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Bifidobacteria are natural inhabitants of the human gastrointestinal tract and have been widely used as functional foods in different products. During industrial processing, bacterial cells undergo several stresses that can limit large-scale production and stability of the final product. To better understand the stress-response mechanisms of bifidobacteria, microarrays were used to obtain a global transcriptome profile of Bifidobacterium longum NCC2705 exposed to a heat shock treatment at 50°C for 3, 7 and 12 min. Gene expression data highlighted a profound modification of gene expression, with 46% of the genes being altered. This analysis revealed a slow-down of Bi. longum general metabolic activity during stress with a simultaneous activation of the classical heat shock stimulon. Moreover, the expression of several genes with unknown function was highly induced under stress conditions. Three of these were conserved in other bacteria species where they were also previously shown to be induced by high temperature, suggesting their widespread role in the heat stress response. Finally, the implication of the trans-translation machinery in the response of Bi. longum cells to heat shock was suggested by the induction of the gene encoding the tmRNA-associated small protein B (SmpB) with concomitant high constitutive expression of the tmRNA gene