Collection and morphological sorting of macrofauna
The macrofauna grab collected at each station was transferred to a sealed plastic bin and transported to an onshore sieving station. A portion of each sediment sample was transferred to a 1 mm mesh sieve table and washed gently with running water. Once the sediment was washed through the sieve, macrofauna were gently removed with tweezers and transferred to a 500 ml container containing 95% ethanol. This process was carried out until the entire grab sample from each station was sieved. In cases where excess debris (e.g. ’shellhash’ or rocks) remained on the sieve that might influence the search for macrofauna, this debris was collected into containers with ethanol for further sorting in the lab.
Macrofauna specimens from each grab were sorted into morphospecies. We only analyzed vermiform macrofauna (e.g. Polychaeta), which comprised the majority of total macrofauna abundance across the aquaculture sites. All individuals of each morphospecies from each grab were separated for counting and downstream DNA barcoding. This was done by one person to maximize consistency. Morphospecies were photographed and defined when first encountered to allow recognition across all farms and grabs. Morphospecies were defined with the aid of taxonomic references (Banse & Hobson, 1974; Dutch et al., 2014; Hobson & Banse, 1981) and/or were identified by Leslie Harris from the Los Angeles County Museum via specimen images uploaded to the iNaturalist platform (https://www.inaturalist.org/). After initial sorting of all stations, each morphospecies was re-examined across all farms and stations to confirm consistency of morphospecies assignment. Generally, one or two specimens from each morphospecies at each station were picked for DNA barcoding; however, in cases where morphospecies delineation was less clear, additional specimens were picked for genetic confirmation.