Using fluorescent promoter-reporters to study sugar utilization control in Bifidobacterium longum NCC 2705

Abstract

Bifidobacteria are amongst the first bacteria to colonize the human gastro-intestinal system and have been proposed to play a crucial role in the development of the infant gut since their absence is correlated to the development of diseases later in life. Bifidobacteria have the capacity to metabolize a diverse range of (complex) carbohydrates, reflecting their adaptation to the lower gastro-intestinal tract. Detailed understanding of carbohydrate metabolism regulation in this genus is of prime importance and availability of additional genetic tools easing such studies would be beneficial. To develop a fluorescent protein-based reporter system that can be used in B. longum NCC 2705, we first selected the most promising fluorescent protein out of the seven we tested (i.e., mCherry). This reporter protein was then used to study the carbohydrate mediated activation of PBl1518 and PBl1694, two promoters respectively predicted to be controlled by the transcriptional factors AraQ and AraU, previously suggested to regulate arabinose utilization and proposed to also act as global transcriptional regulators in bifidobacteria. We confirmed that in B. longum NCC 2705 the AraQ controlled promoter (PBl1518) is induced strongly by arabinose and established that the AraU controlled promoter (PBl1694) was mostly induced by the hexoses galactose and fructose. Combining the mCherry reporter system with flow cytometry, we established that NCC 2705 is able to co-metabolize arabinose and glucose while galactose was only consumed after glucose exhaustion, thus illustrating the complexity of different carbohydrate consumption patterns and their specific regulation in this strain.

Genomic and Biochemical Characterization of Bifidobacterium pseudocatenulatum JCLA3 Isolated from Human Intestine

“…Bifidobacteria are believed to play an important role in carbohydrate fermentation in the colon. They can indeed ferment various complex carbon sources, such as gastric mucin, xylo-oligosaccharides, (trans)-galactooligosaccharides, soybean oligosaccharides, malto-oligosaccharides, fructooligosaccharides, pectin, and other plant derived-oligosaccharides, although the ability to metabolize carbohydrates is species-and strain-dependent and via bifid-shunt such have been reported in Bifidobacterium NCC2705 [34].…”

The Pleiotropic Effects of Carbohydrate-Mediated Growth Rate Modifications in Bifidobacterium longum NCC 2705

“…For example, glucose was demonstrated in B. longum to repress the production of Frk , a fructokinase required for the catabolism of fructose [ 17 ]. Recently, we demonstrated that glucose and galactose are sequentially metabolized in B. longum NCC 2705 and that the presence of residual glucose inhibited the consumption of galactose in that strain [ 18 ]. In B. lactis , glucose was shown to repress the expression of sucrose utilization genes [ 19 ].…”