A Helicobacter pylori whole-genome DNA microarray was constructed to study expression profiles of H. pylori in response to a sudden temperature transfer from 37°C to 20°C. The expression level of the genome at each of four time points (15, 30, 60, and 120 min) after temperature downshift was compared with that just before cold treatment. Globally, 10.2% (n=167) of the total predicted H. pylori genes (n=1636) represented on the microarray were significantly differentially expressed (p<0.05) over a 120 min period after shift to low temperature. The expression profiles of the differentially expressed genes were grouped, and their expression patterns were validated by quantitative real-time PCR. Up-regulated genes mainly included genes involved in energy metabolism and substance metabolism, cellular processes, protein fate, ribosomal protein genes, and hypothetical protein genes, which indicate the compensational responses of H. pylori to temperature downshift. Those genes play important roles in adaption to temperature downshift of H. pylori. Down-regulation of DNA metabolism genes and cell envelope genes and cellular processes genes may reflect damaged functions under low temperature, which is unfavorable to bacterial infection and propagation. Overall, this time-course study provides new insights into the primary response of H. pylori to a sudden temperature downshift, which allow the bacteria to survive and adapt to the new host environment.