Initial Analysis Of The Whole Genome Sequence Of Bacillus Thuringiensis Strain YBT-1520

Bacillus thuringiensis strain YBT-1520, which is highly toxic to lepidopteran pests(e.g., Helicovepa armigera and Plutella xylostella), was isolated by our lab and protectedby patent (Chinese patent number: ZL 95 106749.4). In this thesis, we reported thesequence determination and initial analysis of the genome of this strain. The resultsrevealed that this genome consisted of one chromosome and nine plasmids. Thechromosome was 5,401,320 bp in length and harbored 5,556 open reading frames (ORFs).The total length of the 9 plasmids was 401,376 bp and the total number ofplasmid-encoded ORFs was 395.The YBT-1520 chromosome contained a good few genes encoding putativevirulence factors, such as enterotoxin, phospholipase, chitinase and immune inhibitor A,which may partly account for the high toxicity of YBT-1520. To get further informationabout the impact of gene regulation on the virulence to the insect pests, we investigatedall the chromosome-encoded two-component regulatory systems and genes regulated bythe pleiotropic regulator PlcR. Our analysis indicated that the chromosome encoded 29putative two-component regulatory systems, four of which were suggested to be directlyinvolved in pathogenicity. The potential PlcR regulon included 7 genes encodingvirulence factors and some other genes which may indirectly affect the overallpathogenicity of YBT-1520.Through various analysis concerning chromosome comparison, such as the analysisof the distribution of the homologous genes and the most similar genes, the comparison ofall chromosome-encdoed proteins and the analyses based on COG database and KEGGdatabase, respectively, we found that the chromosome of YBT-1520 was similar to thoseof Bacillus cereus strain ATCC 14579, Bacillus anthracis strain Ames and Bacillusthuringiensis strain 97-27, which indicated that these four representative strains in the B.cereus group of organisms may share a common and relatively short evolutionary history.The analysis results also indicated that in these strains, some differences among severalgenes may result in totally different phenotypes. In addition, we identified allchromosome-encoded genes specific for YBT-1520 (with regard to the strains of B.cereus group of organisms), yet the analysis based on COG database suggested that mostof them are associated with hypothetical proteins or mobile genetic elements.Analysis of the plasmids of YBT-1520 also revealed some interesting features. Mostnotably, plasmid pBMB131 harbored four genes (cry1Aa, cry1Ia, cry2Aa and cry2Ab)encoding insecticidal crystal proteins and one gene (vip3Aa) coding for vegetative insecticidal protein. Two of these genes (cry1Ia, vip3Aa) were found for the first time inYBT-1520. Moreover, plasmid comparison suggested that B. cereus plasmid pBc10987,B. anthracis plasmid pXO1, B. thuringiensis plasmid pBtoxis and pBMB131 may have acommon ancestor, and pBMB131 was likely to contain a putative pathogenicity islandencoding all the virulent factors located on pBMB131. These results provided a goodfoundation for the research of virulence plasmid evolution and the identification of newvirulence factors. The plasmid pBMB95 possessed one copy of cry1Ac and pBMB69contained one copy of cry1Ab, which indicate that these two plasmids also madesubstantial contributions to the pathogenicity of YBT-1520. The plasmid pBMB131 andpBMB67 both carried a potential operon whose product is analogous to the Rap-Phrcassettes from Bacillus subtilis. This observation suggested that these two plasmids mayplay a role in the regulation of several cecullar processs pertaining to the pathogenicity ofYBT-1520, such as sporulation and exoprotease production. Plamid pBMB67 also housedseveral genes putatively involved in conjugation. The analysis with respect to these genesmay lead to a better understanding of the conjugation systems of Gram-positive bacteria.Additionally, we found many other plasmid-encoded genes that may affect the overallpathogenicity of YBT-1520 against insect pests. The presences of all these plasmids werethe main source for the pathogenicity of YBT-1520.