A Preliminary Study On The Prevalence Of Yersinia Enterocolitis And Comparative Genomics Of Pathogenic Yersinia In Children Under 5 Years Old In China

Objectives:To investigate the prevalence of Yersinia enterocolitica infection and its prevalent type in children ? 5 years of age in China.To understand the demographic and clinical characteristics of infection cases and to provide scientific basis for clinical diagnosis and strategies of prevention and control of Y.enterocolitica infection.Methods:Based on the microbiological diagnosis,the study design is composed of general investigation and special investigation.In general investigation,microbiological diagnosis of Y.enterocolitica infection is conducted at sentinel hospitals from 10 provinces in China.Cases of acute diarrhea ? 5 years of age is taken stool samples to look for pathogenic strains of Y.enterocolitica,which is examined,isolated and typed.The special investigation was conducted on the basis of general investigation,besides microbiological diagnosis,temporal distribution,population distribution and clinical features of Y.enterocolitica infection were analyzed.The special investigation was carried out through one of the sentinel hospitals in general investigation and another sentinel hospital set up for collection of adult diarrhea cases.Results:1.General investigation A total of 7304 cases of acute diarrhea in children ? 5 years old were enrolled from the 10 provinces,from which 43 Y.enterocolitica infection cases were diagnosed in 2010-2015.The average infection rate of diarrhea cases in children ? 5 years old was 0.59%,with 3/O:3 being prevalent bioserotype(41/43).Male cases accounted for about 2/3(27/43)of Y.enterocolitica infections cases and about 70%(30/43)Yersiniosis cases were under 2 years old.A total of 27 infection cases took fecal microscopy and 85.19%(23/27)were positive for fecal leukocytes(WBC).Three cases showed long-term excretion of 3/0:3 Y.enterocolitica and the longest duration last as long as 3 months.Most pathogenic Y.enterocolitica isolates(36/43)from diarrhea children shared same PFGE patterns as strains isolated from pigs or dogs.A few children isolates did not find a consistent pattern in animal strains.Most(30/36)children isolates were found indistinguishable with pig or canine strains for PFGE patterns in same province,the rest(6/36)were identical to those from other provinces.There is no cross in PFGE patterns between the rarely isolated 4/0:3 strains and prevalent 3/O:3 strains in China.2.Special investigationA total of 2127 cases<5 years old with diarrhea and 1904 cases ?18 years old with diarrhea were included in special investigation from 2011-2015.Respectively,13 children cases and 2 adult cases were isolated with pathogenic Y.enterocolitica strains.The infection rate of children(0.61%,13/2127)was higher than that of adult(0.11%,2/1904).Both children and adult infection cases had one 2/0:9 infection,the rest cases were 3/0:3 infection.Neither children nor adult diarrhea cases found age,sexual or seasonal difference in Y.enterocolitica infection rate.Diarrhea children with fecal WBC had higher infection rate(1.79%)of pathogenic Y.enterocolitica than that of children without fecal WBC(0.00%),diarrhea children with fever had higher infection rate(2.07%)than those without fever(0.42%).The rate of fever in diarrhea children infected with pathogenic Y.enterocolitica was higher than diarrhea children.The positive rate of fecal WBC in diarrhea children infected with pathogenic Y.enterocolitica was higher than diarrhea children.Pathogenic Y.enterocolitica infection of adult had not shown association with fever or fecal WBC.Isolation rate of biotype 1A Y.enterocolitica of diarrhea adult(1.52%,29/1904)was higher than diarrhea children(0.28%,6/2127).Most adult isolated strains(20/29)and all children isolated strains(6/6)carried gene ystB.The isolation rate of biotype 1A strains in diarrhea children ? 2 years old was higher than that of children<2 years old;for diarrhea children under 5 years old,the isolation rate was higher in summer than autumn.The isolation rate of biotype 1A strains did not show age or seasonal differences in diarrhea children under 5 years old.The isolation rate did shown association with fever or fecal WBC in diarrhea children or adult.Conclusions:Pathogenic Y.enterocolitica infection rate of children ? 5 years of age in China is no less than that of developed countries,with 3/O:3 being prevalent bioserotype.Fecal WBC and fever are clinical characteristic in this population with Y.enterocolitica infection.Slaughter pigs and domestic canines are closely related to the Y.enterocolitica infection of children ? 5 years of age.Objectives:To confirm the phylogenetic relationships of Y.enterocolitica and Y.pseudotuberculosis strains representing different phenotype in China,compared with other complete genome sequenced strains of pathogenic Yersinia from NCBI.In particular,focuse on the Y.enterocolitica strains of ail+ystB+ phenotype within the three pathogenic Yersinia strains.Methods:The genome of representative Y.enterocolitica and Y.pseudotuberculosis strains were sequenced by Single Molecule,Real-Time(SMRT)technology.Genome assembly,component prediction,function annotation were conducted accordingly.Comparative genomic analysis included Core-pan analysis and SNP analysis of the sequenced strains and NCBI complete genome sequenced strains of pathogenic Yersinia.Results:1.Phylogenetic analysis based on core genomea.Y.enterocolitica,Y.pseudotuberculosis and Y.pestis strains had no cross on phylogenetic tree based on core genome(amino acid sequence of 1419 single-copy core gene).The only sequenced Y.pseudotuberculosis strain clustered to same branch but was located outside of all Y.pestis strains from NCBI.This strain is closest to Y.pestis than other Y.pseudotuberculosis strains.b.Within Y.enterocolitica,clustering of typically pathogenic strains showed specific pattern for O serotypes.c.Within Y.enterocolitica,non-pathogenic strains and strains harboring both virulence genes of pathogenic strains(ail)and non-pathogenic strains(ystB)clustered seperately to different branches.2.Phylogenetic analysis based on SNPContrary to phylogenetic tree based on core genome,the differences were:a.The only sequenced Y.pseudotuberculosis strain clustered to different branch of Y.pestis strains,but to same branches of all the Y.pseudotuberculosis strains.b.Within branch of typically pathogenic Y.enterocolitica strains,one 0:9 serotype strain QH2011-2-63B was located outside of all the other pathogenic strains of serotype 0:3 and 0:9.c.Non-pathogenic Y.enterocolitica strains were clustered within ail+ystB+ strains and those isolated with ail+ystB+ strains but only haboring ystB.Conclusions:1.Within Y.enterocolitica strains,based on the branchs of typically pathogenic Y.enterocolitica strains and branch of non-pathogenic Y.enterocolitica strains,a new branch of atypical Y.enterocolitica strains harboring both virulence genes of pathogenic strains(ait)and non-pathogenic strains(ystB)emerged,which had closer phylogenetic relationship to non-pathogenic Y.enterocolitica strains.2.The only sequenced Y.pseudotuberculosis strain in this study had closer phylogenetic relationship to Y.pestis strains than other complete genome sequenced Y.pseudotuberculosis strains from NCBI.