期刊
CLINICA CHIMICA ACTA
卷 357, 期 1, 页码 55-64出版社
ELSEVIER
DOI: 10.1016/j.cccn.2005.02.004
关键词
bile acid 7 alpha-dehydroxylation; colon cancer; PCR; sulfate-reducing bacteria; methanogens; lactobacillus
资金
- NIDDK NIH HHS [P01DK38030] Funding Source: Medline
Background: Mounting evidence suggests a relationship between bacterial metabolism of certain dietary and endogenous factors and the development of colorectal cancer. Deoxycholic acid (DCA) is a well studied co-carcinogen and biotransformation product of 7 alpha-dehydroxylating Clostridia. H2S is a cytotoxic metabolite produced primarily by sulfate-reducing bacteria (SRB). The production of methane indicates low levels of active SRB. Lactic acid bacteria (LAB) have received attention recently due to their putative anti-tumor properties. Method: Human stool was spiked with pure cultures of bacteria and diluted in several enriched media. Each dilution titer was analyzed for the presence of the organism by PCR and biochemical assays. Duplicate stool aliquots were stored under various conditions for a 1 month period at -20 degrees C to test viability and detection. Results: Growth and enumeration of each spiked organism was confirmed by PCR and biochemical assays. The combination of bead beating and chemical lysis steps produced the greatest DNA yields. PCR assays detected as low as 75 fg target DNA. The ability to detect Methanobrevibacter smithii, and Desulfovibrio vulgaris by either PCR or biochemical assay declined significantly after storage at -20 degrees C for 1 month. Conclusions: Accurate detection and quantification of each bacterium using the described methods resulted when stool was processed immediately after collection. Storage of some members of the gut flora results in decrease in or loss of viability. (c) 2005 Elsevier B.V. All rights reserved.
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