Confirmation and Sequence analysis of N gene of PPRV in South Xinjiang, China / Confirmação e análise de sequência do gene N de PPRV em Xinjiang do Sul, China

In China, Peste des petits ruminants (PPR) was officially first reported in 2007. From 2010 until the outbreak of 2013, PPRV infection was not reported. In November 2013, PPRV re-emerged in Xinjiang and rapidly spread to 22 P/A/M (provinces, autonomous regions and municipalities) of China. In the study, suspected PPRV-infected sheep in a breeding farm of South Xinjiang in 2014 were diagnosed and the characteristics of complete sequence of N protein gene of PPRV was analyzed. The sheep showed PPRV-infected signs, such as fever, orinasal secretions increase, dyspnea and diarrhea, with 60% of morbidity and 21.1% of fatality rate. The macroscopic lesions after autopsy and histopathological changes were observed under light microscope including stomatitis, broncho-interstitial pneumonia, catarrhal hemorrhagic enteritis and intracytoplasmic eosinophilic inclusions in multinucleated giantcell in lung. The formalin-fixed mixed tissues samples were positive by nucleic acid extraction and RT-PCR detection. The nucleotide of N protein gene of China/XJNJ/2014 strain was extremely high homology with the China/XJYL/2013 strain, and the highest with PRADESH_95 strain from India in exotic strains. Phylogenetic analysis based on complete sequence of N protein gene of PPRV showed that the China/XJNJ/2014 strain, other strain of 2013-2014 in this study and Tibetan strains all belonged to lineage Ⅳ, but the PPRV strains of 2013-2014 in this study and Tibetan strains were in different sub-branches.(AU)

Na China, Peste des petits ruminants (PPR) foi relatado oficialmente em 2007. De 2010 até o surto de 2013, não houve relato de infecção por PPRV. Em Novembro de 2013, PPRV ressurgiu em Xinjiang e rapidamente se espalhou para 22 P/A/M (províncias, regiões autônomas e municípios) da China. No estudo, ovelhas com suspeita de infecção por PPRV em uma fazenda de reprodução no sul de Xinjiang form diagnosticadas em 2014 e as características da sequência completa da proteína N do gene do PPRV foi analisada. As ovelhas tinham sinais de infecção pelo PPRV, como febre, aumento de secreções oro-nasais, dispneia e diarreia, com 60% de morbidade e 21.1% de fatalidade. As lesões macroscópicas após mudanças histopatológicas foram observadas sob microscópio, incluindo estomatite, pneumonia bronco-intersticial, enterite hemorrágica catarral e inclusões eosinofílicas intracitoplasmáticas em células gigantes multinucleares no pulmão. As amostras de tecido fixadas em formalina testaram positivo para detecção de RT-PCR por extração de ácido nucleico. Os nucleotídeos da proteína N do gene da linhagem China/XJNJ/2014 apresentou extrema homologia com o China/XJYL/2013, e homologia ainda maior com a variedade PRADESH-95 da Índia. Análise filogenética baseada na sequencia completa da proteína N do gene de PPRV mostrou que as variedades China/XJNJ/2014, outra de 2013-2014 mostrada nesse estudo e as Tibetanas todas pertenciam à linhagem Ⅳ, mas as PPRV de 2013-2014 nesse estudo e as Tibetanas estavam em diferentes agrupamentos.(AU)

Prevalence of peste des petits ruminants among sheep and goats in India

This study measured the clinical prevalence of peste des petits ruminants (PPR) among sheep and goats in India between 2003 and 2009 by analyzing clinical samples from suspected cases of PPR that were submitted to the Rinderpest and Allied Disease Laboratory, Division of Virology, IVRI, Mukteswar for PPR diagnosis. PPR outbreaks were confirmed by detecting PPR virus (PPRV)-specific antigen in the clinical samples. Clinical samples (blood, nasal swabs, spleen, lymph node, kidney, liver, intestine, and pooled tissue materials) were taken from a total of 592 sheep and 912 goats in different states of India and screened for the presence of PPRV antigen using a monoclonal antibody-based sandwich ELISA kit. A total of 20, 38, and 11 laboratory-confirmed PPR outbreaks occurred among sheep, goat, and combined sheep and goat populations, respectively. Our findings provide evidence of widespread PPR endemicity in India. The underlying reasons could be variations in husbandry practices in different geographical regions, agro-climatic conditions, and livestock migration. Furthermore, decrease in the number of PPR outbreaks over time might be due to the effectiveness of current live PPR vaccines and timely vaccination of target species. Vaccination against PPR has been practiced in India since 2002 to control this disease.

Sample type is vital for diagnosing infection with peste des petits ruminants virus by reverse transcription PCR

Peste des petits ruminants (PPR) diagnosis from suspected samples from sheep and goats was carried out. Buffy coat, tissues, and oculo-nasal swabs were analyzed using nucleoprotein (NP3/NP4) and fusion protein (F1/F2) gene primers, respectively. Analysis of the sample types and primer set revealed that buffy coat are the best type of samples for PPR diagnosis and the use of two set of primers will increase the number of positives.

Peste des petits ruminants virus detected in tissues from an Asiatic lion (Panthera leo persica) belongs to Asian lineage IV

In this study, peste des petits ruminants virus (PPRV) was detected in frozen pooled tissue samples from a dead Asiatic lion (Panthera leo persica). The samples were negative for canine distemper virus and positive for PPRV nucleic acids when tested with one-step RT-PCR using the appropriate virus-specific primers. Subsequent amplification, cloning, and sequencing of the partial nucleocapsid, matrix, and fusion genes confirmed the presence of PPRV nucleic acid. Comparative sequence and phylogenetic analyses of the structural genes of the isolated virus confirmed that the virus belonged to Asian lineage IV and was closely related to PPRV circulating in India.

The response of bronchial associated lymphoid tissue to intratracheal administration of peste des petit ruminants virus and its Co-Infection with mannheimia hemolytica in goats / Respuesta del tejido linfoide asociado con bronquios a la administración intratraqueal del virus peste de pequeños rumiantes y su coinfección con Mannheimia hemolytica en cabras

The morphometry of bronchial associated lymphoid tissue (BALT) and its possible exploration as to the control of common viral and bacterial pneumonia and their co-infection in Sub-Saharan goats is scanty in literatures. This investigation utilizes archival tissues from study that involved 35 apparently healthy West Africa Dwarf goats which were divided into groups PPRV and PPRV+MH with 15 goats each while 5 goats served as control. PPRV goats were infected intratracheally with 1ml of pure cultured 106.5 TCID50 PPR Pestes des petit Ruminant virus (PPRV) grown in baby hamster kidney cell lines and PPRV+MH with 1 ml of PPRV and a week later 1ml of MH (Mannheimia hemolytica) A2. The BALT number, type and cellularity were determined using standard methods with the use of an image analyzer. Student t-test was used to test for significant differences. The results showed that the average number of BALT was significantly more in PPRV+MH infection than that of PPRV infection i.e. 6.20+/-3.63 for PPRV+MH and 4.11+/-2.92 for PPRV with more nodular types of BALT (8.01+/-1.83) than the aggregates type (4.3+/-1.50) while the average number of lymphocytes in the nodular, aggregate BALT significantly increased with 82.25+/-15.95, 56.00+/-7.28 in PPRV to 103.33+/-44.10, 66.00+/-7.93 in PPRV+MH respectively. There was an irregular pattern in the surface Area (SA) and Perimeter (P) of BALT following infections. The increase in average number of BALT and lymphocytes in PPRV+MH infection than that of PPRV infection showed that the development of BALT and the number of lymphocytes in BALT could be used to assess the mucosal immunity following intratracheal administration of PPRV and MH in goats. This is however the first study that described the morphometry of bronchial associated lymphoid tissue in experimental PPRV and PPRV+MH infection in Sub-Saharan goats.

Es escaso en la literatuta el conocimiento de la morfometría del tejido linfoide asociado a bronquios (BALT) y su posible exploración para el control de la neumonía viral común, pneumonia bacteriana y su co-infección en el ganado caprino subsahariano. Esta investigación utilizó tejidos archivados pertenecientes a 35 cabras enanas del oeste africano aparentemente sanas, que fueron divididas en grupos PPRV y PPRV+MH con 15 cabras cada uno, mientras que 5 cabras sirvieron como control. Las cabras PPRV fueron infectadas por vía intratraqueal con 1 ml de cultivo puro 106,5 TCID50 PPR virus Peste de Pequeños Rumiantes (PPRV) cultivados en líneas celulares renales de hámsters recien nacidos y PPRV+MH con 1 ml de PPRV y una semana después de 1 ml de MH (Mannheimia haemolytica) A2. El número de BALT, tipo y la celularidad se determinaron utilizando métodos estándar con el uso de un analizador de imágenes. Se utilizó la prueba t de Student para probar las diferencias significativas. Los resultados mostraron que el número promedio de BALT fue significativamente mayor en la infección PPRV+MH que la infección PPRV, es decir, 6,20+/-3,63 para PPRV+MH y 4,11+/-2,92 para PPRV con un BALT principalmente de tipo nodular (8,01+/-1,83) que de tipo agregado (4,3+/-1,50), mientras que el número promedio de linfocitos en el BALT nodular y agregado aumentó significativamente con 82,25+/-15,95 y 56,00+/-7,28 en PPRV y, 103,33+/-44,10 y 66,00+/-7,93 en PPRV+MH respectivamente. No hubo un patrón irregular en el área de superficie y perímetro del BALT después de las infecciones. El aumento en el número promedio de BALT y linfocitos en la infección PPRV+MH comparado con la de infección PPRV mostró que el desarrollo de BALT y el número de linfocitos en BALT podría ser utilizado para evaluar la inmunidad de la mucosa después de la administración intratraqueal de PPRV+MH en cabras. Este es el primer estudio que describe la morfometría del tejido linfoide asociado a bronquios en la...

The Gut and Lung Morphometry in Experimental and Natural Lineage 1 Variant of Peste des Petits Ruminants Virus Infection in Nigerian Goats / Morfometría Intestinal y Pulmonar en la Infección Natural y Experimental del Virus Peste des petits ruminants del linaje 1 en Cabras de Nigeria

The lung and gut morphometry in both natural and experimental Peste de petit ruminant (PPR) virus which are scanty in literature hence the need for this study. The goats that were submitted for necropsy in the Department of Veterinary Pathology University of Ibadan between 2009 and 2010 and the gross pathological diagnosis were PPR were enrolled in this study. The degree of pneumonia as a percentage of the total lung volume was estimated using standard methods. The gut morphometry of goats experimentally infected with PPR virus was also used. Student "T" test was used for the test of significance in evaluating the effect of age, sex and the lung consolidation pattern in natural PPR and analysis of the gut morphometry. Complicated PPR had significant higher pulmonary consolidation when compared with the uncomplicated PPR (p< 0.05). The pulmonary consolidation was significantly higher on the right lung with a mean percentage value of 6.54 than the left lung (p< 0.05). The caudal lobe was more consolidated than the cranial and middle lobes in natural PPR. The pulmonary consolidation was more in goats less than a year, while the buck had a significantly higher pulmonary consolidation than the does (p< 0.05). There was no significant difference in the mean length of the villi and width of the villi of PPR virus infected goats when compared to the control, however a significant difference was observed in the cryptal depth (p< 0.05). There was a significant difference in the mean villi length and cryptal depth of goats with complicated PPR (Mannheimia hemolytica) infected goats (p< 0.05) relative to the control. From this study, it showed that most natural PPR were complicated with bacteria and this complication may have contributed to the fatality associated with PPR especially those caused by lineage 1 viruses. This study also showed that secondary bacterial involvement in course of PPR affect the gut morphometry and that could account for the severity of intestinal lesion commonly observed with field PPR in Nigerian goats.

La morfometría del pulmón y el intestino en la infección del virus Peste des petits ruminants (PRR) de forma natural así como experimental es escaza en la literatura, de ahí la necesidad de este estudio. Fueron incluidas en este estudio las cabras que fueron sometidas a autopsia en el Departamento de Patología Veterinaria de la Universidad de Ibadan entre 2009 y 2010, con diagnóstico patológico macroscópico de PPR. El grado de neumonía como porcentaje del volumen pulmonar total fue estimado mediante los métodos estándar. También fue determinada la morfometría del intestino de las cabras infectadas experimentalmente con el virus PPR. Se utilizó la prueba "T" de Student para determinar la significancia en la evaluación de los efectos de edad, sexo, patrón de consolidación pulmonar en PPR natural y análisis de la morfometría intestinal. La PPR complicada tuvo una consolidación pulmonar altamente significativa en comparación con la PPR no complicada (p <0,05). La consolidación pulmonar fue significativamente mayor en el pulmón derecho, con un valor porcentaje promedio de 6,54 en comparación al pulmón izquierdo (p <0,05). El lóbulo caudal fue más consolidado que los lóbulos craneal y medio en presencia del PPR natural. La consolidación pulmonar fue más frecuente en caprinos menores de un año, mientras que los machos cabríos tuvieron una consolidación pulmonar significativamente más alta (p <0,05). No hubo diferencias significativas en la longitud y ancho promedio de las vellosidades en cabras infectadas con PPR en comparación con el control, pero se observó una diferencia significativa en la profundidad de las criptas (p <0,05). Hubo diferencia significativa en la longitud de las vellosidades y la profundidad media de las criptas en las cabras infectadas con PPR complicada (Mannheimia haemolytica) (p <0,05) en relación al control. A partir de este estudio, se demostró que las infecciones con PPR natural se complicaron con bacterias, y estas complicaciones pueden haber contribuido a la mortalidad asociada el PPR, especialmente las causadas por el virus del linaje 1. Este estudio también mostró que la participación bacteriana secundaria en el curso de la PPR afecta la morfometría intestinal y que podría dar cuenta de la gravedad de la lesión intestinal observada comúnmente en la infección de PPR en cabras de Nigeria.

Sequence analysis of the phosphoprotein gene of peste des petits ruminants virus of Chinese origin / 病毒学报

The nucleotide sequences of P gene from a field strain of peste des petits ruminants virus (PPRV) ("China/Tib/Gej/07-30") was firstly determined. The P gene is 1,655 nucleotides long with two overlapping open reading frames (ORFs). The first ORF is 1530 nucleotides long and would produce P protein of 509 amino acid residues. The second ORF is 534 nucleotides long and would produce C protein of 177 amino acid residues. The first ORF produces a second mRNA transcript of 897 nucleotides long with an extra G nucleotide at position 751. Translation from this mRNA would produce V protein of 298 amino acid residues. The nucleotide and deduced amino acid sequence were compared with the homologous region of other PPRV isolates. At the amino acid level, the "China/Tib/Gej/07-30" shares homology of 86.10%-97.3%, 84.3%-94.9%, and 82.9%-96.3% for P, C, and V proteins respectively. Several sequence motifs in the P genes were identified on the basis of conservation in the PPRVs and the morbilliviruses.

The pattern of distribution of pneumonic consolidation in experimental peste des petits ruminants virus (PPRV) and/or Mannheimia Hemolytica Infection in West African dwarf goats / Patrón de distribución de la consolidación pneumónica en infección experimental con virus peste des petits ruminants (PPRV) y/o mannheimia hemolítica en cabras enanas del Oeste Africano

The study into the pattern of distribution of the lung consolidation associated with common viral and bacterial pneumonia and their co-infection in subsaharan goats is scanty in literatures. Fifty apparently healthy West Africa Dwarf goats (WAD) six months of age were used for the experiment. The animals were divided into groups A, B, and C with 15 goats each while 5 goats served as control. Group A goats infected with 1ml of pure culture (1 X 109 CFU) of Mannheimia haemolytica MH A2, while group B with 1ml of pure cultured 106.5 TCID50 PPR virus grown in Baby hamster kidney cell lines and group C with 1 ml of PPRV and a week later 1ml of MH A2. The degree of consolidation or pneumonia as a percentage of the total lung volume was determined by visual observation, palpation and measurement of the lesion which is estimated as a percentage of each lobe. Student t-test were used to test for significant differences. The right lungs have a higher lung consolidation percentage than the left in all the treatment groups. The accessory lobe was affected in the PPRV group. The MH group has the highest lung consolidation percentage (10.1 percent). The PPRV 1-28dpi has the lowest consolidation percentage (1.06 percent). There is significant difference in the consolidation percentage and mortality between MH, PPR+MH, PPRV 28-45 dpi and PPRV 1-28dpi (P<0.05). This observation further show that the right lung and the anterior lobes were more affected in experimental viral and bacterial respiratory pathogen and their co-infection as the trachea birfucation is first to the right and the distance between the right and the left birfucation was 1.5 +/- 0.35cm. It is the first study that describes and compare the pattern of distribution and morphometry of pneumonia in experimental PPRV, MH and PPRV+MH infections in goats.

El estudio sobre el patrón de distribución de la consolidación pulmonar asociada con neumonía virales y bacterianas comunes y sus co-infección en cabras Subsaharianas, es escasa en la literatura. Cincuenta cabras enanas de África occidental (WAD) aparentemente sanas de seis meses de edad fueron utilizados para el experimento. Los animales se dividieron en grupos A, B y C con 15 cabras cada uno mientras que el 5 cabras sirvió como control. Grupo A cabras infectadas con 1 ml de cultivo puro (1 X 109 UFC) de Mannheimia haemolytica MH A2, mientras que el grupo B con 1 ml de cultivo puro 10 6,5 DICT50 PPR cultivado en líneas celulares de riñón de crías de hámsters y el grupo C con 1 ml de PPRV y un semana después de 1 ml de MH A2. El grado de consolidación o neumonía como porcentaje del volumen pulmonar total se determinó por observación visual, palpación y la medición de la lesión que se estima como un porcentaje de cada lóbulo. El test t de Student se utilizaron para probar las diferencias significativas. El pulmón derecho tiene un porcentaje de consolidación pulmonar superior a izquierdo en todos los grupos de tratamiento. El lóbulo accesorio se vio afectado en el grupo de PPRV. El grupo MH tiene el porcentaje más alto de consolidación pulmonar (10,1 por ciento). El PPRV 1-28dpi tiene el menor porcentaje de consolidación (1,06 por ciento). No hay diferencia significativa en el porcentaje de consolidación y la mortalidad entre MH, MH + PPR, PPRV 28-45 dpi y PPRV 1-28dpi (P <0,05). Esta observación muestra además que el pulmón derecho y los lóbulos anteriores se vieron más afectados en infecciones respiratorias patógenas experimentales con agentes virales y bacterianos y su co-infección como la bifurcación traqueal es primero a la derecha y la distancia entre la derecha y la bifurcación izquierda fue de 1,5 +/- 0,35 cm. Es el primer estudio que describe y compara el patrón de distribución y la morfometría de las neumonías en PPRV experimentales, MH y MH + PPRV...

Sequence analysis of the matrix protein and fusion protein genes of a field peste des petits ruminants virus strain from Tibet, China / 病毒学报

The nucleotide sequences of M and F genes from a field strain of peste des petits ruminants virus (PPRV) ("China/Tib/Gej/07-30") was firstly determined. The M gene was 1 483 nucleotides in length with a single open reading frame (ORF), encoding a protein of 335 amino acids. The F gene was 2411 nucleotides in length, encoding a protein of 546 amino acids. The resulting nucleotide sequence and the deduced amino acid sequences were compared with the homologous regions of other PPRV isolates. The nucleotide sequences of M and F genes of the "China/Tib/Gej/07-30" was 92.4%-97.7% and 85.5%-96.1% identical to other PPRV isolates, respectively, while a homology of 97.0%-98.2% and 94.3%-98.2% could be observed at the amino acids level respectively. Several sequence motifs in the M and F genes had been identified on the basis of conservation in the PPRVs and the morbilliviruses. The 3' untranslated region of M gene was 443 nucleotides in length with 82.4%-93.5% identical to other PPRV isolates. The 5' untranslated region of F gene was 634 nucleotides in length with 76.2%-91.7% identical to other PPRV isolates.

Construction and sequencing of full-length cDNA of peste des petits ruminants virus / 病毒学报

To develop a reverse genetics system of Peste des petits ruminants virus(PPRV), five pairs of oligonucleotide primers were designed on the basis of the full-length genomic sequence of PPRV Nigeria 75/ 1 strain. Using RT-PCR technique, five over-lapping cDNA fragments, designated as JF1, JF2, JF3, JF4 and JF5, respectively, were amplified, followed by cloning into pcDNA3.1(+)vector. An AscI restriction enzyme site and a T7 promoter sequence were introduced immediately upstream of 5'-end, while a PacI restriction enzyme site was engineered downstream of 3'-end. Using pok12 as a plasmid vector, the full-length cDNA clone pok12-PPRV of Nigeria 75/1 was assembled by connecting the five cDNA fragments via the unique restriction endonuclease site of PPRV genome. The resultant nucleotide sequence of the PPRV Nigeria 75/1 strain in the study was compared with other members of genus morbillivirus, and phylogenetic analysis was used to examine the evolutionary relationships. The results showed that PPRV Nigeria 75/ 1 was antigenically closely related to Rinderpest virus and Measles virus. Successful construction of full-length cDNA clone of PPRV Nigeria 75/1 strain lays the basis rescuing PPRV effectively and enables further research of PPRV at molecular level.

Genome sequencing and analysis of a peste des petits ruminants virus isolate, China/Tib/07 / 病毒学报

Peste des petits ruminants virus is a member of Morbillivirus Paramyxoviridae. The complete genome of a Peste des petits ruminants virus (PPRV) isolate, China/Tib/07 was sequenced and molecular characteristics was analyzed. The internal sequences of the virus genome were amplified by RT-PCR with primers designed according to the published data in GenBank, while the sequences of the 3' and 5' ends of the genome were determined by RACE. Amplification products were directly sequenced,assembled and analyzed with DNAStar4.0. Results showed that China/Tib/07 genome consisted of 15 948 nucleotides in length, encoding six structural proteins and two non-structural proteins just like other known PPRV genomes. Phylogenetically, the virus genome shared homology of 91.6%-98.1% with Southwest Asian isolates among PPRV strains and the highest homology of 64.3% with rinderpest virus among morbillivirus members.

Recombinant goat pox virus expressing PPRV H protein / 生物工程学报

The purpose of the study is to construct recombinant goat pox virus (GPV) expressing Peste des petits ruminants virus (PPRV) H protein, and to evaluate the immunization effect. Recombinant GPV containing PPRV H gene (rGPV-PPRV-H) was selected and purified by gpt and eGFP utilizing plaque purification, and the final selected recombinant GPV was proved to be purified by PCR. Immunofluorescence and Western blotting showed that the recombinant virus could express H protein of PPRV while infecting lamb testis cells. Six goats were immunized with 2 x 10(6) PFU rGPV-PPRV-H through intradermal injection, and were immunized for the second time at 28 days with the same dose recombinant virus after first immunization. Serum was collected after immunization, and was analyzed for the neutralization antibodies. 21 days after first immunization, the neutralization antibodies of GPV were 40, 80, > or = 80, > or = 80, 40, > or = 80 in turn, and neutralization antibodies of PPRV were 80, 80, 80, 80, 40, 40, 10 in turn; 14 days after second immunization, the neutralization antibodies of GPV were all > or = 80, and the neutralization antibodies of PPRV were > 80, 80, > 80, 80, 80 and 40 in turn. This study established a foundation for the industrialization of the PPRV recombinant GPV vaccine.

Sequence analysis of the nucleocapsid gene and genome promoter region of peste des petits ruminants virus of Chinese origin / 病毒学报

The N gene and genome promoter nucleotide sequence of a Chinese Peste des petits rumiants virus (PPRV) ("China/Tib/Gej/07-30") was firstly determined. The length of N gene was 1689 nucleotides with a single open reading frame (ORF). The nucleotide and deduced amino acid sequence was compared with the homologous region of other PPRV isolates. The nucleotide sequence of the "China/Tib/Gej/07-30" was 91.7%-97.6% identical to other PPRV isolates, while a homology of 94.9%-98.5% could be observed at the amino acids level. The N gene encoded a protein of 525 amino acids. Several sequence motifs were identified on the basis of conservation in the PPRVs and the morbilliviruses. The genome length of promoter region was 107 nucleotides with 91.8%-98.2% identity to other PPRV isolates. Phylogenetic analysis showed that the "China/Tib/Gej/07-30" belonged to the Asian lineage.