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使用实时荧光定量PCR解析婴儿肠道双歧杆菌构建规律

Analyzing Colonization of Bifidobacteria in Infants with Real-time Fluorescent Quantitative PCR

  • 摘要: 目的 通过非培养的分子生物学方法检测婴儿早期肠道双歧杆菌的菌种组成及其数量变化,解析婴儿肠道双歧杆菌种群构建及菌种组成变化的规律及其可能的影响因素。方法 收集2013年3~4月16例正常足月新生儿第1次,第2、4、7、10、14、28天,3个月,6个月,1岁共10次粪便,使用实时荧光定量PCR和双歧杆菌属及种特异性引物对收集的粪便中的双歧杆菌属及人体肠道特有的8个双歧杆菌菌种进行定性及定量分析。结果 共收集婴儿粪便样品136份,双歧杆菌检出率为93.4%(127/136),湿便中菌数为10 5~10 11 CFU/g;第1次胎便双歧杆菌检出率为83.3%(湿便中菌数约为10 5 CFU/g);出生后14 d内双歧杆菌检出率及菌量在相对低的水平,被检出的双歧杆菌为1种或2种;出生28 d及以后双歧杆菌检出率增至100%,湿便中菌量增至108 CFU/g以上;出生后28 d至6月龄多数婴儿、1岁时全部婴儿粪便中被检出双歧杆菌增至3种;双歧杆菌各菌种的检出率为B. breve (92.1%)、B. infantis(66.1%)、B. catenulatum(59.8%)、B. bifidum(25.2%)、B. longum(24.4%)、B. dentium(13.4%)、B. angulatum(5.5%)、B. adolescentis(1.6%)。结论 受试婴儿出生后到1岁其肠道双歧杆菌数量、菌种组成及其菌种多样性均随时间出现显著的变化,是受试婴儿肠道双歧杆菌种群构建的重要时期;同时发现受试婴儿双歧杆菌的菌种群构建整体上延迟、菌种组成上出现异常,推测是和受试婴儿的生产方式、喂养情况及其他各种环境因素密切相关。

     

    Abstract: Objective In order to know how intestinal Bifidobacteria community could be built in the infants and whether the environmental factors could affect them, the present study was conducted to characterizethe species composition and trace the quantitative changes of intestinal Bifidobacteria of the infants in their early stages with non-culture dependent molecular method. The possible association of Bifidobacteria community of the infants with their health was also discussed. Methods Total 16 of full-term newborn infants born between March and April 2013 were recruited for the present study. Fecal samples were collected from them at 1 day, 2 days, 4 days, 7 days, 10 days, 14 days, 28 days, 3 months, 6 months and 1 year after birth. Real-time fluorescent quantitative PCR with genus and species specific premiers was used to detect Bifidobacteria and 8 predominate species in human intestine qualitatively and quantitatively present in these collected fecal samples. Results Total 136 fecal sample were collected and Bifidobacteria were detected from 93.4% (127/136) of them with the concentration of 1.0×10 5 to 1.0×10 11 CFU/g. Bifidobacteria were found in 83.3% of the fecal samples collected from the first day after birth with more than about 10 5 CFU/g. However, Bifidobacteria were detected relative low until 14 days and were taxonomically belonged only to one or two species. Bifidobacteria were found in almost 100% of the fecal samples

     

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