Tannin resistance strategies of Escherichia coli in anaerobic conditions

Gene expression profiles of Escherichia coli, grown anaerobically, with or without Acacia mearnsii (Black wattle) extract were compared to identify tannin-resistance strategies. The cell envelope stress protein, spy, and the multidrug transporter-encoding mdtABCD, both under the control of the BaeSR two-component regulatory system, were significantly up-regulated in the presence of tannins. BaeSR mutants were more tannin-sensitive than their wild-type counterparts. Keywords: tannin resistance E. coli BW13711 cells were grown anaerobically in continuous culture (d = 0.14h-1) in the presence and absence of Acacia mearnsii (Black Wattle) tannin extract (WTE) at 37°C. Under both steady state conditions, E. coli reached an OD600 of ~0.2. The pH at steady-state was 5.7 and 5.3 without and with WTE, respectively. These conditions indicated that the presence of tannins in the medium did not inhibit the growth of E. coli. E. coli BW13711 cells were harvested from continuous culture after steady-state conditions were reached. Duplicate samples were collected after an additional four volume turnover of the culture medium and total RNA was isolated from these samples using the RNeasy mini kit (Qiagen Inc., Valencia, CA). The presence of residual DNA was checked by PCR using the primers for amplification of cca. If necessary, the remaining DNA was removed by RQ1 RNase-Free DNase (Promega corp, Madison, WI) according to the manufacturer’s instructions. E. coli whole genome transcriptional profiling was performed by hybridization on the GeneChip® E. coli Antisense Genome Array (Affymetrix Inc., Santa Clara, CA) according to the manufacturer’s instructions. Microarray data were analyzed using MicroArray Suite 5.0 (Affymetrix Inc.) and GeneSpring 5.0 (Silicon Genetics, Redwood City, CA) software. The 95% confidence interval of the WTE array duplicates demonstrated that genes >1.9 times up- or down-regulated could be considered as significantly differentially regulated. After removing significantly up- or down-regulated genes which had large variability in expression on the duplicate arrays, it was found that in total 1,498 genes were up- or down-regulated