RESUMEN
The present study conducted serosurveillance for the presence of antibody to pandemic influenza A (H1N1) 2009 virus (H1N1pdm virus) in archival serum samples collected between 2009 and 2013 from 317 domestic elephants living in 19 provinces situated in various parts of Thailand. To obtain the most accurate data, hemagglutination-inhibition (HI) assay was employed as the screening test; and sera with HI antibody titers ≥20 were further confirmed by other methods, including cytopathic effect/hemagglutination based-microneutralization (microNT) and Western blot (WB) assays using H1N1pdm matrix 1 (M1) or hemagglutinin (HA) recombinant protein as the test antigen. Conclusively, the appropriate assays using HI in conjunction with WB assays for HA antibody revealed an overall seropositive rate of 8.5% (27 of 317). The prevalence of antibody to H1N1pdm virus was 2% (4/172) in 2009, 32% (17/53) in 2010, 9% (2/22) in 2011, 12% (1/8) in 2012, and 5% (3/62) in 2013. Notably, these positive serum samples were collected from elephants living in 7 tourist provinces of Thailand. The highest seropositive rate was obtained from elephants in Phuket, a popular tourist beach city. Young elephants had higher seropositive rate than older elephants. The source of H1N1pdm viral infection in these elephants was not explored, but most likely came from close contact with the infected mahouts or from the infected tourists who engaged in activities such as elephant riding and feeding. Nevertheless, it could not be excluded that elephant-to-elephant transmission did occur.
Asunto(s)
Animales Domésticos , Elefantes , Subtipo H1N1 del Virus de la Influenza A/aislamiento & purificación , Infecciones por Orthomyxoviridae/epidemiología , Infecciones por Orthomyxoviridae/veterinaria , Animales , Anticuerpos Antivirales/sangre , Subtipo H1N1 del Virus de la Influenza A/inmunología , Infecciones por Orthomyxoviridae/virología , TailandiaRESUMEN
Chlamydiosis, caused by Chlamydiaceae, is a zoonotic disease found in humans and several species of animals, including reptiles and amphibians. Although chlamydiosis in saltwater crocodiles has been previously reported in South Africa and Papua New Guinea, the reported strains have not been identified or confirmed. Therefore, the main aim of this study was to sequence and characterize Chamydiaceae isolated from Siamese crocodiles. Results showed the 16S ribosomal (r) RNA and the 16S/23S rRNA gene of the crocodile isolates were closely related to the genus Chlamydophila with matched identity greater than 98%. The phylogenetic tree constructed from the 16S/23S rRNA gene showed the crocodile cluster diverges far from Cp. caviae with a 100% bootstrap value. The tree based on the ompA gene loci distinguished the crocodile strains into genotypes I, II, and III. The present study is the first report on Chlamydophila detected in Siamese crocodiles that is genetically distinct from the known species of Chlamydiaceae.
Asunto(s)
Caimanes y Cocodrilos , Infecciones por Chlamydophila/veterinaria , Chlamydophila/genética , Chlamydophila/aislamiento & purificación , Proteínas de Reptiles/genética , Animales , Infecciones por Chlamydophila/epidemiología , Infecciones por Chlamydophila/microbiología , Datos de Secuencia Molecular , Filogenia , Reacción en Cadena de la Polimerasa/veterinaria , ARN Ribosómico 16S/genética , ARN Ribosómico 23S/genética , Análisis de Secuencia de Proteína/veterinaria , Tailandia/epidemiologíaRESUMEN
This study describes a proof-of-concept study on the use of small interfering RNA (siRNA)-immunoliposomes as a therapeutic agent against H5N1 influenza virus infection. siRNA specific for influenza virus nucleoprotein (NP) mRNA was employed as the key antiviral agent to inhibit viral replication in this study. A humanized single-chain Fv antibody (huscFv) against the hemagglutinin (HA) of H5N1 highly pathogenic avian influenza virus (HPAI) was used as the targeting molecule to HA of H5N1 virus, which is abundantly expressed on the surface of infected cells (the HA target cells). The huscFv was applied to cationic polyethylene glycol-conjugated 3ß-[N-(N',N'-dimethylaminoethane) carbamoyl] cholesterol-dioleoylphosphatidyl ethanolamine (PEGylated DC-Chol-DOPE) liposomes to generate immunoliposomes for siRNA delivery. The immunoliposomes were shown to specifically bind HA-expressing Sf9 cells and demonstrated enhanced siRNA transfection efficiency. The siRNA transfection efficiency was significantly reduced after preincubation of the HA target cells with an excess amount of free huscFv. These results therefore demonstrated that the enhanced siRNA delivery by use of immunoliposomes was mediated via targeting by huscFv. Furthermore, the siRNA silencing effect was more pronounced when the immunoliposomes were administered 6 to 12 h post-H5N1 infection in MDCK cells compared with the nontargeted liposomes. This proof-of-concept study may contribute to the future design and development of an siRNA delivery system for combating viral infectious diseases in humans.
Asunto(s)
Subtipo H5N1 del Virus de la Influenza A/patogenicidad , Liposomas/química , ARN Interferente Pequeño/química , ARN Interferente Pequeño/genética , Anticuerpos de Cadena Única/química , Animales , Antivirales/química , Antivirales/farmacología , Línea Celular , Supervivencia Celular , Perros , Citometría de Flujo , Subtipo H5N1 del Virus de la Influenza A/efectos de los fármacos , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
This investigation detailed the clinical disease, gross and histologic lesions in juvenile openbill storks (Anastomus oscitans) intranasally inoculated with an avian influenza virus, A/chicken/Thailand/vsmu-3 (H5N1), which is highly pathogenic for chickens. High morbidity and mortality were observed in openbill storks inoculated with HPAI H5N1 virus. Gross lesions from infected birds were congestion and brain hemorrhage (10/20), pericardial effusions, pericarditis and focal necrosis of the cardiac muscle (2/20), pulmonary edema and pulmonary necrosis, serosanguineous fluid in the bronchis (16/20), liver congestion (6/20), bursitis (5/20), subcutaneous hemorrhages (2/20) and pinpoint proventiculus hemorrhage (2/20). Real time RT-PCR demonstrated the presence of viral RNA in organs associated with the lesions: brain, trachea, lungs, liver, spleen and intestines. Similar to viral genome detection, virus was also isolated from these vital organs. Antibodies to influenza virus detected with a hemagglutination inhibition test, were found only in the openbill storks who died 8 days post-inoculation.
Asunto(s)
Subtipo H5N1 del Virus de la Influenza A , Gripe Aviar/fisiopatología , Animales , Aves , Susceptibilidad a Enfermedades , Pruebas de Hemaglutinación , Humanos , ARN Viral , Reacción en Cadena en Tiempo Real de la PolimerasaRESUMEN
Elephant endotheliotropic herpesvirus 1 (EEHV1) can cause fatal hemorrhagic disease in Asian elephants (Elephas maximus). Several studies have described this virus as a major threat to young Asian elephants. A SYBR Green I-based real-time polymerase chain reaction (PCR) was developed to identify EEHV1 on trunk swabs and necropsied tissues. Two of 29 (6.9%) trunk swab samples from healthy Asian elephants were positive for EEHV1. The viruses were analyzed and classified as EEHV1A based on 231 nucleotides of the terminase gene. Necropsied spleen and heart tissue showed the highest level and second highest levels of DNA virus copy accumulation, respectively. The detection limit of the test was 276 copies/µl of DNA. There was no cross-reaction with other mammalian herpesviruses, such as herpes simplex virus 1 and equine herpesvirus 2. Inter- and intra-assay showed low coefficients of variation values indicating the reproducibility of the test. The results indicated that the test can be practically used for epidemiological study, clinical diagnosis, and management and control of EEHV1.
Asunto(s)
Elefantes/virología , Infecciones por Herpesviridae/veterinaria , Herpesviridae/aislamiento & purificación , Técnicas de Diagnóstico Molecular/métodos , Reacción en Cadena en Tiempo Real de la Polimerasa/métodos , Medicina Veterinaria/métodos , Virología/métodos , Estructuras Animales/virología , Animales , Benzotiazoles , ADN Viral/genética , Diaminas , Infecciones por Herpesviridae/diagnóstico , Datos de Secuencia Molecular , Compuestos Orgánicos/metabolismo , Quinolinas , Sensibilidad y Especificidad , Análisis de Secuencia de ADN , Coloración y Etiquetado/métodosRESUMEN
Beak and feather disease virus (BFDV) is a causative agent of psittacine beak and feather disease. Genome sequences of BFDVs isolated from Thailand have not hitherto been reported. The whole genomes of 17 BFDV isolates, obtained from 12 psittacine genera, were amplified and subjected to direct sequencing revealing a length ranging from 1,990 to 2,015 nucleotides. The predicted open reading frames (ORFs) in the viral genome varied from four to six. Only ORF1, ORF2, and ORF5 were found in all isolates. Deduced amino acid sequences of BFDV ORF2 were used to construct a phylogenetic tree. The phylogram grouped BFDV into ten clusters, which showed either host species relationship or regional restriction. The Thai isolates, were grouped into three clusters, cluster I, II, and V. Cluster I and II showed restricted geographical region to Thailand, and cluster II also showed a close relationship with BFDV isolated from Australia. Cluster V demonstrated neither restricted region nor species specificity of birds. In this cluster, there was an insertion of 16 nucleotides at non coding region of all BFDV isolates. The genetic information obtained from this study can be used to help understand BFDV diversity and evolution in Thailand.
Asunto(s)
Infecciones por Circoviridae/veterinaria , Circovirus/genética , Psittaciformes/virología , Animales , Infecciones por Circoviridae/genética , ADN Viral , Plumas/virología , Genoma Viral , Humanos , Sistemas de Lectura Abierta , Reacción en Cadena de la Polimerasa , Análisis de Secuencia de ADN , TailandiaRESUMEN
The investigation of ectoparasitic fauna on birds, and volant and nonvolant small mammals at Srinakarin Dam, Kanchanaburi Province, Thailand was carried out under a national biodiversity and disease surveillance program for four consecutive months: January, February, May and June 2009. A total of 122 animals, comprised of 15 species of birds, 9 species of volant small mammals and 8 species of non-volant small mammals were examined for ectoparasite infestation. Of these animals, 1 genus of hard ticks (Ixodidae), 2 species of mesostigmatid mites (Laelapidae), 4 genera in three families of astigmatid mites (Proctophyllodidae, Pteronyssidae and Trouessartiidae), 4 species in three families of lice (Philopteridae, Polyplacidae and Trichodectidae) and 2 families of batflies (Nycteribiidae and Streblidae) were collected. This is the first survey conducted to determine ectoparasites infesting birds and small mammals living in the reserved forest of Srinakarin Dam, Thailand. A lower infestation rate of ectoparasites was observed in mammals, ranging from 3.5% to 10.3% than birds, with infestation rates between 7.3% and 34.2%. No major potential health risks to people who lived in this area were found.
Asunto(s)
Aves/parasitología , Infestaciones Ectoparasitarias/veterinaria , Mamíferos/parasitología , Animales , Infestaciones Ectoparasitarias/epidemiología , Tailandia/epidemiologíaRESUMEN
A double-antigen sandwich ELISA was developed for the detection of antibodies to influenza A viruses. A recombinant nucleoprotein (rNP) of influenza A virus was used as a capture antigen and an HRP-conjugate for detecting the antibodies. A total of 125 serum samples from birds of different species including chickens, geese, open-billed storks, Khaki Campbell ducks, lesser whistling ducks, and pigeons with known antibodies were tested by ELISA. The sensitivity and the specificity of ELISA were found to be 98% and 97.3%, respectively. The assay was able to detect the presence of influenza A antibodies as early as the fourth day post-inoculation in ducks infected experimentally with influenza A (H5N1) virus. Excellent agreement (97.6%) was obtained between this sandwich ELISA and the hemagglutination inhibition (HI) tests (kappa=0.95). The double-antigen sandwich ELISA correlated well with a commercial avian influenza (AI) multispecies ELISA and was slightly more sensitive than the AI multispecies ELISA. These findings indicate that the double-antigen sandwich ELISA based on rNP may offer an effective screening method for serodiagnosis of influenza A virus. The double-antigen sandwich ELISA also enables the detection of antibodies to influenza A viruses in different species without the need for species-specific secondary antibodies.
Asunto(s)
Anticuerpos Antivirales/sangre , Antígenos Virales , Enfermedades de las Aves/diagnóstico , Ensayo de Inmunoadsorción Enzimática/métodos , Virus de la Influenza A/inmunología , Gripe Aviar/diagnóstico , Enfermedades de las Aves de Corral/diagnóstico , Proteínas de Unión al ARN , Proteínas del Núcleo Viral , Animales , Antígenos Virales/genética , Aves , Pruebas de Inhibición de Hemaglutinación , Proteínas de la Nucleocápside , Aves de Corral , Proteínas de Unión al ARN/genética , Proteínas Recombinantes/genética , Sensibilidad y Especificidad , Estadística como Asunto , Proteínas del Núcleo Viral/genéticaRESUMEN
A survey of ectoparasites on rodents was carried out bimonthly from April 2008 to March 2009 in 3 districts of Sukhothai Province, northern Thailand. A total of 130 rodents comprising 8 species of hosts were captured and examined for ectoparasites. The hosts examined were Bandicota indica, Bandicota savilei, Rattus losea, Rattus rattus, Rattus exulans, Rattus norvegicus, Menetes berdmorei and Tamiops mcclellandii. Ninety-seven ectoparasites were collected: 1 species of tick (Hemaphysalis bandicota), 2 species of mites (Laelaps nuttali and Laelaps echidninus), and 1 species of flea (Xenopsylla cheopis) were identified. The infestation rates by ticks, mites and fleas on the rodents were 0.77, 5.38 and 6.15%, respectively. Monitoring the rodent population and their ectoparasites is important for future planning of prevention and control of zoonotic diseases in the area.
Asunto(s)
Infestaciones Ectoparasitarias/veterinaria , Enfermedades de los Roedores/parasitología , Animales , Reservorios de Enfermedades , Infestaciones Ectoparasitarias/epidemiología , Infestaciones Ectoparasitarias/parasitología , Ácaros , Enfermedades de los Roedores/epidemiología , Siphonaptera , Tailandia/epidemiología , GarrapatasRESUMEN
A multiplex polymerase chain reaction (PCR) was developed for the detection of feline hemotropic mycoplasmas which simultaneously differentiates infections of Mycoplasma haemofelis (Mhf), Candidatus Mycoplasma haemominutum (CMhm) and Candidatus Mycoplasma turicensis (CMtc) in feline blood and spleen. These organisms are responsible for the cause of various pathogenicity of feline infectious anemia. These infections are difficult to be detected by microscopic examination, the most commonly used method for general laboratory diagnoses. Specific primers were designed by selected consensus 16S rDNA sequences of three distinct species. The multiplex PCR assay developed in this study was sensitive and specific with detection limit 100 copies/microl DNA of Mhf and CMhm and 10 copies/microl DNA of CMtc. No amplicons could be amplified for other blood parasites or bacterial pathogens. This multiplex PCR will allow studies of pathogenicity and the monitoring of clinical treatment.
Asunto(s)
Enfermedades de los Gatos/microbiología , Infecciones por Mycoplasma/veterinaria , Mycoplasma/clasificación , Bazo/microbiología , Animales , Enfermedades de los Gatos/sangre , Gatos , Mycoplasma/genética , Infecciones por Mycoplasma/sangre , Infecciones por Mycoplasma/microbiología , Reacción en Cadena de la Polimerasa , Análisis de Secuencia de ADNRESUMEN
This study was conducted to investigate space and time clusters of highly pathogenic avian influenza A (H5N1) virus infection and to determine risk factors at the subdistrict level in Thailand. Highly pathogenic avian influenza A (H5N1) was diagnosed in 1890 poultry flocks located in 953 subdistricts during 2004-2007. The ecologic risk for H5N1 virus infection was assessed on the basis of a spatial-based case-control study involving 824 case subdistricts and 3296 control subdistricts from 6 study periods. Risk factors investigated in clustered areas of H5N1 included human and animal demographic characteristics, poultry production systems, and wild birds and their habitats. Six variables remained statistically significant in the final model: flock density of backyard chickens (odds ratio [OR], 0.98), flock density of fighting cocks (OR, 1.02), low and high human density (OR, 0.60), presence of quail flocks (OR, 1.21), free-grazing duck flocks (OR, 2.17), and a poultry slaughterhouse (OR, 1.33). We observed a strong association between subdistricts with H5N1 virus-infected poultry flocks and evidence of prior and concomitant H5N1 infection in wild birds in the same subdistrict.
Asunto(s)
Subtipo H5N1 del Virus de la Influenza A , Gripe Humana/epidemiología , Gripe Humana/virología , Animales , Análisis por Conglomerados , Brotes de Enfermedades , Exposición a Riesgos Ambientales , Humanos , Gripe Aviar/epidemiología , Gripe Aviar/virología , Oportunidad Relativa , Aves de Corral , Factores de Riesgo , Tailandia/epidemiologíaRESUMEN
A multiplex polymerase chain reaction (PCR) has been developed for simultaneous detection of canine blood parasites, Ehrlichia canis, Babesia spp and Hepatozoon canis, from blood samples in a single reaction. The multiplex PCR primers were specific to E. canis VirB9, Babesia spp 16S rRNA and H. canis 16S rRNA genes. Specificity of the amplicons was confirmed by DNA sequencing. The assay was evaluated using normal canine and infected blood samples, which were detected by microscopic examination. This multiplex PCR offers scope for simultaneous detection of three important canine blood parasites and should be valuable in monitoring parasite infections in dogs and ticks.
Asunto(s)
Apicomplexa/genética , Babesia/genética , Babesiosis/veterinaria , Enfermedades de los Perros/diagnóstico , Ehrlichia canis/genética , Ehrlichiosis/veterinaria , Reacción en Cadena de la Polimerasa/veterinaria , Animales , Apicomplexa/aislamiento & purificación , Babesia/clasificación , Babesia/aislamiento & purificación , Babesiosis/diagnóstico , Babesiosis/genética , Babesiosis/parasitología , Secuencia de Bases , Cartilla de ADN/genética , ADN Protozoario/sangre , ADN Protozoario/genética , Enfermedades de los Perros/genética , Enfermedades de los Perros/parasitología , Perros , Ehrlichia canis/aislamiento & purificación , Ehrlichiosis/diagnóstico , Ehrlichiosis/genética , Ehrlichiosis/microbiología , Genes Bacterianos , Datos de Secuencia Molecular , Reacción en Cadena de la Polimerasa/métodos , Polimorfismo de Longitud del Fragmento de Restricción , ARN Ribosómico 16S/genética , Sensibilidad y Especificidad , Análisis de Secuencia de ADNRESUMEN
In the generation of flavivirus particles, an internal cleavage of the envelope glycoprotein prM by furin is required for the acquisition of infectivity. Unlike cleavage of the prM of other flaviviruses, cleavage of dengue virus prM is incomplete in many cell lines; the partial cleavage reflects the influence of residues at furin nonconsensus positions of the pr-M junction, as flaviviruses share basic residues at positions P1, P2, and P4, recognized by furin. In this study, viruses harboring the alanine-scanning and other multiple-point mutations of the pr-M junction were generated, employing a dengue virus background that exhibited 60 to 70% prM cleavage and a preponderance of virion-sized extracellular particles. Analysis of prM and its cleavage products in viable mutants revealed a cleavage-suppressive effect at the conserved P3 Glu residue, as well as the cleavage-augmenting effects at the P5 Arg and P6 His residues, indicating an interplay between opposing modulatory influences mediated by these residues on the cleavage of the pr-M junction. Changes in the prM cleavage level were associated with altered proportions of extracellular virions and subviral particles; mutants with reduced cleavage were enriched with subviral particles and prM-containing virions, whereas the mutant with enhanced cleavage was deprived of these particles. Alterations of virus multiplication were detected in mutants with reduced prM cleavage and were correlated with their low specific infectivities. These findings define the functional roles of charged residues located adjacent to the furin consensus sequence in the cleavage of dengue virus prM and provide plausible mechanisms by which the reduction in the pr-M junction cleavability may affect virus replication.