X-rays and invisible sand: two new methods for designing burrowing behavioral experiments with juvenile unionoids

Unionoids are in global decline, which may be associated with their complex life cycle. Their juveniles are unique because while hidden (burrowed deeply in bottom sediments) they undergo critical anatomical changes (also developing a characteristic juvenile shell sculpture). Currently, the juveniles’ period of life is believed to be both the least known and one of the most vulnerable—thus the possibility of obtaining any biological knowledge is essential for establishing conservation strategies and addressing functional or evolutionary questions. I propose two new methods for visualization of the burrowing behavior of unionoid juveniles within deposits that are cheap and easy: (1) laminated deposits of quartz–aragonite sand for time-stepped X-ray images of bivalve traces, and (2) silica gel serving as 'invisible sand' for direct observations and video recording of behavior within sediments. Both deposits in a pilot study were accepted by the juvenile unionoids—they were stable enough and penetrable, with no observable signs of harmful effects on animals’ behavior during trials. In both, juveniles were clearly visible, settled within the top 1 cm layer of deposits. Both methods are promising tools for future in situ within the deposits research on the biology of this much unexplored and vulnerable unionoids' life stage.