Population Dynamics of Bifidobacterium Species in Human Feces during Raffinose Administration Monitored by Fluorescence In Situ Hybridization-Flow Cytometry

Abstract

The population dynamics of bifidobacteria in human feces during raffinose administration were investigated at the species level by using fluorescence in situ hybridization (FISH) coupled with flow cytometry (FCM) analysis. Although double-staining FISH-FCM using both fluorescein isothiocyanate (FITC) and indodicarbocyanine (Cy5) as labeling dyes for fecal samples has been reported, the analysis was interfered with by strong autofluorescence at the FITC fluorescence region because of the presence of autofluorescence particles/debris in the fecal samples. We circumvented this problem by using only Cy5 fluorescent dye in the FISH-FCM analysis. Thirteen subjects received 2 g of raffinose twice a day for 4 weeks. Fecal samples were collected, and the bifidobacterial populations were monitored using the established FISH-FCM method. The results showed an increase in bifidobacteria from about 12.5% of total bacteria in the prefeeding period to about 28.7 and 37.2% after the 2-week and 4-week feeding periods, respectively. Bifidobacterium adolescentis, the Bifidobacterium catenulatum group, and Bifidobacterium longum were the major species, in that order, at the prefeeding period, and these bacteria were found to increase nearly in parallel during the raffinose administration. During the feeding periods, indigenous bifidobacterial populations became more diverse, such that minor species in human adults, such as Bifidobacterium breve, Bifidobacterium bifidum, Bifidobacterium dentium, and Bifidobacterium angulatum, proliferated. Four weeks after raffinose administration was stopped, the proportion of each major bifidobacterial species, as well as that of total bifidobacteria, returned to approximately the original values for the prefeeding period, whereas that of each minor species appeared to differ considerably from its original value. To the best of our knowledge, these results provide the first clear demonstration of the population dynamics of indigenous bifidobacteria at the species level in response to raffinose administration.is a nondigestible oligosaccharide that is widely distributed in many plants, such as sugar beet, cane, cabbage, potato, grape, wheat, barley, corn, and the seeds of many legumes (20,21). In Hokkaido, Japan, raffinose is an important agricultural product that is extracted from sugar beets as a by-product of sugar processing. Several prebiotic effects in humans have been reported for this oligosaccharide, including reduction of fecal ammonia and indole (18), improvement of defecation frequency (18), and increased cell numbers among indigenous bifidobacteria (5). Increases in bifidobacterial populations in rats (8) and the suppression of T helper 2 cell-mediated immune responses in mice (19) have also been reported.To obtain a scientific basis for the use of raffinose as a prebiotic, a precise determination of its effects on microbiotic composition in the human intestine, especially on the bifidobacterial population at the species level, is required. Because the majority of microbiota in the huma...

Pathogen destruction and solids decomposition in composting latrines: study of fundamental mechanisms and user operation in rural Panama

“…The pH of human faeces is neutral, ranging from pH 6.6 to 7.0 (Dinoto et al 2006). The two most commonly observed desiccants in our study have literature pH values ranging from 9.4 to11.3 (wood ash) and 4.5 to 7.8 (sawdust) ).…”

Development of a Double-Crossover Markerless Gene Deletion System in Bifidobacterium longum: Functional Analysis of the α-Galactosidase Gene for Raffinose Assimilation

“…3A). The ␣-Gal-encoding gene was selected as the target because ␣-Gal hydrolyzes raffinose, which is known to increase bifidobacterial populations in the intestinal tract of rats and humans (9,10). As shown in Table 3, three genes annotated as ␣-Gal were found in the draft genome sequence of B. longum 105-A (Yoshikawa et al, unpublished).…”

Prebiotic Effect of Lacto-N-biose I on Bifidobacterial Growth

“…Non-digestible food ingredients that promote the growth and/or activity of probiotics are termed prebiotics. Several oligosaccharides, such as raffinose, 2,3) fructo-oligosaccharide, 4,5) and galacto-oligosaccharide, [6][7][8] are known to function as prebiotics for bifidobacteria, and thus are specifically termed bifidus (or bifidogenic) factors.…”