Genome Sequencing And Analysis Of Mycoplasma Iowae And Mycoplasma Synoviae

Mycoplasma iowae is considered to be an special Mycoplasma, characterized by its resistance to heat, antibiotic and bile salts. The pathogen causing deaths of turkey embryos brought serious destroy to the turkey industry. Mycoplasma synoviae was isolated from chicken with synovitis which could cause upper respiratory disease and synovitis in chickens and turkeys.Roche454GX FLX technology was used in this work for genome sequencing of two different mycoplasmas:Mycoplasma iowae and Mycoplasma synoviae which present significant difference in virulence. The article emphasized on the information about genome structure, evolution relationships and virulence island of the two Mycoplasmas. Then, we found an optimizing condition for electro transformation into M. synoviae GX11-T with Tn4001and obtained a few mutants. The details are shown as follows:1. Genome sequencing and analysis of Mycoplasma iowae strain695The genome of M. iowae695is about1195,147bp with20%G+C content. A total of947open-reading frames were predicted with an average length of1089bp. The genome includes2rRNA and29tRNA genes. Analysed the metabolic pathways, usage frequency of amino acid codon, signal transduction systems, pseudogenes and the protein interaction networks, we compared all the Mycoplasma genomes which had been sequenced and found that only Mycoplasma iowae strain695had arbutin and salicin metabolism in the Carbohydrate metabolism pathway which seemed to be unique among all the avian mycoplasmas. We also found a two-component system in Mycoplasma iowae strain695, though two-component system had never been reported in mycoplasmas before. The major components of two-component system are histidine kinase(695.orf0477) and response regulator(695.orf0285). Additionally, the excinuclease ABC subunit A of M. iowae, with57%identity of the UvrA of Listeria monocytogenes may be important for survival in bile salts since the deletion of uvrA resulted in a significant impaired growth of Listeria monocytogenes in as little as0.3%bile salts.2. Genome sequencing and analyzing of Mycoplasma synoviae strain GX11-TThe genome of M. synoviae GX11-T is about738,351bp with28.34%G+C content. A total of679open-reading frames were predicted with an average length of981bp. And the genome includes2rRNA and32tRNA genes. Analysed the metabolic pathways, usage frequency of amino acid codon, signal transduction systems, pseudogenes and the protein interaction networks, we found34pseudogenes and5of34pseudogenes were predicted involving in phase shift of vlhA adhesin family. Additionally, we found the virulence island of M. synoviae GX11-T encoding hemagglutinin which was considered to belong to vlhA adhesin family. This result explains why M. synoviae GX11-T has a higher virulence.3. Phylogenetic analysis and gene family clusteringAnalysed the M.iowae695and M.synoviae GX11-T genomes compared with other Mycoplasma, the results of phylogenetic tree based on16SrDNA and ftsY indicated that Mycoplasma iowae had close genetic relationship with Mycoplasma gallisepticum; while Mycoplasma synoviae had close genetic relationship with Mycoplasma bovis, Mycoplasam hyorhinis and Mycoplasma hyopneumoniae. The genome sequence between Mycoplasma synoviae GX11-T and Mycoplasma synoviae53were90%identities. And comparing the two genome sequence, we found8genes only presented in Mycoplasma synoviae GX11-T, including a putative phase stift hemagglutinin gene. Besides, the analysis of the gene family clusters between three chicken Mycoplasma species indicated that Mycoplasam iowae695had43unique gene families, while Mycoplasma synoviae GX11-T had87unique gene families.4. Genetic transformation of Mycoplasma synoviae GX11-TIn this part, we studied about the optimizing conditions of genetic transformation with Tn4001into Mycoplasma synoviae. The transformation efficiency of Mycoplasma synoviae in different conditions (such as:voltage, resistance, proportion of DNA/Cell) was detected. We proofed the conditions of electroporation which were applicable for Mycoplasma synoviae. Five mutants of Mycoplasam synoviae GX11-T with gentamicin-resistance were gained.