| Objective: Clostridium difficile(CD) is a kind of intestinal obligate anaerobic bacillus, including toxigenic strains and non-toxigenic strains.They can asymptomatic colonize in the healthy population. Long-term using of antibiotics, or using of immunosuppressants etc. can induce environmental change of intestinal flora causing difficult C. difficile infection(CDI) and ensuing difficult clostridium cid-associated diarrhea(CDAD) and other symptoms. CD can long-term asymptomatic colonized in healthy infants and young children, and colonization rate and determinants of colonization and strain genotypes are largerly different in various countries and regions.Knowledge of colonization determinants and strain characteristics is limited. We studied the dynamics of C. difficile colonization in healthy infants from the community of hebei handan area. Determinants of colonization and strain genotypes were also determined in a cohort of infants(age, 0–3 years). These enriched the data of CD colonization in health infant from hebei province, providing evidence for clinical in prevention and treatment strategies of disease.Methods:1 A 1-point screening study involving 1098 healthy infants(age, 0–3 years) from handan city,hebei province was performed. C. difficile isolates were typed by polymerase chain reaction ribotyping and analyzed for the presence of toxin genes. The minimum inhibitory concentrations(MICs) of 14 antimicrobial agents, including vancomycin, metronidazole, fidaxomicin, rifampicin, rifaximin, levofloxacin, ciprofloxacin, tetracycline, erythromycin, clindamycin, ceftriaxone, ceftazidime, meropenem, and chloramphenicol were determined using the agar dilution method recommended by the Clinical and Laboratory Standards Institute(CLSI).2 A 1-year follow-up study involving 29 healthy infants was performed to determine the incidence and kinetics of intestinal C. difficile colonization. The C. difficile isolates were typed by polymerase chain reaction ribotyping and analyzed for the presence of toxin genes, too.Results: 1 In the one-point screening study, there are a total of 250(22.77%) carried C. difficile from 1098 healthy infants(0 to 3 years old),with 138(12.57%) carrying a toxigenic isolate. All toxigenic strains were tcd A+tcd B+cdt A-cdt B-, except one strain was tcd A+tcd B+cdt A+cdt B+, positive for binary toxin genes. All strains belonged to 15 different PCR ribotypes: the largest number is HB3(19.6% of positive samples), followed by HB9(14%), HB1(13.6%), HB2(10.8%) etc.. Interestingly, no infant carried the common epidemic 027 or 078 strains; 2 All isolates were sensitive to the metronidazole, vancomycin, fidaxomicin. In addition, 2% of the Clostridium difficile isolates were resistant to meropenem, and 6.8% to chloramphenicol. However, differen levels resistance to the rest 9 antimicrobial agents were found, the resistance rates were 99.2%(clindamycin), 96.8%(ceftazidime), 94.4%(erythromycin), 88.0%(ciprofloxacin), 63.2%(rifampicin), 63.2%(rifaximin), 54.4%( tetracycline), 51.6%(levofloxacin), 44.8%(ceftriaxone), respectively; 3 During the follow-up study, 20 infants acquired C. difficile during the first year of life from 29 enrolled infants, and the colonization rate is 69%. A total of 437 fecal samples were obtained. 111(25.4%) samples contained C. difficile, and the strain was toxigenic in 79(71.2%). 4 infants were colonized in the first month of life, while the other 16 were colonized later. C. difficile colonization was mostly discontinuous throughout the follow-up period(infants1、2、3、6、9、10、11、13、16、17、18、19). In 5 infants(infants 4、5、7、8、12), carriage was discontinuous, and in 3(infants 14,15 and 20), it was instantaneous colonization; 4 Eight different PCR ribotypes were identified among the 111 C. difficile isolates obtained during the follow-up study. Toxigenic isolates belonged to the following PCR ribotypes: HB42(n=26), HB1(n=16),HB52(n=12), HB3(n=15), HB6(n=5) and HB5(n=5). All toxigenic strains were tcd A+tcd B+cdt A-cdt B-, and none possessed the genes encoding the binary toxin. The following 2 PCR ribotypes were exhibited by nontoxigenic strains: HB9(n=14), HB2(n=18). Infants were most frequently colonized with a single clone of C. difficile. Infants 8, the C. difficile isolate was HB3 at the time of 2 months of life, and it was HB-42 later in the fecal samples. The C. difficile isolates from the infants 11 were HB1 and HB9 respectively, and in infants 15 C. difficile isolates were HB1 and HB52 respectively, and in infants 16 C. difficile isolates were HB1 and HB3 respectively.Conclusions:1 In the one-point screening study, the Clostridium difficile colonization rate in 0-3 years old healthy infants in Handan city Hebei province was slightly lower than that reported abroad, while the toxingenic strains colonization rate was similar compared with the internal reports. The statistics analysis results showed that the pregnant period, using of antibiotics recently and hospitalization history recently may be the determinants of colonization factors, what reported similar with the results at home and abroad. The Clostridium difficile strains’ PCR-ribotyping results show that the advantages of types were HB3 and HB9. Some types were same with the types isolated from hospitalized children and adults in Hebei province. This suggests that it si possible that the Clostridium difficile spreads in these individuals.2 In the one-point screening study, the resistance of C. difficile strains to 14 antimicrobial agents results show that they have a relatively high level resistance to rifomycins compared with several reports in China. The reason for this result may be due to the application of antibiotics in the location of sample collection. The resistances of rest other antimicrobial agents are similar with other reports.3 In the follow-up survey, C. difficile colonization rate in infants is relatively high compared with the rate of 0-12 month infants in one-point screening study. The difference was statistically significant, indicating that the only charge a stool specimens will reduce the overall CD colonization rate. C. difficile colonization appears mainly as an age-dependent process, and the specific relationship needs to further study to prove it. Toxigenic C. difficile isolates in infants are dominant type as long-term colonization isolates. Pathogenic strains circulate in asymptomatic infants from the community, who represent a potential reservoir of pathogenic strains. |
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