FISH-Flow Cytometry: A High-Throughput Molecular Ecological Analysis of Intestinal Microbiota

Abstract

Until recently, ecological analysis of human intestinal microbiota had been conducted with culture-dependent methods. However, the majority of our intstinal bacteria remain not-yet cultured. Many culture-independent, molcular ecological methods have been developed to investigate real compositions of microbiota in the last decade. Among them, fluorescence in situ hybridization (FISH) is one of the most widely used methods，though it is time-consuming and requires laborious counting work of th detected bacteria. Development of FISH coupled with flow cytometry (FISH-FCM) has circumvented this problem. Since the development of FISH-FCM, it has been used in several fields of microbiology such as environmental microbiology and clinical microbiology. It is now being applied to more complex biological sample，such as feces. These days, intestinal microbiota analyses with FISH-FCM are being conducted not only at group or genus level, but also at the species level. Recently, we have succeeded in th developmentof a FISH-FCM method that is applicable to fecal samples containing high amounts of autofluorescnt particles. Further development of specific probes for the bacteria of intrestwill contribute to th illustration of the real picture of our intestinal microbiota.