Identification of adaptive mechanisms leading to reduced antibiotic susceptibility in bacterial biofilms using experimental evolution. Part 3: Pseudomonas aeruginosa floating biofilms in synthetic cystic fibrosis medium

Pseudomonas aeruginosa was repeatedly and intermittently exposed to tobramycin. Bacteria were grown in synthetic cystic fibrosis medium in wells of a 96-well microtiter plate. After 24 hours, more medium with or without tobramycin was added. After another 24 hours of incubation, a subsample of the well content was used to inoculate fresh synthetic cystic fibrosis medium in a 96-well microtiter plate. This was repeated for a total of 15 cycles. Evolved lineages were then DNA-sequenced to screen for genome changes.

铜绿假单胞菌（Pseudomonas aeruginosa）被反复间歇性暴露于妥布霉素（tobramycin）环境中。实验菌株于96孔微孔板（96-well microtiter plate）的孔内，在合成囊性纤维化培养基（synthetic cystic fibrosis medium）中培养。培养24小时后，向各孔补加含有或不含妥布霉素的新鲜培养基。继续孵育24小时后，吸取孔内培养物的子样本，用于接种新的96孔微孔板内的新鲜合成囊性纤维化培养基。上述流程共计重复15个循环。随后对演化得到的菌株谱系进行DNA测序，以筛选基因组变异。