Pathogenicity of an FAdV-4 isolate to chickens and its genomic analysis / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

Fowl adenovirus serotype 4 (FAdV-4) strain SD1511 was isolated from chickens with severe inclusion body hepatitis and hydropericardium syndrome in Shandong Province, China. The isolate was cultured in primary chicken embryo kidney cells. A study of pathogenicity indicated that SD1511 readily infected 7-35-d-old chickens by intramuscular injection and intranasal and oral routes, causing 50%-100% mortality. The 35-d-old chickens suffered more severe infection than 7- and 21-d-old chickens with mortality highest in the intramuscular injection group. The serum from surviving chickens showed potent viral neutralizing capability. The complete genome of SD1511 was sequenced and analyzed. The strain was found to belong to the FAdV-4 cluster with more than 99% identity with the virulent FAdV-4 strains isolated in China in recent years except for some distinct variations, including deletions of open reading frame 27 (ORF27), ORF48, and part of ORF19. Our findings suggest that SD1511 might be used as a prototype strain for the study of pathogenesis and vaccine development.

Assessment of the pathogenicity of cell-culture-adapted Newcastle disease virus strain Komarov

Newcastle disease vaccines hitherto in vogue are produced from embryonated chicken eggs. Egg-adapted mesogenic vaccines possess several drawbacks such as paralysis and mortality in 2-week-old chicks and reduced egg production in the egg-laying flock. Owing to these possible drawbacks, we attempted to reduce the vaccine virulence for safe vaccination by adapting the virus in a chicken embryo fibroblast cell culture (CEFCC) system. Eighteen passages were carried out by CEFCC, and the pathogenicity was assessed on the basis of the mean death time, intracerebral pathogenicity index, and intravenous pathogenicity index, at equal passage intervals. Although the reduction in virulence demonstrated with increasing passage levels in CEFCC was encouraging, 20% of the 2-week-old birds showed paralytic symptoms with the virus vaccine from the 18^th(final) passage. Thus, a tissue-culture-adapted vaccine would demand a few more passages by CEFCC in order to achieve a complete reduction in virulence for use as a safe and effective vaccine, especially among younger chicks. Moreover, it can be safely administered even to unprimed 8-week-old birds.

Isolation and phylogenetic analysis of hemagglutinin gene of H9N2 influenza viruses from chickens in South China from 2012 to 2013

As part of our ongoing influenza surveillance program in South China, 19 field strains of H9N2 subtype avian influenza viruses (AIVs) were isolated from dead or diseased chicken flocks in Guangdong province, South China, between 2012 and 2013. Hemagglutinin (HA) genes of these strains were sequenced and analyzed and phylogenic analysis showed that 12 of the 19 isolates belonged to the lineage h9.4.2.5, while the other seven belonged to h9.4.2.6. Specifically, we found that all of the viruses isolated in 2013 belonged to lineage h9.4.2.5. The lineage h9.4.2.5 viruses contained a PSRSSRdownward arrowGLF motif at HA cleavage site, while the lineage h9.4.2.6 viruses contained a PARSSRdownward arrowGLF at the same position. Most of the isolates in lineage h9.4.2.5 lost one potential glycosylation site at residues 200-202, and had an additional one at residues 295-297 in HA1. Notably, 19 isolates had an amino acid exchange (Q226L) in the receptor binding site, which indicated that the viruses had potential affinity of binding to human like receptor. The present study shows the importance of continuing surveillance of new H9N2 strains to better prepare for the next epidemic or pandemic outbreak of H9N2 AIV infections in chicken flocks.

Detection and molecular characterization of infectious bronchitis virus isolated from recent outbreaks in broiler flocks in Thailand

Thirteen field isolates of infectious bronchitis virus (IBV) were isolated from broiler flocks in Thailand between January and June 2008. The 878-bp of the S1 gene covering a hypervariable region was amplified and sequenced. Phylogenetic analysis based on that region revealed that these viruses were separated into two groups (I and II). IBV isolates in group I were not related to other IBV strains published in the GenBank database. Group 1 nucleotide sequence identities were less than 85% and amino acid sequence identities less than 84% in common with IBVs published in the GenBank database. This group likely represents the strains indigenous to Thailand. The isolates in group II showed a close relationship with Chinese IBVs. They had nucleotide sequence identities of 97-98% and amino acid sequence identities 96-98% in common with Chinese IBVs (strain A2, SH and QXIBV). This finding indicated that the recent Thai IBVs evolved separately and at least two groups of viruses are circulating in Thailand.

Pathogenicity and antigenicity of a new variant of Korean nephropathogenic infectious bronchitis virus

Despite the existence of an active vaccination program, recently emerged strains of nephropathogenic infectious bronchitis virus (IBV) in Korea have caused significant economic losses in the poultry industry. In this study, we assessed the pathogenic and antigenic characteristics of a K-IIb type field strain of IBV that emerged in Korea since 2003, such as Kr/Q43/06. Specific pathogen free 1-week-old chickens exhibited severe respiratory symptoms (dyspnea) and nephropathogenic lesions (swollen kidneys with nephritis and urate deposits) following challenge with the recent IBV field strain. The antigenic relatedness (R value), based on a calculated virus neutralization index, of the K-IIb type field strain and K-IIa type strain KM91 (isolated in 1991) was 30%, which indicated that the recent strain, Kr/Q43/06, is a new variant that is antigenically distinct from strain KM91. This report is the first to document the emergence of a new antigenic variant of nephropathogenic IBV in chicken from Korea.

Torque teno virus (TTV) is prevalent in Brazilian non-human primates and chickens (Gallus gallus domesticus) / Torque teno virus (TTV) es prevalente en primates no humanos brasileños y pollos (Gallus gallus domesticus)

Torque Teno virus (TTV) is an infectious agent of worldwide distribution isolated by the first time as the agent of an acute post-transfusion hepatitis in a patient in Japan. It has been classified into a new floating genus called Anellovirus. Recent studies showed that TTV can also be identified in serum specimens obtained from domesticated farm animals and from non-human primates. To better understand the relationship between TTV and their hosts, a study to detect virus in the serum and whole blood of Brazilian non-human primates and in the plasm of chickens was performed by applying the PCR-UTR-A technique, followed by a genomic sequence and phylogenetic analysis. By nested-PCR-UTR, the DNA of TTV was detected in sera from 4 (5.3 percent) of 75 Cebus apella, 2 (40 percent) of 5 Alouatafusca, 1 (20 percent) of 5 Alouata caraya, 1 (5.2 percent) of 19 Callithrixpenicilata, 1 (4 percent) of 25 Callithrixjacchus, 1 (20 percent) of 5 Saimiri sciureus and 1 (25 percent) of 4 Leontopithecus chrysomelas. Phylogenetic analysis revealed that sequences detected in 8 samples clustered with TTV sequences So-TTV2 (Sagüínus oedipus) and At-TTV3 (Aotes Trivirgatus). Three sequences showed similarity with a human Torque Teno Minivirus (TLMV). TTV ORF2 DNA was detected in one sera sample and one whole blood sample of non-human primates and in one plasm sample of chicken. Phylogenetic analysis revealed that the sequences amplified by the ORF2 region show no difference between human, non-human primates and chicken. This is the first report of TTV in Brazilian new world non-human primates and chicken.

Torque Teno virus (TTV) es una agente infeccioso de distribución mundial, aislado por primera vez como el agente de una hepatitis aguda posterior a la transfusión de un paciente en Japón. Se ha clasificado en un nuevo género flotante llamado Anellovirus. Recientes estudios han demostrado que TTV también puede ser identificado en el suero de especímenes obtenidos desde granjas de animales domésticos y desde primates no humanos. Para entender mejor la relación entre la TTV y sus huéspedes, fue realizado un estudio para detectar el virus en el suero y la sangre de primates no humanos brasileños y en el plasma de pollos mediante la aplicación de la técnica PCR-UTR-A, seguida de una secuencia genómica y análisis filogenético. Por medio de PCR-UTR-anidado, el ADN de TTV fue detectado en sueros de 4 de 75 (5,3 por ciento)Cebus apella, 2 de 5 (40 por ciento) Alouata fusca, 1 de 5 (20 por ciento) de Alouata caraya, 1 de 19 (5,2 por ciento) de Callithrixpenicilata, 1 de 25 (4 por ciento) Callithrixjacchus, 1 de 5 (20 por ciento) de Saimiri sciureus y 1 de 4 (25 por ciento) de Leontopithecus chrysomelas. El análisis filogenético reveló secuencias detectadas en 8 muestras agrupadas con TTV secuencias So-TTV2 (Sagüínus oedipus) y At-TTV3 (Aotes Trivirgatus). Tres secuencias mostraron similitud con el Torque Teno Minivirus humano (TLMV). Fue detectado TTV ORF2 ADN en una muestra de suero y una muestra de sangre de primates no-humanos y en una muestra de plasma de pollo. El análisis filogenético reveló que las secuencias amplificadas por la región ORF2 no muestran ninguna diferencia entre humanos, primates no humanos y pollos. Este es el primer informe de nuevos TTV en primates-no humanos brasileños y en pollos.

Research of torque teno virus (TTV) in serum and total blood of Brazilian non-human primates and in chicken plasma (Gallus gallus domesticus) by the PCR N22 region / Investigación de Torque teno virus (TTV) en suero y sangre de primates no humanos brasileños y en plasma de pollo (Gallus gallus domesticus) por PCR en la region N22

Torque teno virus (TTV) is a recently discovered DNA virus that was originally isolated from a Japanese patient (initials, TT) with post-transfusion hepatitis of unknown aetiology. TTV is an circular DNA virus classified recently together with related Torque teño minivirus, into a new genus called Anellovirus. Infection TTV has been detected in a range of non-human primates as well as domestic animals. The purpose of this study was to search TTV in the serum and total blood of Brazilian monkeys and in plasma of domestic chickens by seminested PCR of coding region (N22), followed by a genomic sequence and phylogenetic analysis. No serum sample was amplified. TTV DNA was detected in total blood from 3 (4 percent) out of 75 brown-capuchin (Cebus apella) and from 1 (25 percent) out of 4 golden-headed lion-tamarin (Leontopithecus chrysomelas). Phylogenetic analysis revealed that one sample showed similarity with one sequence of the cotton top tamarin (Saguinus oedipus) (So-TTV2) and with one of the douroucoulis (ão tes trivirgatus) (At-TTV3). Two samples showed similarity with a human Torque Teño Mini Virus (TLMV). The other sample clustered with one sequence of the chimpanzee (Pt-TTV6) and with the human TTV strain TA278. The plasma chicken samples tested were all negative. The amino acid sequences reported in this study are the first obtained in Brazil from total blood of non-human primates naturally infected by TTV.

Torque teno virus (TTV) es un virus de ADN recientemente descubierto que fue inicialmente aislado de un paciente japonés (iniciales TT) después de la transfusión de hepatitis de etiología desconocida. TTV es un virus de ADN circular recientemente clasificado junto con los torque teno minivirus, en un nuevo género llamado Anellovirus. La infección de TTV se ha detectado en una serie de primates no humanos, así como animales domésticos. El objetivo de este estudio fue buscar TTV en el suero y sangre total de monos de Brasil y en el plasma de pollos domésticos, por seminested PCR de la región de codificación (N22), seguido de una secuencia genómica y el análisis filogenético. Las muestras que no eran suero fueron amplificadas. TTV DNA se detectó en sangre total de 3 (4 por ciento) de un total de 75 capuchinos de cabeza dura (Cebus apella) y de 1 (25 por ciento) de un total de 4 tití- león de cabeza dorada (Leontopithecus chrysomelas). El análisis filogenético demostró que una muestra presentaba similitud con una secuencia de Saguinus Edipo (So-TTV2) y con una de Aotes trivirgatus (A-TTV3). Dos muestras mostraron similitud con un torque teno mini virus (TLMV) humano. La otra muestra agrupada con una secuencia de los chimpancés (PT-TTV6) y con el TTV humanos cepa TA278. El análisis de las muestras de plasma de pollo fueron negativas Las secuencias de aminoácidos que se reportan en este estudio son las primeras obtenidas en Brasil de sangre de primates no humanos infectados naturalmente por TTV.

Sequence analysis of the S1 glycoprotein gene of infectious bronchitis viruses: identification of a novel phylogenetic group in Korea

Twelve Korean infectious bronchitis viruses (IBVs) were isolated in the field from chickens suspected of being carriers of infectious bronchitis between 2001 and 2003. The S1 glycoprotein genes of these IBV isolates were amplified by reverse transcriptase-polymerase chain reaction (RTPCR) and analyzed by restriction fragment length polymorphism (RFLP) analysis. These Korean IBV isolates were classified into three groups according to their RFLP patterns obtained using the restriction enzyme HaeIII. Half of the twelve isolates were similar to the KM91 RFLP pattern, which is a common pattern in Korea. Three more isolates were related to the Arkansas strain pattern, but with some unique variations. The other three viruses showed variant RFLP patterns. For a comparison with the published sequences for non-Korean IBV strains, amplified PCR products from the twelve isolates were cloned and sequenced. The Korean IBV field isolates had 71.2-99.7% nucleotide sequence homology with each other and 45.9-80.7% nucleotide sequence homology with non-Korean IBV strains. With respect to the deduced amino acid sequence, the Korean IBV isolates had 71.5-99.3% similarity with each other and 44.9-80.3% similarity with non-Korean IBV strains. Phylogenetic tree analysis revealed that some of the IBV isolates appear to belong to a new group, different from the non-Korean IBV strains or from previously isolated Korean IBV strains. Specifically, the new Korean IBV isolates K10217-03, K3-3 and K1255-03 represented a separate group. These findings suggest that the Korean IBVs appear to be continuously evolving.

Isolation of Subgroup J Avian Leukosis Virus in Korea

Two subgroup J avian leukosis viurses (ALVs) were isolated from broiler breeder flocks, in which myeloid leukosis had occurred. The isolates could be classified as subgroup J ALV. by the positive reaction in polymerase chain reaction (PCR) with primers specific for subgroup J ALV. Two isolates replicated in chicken embryo fibroblast (CEF) cells from the alv6 chicken line in which cells are resistant to subgroup A and E ALVs. In in vitro serum neutralization tests with other subgroup ALVs including ADOL-Hc1, the prototype of subgroup J ALVs isolated in the United States of America, two isolates were partially neutralized by antibody to ADOL-Hc1, indicating that Korean isolates and ADOL-Hc1 may be antigenically related, but not identical. When the PCR was done with a primer pair designed to amplify genes of E element and long terminal repeat of proviral DNA, the PCR product size of one isolate (KOAL-PET) was smaller than that of ADOL-Hc1, suggesting that some sequences in these regions are deleted.

Caracterización clínico-patológica de tres aislamientos del virus de la infección de la bolsa de Fabricio, obtenidos en granjas comerciales de pollo de engorda en México / Clinical and pathological characterization of three isolates of infectious bursal disease virus obtained from commercial broiler flocks in Mexico

Se aislaron tres virus de la infección de la bolsa de Fabricio (VIBF) a partir de muestras obtenidas de muestras en granjas avícolas en México, aquéllos se denominaron H, P y G. Se determinaron sus propiedades antigénicas mediante la técnica de ELISA por captura de antígenos, utilizando los anticuerpos monoclonales (Mab) B-29, R-63, BK-9, B-69 y 57. Se evaluó su virulencia en aves inoculadas mediante la observación de signos clínicos, mortalidad, proporción del peso bursal con el peso corporal, lesiones microscópicas, persistencia de los virus en tejidos linfoides, serología contra VIBF y medición de la respuesta inmune hacia Brucella abortus y eritrocitos de ovino. Los virus P y G presentaron los epítopes reconocidos por lo B-29, R-63 y B-69, que están presentes en las cepas clásicas de VIBF, el virus H no pudo ser reconocido por ningún Mab. Los tres aislamientos indujeron la presentación subclínica de la IBF en las aves inoculadas, se observó marcada atrofia bursal con deplesión linfoide severa, persistencia del VIBF en bolsa hasta los catorce días posinoculación y seroconversión de las aves a partir de la segunda semana después de la inoculación. A pesar de que no se detectó plenamente un estado de inmunodepresión mediante la respuesta a los dos antígenos utilizados, no se puede descartar un eventual efecto inmunodepresor por el aislamiento "H". Tampoco se detectaron diferencias en la virulencia de los tres VIBF aislados