Real-time PCR quantitation of clostridia in Feces of autistic children

被引:396
作者
Song, YL
Liu, CX
Finegold, SA
机构
[1] VA Med Ctr W Los Angeles, Res Serv, Los Angeles, CA 90073 USA
[2] VA Med Ctr W Los Angeles, Infect Dis Sect, Los Angeles, CA 90073 USA
[3] Univ Calif Los Angeles, Sch Med, Dept Med, Los Angeles, CA USA
[4] Univ Calif Los Angeles, Sch Med, Dept Microbiol Immunol & Mol Genet, Los Angeles, CA USA
关键词
D O I
10.1128/AEM.70.11.6459-6465.2004
中图分类号
Q81 [生物工程学(生物技术)]; Q93 [微生物学];
学科分类号
071005 ; 0836 ; 090102 ; 100705 ;
摘要
Based on the hypothesis that intestinal clostridia play a role in late-onset autism, we have been characterizing clostridia from stools of autistic and control children. We applied the TaqMan real-time PCR procedure to detect and quantitate three Clostridium clusters and one Clostridium species, C. bolteae, in stool specimens. Group- and species-specific primers targeting the 16S rRNA genes were designed, and specificity of the primers was confirmed with DNA from related bacterial strains. In this procedure, a linear relationship exists between the threshold cycle (C-T) fluorescence value and the number of bacterial cells (CFU). The assay showed high sensitivity: as few as 2 cells of members of cluster 1, 6 cells of cluster XI, 4 cells of cluster XIVab, and 0.6 cell of C. bolteae could be detected per PCR. Analysis of the real-time PCR data indicated that the cell count differences between autistic and control children for C. bolteae and the following Clostridium groups were statistically significant: mean counts of C. bolteae and clusters I and XI in autistic children were 46-fold (P = 0.01), 9.0-fold (P = 0.014), and 3.5-fold (P = 0.004) greater than those in control children, respectively, but not for cluster XIVab (2.6 X 10(8) CFU/g in autistic children and 4.8 X 10(8) CFU/g in controls; respectively). More subjects need to be studied. The assay is a rapid and reliable method, and it should have great potential for quantitation of other bacteria in the intestinal tract.
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页码:6459 / 6465
页数:7
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