After the first phase was completed, each larva was stored in a different 1 L glass jar (10 cm × 10 cm × 10 cm) with a clean substrate. This new substrate was collected in a field with the same composition as the one used in the first phase. Temperature and humidity were kept at an environment level (e.g., 24° and 60%, respectively).
The substrate used in glass jars was checked under a stereomicroscope every 5 days for one month to look for plastic fragments potentially ingested by larvae when still in the EPS box and now egested with feces. Fragments of EPS are easily recognizable for their white color and round shape, and we checked for MP fragments using a stereomicroscope (Nikon C‐LEDS) following the protocol by Gallitelli et al. (2020 (link)), Gallitelli et al. (2021 (link)). During the experiment we followed protocols to prevent sample contamination, considering EPS particles our target. Specifically, we used nitrile gloves, laboratory coats, and steel tools to avoid external contamination (see Gallitelli et al., 2020 (link)).
The substrate used in glass jars was checked under a stereomicroscope every 5 days for one month to look for plastic fragments potentially ingested by larvae when still in the EPS box and now egested with feces. Fragments of EPS are easily recognizable for their white color and round shape, and we checked for MP fragments using a stereomicroscope (Nikon C‐LEDS) following the protocol by Gallitelli et al. (2020 (link)), Gallitelli et al. (2021 (link)). During the experiment we followed protocols to prevent sample contamination, considering EPS particles our target. Specifically, we used nitrile gloves, laboratory coats, and steel tools to avoid external contamination (see Gallitelli et al., 2020 (link)).
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