Effects of biodegradable microplastics on soil microbial communities and activities: Insight from an ecological mesocosm experiment

For soil pH assessment, duplicate samples of six grams of soil from each soil column, at approximately 50% WHC, were mixed with 24 mL of 0.01 M CaCl2 and shaken at 200 rpm for two hours. After allowing samples to settle overnight, pH was measured using a pH meter (WTW pH 7710). Soils were analyzed for microbial activity i.e., C mineralization: four samples from each control (0%), 0.05% and 0.8% of PBAT-BD-MPs treatments were selected and analyzed in 3 – 4 sub-replicates. Microbial activity was characterized by basal respiration (BR) and substrate-induced respiration (SIR) according to ISO (2002) and ISO (1997), respectively. The soil samples were preincubated for seven days in the dark at 21 ± 2 °C to stabilize the soil environment before measurements (Hund-Rinke and Simon, 2008). The equivalents of 10 grams dry soil weight (gdw) were placed in the infusion bottles (150 mL) for BR measurement. The soil was moistened to 60% WHC and kept in the closed jar for two days to stabilize. After two days, the infusion bottles were aerated, hermetically sealed, and kept in the dark at 21 ± 2 °C for 24 h. The amount of carbon dioxide (CO2) was measured on an Aquilent GC 6850 (manual injection, column GasPRO, mobile phase He, detector TCD, SW Clarity). Each sub-replicate was measured two times. The data were expressed as µg CO2-C gdw-1 h-1. The measurement of SIR was conducted similarly to BR, only 5 mg glucose per gdw-1 was added to each aerated infusion bottle at the beginning of the measurement, the bottles were hermetically sealed, and the CO2 was measured after 0, 3, and 6 h. SIR was calculated using the linear regression and expressed as µg CO2-C gdw-1 h-1.