Establishment And Evaluation Of Reference Mass Spectrometry Database And Classification Of Campylobacter Fetus

ObjectivesTo elucidate the phylogenetic status of the new subtype of "Campylobacter.fetus subsp.testudinum",analyze the epidemiology and genomics characteristics of Campylobacter fetus,establish a database for rapid mass spectrometry detection and evaluate Its quality.MethodsAccording to the experimental strains and related standard strains collected in the previous period of the research group:(1)Multi-phase taxonomic identification and typing,and preliminary analysis of genomic characteristics and differential drug resistance and virulence genes by bioinformatics;(2)Whole genome resequencing and average nucleotide identity analysis(ANI)were used as reference standards to establish a reference database of "Campylobacter.fetus subsp.testudinum" to verify its repeatability and accuracy,and to evaluate its clinical application value.Results(1)The microscopic morphology and biochemical reaction of the"Campylobacter.fetus subsp.testudinum" are close to the Campylobacter.fetus subsp.fetus,but with the subspecies of Campylobacter fetus subsp.venerealis is different:The first two 1%glycine growth test is positive,the latter is negative;(2)16SrRNA,RPOB gene sequencing and phylogenetic analysis showed that the "Campylobacter.fetus subsp.testudinum" is close to the Campylobacter fetus subsp.fetus the Campylobacter fetus subsp.venerealis,but the three subspecies are in the "grey area" of the same species,and the same kind of attribution cannot be determined;(3)The average nucleotide identity based on whole genome resequencing Average Nucleotide Identity(ANI)and Average amino acid identity(AAI)(with ANI,AAI similarity ?95%,core homologous gene POGs-OrthoANI ?96%as the same species)."Campylobacter.fetus subsp.testudinum" does not belong to the same species as the Campylobacter fetus subsp.fetus and the Campylobacter fetus subsp.venerealis.(ANI and AAI are<95%).The results of mass spectrometry also show three subtypes.Does not belong to the same species(refer to SARAMS Premium cluster analysis similarity<65%does not belong to the same species);genomic characterization analysis predicts differential CDs(genes encoding proteins)between Campylobacter fetuses and preliminary predictions Its function;(4)OrthoANI analysis:The OrthoANI value of self-built strain and standard strain C.fetus subsp.testudinum 03-427T is:99.30%~99.96%,and it is related to C.fetus subsp.venerealis NCTC10354,The OrthoANI value of C.fetus subsp.fetus 82-40T is 91.05%~92.26%.With reference to OrthoANI>96%as the basis for the determination of the same strain,the self-built strain was finally identified as "Campylobacter.fetus subsp.testudinum".The new super reference library has an accuracy of 100%(consistent with the genome sequencing results),the confidence interval is 82.3%~99.9%,and the same strain is 100%reproducible.ConclusionsThe reclassification of the new subtype of Campylobacter fetus has important epidemiological significance.It is speculated that it is the main type of bloodstream infection caused by Campylobacter in China;its analysis and prediction of genomic characteristics provide future research on virulence and phylogenetic evolution.A good foundation;the establishment and evaluation of the mass spectrometry database fills a gap in the field of microbial mass spectrometry and provides a new platform for rapid diagnosis of clinical infections.