ABSTRACT
The recombinant strain Flu-NS1-124-Omp16 (H5N1) of the influenza virus expressing the brucellar Omp16 gene was constructed on the basis of the technology of reverse genetics for the purpose of developing vector anti-brucellosis vaccine. The obtained recombinant strain is a genetically stable construction. This stability is confirmed by the comparative analysis of the nucleotide sequences of the HA, NA, and NS genes of the recombinant vector virus Flu-NS1-124-Omp16 (H5N1) expressing the Omp16 gene of the Brucella abortus (GenBank: AAA59360.1). The comparative analysis showed that the nucleotide sequence of the NS gene of the first and the fifth passage level of the Flu-NS1-124-Omp16 (H5N1) virus corresponded for 100% to the initial part of 12AAS2TC_124 Omp16g containing the chimera NS1-124-Omp16 in the composition of DNA (deoxyribonucleic acid) plasmids pHW2000. Total identity with HA and NA genes of the strain A/AstanaRG/6:2/2009 (H5N1) was shown by the comparative analysis of the nucleotide sequences of HA and NA genes of the first and the fifth passage level of the recombinant strain Flu-NS1-124-Omp16 (H5N1). The recombinant vector virus Flu-NS1-124-Omp16 (H5N1) expressing the brucella Omp16 gene maintains the genetic stability during 5 passages in 10-day developing chicken embryos.
Subject(s)
Bacterial Outer Membrane Proteins , Brucella abortus/genetics , Genes, Bacterial , Genetic Vectors , Hemagglutinin Glycoproteins, Influenza Virus , Influenza A Virus, H5N1 Subtype , Neuraminidase , Viral Nonstructural Proteins , Viral Proteins , Animals , Bacterial Outer Membrane Proteins/genetics , Bacterial Outer Membrane Proteins/metabolism , Chick Embryo , Genetic Vectors/genetics , Genetic Vectors/metabolism , Hemagglutinin Glycoproteins, Influenza Virus/genetics , Hemagglutinin Glycoproteins, Influenza Virus/metabolism , Influenza A Virus, H5N1 Subtype/genetics , Influenza A Virus, H5N1 Subtype/metabolism , Neuraminidase/genetics , Neuraminidase/metabolism , Viral Proteins/genetics , Viral Proteins/metabolismABSTRACT
The paper describes comparative evaluation of IAVchip DNA microarray, reverse transcription PCR (RT-PCR), and real-time RT-PCR versus virus isolation in chicken embryos and shows their diagnostic effectiveness in detection and subtyping of influenza A virus. The tests were evaluated with use of 185 specimens from humans, animals, and birds. IAVchip DNA microarray demonstrates higher diagnostic effectiveness (99.45%) in early influenza A diagnosis as compared to the real-time PCR (98.38%) and RT-PCR (96.22%), thus showing its clear superiority. Diagnostic sensitivity of IAVchip DNA microarray (100%) exceeds the same of RT-PCR (95.95%) and real-time RT-PCR (97.96%) in the range of estimated confidence intervals. IAVchip DNA microarray and real-time RT-PCR displayed equal diagnostic specificity (98.85%), while diagnostic specificity of RT-PCR was 96.40%. IAVchip DNA microarray has an advantage over the other tests for influenza A diagnosis and virus identification as a more rapid method that allows performing simultaneous detection and subtyping of about tens of specimens within one experiment during 8-10 hours. The developed IAVchip DNA microarray is a general test tool that enables identifying simultaneously 16 hemagglutinin (HA) and 9 neuraminidase (NA) subtypes of influenza A virus and also to screen the influenza A viruses from humans, animals, and birds by M and NP genes.
Subject(s)
Influenza A virus/genetics , Influenza, Human/diagnosis , Influenza, Human/virology , Oligonucleotide Array Sequence Analysis/methods , Orthomyxoviridae Infections/diagnosis , Orthomyxoviridae Infections/virology , Animals , Birds/virology , Comparative Effectiveness Research , Humans , Influenza A virus/classification , Influenza A virus/isolation & purification , Real-Time Polymerase Chain ReactionABSTRACT
The diagnostic oligonucleotide microarray for subtyping of human and animal influenza A viruses (IAVs) was developed. We proposed a simple method of the fluorescent labeling of genomic segments of all known IAVs subtypes, the composition of the hybridization buffer, as well as the software of the data processing. 48 IAVs strains of different subtypes were analyzed using our microarray. All of them were identified, while 45 of 48 strains were unambiguously subtyped.
Subject(s)
Genome, Viral , Influenza A virus/classification , Molecular Typing/methods , Oligonucleotide Array Sequence Analysis/instrumentation , Orthomyxoviridae Infections/virology , RNA, Viral/classification , Software , Animals , Humans , Influenza A virus/genetics , Influenza A virus/isolation & purification , Influenza, Human/diagnosis , Influenza, Human/virology , Lab-On-A-Chip Devices , Orthomyxoviridae Infections/diagnosis , RNA, Viral/geneticsABSTRACT
This report describes the first isolation and characterization of porcine circovirus 2 (PCV2) in the Republic of Kazakhstan. The virus was isolated from a dead piglet that did not exhibit any typical clinical symptoms of porcine circovirus disease at a pig factory in North Kazakhstan oblast (region). The isolated virus belongs to genotype 2 (PCV2) and shares 96.6% sequence homology with one isolate and two strains from China and two French strains in group 1, cluster 1ab and 96.4% homology with two strains isolated in China and one strain from Hungary. Electron microscopy revealed the isolated virus had the typical morphological structure of PCV. This is the evidence of occurrence of porcine circovirus 2 isolation and characterization in Kazakhstan.
ABSTRACT
Peste des petits ruminant (PPR) is endemic in many Asian countries with expansion of the range in recent years including across China during 2013-2014 (OIE, 2014). Till the end of 2014, no cases of PPR virus (PPRV) were officially reported to the Office Internationale des Epizooties (OIE) from Kazakhstan. This study describes for the first time clinicopathological, epidemiological and genetic characterization of PPRV in 3 farm level outbreaks reported for the first time in Zhambyl region (oblast), southern Kazakhstan. Phylogenetic analysis based on partial N gene sequence data confirms the lineage IV PPRV circulation, similar to the virus that recently circulated in China. The isolated viruses are 99.5-99.7% identical to the PPRV isolated in 2014 from Heilongjiang Province in China and therefore providing evidence of transboundary spread of PPRV. There is a risk of further maintenance of virus in young stock despite vaccination of adult sheep and goats, along livestock trade and pastoral routes, threatening both small livestock and endangered susceptible wildlife populations throughout Kazakhstan.