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Genome Sequencing and Comparative Genomics Analysis of Actinobacillus Pleuropneumoniae and Classical Swine Fever Virus

Author: XuZhuoFei
Tutor: ChenHuanChun;ZhouRui;XiongYuanZhu
School: Huazhong Agricultural University
Course: Animal Genetic Breeding and Reproduction
Keywords: Pleuropneumoniae Actinobacillus Classical swine fever virus Genome analysis Comparative genomics Bioinformatics
CLC: S852.61
Type: PhD thesis
Year: 2008
Downloads: 645
Quote: 4
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Porcine contagious pleuropneumonia (Porcine Contagious Pleuropneumonia, PCP) from Bacillus pleuropneumoniae (Actinobacillus pleuropneumoniae, APP) caused by a highly contagious respiratory disease. Classical swine fever (Swine fever, SF), the Europeans called classical swine fever (Classical swine fever, CSF), is caused by swine fever virus (Classical swine fever virus, CSFV), a highly contagious and lethal infectious disease. The diseases caused by these two important pathogenic microorganisms has caused huge economic losses to the pig industry worldwide. With the development of high-throughput sequencing technology, more and more genomic sequences were sequenced complete comparative analysis of the genome sequence provided a new opportunity to explore microbial genetic variation. We bioinformatics analysis methods and strategies, and in-depth comparison and analysis of the above two kinds of the genome sequence of the pathogenic microorganisms, described the relationship between the genome sequence structure and pathogenicity of the pathogen / metabolic mechanisms. The main contents include: 1.APP genome sequencing and analysis, APP according to the different divisions of surface lipopolysaccharide and capsular polysaccharide into at least 15 serotypes, virulence of different serotypes, antigenicity and geographical distribution endless the same. These characteristics of the APP serotype brought difficulties for the development of safe and effective vaccines and simple diagnostic method. APP metabolism and pathogenic molecular mechanism has always been a hot spot of the veterinary profession and the diagnosis of the medical profession. But the lack of information of the APP genome has become a bottleneck restricting the research and technical development of the bacteria. Therefore, the advantage serum complete genome sequence of the 3 type APP resolve our popular, not only for international APP Genome Project \In the present study, we are an advantage in the Chinese popular serotype 3 strain APP JL03 genome sequencing of APP, and for the first time a comprehensive functional annotation and analysis and comparison. JL03 genome sequenced by a length 2,242,062 bp chromosomal composition, predict a total of 2097 protein coding sequence, 6 to the ribosomal rRNA operon and 63 tRNA genes (GenBank Accession No.: CP000687). Additionally, the APP serum 5b L20 Canada sequenced strains length of approximately 2.27 Mbp, containing 2012 CDS: partial sequencing of APP serotype 1 4074 sequence contains 140 connected group (contig), about 2.07 Mbp, containing 2132 CDS. In the three sequenced APP strains, JL03 strain of low virulent strains L20 and 4074 are virulent strain. By of APP three strains between APP pasteurellosis Division species of genome science, and a detailed description of the characteristics of APP metabolism and pathogenicity / virulence. Confirmation APP metabolic pathway genes, confirmed the APP by fermentation and respiration (aerobic and anaerobic respiration) are two ways to generate energy (ATP), and to elucidate the complex metabolic network to provide a comprehensive The genetic basis. JL03 genome identified a characteristic genome Island: assimilation sulfur reducing gene string; FLP flagellar synthesis genes strings, involved in bacterial adhesion; capsular polysaccharide synthesis gene cluster involved in the synthesis of the bacterial capsular polysaccharide; lipopolysaccharide O-antigen gene cluster involved in the synthesis of bacterial O antigen. In the same time, we found that the presence of a 37.7 kb genomic island in the L20 strain on which encodes 8 phage genes, the island is not found in strain, JL03. These genomic island discovery provides a basis for the further study of the mechanisms of the APP serotype diversity. In addition, we describe a set of genes associated with the virulence of bacteria, JL03 strain Some gene missing or different types of genetic mutations. The above analysis confirms the previous understanding of the pathogen virulence determinant, provides a theoretical basis to explain the differences in virulence between different serotypes. In short, the preliminary analysis of the APP genomic sequence analysis of the structures and the encoded protein function not only confirmed the phenotypic characteristics of bacteria of some physiological and biochemical, new research ideas and to explore this important pathogen virulence and metabolic characteristics. Genome sequencing and analysis of CSFV 2.CSFV the flaviviridae Branch Pestivirus genus, only a single serotype, a single positive-strand RNA viruses. In the present study, we have a separation since the China Henan the CSFV SWH/CA/2004 of strains genome sequencing and analysis and comparison. The SWH/CA/2004 genomic cDNA sequence length of 12296 nt (GenBank accession number: DQ127910), NTR is 373 nT in length, and NTR's length is 226 NT and an open reading frame encoding a 3898 amino acid polyprotein precursor. Other have been reported for the CSFV strain SWH/CA/2004 strain genome sequence alignment and evolution analysis revealed that: the different strains of the genome nucleotide similarity between 92.4% -97.9%, the similarity in the amino acid between 96.1% -53.5%. The SWH/CA/2004 with the virulent strain cF114/CA/2001 strains closest genetic distance: far with middle the virulence of GXWZ02/CA/2003 strain genetic distance of 0.013, 0.170. We make full use of existing sources in different parts of the swine fever virus full-length genomic sequence information, reveal the phylogenetic relationships of CSFV, reference materials and study genetic variation and molecular epidemiology of classical swine fever virus foundation. 3 the Flaviviridae RNA polymerase expression suppressed Flaviviridae dependence involved in the composition of the virus replication enzyme complex in RNA polymerase (RNA-dependent RNA polymerase, RdRp), play in the viral replication cycle role. In the present study, the highly conserved MotifV for the three viral RNA polymerase gene region designed corresponding small interfering RNA (small interfering RNA, siRNA), and used to study the role of the siRNA molecule in the process of regulating the expression of RNA polymerase . By fluorescence microscopy, flow cytometry, Western blot detection and semi-quantitative fluorescent RT-PCR and other methods to verify the expression of inhibitory effect. Screening to CSFV siRNA can effectively inhibit RdRp-GFP fusion protein expression in PK-15 cells, is expected to become a candidate vaccine for prevention and immune swine fever virus infected pigs.

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CLC: > Agricultural Sciences > Livestock, animal medicine,hunting,silkworm,bee > Animal Medicine ( Veterinary Medicine) > Basic Veterinary Science > Animal Microbiology ( Veterinary Microbiology, ) > Pathogenic bacteria
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