Study design for age-related dysbiosis study

Synchronized L1 larvae were grown on NGM agar plates seeded with E. coli OP50 at 20°C until they reached the young adult stage. All the experiments started from the young adult stage, which was considered day 0. From days 1 to 10, the worms were transferred to new NGM plates containing E. coli strains at 20°C daily for further experiments. For preparing for HK E. coli OP50, the bacterium cultured overnight in liquid LB medium was treated with 75°C water bath for 90 min (Qi et al., 2017). For preparing for ampicillin-killed E. coli OP50, the bacterium cultured overnight in liquid LB medium was seeded onto NGM plates containing 200 μg/ml ampicillin. Then the OP50 strain was periodically streaked onto ampicillin-containing and ampicillin-free LB plates to confirm that it could not proliferate (Garigan et al., 2002). BW25113 (E. coli K-12 WT) and E. coli K-12 ΔtnaA strain were obtained from the Keio collection (Baba et al., 2006).