Analyzing Plastic Surface Erosion After Microbial Degradation

To observe the plastic surface erosion or decomposition after microbial degradation, we used scanning electron microscopy [39 (link)] (Jeol, JSM-6510, Tokyo, Japan). The SEM sample preparation protocol followed that of the Joint Center for Instruments and Researchers, College of Bio-Resources and Agriculture, NTU. Plastic films cultured with A. fumigatus L30 or A. terreus HC for 30 days were removed from the flask, and each film was cut into 2 pieces and divided into washed and unwashed groups. For the washed group, the plastic film was washed with distilled water to remove the mycelia attached to the plastic surface. The plastic films were immediately soaked in 2.5% (w/v) glutaraldehyde (Sigma–Aldrich Co.) at 4 °C and then shaken overnight for cell fixation. The fixed films were washed with 0.1 M sodium phosphate buffer (pH = 7.3) and postfixed in 1% osmium tetroxide (w/v) in an ice bath for 1 h. The samples were dehydrated gradually with different concentrations of ethanol (30%, 50%, 70%, 85%, 90%, 95%, 100%) for 60 min at each concentration, except for 70% and 100% ethanol for which overnight dehydration was conducted. The dehydrated samples were placed in a critical point dryer for critical spot drying, and subsequently, the samples were coated with gold and observed under a scanning electron microscope (Jeol, JSM-6510) [40 (link)].