral product essential for the induction of innate immunity, leading to the production of type I interferons (IFNs) and the activation of hundreds

  20. DEVELOPMENT OF MOLECULAR METHODS TO DETECT EMERGING VIRUSES

    EPA Science Inventory

    A large number of human enteric viruses are known to cause gastrointestinal illness and waterborne outbreaks. Many of these are emerging viruses that do not grow or grow poorly in cell culture and so molecular detectoin methods based on the polymerase chain reaction (PCR) are be...

  1. Development and evaluation of a Luminex multiplex serology assay to detect antibodies to bovine herpes virus 1, parainfluenza 3 virus, bovine viral diarrhoea virus, and bovine respiratory syncytial virus, with comparison to existing ELISA detection methods.

    PubMed

    Anderson, Steve; Wakeley, Phil; Wibberley, Guy; Webster, Kath; Sawyer, Jason

    2011-03-01

    Detection of circulating antibodies to bovine herpes virus 1 (BHV-1), parainfluenza 3 virus (PI3V), bovine viral diarrhoea virus (BVDV) and bovine respiratory syncytial virus (BRSV) using ELISA is widely used for veterinary diagnostics and surveillance. In this paper, the potential of a multiplex serology test based on Luminex technology, where all antibodies are simultaneously detected in a single assay was investigated. The performance of "in-house" separate ELISAs which use relatively crude lysates of cultured virus as capture antigens, was compared to the multiplex assay where the same antigens were covalently bound to the fluorescent beads used in the Luminex platform. A panel of field serum samples was tested by the multiplex assay in parallel with the separate routine ELISAs to provide a comparison between tests. The BHV-1 and PI3V components of the multiplex test showed similar sensitivities and specificities to the separate "in-house" ELISAs. The performance of the BVDV and BRSV components was less successful and was attributed to relatively low signal strength for these antigens, leading to higher assay variability and a reduced ability to distinguish positive and negative samples compared to the "in-house" ELISAs. The results illustrated that antigens commonly used successfully in ELISAs cannot always be transferred for use in alternative assay systems. The use of recombinant BVDV E2 protein was investigated and was shown to lead to an appreciable increase in signal strength compared to the use of crude BVDV antigen in the Luminex system.

  2. Rapid detection of Ebola virus with a reagent-free, point-of-care biosensor

    SciTech Connect

    Baca, Justin T.; Severns, Virginia; Lovato, Debbie; Branch, Darren W.; Larson, Richard S.

    2015-04-14

    Surface acoustic wave (SAW) sensors can rapidly detect Ebola antigens at the point-of-care without the need for added reagents, sample processing, or specialized personnel. This preliminary study demonstrates SAW biosensor detection of the Ebola virus in a concentration-dependent manner. The detection limit with this methodology is below the average level of viremia detected on the first day of symptoms by PCR. We observe a log-linear sensor response for highly fragmented Ebola viral particles, with a detection limit corresponding to 1.9 × 10⁴ PFU/mL prior to virus inactivation. We predict greatly improved sensitivity for intact, infectious Ebola virus. This point-of-care methodology has the potential to detect Ebola viremia prior to symptom onset, greatly enabling infection control and rapid treatment. This biosensor platform is powered by disposable AA batteries and can be rapidly adapted to detect other emerging diseases in austere conditions.

  3. Rapid detection of Ebola virus with a reagent-free, point-of-care biosensor

    DOE PAGES

    Baca, Justin T.; Severns, Virginia; Lovato, Debbie; Branch, Darren W.; Larson, Richard S.

    2015-04-14

    Surface acoustic wave (SAW) sensors can rapidly detect Ebola antigens at the point-of-care without the need for added reagents, sample processing, or specialized personnel. This preliminary study demonstrates SAW biosensor detection of the Ebola virus in a concentration-dependent manner. The detection limit with this methodology is below the average level of viremia detected on the first day of symptoms by PCR. We observe a log-linear sensor response for highly fragmented Ebola viral particles, with a detection limit corresponding to 1.9 × 10⁴ PFU/mL prior to virus inactivation. We predict greatly improved sensitivity for intact, infectious Ebola virus. This point-of-care methodologymore » has the potential to detect Ebola viremia prior to symptom onset, greatly enabling infection control and rapid treatment. This biosensor platform is powered by disposable AA batteries and can be rapidly adapted to detect other emerging diseases in austere conditions.« less

  4. Type- and subtype-specific detection of influenza viruses in clinical specimens by rapid culture assay.

    PubMed

    Ziegler, T; Hall, H; Sánchez-Fauquier, A; Gamble, W C; Cox, N J

    1995-02-01

    A rapid culture assay which allows for the simultaneous typing and subtyping of currently circulating influenza A(H1N1), A(H3N2), and B viruses in clinical specimens was developed. Pools of monoclonal antibodies (MAbs) against influenza A and B viruses and MAbs HA1-71 and HA2-76, obtained by immunizing mice with the denatured hemagglutinin subfragments HA1 and HA2 of influenza virus A/Victoria/3/75, were used for immunoperoxidase staining of antigens in infected MDCK cells. MAb HA1-71 reacted exclusively with influenza A viruses of the H3 subtype, while MAb HA2-76 reacted with subtypes H1, H3, H4, H6, H8, H9, H10, H11, and H12, as determined with 78 human, 4 swine, and 10 avian influenza virus reference strains subtyped by the hemagglutination inhibition test. To determine if the technique can be used as a rapid diagnostic test, 263 known influenza virus-positive frozen nasal or throat swabs were inoculated into MDCK cells. After an overnight incubation, the cells were fixed and viral antigens were detected by immunoperoxidase staining. Influenza A viruses of the H1 and H3 subtypes were detected in 31 and 113 specimens, respectively. The subtypes of 10 influenza A virus-positive specimens could not be determined because they contained too little virus. Influenza B viruses were detected in 84 specimens, and 25 specimens were negative. We conclude that this assay is a rapid, convenient, non-labor-intensive, and relatively inexpensive test for detecting, typing, and subtyping influenza viruses in clinical specimens. PMID:7714186

  5. DNA Microarray Platform for Detection and Surveillance of Viruses Transmitted by Small Mammals and Arthropods

    PubMed Central

    Khan, Mohd Jaseem; Trabuco, Amanda Cristina; Alfonso, Helda Liz; Figueiredo, Mario Luis; Batista, Weber Cheli; Badra, Soraya Jabur; Figueiredo, Luiz Tadeu; Lavrador, Marco Aurélio

    2016-01-01

    Viruses transmitted by small mammals and arthropods serve as global threats to humans. Most emergent and re-emergent viral agents are transmitted by these groups; therefore, the development of high-throughput screening methods for the detection and surveillance of such viruses is of great interest. In this study, we describe a DNA microarray platform that can be used for screening all viruses transmitted by small mammals and arthropods (SMAvirusChip) with nucleotide sequences that have been deposited in the GenBank. SMAvirusChip was designed with more than 15,000 oligonucleotide probes (60-mers), including viral and control probes. Two SMAvirusChip versions were designed: SMAvirusChip v1 contains 4209 viral probes for the detection of 409 viruses, while SMAvirusChip v2 contains 4943 probes for the detection of 416 viruses. SMAvirusChip was evaluated with 20 laboratory reference-strain viruses. These viruses could be specifically detected when alone in a sample or when artificially mixed within a single sample. The sensitivity of SMAvirusChip was evaluated using 10-fold serial dilutions of dengue virus (DENV). The results showed a detection limit as low as 2.6E3 RNA copies/mL. Additionally, the sensitivity was one log10 lower (2.6E2 RNA copies/mL) than quantitative real-time RT-PCR and sufficient to detect viral genomes in clinical samples. The detection of DENV in serum samples of DENV-infected patients (n = 6) and in a whole blood sample spiked with DENV confirmed the applicability of SMAvirusChip for the detection of viruses in clinical samples. In addition, in a pool of mosquito samples spiked with DENV, the virus was also detectable. SMAvirusChip was able to specifically detect viruses in cell cultures, serum samples, total blood samples and a pool of mosquitoes, confirming that cellular RNA/DNA did not interfere with the assay. Therefore, SMAvirusChip may represent an innovative surveillance method for the rapid identification of viruses transmitted by small

  6. Methodological Guidelines for Accurate Detection of Viruses in Wild Plant Species

    PubMed Central

    Renner, Kurra; Cole, Ellen; Seabloom, Eric W.; Borer, Elizabeth T.; Malmstrom, Carolyn M.

    2016-01-01

    Ecological understanding of disease risk, emergence, and dynamics and of the efficacy of control strategies relies heavily on efficient tools for microorganism identification and characterization. Misdetection, such as the misclassification of infected hosts as healthy, can strongly bias estimates of disease prevalence and lead to inaccurate conclusions. In natural plant ecosystems, interest in assessing microbial dynamics is increasing exponentially, but guidelines for detection of microorganisms in wild plants remain limited, particularly so for plant viruses. To address this gap, we explored issues and solutions associated with virus detection by serological and molecular methods in noncrop plant species as applied to the globally important Barley yellow dwarf virus PAV (Luteoviridae), which infects wild native plants as well as crops. With enzyme-linked immunosorbent assays (ELISA), we demonstrate how virus detection in a perennial wild plant species may be much greater in stems than in leaves, although leaves are most commonly sampled, and may also vary among tillers within an individual, thereby highlighting the importance of designing effective sampling strategies. With reverse transcription-PCR (RT-PCR), we demonstrate how inhibitors in tissues of perennial wild hosts can suppress virus detection but can be overcome with methods and products that improve isolation and amplification of nucleic acids. These examples demonstrate the paramount importance of testing and validating survey designs and virus detection methods for noncrop plant communities to ensure accurate ecological surveys and reliable assumptions about virus dynamics in wild hosts. PMID:26773088

  7. Simple and Portable Magnetic Immunoassay for Rapid Detection and Sensitive Quantification of Plant Viruses

    PubMed Central

    Rettcher, Stefanie; Jungk, Felicitas; Kühn, Christoph; Krause, Hans-Joachim; Nölke, Greta; Commandeur, Ulrich; Fischer, Rainer; Schillberg, Stefan

    2015-01-01

    Plant pathogens cause major economic losses in the agricultural industry because late detection delays the implementation of measures that can prevent their dissemination. Sensitive and robust procedures for the rapid detection of plant pathogens are therefore required to reduce yield losses and the use of expensive, environmentally damaging chemicals. Here we describe a simple and portable system for the rapid detection of viral pathogens in infected plants based on immunofiltration, subsequent magnetic detection, and the quantification of magnetically labeled virus particles. Grapevine fanleaf virus (GFLV) was chosen as a model pathogen. Monoclonal antibodies recognizing the GFLV capsid protein were immobilized onto immunofiltration columns, and the same antibodies were linked to magnetic nanoparticles. GFLV was quantified by immunofiltration with magnetic labeling in a double-antibody sandwich configuration. A magnetic frequency mixing technique, in which a two-frequency magnetic excitation field was used to induce a sum frequency signal in the resonant detection coil, corresponding to the virus concentration within the immunofiltration column, was used for high-sensitivity quantification. We were able to measure GFLV concentrations in the range of 6 ng/ml to 20 μg/ml in less than 30 min. The magnetic immunoassay could also be adapted to detect other plant viruses, including Potato virus X and Tobacco mosaic virus, with detection limits of 2 to 60 ng/ml. PMID:25710366

  8. Simple and portable magnetic immunoassay for rapid detection and sensitive quantification of plant viruses.

    PubMed

    Rettcher, Stefanie; Jungk, Felicitas; Kühn, Christoph; Krause, Hans-Joachim; Nölke, Greta; Commandeur, Ulrich; Fischer, Rainer; Schillberg, Stefan; Schröper, Florian

    2015-05-01

    Plant pathogens cause major economic losses in the agricultural industry because late detection delays the implementation of measures that can prevent their dissemination. Sensitive and robust procedures for the rapid detection of plant pathogens are therefore required to reduce yield losses and the use of expensive, environmentally damaging chemicals. Here we describe a simple and portable system for the rapid detection of viral pathogens in infected plants based on immunofiltration, subsequent magnetic detection, and the quantification of magnetically labeled virus particles. Grapevine fanleaf virus (GFLV) was chosen as a model pathogen. Monoclonal antibodies recognizing the GFLV capsid protein were immobilized onto immunofiltration columns, and the same antibodies were linked to magnetic nanoparticles. GFLV was quantified by immunofiltration with magnetic labeling in a double-antibody sandwich configuration. A magnetic frequency mixing technique, in which a two-frequency magnetic excitation field was used to induce a sum frequency signal in the resonant detection coil, corresponding to the virus concentration within the immunofiltration column, was used for high-sensitivity quantification. We were able to measure GFLV concentrations in the range of 6 ng/ml to 20 μg/ml in less than 30 min. The magnetic immunoassay could also be adapted to detect other plant viruses, including Potato virus X and Tobacco mosaic virus, with detection limits of 2 to 60 ng/ml. PMID:25710366

  9. Patterns of detection of respiratory viruses in nasal swabs from calves in Ireland: a retrospective study.

    PubMed

    O'Neill, R; Mooney, J; Connaghan, E; Furphy, C; Graham, D A

    2014-10-11

    A retrospective analysis was conducted to investigate the prevalence and seasonality of bovine viral diarrhoea virus (BVDV), bovine coronavirus (BoCV), bovine herpesvirus-1 (BoHV-1), bovine respiratory syncytical virus (BRSV) and parainfluenza virus-3 (PI3V) in calves (aged three months and below) in Ireland. Results from real-time PCR testing, including cycle threshold values, conducted on nasal swabs (single or pooled) submitted from 1364 respiratory disease outbreaks between January 1, 2008 and December 31, 2012 were included in this study. One or more viruses were detected in 34.6 per cent of submissions, with BoCV detected most frequently (22.9 per cent), followed by BRSV (11.6 per cent), PI3 V (7.0 per cent), BoHV-1 (6.1 per cent) and BVDV (5.0 per cent). The detection rate of all viruses was higher when pooled multiple swabs were submitted from outbreaks rather than single swabs, with these differences being significant for all except BVDV. Two or more viruses were detected in 39.4 per cent of positive submissions, with BoCV and BRSV most commonly present as one of the two partners in detection. With the exception of BVDV, which was detected all year round, the others showed a clear seasonal pattern, being most commonly detected in winter and spring. PMID:25037889

  10. Individual donor nucleic acid amplification testing for detection of West Nile virus.

    PubMed

    Lee, Dong-Hun; Mathew, John; Pfahler, Wolfram; Ma, Dongling; Valinsky, Jay; Prince, Alfred M; Andrus, Linda

    2005-10-01

    We have developed an economical, high-throughput nucleic acid amplification test (NAT) for blood-borne viruses, suitable for use in the screening of plasma samples from individual blood donors. This assay system includes a semiautomated procedure, using 96-well glass fiber plates for the extraction of viral nucleic acids from plasma and "universal beacon" technology which permits the detection of all genotypes of highly variable viruses (e.g., human immunodeficiency virus and hepatitis C virus). In this detection system, two fluorescent- detection technologies were employed successfully in a single tube: molecular beacon for West Nile virus (WNV) detection using a 6-carboxyfluorescein fluorophore and TaqMan for internal control detection using a VIC fluorophore. To establish proof of concept, we focused on the development of a robust individual donor NAT for WNV. The assay showed no reactivity to 15 other viruses tested or to 420 blood donor samples from the WNV pre-epidemic season. No cross-contamination was observed on an alternating positive-/negative-well test. The sensitivity (limit of detection, 95%) of the assay for WNV is between 3.79 and 16.3 RNA copies/ml, depending on which material was used as a standard. The assay detected all positive blood donation samples identified by the Roche WNV NAT. The assay can be performed qualitatively for screening and quantitatively for confirmation.

  11. Detection of sweet potato virus C, sweet potato virus 2 and sweet potato feathery mottle virus in Portugal.

    PubMed

    Varanda, Carla M R; Santos, Susana J; Oliveira, Mônica D M; Clara, Maria Ivone E; Félix, Maria Rosário F

    2015-06-01

    Field sweet potato plants showing virus-like symptoms, as stunting, leaf distortion, mosaic and chlorosis, were collected in southwest Portugal and tested for the presence of four potyviruses, sweet potato virus C (SPVC), sweet potato virus 2 (SPV2), sweet potato feathery mottle virus (SPFMV), sweet potato virus G (SPVG), and the crinivirus sweet potato chlorotic stunt virus (SPCSV). DsRNA fractions were extracted from symptomatic leaves and used as templates in single and multiplex RT-PCR assays using previously described specific primers for each analyzed virus. The amplified reaction products for SPVC, SPV2 and SPFMV were of expected size, and direct sequencing of PCR products revealed that they correspond to the coat protein gene (CP) and showed 98%, 99% and 99% identity, respectively, to those viruses. Comparison of the CP genomic and amino acid sequences of the Portuguese viral isolates recovered here with those of ten other sequences of isolates obtained in different countries retrieved from the GenBank showed very few differences. The application of the RT-PCR assays revealed for the first time the presence of SPVC and SPFMV in the sweet potato crop in Portugal, the absence of SPVG and SPCSV in tested plants, as well as the occurrence of triple virus infections under field conditions.

  12. Detection of sweet potato virus C, sweet potato virus 2 and sweet potato feathery mottle virus in Portugal.

    PubMed

    Varanda, Carla M R; Santos, Susana J; Oliveira, Mônica D M; Clara, Maria Ivone E; Félix, Maria Rosário F

    2015-06-01

    Field sweet potato plants showing virus-like symptoms, as stunting, leaf distortion, mosaic and chlorosis, were collected in southwest Portugal and tested for the presence of four potyviruses, sweet potato virus C (SPVC), sweet potato virus 2 (SPV2), sweet potato feathery mottle virus (SPFMV), sweet potato virus G (SPVG), and the crinivirus sweet potato chlorotic stunt virus (SPCSV). DsRNA fractions were extracted from symptomatic leaves and used as templates in single and multiplex RT-PCR assays using previously described specific primers for each analyzed virus. The amplified reaction products for SPVC, SPV2 and SPFMV were of expected size, and direct sequencing of PCR products revealed that they correspond to the coat protein gene (CP) and showed 98%, 99% and 99% identity, respectively, to those viruses. Comparison of the CP genomic and amino acid sequences of the Portuguese viral isolates recovered here with those of ten other sequences of isolates obtained in different countries retrieved from the GenBank showed very few differences. The application of the RT-PCR assays revealed for the first time the presence of SPVC and SPFMV in the sweet potato crop in Portugal, the absence of SPVG and SPCSV in tested plants, as well as the occurrence of triple virus infections under field conditions. PMID:26104336

  13. Detection and characterization of enteric viruses in flood water from the 2011 thai flood.

    PubMed

    Ngaosuwankul, Nathamon; Thippornchai, Narin; Yamashita, Akifumi; Vargas, Ronald E Morales; Tunyong, Witawat; Mahakunkijchareon, Yuvadee; Ikuta, Kazuyoshi; Singhasivanon, Pratap; Okabayashi, Tamaki; Leaungwutiwong, Pornsawan

    2013-01-01

    Severe flooding, which is associated with numerous outbreaks of a wide range of infectious diseases, particularly those caused by enteric viruses, occurred in all areas of Thailand in 2011. To determine the prevalence of five human enteric viruses, namely enterovirus, rotavirus (RV), norovirus (NV), hepatitis A virus (HAV), and hepatitis E virus, in the flood water, 100 water samples were collected from flood-damaged areas in central Thailand. Viral RNA was extracted from concentrated samples and analyzed by RT-PCR and sequencing. NV was the most commonly detected pathogen in the tested samples (14%). RV and HAV were detected in 9% and 7% of samples, respectively. This study is the first to detect enteric viral genes in flood water in Thailand. Furthermore, it is the first to detect an NV gene in any type of environmental water in Thailand. These results provide useful information for estimating the risk of flood waterborne viral infection.

  14. Rabies, canine distemper, and canine parvovirus exposure in large carnivore communities from two Zambian ecosystems.

    PubMed

    Berentsen, Are R; Dunbar, Mike R; Becker, Matthew S; M'soka, Jassiel; Droge, Egil; Sakuya, Nicholas M; Matandiko, Wigganson; McRobb, Rachel; Hanlon, Cathleen A

    2013-09-01

    Disease transmission within and among wild and domestic carnivores can have significant impacts on populations, particularly for threatened and endangered species. We used serology to evaluate potential exposure to rabies virus, canine distemper virus (CDV), and canine parvovirus (CPV) for populations of African lions (Panthera leo), African wild dogs (Lycaon pictus), and spotted hyenas (Crocuta crocuta) in Zambia's South Luangwa National Park (SLNP) and Liuwa Plain National Park (LPNP) as well as community lands bordering these areas. In addition, domestic dogs in the study region were evaluated for exposure to CDV and rabies. We provide the first comprehensive disease exposure data for these species in these ecosystems. Twenty-one lions, 20 hyenas, 13 wild dogs, and 38 domestic dogs were sampled across both regions from 2009 to 2011. Laboratory results show 10.5% of domestic dogs, 5.0% of hyenas, and 7.7% of wild dogs sampled were positive for CDV exposure. All lions were negative. Exposure to CPV was 10.0% and 4.8% for hyenas and lions, respectively. All wild dogs were negative, and domestic dogs were not tested due to insufficient serum samples. All species sampled were negative for rabies virus neutralizing antibodies except lions. Forty percent of lions tested positive for rabies virus neutralizing antibodies. Because these lions appeared clinically healthy, this finding is consistent with seroconversion following exposure to rabies antigen. To our knowledge, this finding represents the first ever documentation of rabies virus neutralizing antibodies consistent with rabies exposure that did not lead to clinical disease in free-ranging African lions from this region. With ever-increasing human pressure on these ecosystems, understanding disease transmission dynamics is essential for proper management and conservation of these carnivore species.

  15. Rabies, canine distemper, and canine parvovirus exposure in large carnivore communities from two Zambian ecosystems.

    PubMed

    Berentsen, Are R; Dunbar, Mike R; Becker, Matthew S; M'soka, Jassiel; Droge, Egil; Sakuya, Nicholas M; Matandiko, Wigganson; McRobb, Rachel; Hanlon, Cathleen A

    2013-09-01

    Disease transmission within and among wild and domestic carnivores can have significant impacts on populations, particularly for threatened and endangered species. We used serology to evaluate potential exposure to rabies virus, canine distemper virus (CDV), and canine parvovirus (CPV) for populations of African lions (Panthera leo), African wild dogs (Lycaon pictus), and spotted hyenas (Crocuta crocuta) in Zambia's South Luangwa National Park (SLNP) and Liuwa Plain National Park (LPNP) as well as community lands bordering these areas. In addition, domestic dogs in the study region were evaluated for exposure to CDV and rabies. We provide the first comprehensive disease exposure data for these species in these ecosystems. Twenty-one lions, 20 hyenas, 13 wild dogs, and 38 domestic dogs were sampled across both regions from 2009 to 2011. Laboratory results show 10.5% of domestic dogs, 5.0% of hyenas, and 7.7% of wild dogs sampled were positive for CDV exposure. All lions were negative. Exposure to CPV was 10.0% and 4.8% for hyenas and lions, respectively. All wild dogs were negative, and domestic dogs were not tested due to insufficient serum samples. All species sampled were negative for rabies virus neutralizing antibodies except lions. Forty percent of lions tested positive for rabies virus neutralizing antibodies. Because these lions appeared clinically healthy, this finding is consistent with seroconversion following exposure to rabies antigen. To our knowledge, this finding represents the first ever documentation of rabies virus neutralizing antibodies consistent with rabies exposure that did not lead to clinical disease in free-ranging African lions from this region. With ever-increasing human pressure on these ecosystems, understanding disease transmission dynamics is essential for proper management and conservation of these carnivore species. PMID:23805791

  16. A two-tube multiplex real-time RT-PCR assay for the detection of four hemorrhagic fever viruses: severe fever with thrombocytopenia syndrome virus, Hantaan virus, Seoul virus, and dengue virus.

    PubMed

    Li, Zhifeng; Qi, Xian; Zhou, Minghao; Bao, Changjun; Hu, Jianli; Wu, Bin; Wang, Shenjiao; Tan, Zhongmin; Fu, Jianguang; Shan, Jun; Zhu, Yefei; Tang, Fenyang

    2013-09-01

    The aim of this study was to develop and evaluate a two-tube multiplex real-time RT-PCR assay for the detection and identification of four viral hemorrhagic fever (VHF) pathogens, severe fever with thrombocytopenia syndrome virus (SFTSV), Hantaan virus (HTNV), Seoul virus (SEOV), and dengue virus (DENV), from human clinical samples. The two-tube multiplex real-time RT-PCR assay we developed has a sensitivity of 10 copies/μL for each of the targets, and the performance was linear within the range of at least 10(7) transcript copies. Moreover, we evaluated the specificity of the assay using other virus RNA as template, and found no cross-reactivity. This new assay is able to detect SFTSV, HTNV, SEOV and DENV in two reactions and brings a cost of 40 % compared to separate reactions. Evaluation of this assay with clinical serum samples from laboratory-confirmed patients and healthy donors showed 100 % clinical diagnostic sensitivity and over 99 % specificity. The assay was applied for scanning 346 clinical samples collected from patients admitted to the hospital with suspected VHF and compared with virus isolation and immunofluorescence assay (IFA). The assay indentified 59 SFTSV-, 12 HTNV-, 11 SEOV- and 9 DENV-positive samples and showed higher sensitivity. This assay thus provides a reliable and cost-effective screening tool for early clinical diagnosis of SFTSV, HTNV, SEOV and DENV in the acute phase.

  17. Infectious hematopoietic necrosis virus detected by separation and incubation of cells from salmonid cavity fluid.

    USGS Publications Warehouse

    Mulcahy, D.; Batts, W.N.

    1987-01-01

    Infectious hematopoietic necrosis (IHN) virus is usually detected by inoculating susceptible cell cultures with cavity ("ovarian") fluid (CF) from spawning females. We identified additional adult carriers of virus in spawning populations of steelhead trout (Salmo gairdneri) and sockeye salmon (Oncorhynchus nerka) by collecting nonerythrocytic cells from CF samples by low-speed centrifugation, culturing the cells for at least 7 d at 15 °C, and then testing the culture medium for virus. Virus appeared in the cultured cells from some samples of CF that remained negative during incubation. In additional samples of CF from these species, the virus titer increased in cultured cells compared with the titer in the original CF sample. With chinook salmon (O.tshawytscha), no negative samples converted to positive during incubation, but the virus titer was retained in incubated CF cells, but not in cell-free CF.

  18. Single virus detection by means of atomic force microscopy in combination with advanced image analysis.

    PubMed

    Bocklitz, Thomas; Kämmer, Evelyn; Stöckel, Stephan; Cialla-May, Dana; Weber, Karina; Zell, Roland; Deckert, Volker; Popp, Jürgen

    2014-10-01

    In the present contribution virions of five different virus species, namely Varicella-zoster virus, Porcine teschovirus, Tobacco mosaic virus, Coliphage M13 and Enterobacteria phage PsP3, are investigated using atomic force microscopy (AFM). From the resulting height images quantitative features like maximal height, area and volume of the viruses could be extracted and compared to reference values. Subsequently, these features were accompanied by image moments, which quantify the morphology of the virions. Both types of features could be utilized for an automatic discrimination of the five virus species. The accuracy of this classification model was 96.8%. Thus, a virus detection on a single-particle level using AFM images is possible. Due to the application of advanced image analysis the morphology could be quantified and used for further analysis. Here, an automatic recognition by means of a classification model could be achieved in a reliable and objective manner. PMID:25196422

  19. Host-Encoded Reporters for the Detection and Purification of Multiple Enveloped Viruses

    PubMed Central

    Ketteler, Robin; Tomov, Vesko; Neunkirchner, Alina; Xie, Qiang; Pickl, Winfried F.; Seed, Brian

    2010-01-01

    The identification of host cell factors for virus replication holds great promise for the development of new anti-viral therapies. Recently, high-throughput screening methods have emerged as powerful tools to identify candidate host factors for therapeutic intervention. The development of assay systems suitable for large-scale automated screening is of particular importance for novel viruses with high pathogenic potential for which limited biological information can be developed in a short period of time. This report presents a general enzymatic reporter system for the detection and characterization of multiple enveloped viruses that does not rely on engineering of the virus. Instead, reporter enzymes are incorporated into virus particles by targeting to lipid microdomains in producer cells. The approach allows a variety of human pathogenic enveloped viruses to be detected by sensitive, inexpensive and automatable enzymatic assays. Tagged viruses can be purified quickly and efficiently by a magnetic bead-based capture method. The method allows general detection of enveloped viruses without prior reference to their sequence. PMID:20399809

  20. Detection of plant viruses in mixed infection by a macroarray-assisted method.

    PubMed

    Shimura, Hanako; Furuta, Kazuyoshi; Masuta, Chikara

    2015-01-01

    The protocol for a simple, sensitive, and specific method using a cDNA macroarray to detect multiple viruses is provided. The method can be used even at the production sites for crops, which need a reliable routine diagnosis for mixed infection of plant viruses. The method consists of three steps: RNA extraction, duplex RT-PCR, and "microtube hybridization" (MTH). Biotinylated cDNA probes are prepared using RT-PCR and used to hybridize a nylon membrane containing target viral cDNAs by MTH. Positive signals can be visualized by colorimetric reaction and judged by eyes. We here demonstrate this method to detect asparagus viruses (Asparagus virus 1 and Asparagus virus 2) from latently infected asparagus plants. PMID:25287491

  1. Real-time detection of airborne viruses on a mass-sensitive device

    NASA Astrophysics Data System (ADS)

    Lee, Joonhyung; Jang, Jaesung; Akin, Demir; Savran, Cagri A.; Bashir, Rashid

    2008-07-01

    We present real-time detection of airborne Vaccinia viruses using quartz crystal microbalance (QCM) in an integrated manner. Vaccinia viruses were aerosolized and neutralized using an electrospray aerosol generator, transported into the QCM chamber, and captured by a QCM crystal. The capture of the viruses on the QCM crystal resulted in frequency shifts proportional to the number of viruses. The capture rate varied linearly with the concentration of initial virus suspensions (8.5×108-8.5×1010particles/ml) at flow rates of 2.0 and 1.1l/min. This work demonstrates the general potential of mass sensitive detection of nanoscale biological entities in air.

  2. Detection of two different influenza A viruses using a nitrocellulose membrane and a magnetic biosensor.

    PubMed

    Hong, Hyo-Bong; Krause, Hans-Joachim; Song, Ki-Bong; Choi, Chel-Jong; Chung, Myung-Ae; Son, Sung-Won; Offenhäusser, Andreas

    2011-02-28

    Here we describe a new analytical method for the detection of two influenza A viruses by nitrocellulose membrane and magnetic sensors that employ a special frequency mixing technique. The combination of the nitrocellulose membrane and magnetic bead detection permits a rapid assay procedure and excludes two steps (the development of color and the stop reaction) required for usual immunochemical detection methods such as ELISA. Quantitative virus detection was performed using magnetic beads conjugated with secondary antibody. The results were compared with conventional assay methods and with a dot-blot assay with fluorescence compound (FITC). Under optimum conditions, our new assay procedure is capable of detecting picograms of virus per well. This new method combining the nitrocellulose membrane and magnetic bead detection reduces analytical time and allows stable and repeatable analyses of samples in point-of-care applications.

  3. A mediator embedded micro-immunosensing unit for electrochemical detection on viruses within physiological saline media

    NASA Astrophysics Data System (ADS)

    Pires, Nuno M. M.; Dong, Tao; Yang, Zhaochu; Høivik, Nils; Zhao, Xinyan

    2011-11-01

    To provide a time- and cost-saving alternative to the conventional methods for virus detection in biological media, this work presents an electrochemical micro-immunosensor based on the nickel hexacyanoferrate (NiHCF) redox mediator film coating the interdigitated microelectrodes (IDMEs). By chelation binding with no additional cross-linker, the 6xHis-tagged antibodies were immobilized on a NiHCF film. Secondly, an immunoassay response was enhanced by employing microbeads coated with 6xHis antibody. The electrochemical properties and the stability of the NiHCF film modified IDMEs were evaluated by cyclic voltammetry. The bead-induced impedance variations at the electrode film/electrolyte interface were characterized by electrochemical impedance spectroscopy and verified using FEM simulation. Experiments of virus detection were conducted through targeting the antigens of the vital infectious salmon viruses, such as infectious salmon anaemia virus, infectious pancreatic necrosis virus and salmonid alphavirus subtype 3. The micro-immunosensor exhibited detection limits as low as 10 pg ml-1 and detection sensitivities as high as 57.5 kΩ µM-1 within a physiological saline solution. Tests for multiple antigen-antibody interactions showed good detection specificity, as confirmed by ELISA. By incorporating the microfluidic network, electrochemical impedance micro-immunosensing units can be realized in a fully integrated platform for multiplex virus detection in tissue samples.

  4. Detection of white spot syndrome virus (WSSV) of Penaeus chinensis by in situ hybridization

    NASA Astrophysics Data System (ADS)

    Zhan, Wen-Bin; Wang, Yuan-Hong; Zhang, Zhi-Dong; Hideo, Fukuda

    2000-09-01

    White Spot Syndrome Virus (WSSV) was purified from hemolymph of infected shrimp. After nucleic acid extraction from the purified virus particles, EcoR I-digested fragments of the WSSV genome were cloned; three of these fragments were used as non-radioactive probes labeled with DIG-11-dUTP. The probes hybridized in situ, with sections located in the nuclei of all WSSV-infected tissues. The virus was detected in the gill, stomach, epidermis, and connective tissue and so on, but not detected in healthy shrimp tissues and epithelial cells of hepatopancreatic tubules of diseased shrimp.

  5. Carbon Nanotube Thin Film Biosensors for Sensitive and Reproducible Whole Virus Detection

    PubMed Central

    Mandal, Himadri S.; Su, Zhengding; Ward, Andrew; Tang, Xiaowu (Shirley)

    2012-01-01

    Here, we report the label-free, sensitive, and real-time electrical detection of whole viruses using carbon nanotube thin film (CNT-TF) field effect devices. Selective detection of approximately 550 model viruses, M13-bacteriophage, is demonstrated using a simple two-terminal (no gate electrode) configuration. Chemical gating through specific antibody-virus binding on CNT surface is proposed to be the sensing mechanism. Compared to electrical impedance sensors with identical microelectrode dimensions (no CNT), the CNT-TF sensors exhibit sensitivity 5 orders higher. We believe the reported approach could lead to a reproducible and cost-effective solution for rapid viral identification. PMID:22448194

  6. Comparison of the 1988 and 2002 phocine distemper epizootics in British harbour seal Phoca vitulina populations.

    PubMed

    Lonergan, Mike; Hall, Ailsa; Thompson, Hal; Thompson, Paul M; Pomeroy, Paddy; Harwood, John

    2010-02-17

    In 1988 and 2002 dramatic and well-documented phocine distemper epizootics occurred in Europe. While their progression and impact were remarkably similar and consistent over much of Europe, mortality in the UK varied greatly between and within the 2 epizootics. We use antibody levels in blood samples to show that 51% (Bayesian 95% CI: 41 to 61%) of the individuals alive in 5 UK harbour seal populations at the end of the 1988 epizootic had been exposed to the virus, and that the equivalent figure after the 2002 outbreak was 22% (95% CI: 16 to 30%). Antibody prevalence was significantly higher in females than males after the 2002 epizootic. Combining these estimates with information on reductions in the numbers of animals observed hauled out during surveys of the Wash, Moray Firth, and Orkney populations and a simple epidemiological model, suggests that the differences between the 2 epizootics were primarily due to a 27% (95% CI: 8 to 43%) fall in R0, the basic reproductive rate of the virus. The large geographic variation in population effects observed within the UK during each epizootic appears to have been mainly due to differences in case mortality, with R0 being remarkably similar in all the populations investigated. PMID:20377007

  7. Fatal canine distemper infection in a pack of African wild dogs in the Serengeti ecosystem, Tanzania.

    PubMed

    Goller, Katja V; Fyumagwa, Robert D; Nikolin, Veljko; East, Marion L; Kilewo, Morris; Speck, Stephanie; Müller, Thomas; Matzke, Martina; Wibbelt, Gudrun

    2010-12-15

    In 2007, disease related mortality occurred in one African wild dog (Lycaon pictus) pack close to the north-eastern boundary of the Serengeti National Park, Tanzania. Histopathological examination of tissues from six animals revealed that the main pathologic changes comprised interstitial pneumonia and suppurative to necrotizing bronchopneumonia. Respiratory epithelial cells contained numerous eosinophilic intracytoplasmic inclusion bodies and multiple syncytial cells were found throughout the parenchymal tissue, both reacting clearly positive with antibodies against canine distemper virus (CDV) antigen. Phylogenetic analysis based on a 388 nucleotide (nt) fragment of the CDV phosphoprotein (P) gene revealed that the pack was infected with a CDV variant most closely related to Tanzanian variants, including those obtained in 1994 during a CDV epidemic in the Serengeti National Park and from captive African wild dogs in the Mkomazi Game Reserve in 2000. Phylogenetic analysis of a 335-nt fragment of the fusion (F) gene confirmed that the pack in 2007 was infected with a variant most closely related to one variant from 1994 during the epidemic in the Serengeti National Park from which a comparable fragment is available. Screening of tissue samples for concurrent infections revealed evidence of canine parvovirus, Streptococcus equi subsp. ruminatorum and Hepatozoon sp. No evidence of infection with Babesia sp. or rabies virus was found. Possible implications of concurrent infections are discussed. This is the first molecular characterisation of CDV in free-ranging African wild dogs and only the third confirmed case of fatal CDV infection in a free-ranging pack.

  8. Detection of West Nile virus and tick-borne encephalitis virus in birds in Slovakia, using a universal primer set.

    PubMed

    Csank, Tomáš; Bhide, Katarína; Bencúrová, Elena; Dolinská, Saskia; Drzewnioková, Petra; Major, Peter; Korytár, Ľuboš; Bocková, Eva; Bhide, Mangesh; Pistl, Juraj

    2016-06-01

    West Nile virus (WNV) is a mosquito-borne neurotropic pathogen that presents a major public health concern. Information on WNV prevalence and circulation in Slovakia is insufficient. Oral and cloacal swabs and bird brain samples were tested for flavivirus RNA by RT-PCR using newly designed generic primers. The species designation was confirmed by sequencing. WNV was detected in swab and brain samples, whereas one brain sample was positive for tick-borne encephalitis virus (TBEV). The WNV sequences clustered with lineages 1 and 2. These results confirm the circulation of WNV in birds in Slovakia and emphasize the risk of infection of humans and horses. PMID:27001305

  9. Detection of 12 respiratory viruses by duplex real time PCR assays in respiratory samples.

    PubMed

    Arvia, Rosaria; Corcioli, Fabiana; Ciccone, Nunziata; Della Malva, Nunzia; Azzi, Alberta

    2015-12-01

    Different viruses can be responsible for similar clinical manifestations of respiratory infections. Thus, the etiological diagnosis of respiratory viral diseases requires the detection of a large number of viruses. In this study, 6 duplex real-time PCR assays, using EvaGreen intercalating dye, were developed to detect 12 major viruses responsible for respiratory diseases: influenza A and B viruses, enteroviruses (including enterovirus spp, and rhinovirus spp), respiratory syncytial virus, human metapneumovirus, coronaviruses group I (of which CoV 229E and CoV NL63 are part) and II (including CoV OC43 and CoV HKU1), parainfluenza viruses type 1, 2, 3 and 4, human adenoviruses and human bocaviruses. The 2 target viruses of each duplex reaction were distinguishable by the melting temperatures of their amplicons. The 6 duplex real time PCR assays were applied for diagnostic purpose on 202 respiratory samples from 157 patients. One hundred fifty-seven samples were throat swabs and 45 were bronchoalveolar lavages. The results of the duplex PCR assays were confirmed by comparison with a commercial, validated, assay; in addition, the positive results were confirmed by sequencing. The analytical sensitivity of the duplex PCR assays varied from 10(3) copies/ml to 10(4) copies/ml. For parainfluenza virus 2 only it was 10(5) copies/ml. Seventy clinical samples (35%) from 55 patients (30 children and 25 adults) were positive for 1 or more viruses. In adult patients, influenza A virus was the most frequently detected respiratory virus followed by rhinoviruses. In contrast, respiratory syncytial virus was the most common virus in children, followed by enteroviruses, influenza A virus and coronavirus NL63. The small number of samples/patients does not allow us to draw any epidemiological conclusion. Altogether, the results of this study indicate that the 6 duplex PCR assays described in this study are sensitive, specific and cost-effective. Thus, this assay could be

  10. A bead-based multiplex assay for the detection of DNA viruses infecting laboratory rodents.

    PubMed

    Höfler, Daniela; Nicklas, Werner; Mauter, Petra; Pawlita, Michael; Schmitt, Markus

    2014-01-01

    The Federation of European Laboratory Animal Science Association (FELASA) recommends screening of laboratory rodents and biological materials for a broad variety of bacterial agents, viruses, and parasites. Methods commonly used to date for pathogen detection are neither cost-effective nor time- and animal-efficient or uniform. However, an infection even if silent alters experimental results through changing the animals' physiology and increases inter-individual variability. As a consequence higher numbers of animals and experiments are needed for valid and significant results. We developed a novel high-throughput multiplex assay, called rodent DNA virus finder (rDVF) for the simultaneous identification of 24 DNA viruses infecting mice and rats. We detected all 24 DNA viruses with high specificity and reproducibility. Detection limits for the different DNA viruses varied between 10 and 1000 copies per PCR. The validation of rDVF was done with DNA isolated from homogenised organs amplified by pathogen specific primers in one multiplex PCR. The biotinylated amplicons were detected via hybridisation to specific oligonucleotide probes coupled to spectrally distinct sets of fluorescent Luminex beads. In conclusion, rDVF may have the potential to replace conventional testing and may simplify and improve routine detection of DNA viruses infecting rodents.

  11. Development of an indirect ELISA and immunocapture rt-PCR for Lily virus detection.

    PubMed

    Kim, Jin Ha; Yoo, Ha Na; Bae, Eun Hye; Jung, Yong-Tae

    2012-12-01

    Multiple viruses such as Lily symptomless virus (LSV), Lily mottle virus (LMoV), and cucumber mosaic virus (CMV) are the most prevalent viruses infecting lilies in Korea. Leaf samples and bulbs showing characteristic symptoms of virus infection were collected from Gangwon, Chungnam, and Jeju provinces of Korea in 2008-2011. Coat protein (CP) genes of LSV and LMoV were amplified from collected samples by reverse transcription-polymerase chain reaction (RT-PCR) and cloned into a pET21d(+) expression vector to generate recombinant CPs. The resulting carboxy-terminal His-tagged CPs were expressed in Escherichia coli strain BL21(DE3) by isopropyl-1-thio- beta-D-galactoside induction. The recombinant proteins were purified using Ni-NTA agarose beads, and the purified proteins were used as an immunogen to produce polyclonal antibodies in rabbits. The resulting polyclonal antisera recognized specifically LSV and LMoV from infected plant tissues in Western blotting assays. Indirect enzyme-linked immunosorbent assay and immunocapture RTPCR using these polyclonal antisera were developed for the sensitive, efficient, economic, and rapid detection of Lily viruses. These results suggest that large-scale bulb tests and economic detection of Lily viruses in epidemiological studies can be performed routinely using these polyclonal antisera.

  12. Relationship between airborne detection of influenza A virus and the number of infected pigs.

    PubMed

    Corzo, Cesar A; Romagosa, Anna; Dee, Scott A; Gramer, Marie R; Morrison, Robert B; Torremorell, Montserrat

    2013-05-01

    Influenza A virus infects a wide range of species including both birds and mammals (including humans). One of the key routes by which the virus can infect populations of animals is by aerosol transmission. This study explored the relationship between number of infected pigs and the probability of detecting influenza virus RNA in bioaerosols through the course of an acute infection. Bioaerosols were collected using a cyclonic collector in two groups of 7 week-old pigs that were experimentally infected by exposure with a contact infected pig (seeder pig). After contact exposure, individual pig nasal swab samples were collected daily and air samples were collected three times per day for 8 days. All samples were tested for influenza by real-time reverse transcriptase (RRT)-PCR targeting the influenza virus matrix gene. All pigs' nasal swabs became influenza virus RRT-PCR positive upon exposure to the infected seeder pig. Airborne influenza was detected in 28/43 (65%) air samples. The temporal dynamics of influenza virus detection in air samples was in close agreement with the nasal shedding pattern in the infected pigs. First detection of positive bioaerosols happened at 1 day post contact (DPC). Positive bioaerosols were consistently detected between 3 and 6 DPC, a time when most pigs were also shedding virus in nasal secretions. Overall, the odds of detecting a positive air sample increased 2.2 times for every additional nasal swab positive pig in the group. In summary, there was a strong relationship between the number of pigs shedding influenza virus in nasal secretions and the generation of bioaerosols during the course of an acute infection.

  13. Relationship between airborne detection of influenza A virus and the number of infected pigs

    PubMed Central

    Corzo, Cesar A.; Romagosa, Anna; Dee, Scott; Gramer, Marie; Morrison, Robert B; Torremorell, Montserrat

    2012-01-01

    Influenza A virus infects a wide range of species including both birds and mammals (including humans). One of the key routes by which the virus can infect populations of animals is by aerosol transmission. This study explored the relationship between number of infected pigs and the probability of detecting influenza virus RNA in bioaerosols through the course of an acute infection. Bioaerosols were collected using a cyclonic collector in two groups of 7 week-old pigs that were experimentally infected by exposure with a contact infected pig (seeder pig). After contact exposure, individual pig nasal swab samples were collected daily and air samples were collected three times per day for 8 days. All samples were tested for influenza by real-time reverse transcriptase (RRT)-PCR targeting the influenza virus matrix gene. All pigs' nasal swabs became influenza virus RRT-PCR positive upon exposure to the infected seeder pig. Airborne influenza was detected in 28/43 (65%) air samples. The temporal dynamics of influenza virus detection in air samples was in close agreement with the nasal shedding pattern in the infected pigs. First detection of positive bioaerosols happened at 1 day post contact (DPC). Positive bioaerosols were consistently detected between 3 and 6 DPC, a time when most pigs were also shedding virus in nasal secretions. Overall, the odds of detecting a positive air sample increased 2.2 times for every additional nasal swab positive pig in the group. In summary, there was a strong relationship between the number of pigs shedding influenza virus in nasal secretions and the generation of bioaerosols during the course of an acute infection. PMID:23164957

  14. Relationship between airborne detection of influenza A virus and the number of infected pigs.

    PubMed

    Corzo, Cesar A; Romagosa, Anna; Dee, Scott A; Gramer, Marie R; Morrison, Robert B; Torremorell, Montserrat

    2013-05-01

    Influenza A virus infects a wide range of species including both birds and mammals (including humans). One of the key routes by which the virus can infect populations of animals is by aerosol transmission. This study explored the relationship between number of infected pigs and the probability of detecting influenza virus RNA in bioaerosols through the course of an acute infection. Bioaerosols were collected using a cyclonic collector in two groups of 7 week-old pigs that were experimentally infected by exposure with a contact infected pig (seeder pig). After contact exposure, individual pig nasal swab samples were collected daily and air samples were collected three times per day for 8 days. All samples were tested for influenza by real-time reverse transcriptase (RRT)-PCR targeting the influenza virus matrix gene. All pigs' nasal swabs became influenza virus RRT-PCR positive upon exposure to the infected seeder pig. Airborne influenza was detected in 28/43 (65%) air samples. The temporal dynamics of influenza virus detection in air samples was in close agreement with the nasal shedding pattern in the infected pigs. First detection of positive bioaerosols happened at 1 day post contact (DPC). Positive bioaerosols were consistently detected between 3 and 6 DPC, a time when most pigs were also shedding virus in nasal secretions. Overall, the odds of detecting a positive air sample increased 2.2 times for every additional nasal swab positive pig in the group. In summary, there was a strong relationship between the number of pigs shedding influenza virus in nasal secretions and the generation of bioaerosols during the course of an acute infection. PMID:23164957

  15. Development of a candidate reference material for adventitious virus detection in vaccine and biologicals manufacturing by deep sequencing

    PubMed Central

    Mee, Edward T.; Preston, Mark D.; Minor, Philip D.; Schepelmann, Silke; Huang, Xuening; Nguyen, Jenny; Wall, David; Hargrove, Stacey; Fu, Thomas; Xu, George; Li, Li; Cote, Colette; Delwart, Eric; Li, Linlin; Hewlett, Indira; Simonyan, Vahan; Ragupathy, Viswanath; Alin, Voskanian-Kordi; Mermod, Nicolas; Hill, Christiane; Ottenwälder, Birgit; Richter, Daniel C.; Tehrani, Arman; Jacqueline, Weber-Lehmann; Cassart, Jean-Pol; Letellier, Carine; Vandeputte, Olivier; Ruelle, Jean-Louis; Deyati, Avisek; La Neve, Fabio; Modena, Chiara; Mee, Edward; Schepelmann, Silke; Preston, Mark; Minor, Philip; Eloit, Marc; Muth, Erika; Lamamy, Arnaud; Jagorel, Florence; Cheval, Justine; Anscombe, Catherine; Misra, Raju; Wooldridge, David; Gharbia, Saheer; Rose, Graham; Ng, Siemon H.S.; Charlebois, Robert L.; Gisonni-Lex, Lucy; Mallet, Laurent; Dorange, Fabien; Chiu, Charles; Naccache, Samia; Kellam, Paul; van der Hoek, Lia; Cotten, Matt; Mitchell, Christine; Baier, Brian S.; Sun, Wenping; Malicki, Heather D.

    2016-01-01

    Background Unbiased deep sequencing offers the potential for improved adventitious virus screening in vaccines and biotherapeutics. Successful implementation of such assays will require appropriate control materials to confirm assay performance and sensitivity. Methods A common reference material containing 25 target viruses was produced and 16 laboratories were invited to process it using their preferred adventitious virus detection assay. Results Fifteen laboratories returned results, obtained using a wide range of wet-lab and informatics methods. Six of 25 target viruses were detected by all laboratories, with the remaining viruses detected by 4–14 laboratories. Six non-target viruses were detected by three or more laboratories. Conclusion The study demonstrated that a wide range of methods are currently used for adventitious virus detection screening in biological products by deep sequencing and that they can yield significantly different results. This underscores the need for common reference materials to ensure satisfactory assay performance and enable comparisons between laboratories. PMID:26709640

  16. Calorimetric detection of influenza virus induced membrane fusion.

    PubMed

    Nebel, S; Bartoldus, I; Stegmann, T

    1995-05-01

    Membrane fusion induced by the hemagglutinin glycoprotein of influenza virus has been extensively characterized, but the mechanism whereby the protein achieves the merger of the viral and target membrane lipids remains enigmatic. Various lipid intermediate structures have been proposed, and the energies required for their formation predicted. Here, we have analyzed the enthalpies of fusion of influenza with liposomes by titration calorimetry. If a small sample of virus in a weak neutral pH buffer was added to an excess of liposomes at low pH, a two-component reaction was seen, composed of an exothermic reaction and a slower endothermic reaction. The exothermic reaction was the result of acid-base reactions between the neutral pH virus sample and low pH buffer and low-pH-induced changes in the virus. The endothermic reaction was not observed in the absence of liposomes and much reduced if acid-inactivated virus, which had lost its fusion but not its binding activity, was added to liposomes. The endothermic reaction was more temperature dependent than the exothermic reaction; its pH dependence corresponded with that of fusion and its enthalpy was higher if fusion was more extensive. These data indicate that most of the endothermic reaction was due to membrane fusion. The experimentally determined enthalpy of fusion, 0.6-0.7 kcal per mol of viral phospholipids, is much higher than expected on the basis of current theories about the formation of lipid intermediates during membrane fusion.

  17. Respiratory Virus Detection and Clinical Diagnosis in Children Attending Day Care

    PubMed Central

    Moe, Nina; Pedersen, Bård; Nordbø, Svein Arne; Skanke, Lars Høsøien; Krokstad, Sidsel; Smyrnaios, Anastasios; Døllner, Henrik

    2016-01-01

    Background Respiratory viruses often have been studied in children with respiratory tract infection (RTI), but less knowledge exists about viruses in asymptomatic children. We have studied the occurrence of a broad panel of respiratory viruses in apparently healthy children attending day care, taking into account the influence of possible confounding factors, such as age, clinical signs of respiratory tract infection (RTI), location (day-care section) and season. Methods We have studied 161 children in two day-care centers, each with separate sections for younger and older children, during four autumn and winter visits over a two-year period. A total of 355 clinical examinations were performed, and 343 nasopharyngeal samples (NPS) were analyzed by semi-quantitative, real-time, polymerase chain reaction (PCR) tests for 19 respiratory pathogens. Result Forty-three percent of all NPS were PCR-positive for ≥ 1 of 13 virus species, with high species variation during visits. Rhinovirus 26% (88/343 NPS), enterovirus 12% (40/343) and parechovirus 9% (30/343) were detected in every visit, and the rates varied in relation to age, day-care section and season. Ten other viruses were detected in ≤ 3% of the NPS. Generally, viruses occurred together in the NPS. In 24% (79/331) of the clinical examinations with available NPS, the children had clear signs of RTI, while in 41% (135/331) they had mild signs, and in 35% (117/331) the children had no signs of RTI. Moreover, viruses were found in 70% (55/79) of children with clear signs of RTI, in 41% (55/135) with mild signs and in 30% (35/117) without any signs of RTI (p < 0.001). Conclusions Positive PCR tests for respiratory viruses, particularly picornaviruses, were frequently detected in apparently healthy children attending day care. Virus detection rates were related to age, presence of clinical signs of RTI, location in day care and season. PMID:27433803

  18. Application of immunoassay of encephalomyocarditis virus in cell culture with enzyme-labeled virus-specific monoclonal antibodies for rapid detection of virus, neutralizing antibodies, and interferon.

    PubMed

    Vlaspolder, F; Harmsen, T; van Veenendaal, D; Kraaijeveld, C A; Snippe, H

    1988-12-01

    Encephalomyocarditis virus (EMCV)-specific monoclonal antibody UM 21.1 labeled with horseradish peroxidase was used to detect EMCV in L-cell monolayers. This direct enzyme immunoassay of EMCV, performed in wells of 96-well plates, could be applied for various purposes, such as early detection of virus multiplication, determination of 50% tissue culture infective doses, and rapid titration of interferon and EMCV-neutralizing antibodies. Multiplication of EMCV is indicated by a rapid increase of the absorbance values measured against EMCV-infected L cells starting as early as 4.5 h after virus inoculation. The early rise of absorbance (i.e., virus multiplication) is inhibited by interferon, allowing its rapid titration. Preincubation of the virus inoculum with neutralizing antibodies also yielded decreased absorbance values. With the latter enzyme immunoassay for neutralizing antibodies, performed after an infection period of 8 h, antibody titers measured were comparable to those obtained with a conventional plaque reduction test. We assume that similar assays could be developed for other picornaviruses (e.g., polioviruses).

  19. Optimization of the virus concentration method using polyethyleneimine-conjugated magnetic beads and its application to the detection of human hepatitis A, B and C viruses.

    PubMed

    Uchida, Eriko; Kogi, Mieko; Oshizawa, Tadashi; Furuta, Birei; Satoh, Koei; Iwata, Akiko; Murata, Mitsuhiro; Hikata, Mikio; Yamaguchi, Teruhide

    2007-07-01

    To enhance the sensitivity of virus detection by polymerase chain reaction (PCR) and reverse transcription PCR (RT-PCR), a novel virus concentration method using polyethyleneimine (PEI)-conjugated magnetic beads was developed in our previous study. However, several viruses could not be concentrated by this method. In this paper, the conditions of virus concentration were optimized to concentrate a wide range of viruses more efficiently. The PEI beads adsorbed viruses more efficiently than other cationic polymers, and the optimum virus concentration was obtained under weak acidic conditions. Mass spectrometric analysis revealed that several serum proteins, such as complement type 3, complement type 4 and immunoglobulin M (IgM), were co-adsorbed by the PEI beads, suggesting that the beads may adsorb viruses not only by direct adsorption, but also via immune complex formation. This hypothesis was confirmed by the result that poliovirus, which PEI beads could not adsorb directly, could be concentrated by the beads via immune complex formation. On the other hand, hepatitis A (HAV) and hepatitis C (HCV) viruses were adsorbed directly by PEI beads almost completely. Like poliovirus, hepatitis B virus (HBV) was concentrated efficiently by the addition of anti-HBV IgM. In conclusion, virus concentration using PEI beads is a useful method to concentrate a wide range of viruses and can be used to enhance the sensitivity of detection of HAV, HBV and HCV.

  20. Development of a multiplexed PCR detection method for Barley and Cereal yellow dwarf viruses, Wheat spindle streak virus, Wheat streak mosaic virus and Soil-borne wheat mosaic virus.

    PubMed

    Deb, Mahua; Anderson, Joseph M

    2008-03-01

    Barley and Cereal yellow dwarf viruses (B/CYDVs), Wheat spindle streak mosaic (WSSMV), Soil-borne wheat mosaic virus (SBWMV) and Wheat streak mosaic virus (WSMV) constitute the most economically important group of wheat viruses. In this paper, a multiplex reverse transcription polymerase chain reaction (M-RT-PCR) method was developed for the simultaneous detection and discrimination of eight viruses: five strains of B/CYDVs, WSSMV, SBWMV and WSMV. The protocol uses specific primer sets for each virus producing five distinct fragments 295, 175, 400, 237, and 365 bp, indicating the presence of two strains of BYDVs, -PAV, -MAV, CYDV-RPV and two unassigned Luteoviridae BYDV-SGV and -RMV, respectively. This system also readily detected WSSMV, SBWMV and WSMV specific amplicons at 154, 219 and 193 bp, respectively. The amplification specificity of these primers was tested against a range of field samples from different parts of United States. The protocol also utilizes fluorescently tagged primers that can streamline the detection of each virus through capillary electrophoresis. This study fulfills the need for a rapid and specific wheat virus diagnostic tool that also has the potential for investigating the epidemiology of these viral diseases.

  1. Infectious laryngotracheitis virus (ILTV) vaccine intake evaluation by detection of virus amplification in feather pulps of vaccinated chickens.

    PubMed

    Davidson, I; Raibshtein, I; Altori, A; Elkin, N

    2016-03-18

    Infectious laryngotracheitis (ILT) is a respiratory disease of poultry caused by an alphaherpesvirus, ILTV. The live vaccine is applied worldwide by drinking water or by the respiratory route, and by the vent application in Israel. No system of direct evaluation of the efficacy of vaccination exists today, except of antibody elicitation, which is an indirect indication of vaccination intake and might happen due to environment exposure. We suggest for the first time an assay for evaluating the accuracy of the vaccination process by spotting the spread of the live vaccine systemically, namely by virus detection in the feather shafts of the vaccinated birds. The feathers are particularly beneficial as they are easy to collect, non-lethal for the bird, therefore advantageous for monitoring purposes. Moreover, the continuous survey of the vaccine virus unveiled the different kinetics of viremia by the different vaccination routes; while after the vent vaccination the systemic viremia peaks during the first week afterwards, after two consecutive vaccine administration by drinking water with 6 day interval, the vireamia peaks only after the second administration. A robust amplification was needed because the vaccine ILTV was present in the bird in minute quantities compared to the wild-type virus. For the vaccine virus identification in feather shafts a nested real-time PCR for the TK ILTV gene was developed. The sensitivity of detection of the nested rtPCR was greater by 1000 compared to conventional nested PCR and 10 times that real-time PCR. PMID:26784685

  2. Infectious laryngotracheitis virus (ILTV) vaccine intake evaluation by detection of virus amplification in feather pulps of vaccinated chickens.

    PubMed

    Davidson, I; Raibshtein, I; Altori, A; Elkin, N

    2016-03-18

    Infectious laryngotracheitis (ILT) is a respiratory disease of poultry caused by an alphaherpesvirus, ILTV. The live vaccine is applied worldwide by drinking water or by the respiratory route, and by the vent application in Israel. No system of direct evaluation of the efficacy of vaccination exists today, except of antibody elicitation, which is an indirect indication of vaccination intake and might happen due to environment exposure. We suggest for the first time an assay for evaluating the accuracy of the vaccination process by spotting the spread of the live vaccine systemically, namely by virus detection in the feather shafts of the vaccinated birds. The feathers are particularly beneficial as they are easy to collect, non-lethal for the bird, therefore advantageous for monitoring purposes. Moreover, the continuous survey of the vaccine virus unveiled the different kinetics of viremia by the different vaccination routes; while after the vent vaccination the systemic viremia peaks during the first week afterwards, after two consecutive vaccine administration by drinking water with 6 day interval, the vireamia peaks only after the second administration. A robust amplification was needed because the vaccine ILTV was present in the bird in minute quantities compared to the wild-type virus. For the vaccine virus identification in feather shafts a nested real-time PCR for the TK ILTV gene was developed. The sensitivity of detection of the nested rtPCR was greater by 1000 compared to conventional nested PCR and 10 times that real-time PCR.

  3. How Does Sampling Methodology Influence Molecular Detection and Isolation Success in Influenza A Virus Field Studies?

    PubMed

    Latorre-Margalef, Neus; Avril, Alexis; Tolf, Conny; Olsen, Björn; Waldenström, Jonas

    2016-02-01

    Wild waterfowl are important reservoir hosts for influenza A virus (IAV) and a potential source of spillover infections in other hosts, including poultry and swine. The emergence of highly pathogenic avian influenza (HPAI) viruses, such as H5N1 and H5N8, and subsequent spread along migratory flyways prompted the initiation of several programs in Europe, North America, and Africa to monitor circulation of HPAI and low-pathogenicity precursor viruses (low-pathogenicity avian influenza [LPAI] viruses). Given the costs of maintaining such programs, it is essential to establish best practice for field methodologies to provide robust data for epidemiological interpretation. Here, we use long-term surveillance data from a single site to evaluate the influence of a number of parameters on virus detection and isolation of LPAI viruses. A total of 26,586 samples (oropharyngeal, fecal, and cloacal) collected from wild mallards were screened by real-time PCR, and positive samples were subjected to isolation in embryonated chicken eggs. The LPAI virus detection rate was influenced by the sample type: cloacal/fecal samples showed a consistently higher detection rate and lower cycle threshold (Ct) value than oropharyngeal samples. Molecular detection was more sensitive than isolation, and virus isolation success was proportional to the number of RNA copies in the sample. Interestingly, for a given Ct value, the isolation success was lower in samples from adult birds than in those from juveniles. Comparing the results of specific real-time reverse transcriptase (RRT)-PCRs and of isolation, it was clear that coinfections were common in the investigated birds. The effects of sample type and detection methods warrant some caution in interpretation of the surveillance data. PMID:26655759

  4. How Does Sampling Methodology Influence Molecular Detection and Isolation Success in Influenza A Virus Field Studies?

    PubMed Central

    Avril, Alexis; Tolf, Conny; Olsen, Björn

    2015-01-01

    Wild waterfowl are important reservoir hosts for influenza A virus (IAV) and a potential source of spillover infections in other hosts, including poultry and swine. The emergence of highly pathogenic avian influenza (HPAI) viruses, such as H5N1 and H5N8, and subsequent spread along migratory flyways prompted the initiation of several programs in Europe, North America, and Africa to monitor circulation of HPAI and low-pathogenicity precursor viruses (low-pathogenicity avian influenza [LPAI] viruses). Given the costs of maintaining such programs, it is essential to establish best practice for field methodologies to provide robust data for epidemiological interpretation. Here, we use long-term surveillance data from a single site to evaluate the influence of a number of parameters on virus detection and isolation of LPAI viruses. A total of 26,586 samples (oropharyngeal, fecal, and cloacal) collected from wild mallards were screened by real-time PCR, and positive samples were subjected to isolation in embryonated chicken eggs. The LPAI virus detection rate was influenced by the sample type: cloacal/fecal samples showed a consistently higher detection rate and lower cycle threshold (Ct) value than oropharyngeal samples. Molecular detection was more sensitive than isolation, and virus isolation success was proportional to the number of RNA copies in the sample. Interestingly, for a given Ct value, the isolation success was lower in samples from adult birds than in those from juveniles. Comparing the results of specific real-time reverse transcriptase (RRT)-PCRs and of isolation, it was clear that coinfections were common in the investigated birds. The effects of sample type and detection methods warrant some caution in interpretation of the surveillance data. PMID:26655759

  5. Comparison of conventional and molecular detection of respiratory viruses inhematopoietic cell transplant recipients

    PubMed Central

    Kuypers, J.; Campbell, A.P.; Cent, A.; Corey, L; Boeckh, M.

    2009-01-01

    Background Sensitive detection of respiratory viruses is important for early diagnosis of infection in patients following hematopoietic cell transplantation (HCT). To evaluate the relative sensitivity of respiratory virus detection in specimens from HCT recipients, we compared the results of conventional and quantitative molecular methods. Methods We tested 688 nasal wash samples collected prospectively from 131 patients during the first 100 days after HCT by viral culture, fluorescent antibody staining (FA), and real-time quantitative reverse transcription-polymerase chain reaction (PCR) assay for detection of respiratory syncytial virus (RSV), influenza virus types A (FluA) and B (FluB), and parainfluenza virus types 1 (PIV1) and 3 (PIV3). Testing for human metapneumovirus (MPV) was performed only by PCR. Data regarding 10 respiratory symptoms were collected with each sample. Results By any method 37 specimens were positive for a respiratory virus; 34 were positive by PC, 15 by culture, and 6 by FA. Four specimens were positive by all 3 methods (3 RSV, 1 FluA). One specimen was positive for PIV1, and 2 were positive for rhinovirus by culture alone. Specimens positive by PCR alone included 2 RSV, 2 PIV1, 8 PIV3, and 8 MPV. In 10 specimens positive for RSV, PIV, or influenza virus collected from patients reporting no respiratory symptoms, 9, 4, and 1 specimen were positive by PCR, culture, and FA respectively. Overall, specimens positive only by PCR had significantly fewer viral copies/mL (mean log10=4.32) than specimens positive by both PCR and culture (mean log10=5.75; p=0.002) or PCR and FA (mean log10=6.83; p<0.001). Conclusions FA testing alone did not detect a significant proportion of respiratory virus positive samples in HCT recipients, especially in patients with no respiratory symptoms and patients with PIV detection. PCR increased the yield of positive specimens 2 times relative to culture and more than 4 times relative to FA. Detection of respiratory

  6. SimulFluor Respiratory Screen for Rapid Detection of Multiple Respiratory Viruses in Clinical Specimens by Immunofluorescence Staining

    PubMed Central

    Landry, Marie L.; Ferguson, David

    2000-01-01

    A new rapid direct immunofluorescence assay (DFA) respiratory screen reagent for detection of seven common respiratory viruses (respiratory syncytial virus [RSV], influenza A and B viruses, parainfluenza virus types 1 to 3, and adenovirus) was compared with standard single or dual DFA reagents and culture. In total, 1,531 respiratory samples were adequate for testing with both SimulFluor Respiratory Screen (RS) reagent (Chemicon International, Temecula, Calif.) and single or dual DFA reagents. The RS DFA reagent detected 367 (98.4%) and single or dual DFA reagents detected 368 (98.7%) of 373 DFA-positive samples. In addition, the RS DFA reagent was equivalent to or better than culture for detection of all viruses except adenovirus. Only 15 of 799 (1.9%) RS-negative samples inoculated into cell cultures yielded respiratory virus isolates (one RSV, five influenza A virus, two influenza B virus, one parainfluenza virus, and six adenovirus). Sixty-six other virus isolates (13 rhinovirus, 24 cytomegalovirus, 28 herpes simplex virus type 1, and 1 enterovirus) were also recovered in culture. With cytospin preparation of slides, only 7.5% of samples submitted were deemed inadequate for DFA. The availability of a rapid DFA screening reagent for detection of multiple common respiratory viruses within 1 to 2 h of sample collection should be of great benefit in terms of patient management and infection control. PMID:10655371

  7. Magnetic beads-based electrochemical immunosensor for detection of pseudorabies virus antibody in swine serum.

    PubMed

    Li, Fang; Zhou, Rui; Zhao, Kaihong; Chen, Huanchun; Hu, Yonggang

    2011-12-15

    A novel magnetic electrochemical immunosensor has been developed for the detection of pseudorabies virus antibody in swine serum. The magnetic glass carbon electrode was fabricated to manipulate magnetic beads for the direct sensing applications. Magnetic beads were employed as the platforms for the immobilization and immunoreaction process, and gold nanoparticles were chosen as electroactive labels for the electrochemical detection. The parameters concerning the assay strategy were carefully investigated. Under the optimal conditions, the linear response range of pseudorabies virus antibody dilution ratio (standard positive serum) was 1:250 to 1:1000 with a detection limit of 1:1000. Finally, this developed immunoassay method was successfully applied in the detection of pseudorabies virus antibody in swine serum, and had a good diagnostic accordance in comparison with ELISA.

  8. Corvidae feather pulp and West Nile virus detection

    USGS Publications Warehouse

    Docherty, D.E.; Romaine Long, R.; Griffin, Katie M.; Saito, E.K.

    2004-01-01

    We evaluated cloacal swab, vascular pulp of flight feather, and kidney and spleen pool samples from carcasses of members of the family Corvidae as sources of West Nile virus (WNV). The cloacal swab, kidney and spleen pool, and feather pulp were the source of WNV in 38%, 43%, and 77%, respectively, of the carcasses.

  9. Molecular Detection of Some Strawberry Viruses in Egypt

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Strawberry plants exhibiting distinct virus-like symptoms (stunting, mottling, yellowing, vein clearing, vein necrosis and vein banding) were collected from strawberry production fields and nurseries in Qalubia Governorate, Egypt (about 20 km north of Cairo). Plants of 'Festival' and 'Sweet Charlie'...

  10. Detection by immunofluorescence of avian myeloblastosis virus reverse transcriptase.

    PubMed

    Karmysheva, V Y; Graevskaya, N A; Sito, A F; Sushko, L N

    1976-08-01

    Peripheral blood cells of avian myeloblastosis virus (AMV-(infected chickens were examined at various intervals post infection by immunofluorescence. AMV revertase was identified in pro- and myeloblasts; it was localized mainly in the perinuclear zone or throughout the cytoplasm. No revertase was found in erythrocytes or granulocytes. Blood cells from uninfected chickens of man contained no revertase.

  11. Detection and survey of coffee ringspot virus in Brazil.

    PubMed

    Ramalho, T O; Figueira, A R; Wang, R; Jones, O; Harris, L E; Goodin, M M

    2016-02-01

    Coffee ringspot virus (CoRSV) a member of the proposed genus "Dichorhavirus", was surveyed on commercial and research farms spanning an area responsible for the majority of Coffea arabica production in Brazil. Virus-infected plants were found at one hundred percent of locations (n = 45) sampled. All cultivars, regardless of cherry color, were found to serve as hosts, suggesting that there is limited resistance in commercially employed germplasm. Reverse transcription PCR analysis revealed that the virus is contained within symptomatic lesions, with little systemic spread throughout leaves. Phylogenetic analysis based on the ORF1 (nucleocapsid) gene identified a strong geo-spatial relationship among isolates, which clustered into three clades. Despite low genetic diversity among isolates, variation in symptom expression was observed in the experimental host Chenopodium quinoa. Our analyses support the hypothesis that the spread of CoRSV is constrained by the clonal expansion of thelytokous populations of Brevipalpus phoenicis. The widespread occurrence of this virus suggests that it is much more prevalent than previously thought. PMID:26553342

  12. Potential virus detection and intervention methods for molluscan shellfish

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Norovirus is the number one cause of foodborne illness in the Unites States, causing an estimated 9 million cases/yr. Hepatitis A is uncommon in the US but can result in serious illness. Bivalve shellfish are efficient bioconcentrators of these viruses from contaminated growing waters. Consequentl...

  13. Use of bacteriophage particles displaying influenza virus hemagglutinin for the detection of hemagglutination-inhibition antibodies.

    PubMed

    Domm, William; Brewer, Matthew; Baker, Steven F; Feng, Changyong; Martínez-Sobrido, Luis; Treanor, John; Dewhurst, Stephen

    2014-03-01

    Bacteriophage lambda capsids provide a flexible molecular scaffold that can be engineered to display a wide range of exogenous proteins, including full-length viral glycoproteins produced in eukaryotic cells. One application for such particles lies in the detection of virus-specific antibodies, since they may obviate the need to work with infectious stocks of highly pathogenic or emerging viruses that can pose significant biosafety and biocontainment challenges. Bacteriophage lambda capsids were produced that displayed an insect-cell derived, recombinant H5 influenza virus hemagglutinin (HA) on their surface. The particles agglutinated red blood cells efficiently, in a manner that could be blocked using H5 HA-specific monoclonal antibodies. The particles were then used to develop a modified hemagglutinination-inhibition (HAI) assay, which successfully identified human sera with H5 HA-specific HAI activity. These results demonstrate the utility of HA-displaying bacteriophage capsids for the detection of influenza virus-specific HAI antibodies.

  14. DNA-Dependent RNA Polymerase Detects Hidden Giant Viruses in Published Databanks

    PubMed Central

    Sharma, Vikas; Colson, Philippe; Giorgi, Roch; Pontarotti, Pierre; Raoult, Didier

    2014-01-01

    Environmental metagenomic studies show that there is a “dark matter,” composed of sequences not linked to any known organism, as determined mainly using ribosomal DNA (rDNA) sequences, which therefore ignore giant viruses. DNA-dependent RNA polymerase (RNAP) genes are universal in microbes and conserved in giant viruses and may replace rDNA for identifying microbes. We found while reconstructing RNAP subunit 2 (RNAP2) phylogeny that a giant virus sequenced together with the genome of a large eukaryote, Hydra magnipapillata, has been overlooked. To explore the dark matter, we used viral RNAP2 and reconstructed putative ancestral RNAP2, which were significantly superior in detecting distant clades than current sequences, and we revealed two additional unknown mimiviruses, misclassified as an euryarchaeote and an oomycete plant pathogen, and detected unknown putative viral clades. We suggest using RNAP systematically to decipher the black matter and identify giant viruses. PMID:24929085

  15. [Real-time PCR Detection Method for the Reston Subtype of the Ebola Virus].

    PubMed

    Xu, Lili; Bao, Linlin; Gu, Songzhi; Qin, Chuan

    2015-05-01

    We aimed to develop a real-time polymerase chain reaction (PCR) detection method for the Reston subtype of the Ebola virus. The NP gene of the Reston subtype of the Ebola virus was selected as the detection object. Sequences of different subtypes of Ebola viruses were aligned using Clustal W software. The most unique and conserved regions of the Reston subtype of the Ebola virus were recruited as candidate sequences for specific primers. Primer Express and Primer Premier 5. 0 software were used to filter the optimal pair of primers for detection. Real-time PCR was carried out using optimized parameters and positive DNA prepared by serial (tenfold) dilution of a recombinant plasmid and by plotting a standard curve. In addition, the reproducibility, accuracy, and specificity of the assay were tested. Results showed that the sensitivity of detection of the Reston subtype of the Ebola virus by real-time PCR could reached 10(2) copies/microL. The linear relationship (R2) reached 0.997, the slope of the standard curve was -0.3101, and amplification efficiency was 110.145%. A sharp and narrow melting peak appeared at 79.94 degrees C for all standards in different dilutions. In conclusion, a fast and sensitive real-time PCR detection system for the Reston subtype of the Ebola virus was developed. This system could be used as a supplementary diagnostic and monitoring approach for basic and clinical studies on the Reston subtype of the Ebola virus. The detection system does not require expensive technology or specialist operators. PMID:26470534

  16. Simultaneous detection of papaya ringspot virus, papaya leaf distortion mosaic virus, and papaya mosaic virus by multiplex real-time reverse transcription PCR.

    PubMed

    Huo, P; Shen, W T; Yan, P; Tuo, D C; Li, X Y; Zhou, P

    2015-12-01

    Both the single infection of papaya ringspot virus (PRSV), papaya leaf distortion mosaic virus (PLDMV) or papaya mosaic virus (PapMV) and double infection of PRSV and PLDMV or PapMV which cause indistinguishable symptoms, threaten the papaya industry in Hainan Island, China. In this study, a multiplex real-time reverse transcription PCR (RT-PCR) was developed to detect simultaneously the three viruses based on their distinctive melting temperatures (Tms): 81.0±0.8°C for PRSV, 84.7±0.6°C for PLDMV, and 88.7±0.4°C for PapMV. The multiplex real-time RT-PCR method was specific and sensitive in detecting the three viruses, with a detection limit of 1.0×10(1), 1.0×10(2), and 1.0×10(2) copies for PRSV, PLDMV, and PapMV, respectively. Indeed, the reaction was 100 times more sensitive than the multiplex RT-PCR for PRSV, and 10 times more sensitive than multiplex RT-PCR for PLDMV. Field application of the multiplex real-time RT-PCR demonstrated that some non-symptomatic samples were positive for PLDMV by multiplex real-time RT-PCR but negative by multiplex RT-PCR, whereas some samples were positive for both PRSV and PLDMV by multiplex real-time RT-PCR assay but only positive for PLDMV by multiplex RT-PCR. Therefore, this multiplex real-time RT-PCR assay provides a more rapid, sensitive and reliable method for simultaneous detection of PRSV, PLDMV, PapMV and their mixed infections in papaya.

  17. Simultaneous detection of papaya ringspot virus, papaya leaf distortion mosaic virus, and papaya mosaic virus by multiplex real-time reverse transcription PCR.

    PubMed

    Huo, P; Shen, W T; Yan, P; Tuo, D C; Li, X Y; Zhou, P

    2015-12-01

    Both the single infection of papaya ringspot virus (PRSV), papaya leaf distortion mosaic virus (PLDMV) or papaya mosaic virus (PapMV) and double infection of PRSV and PLDMV or PapMV which cause indistinguishable symptoms, threaten the papaya industry in Hainan Island, China. In this study, a multiplex real-time reverse transcription PCR (RT-PCR) was developed to detect simultaneously the three viruses based on their distinctive melting temperatures (Tms): 81.0±0.8°C for PRSV, 84.7±0.6°C for PLDMV, and 88.7±0.4°C for PapMV. The multiplex real-time RT-PCR method was specific and sensitive in detecting the three viruses, with a detection limit of 1.0×10(1), 1.0×10(2), and 1.0×10(2) copies for PRSV, PLDMV, and PapMV, respectively. Indeed, the reaction was 100 times more sensitive than the multiplex RT-PCR for PRSV, and 10 times more sensitive than multiplex RT-PCR for PLDMV. Field application of the multiplex real-time RT-PCR demonstrated that some non-symptomatic samples were positive for PLDMV by multiplex real-time RT-PCR but negative by multiplex RT-PCR, whereas some samples were positive for both PRSV and PLDMV by multiplex real-time RT-PCR assay but only positive for PLDMV by multiplex RT-PCR. Therefore, this multiplex real-time RT-PCR assay provides a more rapid, sensitive and reliable method for simultaneous detection of PRSV, PLDMV, PapMV and their mixed infections in papaya. PMID:26666186

  18. Detection of Evolutionarily Distinct Avian Influenza A Viruses in Antarctica

    PubMed Central

    Vijaykrishna, Dhanasekaran; Butler, Jeffrey; Baas, Chantal; Maurer-Stroh, Sebastian; Silva-de-la-Fuente, M. Carolina; Medina-Vogel, Gonzalo; Olsen, Bjorn; Kelso, Anne; Barr, Ian G.; González-Acuña, Daniel

    2014-01-01

    ABSTRACT Distinct lineages of avian influenza viruses (AIVs) are harbored by spatially segregated birds, yet significant surveillance gaps exist around the globe. Virtually nothing is known from the Antarctic. Using virus culture, molecular analysis, full genome sequencing, and serology of samples from Adélie penguins in Antarctica, we confirmed infection by H11N2 subtype AIVs. Their genetic segments were distinct from all known contemporary influenza viruses, including South American AIVs, suggesting spatial separation from other lineages. Only in the matrix and polymerase acidic gene phylogenies did the Antarctic sequences form a sister relationship to South American AIVs, whereas distant phylogenetic relationships were evident in all other gene segments. Interestingly, their neuraminidase genes formed a distant relationship to all avian and human influenza lineages, and the polymerase basic 1 and polymerase acidic formed a sister relationship to the equine H3N8 influenza virus lineage that emerged during 1963 and whose avian origins were previously unknown. We also estimated that each gene segment had diverged for 49 to 80 years from its most closely related sequences, highlighting a significant gap in our AIV knowledge in the region. We also show that the receptor binding properties of the H11N2 viruses are predominantly avian and that they were unable to replicate efficiently in experimentally inoculated ferrets, suggesting their continuous evolution in avian hosts. These findings add substantially to our understanding of both the ecology and the intra- and intercontinental movement of Antarctic AIVs and highlight the potential risk of an incursion of highly pathogenic AIVs into this fragile environment. PMID:24803521

  19. Canine distemper outbreak in raccoons suggests pathogen interspecies transmission amongst alien and native carnivores in urban areas from Germany.

    PubMed

    Rentería-Solís, Zaida; Förster, Christine; Aue, Angelika; Wittstatt, Ulrich; Wibbelt, Gudrun; König, Matthias

    2014-11-01

    From December 2012 to May 2013, an outbreak occurred among urban wild carnivores from Berlin. We collected 97 free-ranging raccoons from the city area. PCR assays, histopathology and immunohistochemistry confirmed canine distemper virus (CDV) infection in 74 raccoons. Phylogenetic analysis of haemagglutinin gene fragments (1767 nucleotides) of CDV isolated from four raccoons showed close relation to CDV isolates from foxes from Germany and a domestic dog from Hungary; all belonging to the "Europe" lineage of CDV. These study results suggest an inter-species transmission of CDV as the origin for the outbreak among the raccoon population. Implications for domestic pets and suggested interspecies transmission between urban wildlife and raccoons are discussed. This is the first major outbreak of CDV amongst free-ranging raccoons in Europe.

  20. Duration of immunity in red wolves (Canis rufus) following vaccination with a modified live parvovirus and canine distemper vaccine.

    PubMed

    Anderson, Kadie; Case, Allison; Woodie, Kathleen; Waddell, William; Reed, Holly H

    2014-09-01

    There is growing information available regarding duration of immunity for core vaccines in both domestic and nondomestic species. Vaccination protocols in nondomestic canids have frequently followed guidelines developed for the domestic dog; however, these protocols can be inappropriate for nondomestic canids such as the African wild dog (Lycaon pictus), leaving some animals susceptible to infectious disease and others at risk for contracting vaccine-induced disease. In this study, red wolves (Canis rufus) were vaccinated against canine distemper virus (CDV) and canine parvovirus (CPV) and vaccination titers were followed annually for 3 yr. One hundred percent of wolves developed and maintained a positive titer to CDV for 3 yr and 96.9% of wolves developed and maintained a positive titer to CPV for 3 yr. Seroconversion for canine adenovirus was sporadic. The results of this study support decreasing the frequency of vaccine administration in the red wolf population to a triennial basis. PMID:25314821