Initi Al Analysis Of The Whole Genome Sequence Of Brucella RB51Strain

Brucellosis， caused by a group of Gram-negative bacteria of the genus Brucella， is one ofthe major reproductive disorders causing allergic infectious zoonotic diseases and resultingin huge economic losses and serious public health problem。 Currently， vaccination is stillthe main means of prevention and control of brucellosis。 RB51is the most widely usedvaccine in North America and initially generated by smooth Brucella abortus2308afterrepeated passaging in vitro and screen of rifampicin and penicillin。 RB51is very stable R-type strain which is the most widely used vaccine applicated in the current NorthAmerican。 Comparing to Brucella abortus2308， IS711sequence is inserted in RB51wboAgene （encoding glycosyltransferase）。 A large number of experiments have proved thatexcept wboA gene there are a lot of other gene mutation in RB51strain。 In order to explorethe treatment of brucellosis， diagnosis， vaccine development and provide a theoreticalbasis。 this study is designed to research the complete genome sequence of strain RB51through the application of sequencing。 The main results are as follows:Objective: microbial genome research has a crucial role in the development ofpathogenicity， prevention and treatment。 Through genome research， each functional geneand the relationship between genes can be clear which is useful for the vaccine development。In addition， through the whole genome sequencing， we can find fully parallel geneticdifferences between species， providing data for the research of microbial evolution。 Thisstudy is committed to indicate RB51genome information and the main functions of its genes。Through the comparison with the original strain S2308， the activity and the virulence genesare determined， indicating the research direction for the development of brucella vaccine。Methods: The depth of the new generation sequencing （hiseq2000） was used to detectthe RB51whole genome sequence。 Rapid analysis software （SOAPdenovo， Glimmer， etc）are used to rationally classify the sequence information obtained by sequencing andannotation。 The difference between functional genes are gotten by comparing with S2308.Results:1. Genome-wide overview: Through gene prediction， repetitive sequencepredicted， non-coding RNA prediction and other methods to obtain the composition of thestrains sequenced genome。 Genome analysis found that RB51-1genome contains3，283genes， a total length of2，797，347bp， the average length of852bp， accounting for85.91%of the full length genome。 A total of80tandem repeat sequences， a total lengthof6，564bp， accounting for0.2016%of the total length of the genome，54of smallsatellite sequences，12microsatellite sequences，49tRNA and3rRNA。2. Functionalannotation of genomes: There are8major databases for the annotation of the RB51genomein my experiment。（1） The GO database annotation results: GO database for thecorresponding gene， wherein one is associated with the composition of cytologycomponent， another is with molecular function， other is corresponding biologicalpathways。 Indicating that this genomic genes involved in60%of the biological componentconstitute and only10%of the genes involved in the expression of biological pathways，thereby illustrate the real impact of functional and metabolic pathway genes accounted forless;（2） KEGG database annotation results: There are2131genes involved in metabolicpathways，distributed in147metabolic pathways;（3） SwissPort database annotation results: a total of1501genes can correspond to determine the source of homologous species andclearly the corresponding to these genes protein information;（4） COG database annotationresults: Getting2570corresponding genes， this experiment clearly information such genesand the focal point with22categories COG species， thereby infer the function of thesehaplotype sequences;（5） Pathogen host interaction database （PHI） annotation results:20genes can be expected correspondence， indicating that the interaction between the20genesof the genome and host;（6） Bacterial pathogen virulence factors （VFDB） annotation results:75genes are expected to correspond to illustrate the genome containing75genes are relatedwith virulence;（7） Resistance gene （ARDB） note: A resistance associated with thecorresponding genes can clear the name of the Department of the gene and the correspondingtypes of antibiotics。（8） NR database annotation results: By comparison with the NRdatabase，a total of3261genes can be corresponding to provide data to support the forecastthe late genes， and study the function and significance of these genes。3. Comparativegenomic analysis:（1） Compared with the reference sequence B.abortus2308， RB51-1genome were found135SNP， which synonymous mutations26，89non-synonymousmutations。 B.abortus2308with the reference sequence for comparison， RB51-1genomicinsertion mutations were found in five， three deletion mutants， CDS area is not InDeloccurred;（2） In the mutant of SNP and InDel sites， there is a SNP loci and two InDel locimay relate with the toxicity of RB51-1（Table5-3，5-4） and these genes can be consisted withVFDB;（3） Found in Scaffold4of RB51-1a inserted sequence， the site of insertion is122559-129801， length5242bp。 This sequence located in the B.abortus2，308，881，899bp and the fragment contains six predicted genes， of which two with nitritemetabolism-related， three and sucrose metabolism-related， and one related totranscriptional regulator;（4） In the protein sequences of YP₄13569.1， there are six copyof long tandem repeat and the phenomenon of the tandem repeat deletion in the toxic strainsB.abortus2308and B.abortus9-941which also appear in the virulent strains of other host，so whether the loss of the strains associate with the virulence of strains or not also needfurther validation experiments。Conclusion: In this study， genomic information of RB51strain was completelyannotated and analysed。 Comparing with the strain S2308we found the genes related withthe virulence and viability。 That is useful for later development of vaccines and brucellosisprevention。...