Identification of isolated viral strains of atypical avian influenza using molecular methods of virological diagnostics
Identifikacija izolovanih sojeva virusa atipične kuge živine primenom molekularnih metoda virusološke dijagnostike
2008
Authors
Vidanović, DejanŠekler, Milanko
Vasković, Nikola
Žarković, Aleksandar
Matović, Kazimir
Milić, Nenad
Nišavić, Jakov
Article (Published version)
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In addition to the implementation of standard methods of virological diagnostics used for the isolation of the Newcastle disease virus from suspect material, as well as for its identification, nowadays there is increasing use of molecular diagnostic methods, primarily reverse transcription polymerase chain reaction (RT-PCR) and the sequencing method. The objective of this work was to examine possibilities for the implementation of the above methods in the diagnosis of poultry infection caused by the Newcastle disease virus. The presence of hem agglutination antigens for the Newcastle disease virus was established in samples of allantoises liquid from 62 poultry embargoed eggs 72 h after inoculation, whose titers ranged from 1:16 to 1:2048, while the hem agglutination inhibition test (HI test) with the implementation of a referent immuno serum against the given cause provided the identification of isolated viruses in serum dilutions of 1:128 to 1:1024. The RT-PCR method and the PCR esta...blished that in eight examined samples one fragment each of viral RNA is formed in agars gel of a size of 254bp, which is characteristic for the Newcastle disease virus genome according to its nucleotide sequence. On the grounds of a comparative analysis of RNA sequences obtained from eight isolated NDV strains and the genome sequences of referent atypical poultry influenza viral strains using Mega 40 and BLAST programmers, it was established that the isolated strains of the Newcastle disease virus were highly virulent.
Pored primene standardnih metoda virusološke dijagnostike koje se koriste za izolovanje virusa Newcastle bolesti iz suspektnog materijala kao i za njegovu identifikaciju, danas su sve više u upotrebi i molekularne metode dijagnostike i to pre svega lančana reakcija polimeraze (RT-PCR) i metoda sekvenciranja. Cilj ovog rada bio je ispitivanje mogućnosti primene navedenih metoda u dijagnostici infekcije živine izazvane virusom Newcastle bolesti. Prisustvo hemaglutinacionih antigena virusa Newcastle bolesti ustanovljeno je u uzorcima alantoisne tečnosti od 62 embrionirana jaja živine posle 72h od inokulacije čiji su se titri kretali od 1:16 do 1:2048, dok je testom heminhibicije (HI testom) uz primenu referentnog imunog seruma protiv navedenog uzročnika izvršena identifikacija izolovanih virusa u razređenjima seruma od 1:128 do 1:1024. Metodom reverzne transkripcije (RT-PCT) i lančane reakcije polimeraze (PCR) ustanovljeno da se kod osam ispitivanih uzoraka formira po jedan fragment virus...ne RNK u gelu agaroze veličine od 254bp koji je po sekvenci nukleotida karakterističan za genom virusa Newcastle bolesti. Na osnovu uporedne analize sekvenci RNK dobijenih od osam izolovanih sojeva virusa NDV i sekvenci genoma referentnih sojeva virusa atipične kuge živine uz korišćenje Mega 40 i BLAST programa, utvrđeno je da su izolovani sojevi virusa Newcastle bolesti visoko virulentni.
Keywords:
Newcastle disease virus / embargoed eggs / hemagglutination test / hemagglutination inhibition test / RT-PCR / sequencing / virus Newcastle bolesti / embrionirana jaja / test hemaglutinacije / test inhibicije hemaglutinacije / RT-PCR / sekvenciranjeSource:
Veterinarski Glasnik, 2008, 62, 3-4, 167-177Publisher:
- Univerzitet u Beogradu - Fakultet veterinarske medicine, Beograd
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Fakultet veterinarske medicineTY - JOUR AU - Vidanović, Dejan AU - Šekler, Milanko AU - Vasković, Nikola AU - Žarković, Aleksandar AU - Matović, Kazimir AU - Milić, Nenad AU - Nišavić, Jakov PY - 2008 UR - https://vet-erinar.vet.bg.ac.rs/handle/123456789/518 AB - In addition to the implementation of standard methods of virological diagnostics used for the isolation of the Newcastle disease virus from suspect material, as well as for its identification, nowadays there is increasing use of molecular diagnostic methods, primarily reverse transcription polymerase chain reaction (RT-PCR) and the sequencing method. The objective of this work was to examine possibilities for the implementation of the above methods in the diagnosis of poultry infection caused by the Newcastle disease virus. The presence of hem agglutination antigens for the Newcastle disease virus was established in samples of allantoises liquid from 62 poultry embargoed eggs 72 h after inoculation, whose titers ranged from 1:16 to 1:2048, while the hem agglutination inhibition test (HI test) with the implementation of a referent immuno serum against the given cause provided the identification of isolated viruses in serum dilutions of 1:128 to 1:1024. The RT-PCR method and the PCR established that in eight examined samples one fragment each of viral RNA is formed in agars gel of a size of 254bp, which is characteristic for the Newcastle disease virus genome according to its nucleotide sequence. On the grounds of a comparative analysis of RNA sequences obtained from eight isolated NDV strains and the genome sequences of referent atypical poultry influenza viral strains using Mega 40 and BLAST programmers, it was established that the isolated strains of the Newcastle disease virus were highly virulent. AB - Pored primene standardnih metoda virusološke dijagnostike koje se koriste za izolovanje virusa Newcastle bolesti iz suspektnog materijala kao i za njegovu identifikaciju, danas su sve više u upotrebi i molekularne metode dijagnostike i to pre svega lančana reakcija polimeraze (RT-PCR) i metoda sekvenciranja. Cilj ovog rada bio je ispitivanje mogućnosti primene navedenih metoda u dijagnostici infekcije živine izazvane virusom Newcastle bolesti. Prisustvo hemaglutinacionih antigena virusa Newcastle bolesti ustanovljeno je u uzorcima alantoisne tečnosti od 62 embrionirana jaja živine posle 72h od inokulacije čiji su se titri kretali od 1:16 do 1:2048, dok je testom heminhibicije (HI testom) uz primenu referentnog imunog seruma protiv navedenog uzročnika izvršena identifikacija izolovanih virusa u razređenjima seruma od 1:128 do 1:1024. Metodom reverzne transkripcije (RT-PCT) i lančane reakcije polimeraze (PCR) ustanovljeno da se kod osam ispitivanih uzoraka formira po jedan fragment virusne RNK u gelu agaroze veličine od 254bp koji je po sekvenci nukleotida karakterističan za genom virusa Newcastle bolesti. Na osnovu uporedne analize sekvenci RNK dobijenih od osam izolovanih sojeva virusa NDV i sekvenci genoma referentnih sojeva virusa atipične kuge živine uz korišćenje Mega 40 i BLAST programa, utvrđeno je da su izolovani sojevi virusa Newcastle bolesti visoko virulentni. PB - Univerzitet u Beogradu - Fakultet veterinarske medicine, Beograd T2 - Veterinarski Glasnik T1 - Identification of isolated viral strains of atypical avian influenza using molecular methods of virological diagnostics T1 - Identifikacija izolovanih sojeva virusa atipične kuge živine primenom molekularnih metoda virusološke dijagnostike VL - 62 IS - 3-4 SP - 167 EP - 177 DO - 10.2298/VETGL0804167V ER -
@article{ author = "Vidanović, Dejan and Šekler, Milanko and Vasković, Nikola and Žarković, Aleksandar and Matović, Kazimir and Milić, Nenad and Nišavić, Jakov", year = "2008", abstract = "In addition to the implementation of standard methods of virological diagnostics used for the isolation of the Newcastle disease virus from suspect material, as well as for its identification, nowadays there is increasing use of molecular diagnostic methods, primarily reverse transcription polymerase chain reaction (RT-PCR) and the sequencing method. The objective of this work was to examine possibilities for the implementation of the above methods in the diagnosis of poultry infection caused by the Newcastle disease virus. The presence of hem agglutination antigens for the Newcastle disease virus was established in samples of allantoises liquid from 62 poultry embargoed eggs 72 h after inoculation, whose titers ranged from 1:16 to 1:2048, while the hem agglutination inhibition test (HI test) with the implementation of a referent immuno serum against the given cause provided the identification of isolated viruses in serum dilutions of 1:128 to 1:1024. The RT-PCR method and the PCR established that in eight examined samples one fragment each of viral RNA is formed in agars gel of a size of 254bp, which is characteristic for the Newcastle disease virus genome according to its nucleotide sequence. On the grounds of a comparative analysis of RNA sequences obtained from eight isolated NDV strains and the genome sequences of referent atypical poultry influenza viral strains using Mega 40 and BLAST programmers, it was established that the isolated strains of the Newcastle disease virus were highly virulent., Pored primene standardnih metoda virusološke dijagnostike koje se koriste za izolovanje virusa Newcastle bolesti iz suspektnog materijala kao i za njegovu identifikaciju, danas su sve više u upotrebi i molekularne metode dijagnostike i to pre svega lančana reakcija polimeraze (RT-PCR) i metoda sekvenciranja. Cilj ovog rada bio je ispitivanje mogućnosti primene navedenih metoda u dijagnostici infekcije živine izazvane virusom Newcastle bolesti. Prisustvo hemaglutinacionih antigena virusa Newcastle bolesti ustanovljeno je u uzorcima alantoisne tečnosti od 62 embrionirana jaja živine posle 72h od inokulacije čiji su se titri kretali od 1:16 do 1:2048, dok je testom heminhibicije (HI testom) uz primenu referentnog imunog seruma protiv navedenog uzročnika izvršena identifikacija izolovanih virusa u razređenjima seruma od 1:128 do 1:1024. Metodom reverzne transkripcije (RT-PCT) i lančane reakcije polimeraze (PCR) ustanovljeno da se kod osam ispitivanih uzoraka formira po jedan fragment virusne RNK u gelu agaroze veličine od 254bp koji je po sekvenci nukleotida karakterističan za genom virusa Newcastle bolesti. Na osnovu uporedne analize sekvenci RNK dobijenih od osam izolovanih sojeva virusa NDV i sekvenci genoma referentnih sojeva virusa atipične kuge živine uz korišćenje Mega 40 i BLAST programa, utvrđeno je da su izolovani sojevi virusa Newcastle bolesti visoko virulentni.", publisher = "Univerzitet u Beogradu - Fakultet veterinarske medicine, Beograd", journal = "Veterinarski Glasnik", title = "Identification of isolated viral strains of atypical avian influenza using molecular methods of virological diagnostics, Identifikacija izolovanih sojeva virusa atipične kuge živine primenom molekularnih metoda virusološke dijagnostike", volume = "62", number = "3-4", pages = "167-177", doi = "10.2298/VETGL0804167V" }
Vidanović, D., Šekler, M., Vasković, N., Žarković, A., Matović, K., Milić, N.,& Nišavić, J.. (2008). Identification of isolated viral strains of atypical avian influenza using molecular methods of virological diagnostics. in Veterinarski Glasnik Univerzitet u Beogradu - Fakultet veterinarske medicine, Beograd., 62(3-4), 167-177. https://doi.org/10.2298/VETGL0804167V
Vidanović D, Šekler M, Vasković N, Žarković A, Matović K, Milić N, Nišavić J. Identification of isolated viral strains of atypical avian influenza using molecular methods of virological diagnostics. in Veterinarski Glasnik. 2008;62(3-4):167-177. doi:10.2298/VETGL0804167V .
Vidanović, Dejan, Šekler, Milanko, Vasković, Nikola, Žarković, Aleksandar, Matović, Kazimir, Milić, Nenad, Nišavić, Jakov, "Identification of isolated viral strains of atypical avian influenza using molecular methods of virological diagnostics" in Veterinarski Glasnik, 62, no. 3-4 (2008):167-177, https://doi.org/10.2298/VETGL0804167V . .