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Chapter 7 : Sequence-Based Methods for Investigating Intestinal Microbes
This chapter focuses on the use of microbial nucleic acid sequences for the detection, localization, and characterization of microbes in the human intestine, with emphasis on cultivation-resistant pathogens and commensals. Organisms are recovered from diseased tissues by inoculating axenic media and classified based on their ability to grow, using defined substrates at particular temperatures and atmospheres. Having a cultivated microbe is again critical for making antibodies for serological or immunohistochemical detection of infection. Fluorescence in situ hybridization can be used to localize microbes to cells in the intestine by targeting microbial nucleic acid sequences with fluorescently labeled probes and visualizing them with fluorescence microscopy. Leser and colleagues performed an exhaustive study of the bacterial flora of pig intestines using PCR of bacterial 16S rDNA with cloning and sequence analysis. The microbial cause of Whipple’s disease was first revealed by studies showing that this fatal disease could be cured with antibiotics. In 1997, Schoedon and colleagues reported the successful propagation of Tropheryma whippelii in human macrophages by using cytokines to deactivate intracellular killing of bacteria. Cyclospora cayetanensis infection causes prolonged diarrhea and is treated with trimethoprim-sulfamethoxazole.
Key Concept Ranking
- Reverse Transcriptase PCR