Genetic Analysis Of Novel Capsular Polysaccharide Synthesis Gene Loci And The Development Of High-throughput Novel Capsular Gene Typing In Streptococcus Suis

Streptococcus suis(S.suis)is an important zoonotic agent.Capsular polysaccaride(CPS),which plays a key role in the process of anti-phagocytosis and is encoded by cps cluster in genome,is one of important pathogenic factors in S.suis.According to the difference of CPSs antigenicity,33 serotypes were assigned in S.suis.Agglutination and co-agglutination tests are traditionally used to serotype S.suis strains.However,multiple researches showed that some S.suis strains could not be typed by above methods,named non-serotypeable strains.In the genome era,the draft genomes of 33 serotype reference strains have been sequenced by high-throughput Illumina sequencing technology.Sequence analysis of cps confirmed that CPSs of S.suis are synthesized by the Wzx/Wzy pathway and wzy genes are critical in the pathway.Recently,11 novel cps loci(NCL1 to 10,Chz)have been found continually by the comparative analysis of wzy genes.In the first part of this study,a collection of non-serotypeable strains isolated from China and Canada were sequenced by Illumina sequencing and their cps loci were analyzed.Ten novel cps loci(NCL11 to 20)were found.Based on previous studies,we tentatively concluded that these 10 NCLs represent novel serotypes.In addition,those non-serotypeable strains assigned to known serotypes by 32-plex Luminex assay were also sequenced by Illumina sequencing.The analysis of cps sequence indicated that large scales of insertions,deletions,replacements,frameshift mutations within one gene or complete deletion of cps locus were found in these cps loci.Our results revealed the reason why these strains cannot be typed by antiserum.Luminex xMAP system is a suspension array platform based on multiple microspheres.A 32-plex Luminex system has been developed to detect 33 S.suis serotypes simultaneously.In the second part of this study,we developed a 22-plex Luminex assay which is able to simultaneously detect all 21 novel cps loci.This assay provides a rapid,effective and convenient tool to facilitate the research of non-serotypeable S.suis strains.