Biofluorescence in decapods has been reported for several taxa. However, few studies have carried out a detailed examination under controlled conditions to describe fluorescence patterns and how it varies between individuals and under different conditions. Red king crabs Paralithodes camtschaticus ( RKC) are an introduced species to Norway in the 1960s (Lorentzen et. al. 2018) that are routinely kept in long-term holding facilities prior to live shipping (Mota et. al., 2021). The aim of this study was to evaluate whether P. camtchaticus biofluoresces and to document the dynamics of fluorescence in the species under controlled conditions . The specific objectives for this study are to determine whether detectable changes in fluorescence levels occur in either the hemolymph or exoskeleton of P. camtchaticus.
The study of biofluorescence in male RKC was divided into two groups: hemolymph and exoskeleton. RKC (N=19) were sampled for hemolymph before and after they were transported and stored overnight in 20-liter styrofoam boxes. Each RKC had ~1. 5 mL of hemolymph sampled and analyzed with a Duetta fluorescence and absorbance spectrometer . Fluorescence emissions were recorded in the EZ Spec Software . Each sample was recorded with excitation wavelengths from 250 to 350 nm with a 5 nm interval and emission wavelengths from 400 to 800 nm with a 0.5 nm spectral resolution.
The exoskeleton analysis scanned RKC (N=10 juvenile, 8 adults ) on a conveyor belt illuminated with ultraviolet (~ 400 nm) and royal blue (~445 nm) LED lighting . After hyperspectral scanning, the crabs were placed overnight for post shipping recovery in an aerated 1000-liter holding tank at 6⁰ C overnight . All hyperspectral images were taken with the HySpex VNIR-1800 hyperspectral camera . Raw data files collected with hyperspectral scanning were radiometrically calibrated with HySpex Rad v2.5 to produce radiance values in units of . The radiometrically calibrated data was uploaded for analysis in Breeze hyperspectral imaging software and inspected visually for fluorescence signals.
Biofluorescence was recorded in both the exoskeleton and the hemolymph in the species . We found that RKC show great variability in biofluorescence and that most displayed a green biofluorescence in their exoskeleton (~500 nm), with the greatest intensity occurring in the eye stalks and the cervical groove of the carapace. The arthrodial joints between the limbs fluoresced in red (~680 nm). Hyperspectral analysis detected both green and red fluorescence in juvenile (N=10) and adult (N=8) crabs. A decrease in fluorescence occurred in the eye stalks after a post transportation recovery period while the cervical groove remained constant (Figure 1B). After applying a live transport simulation to a second RKC group (n=19), we found a significant increase (Anova ; p = 0.008 ) in the hemolymph fluorescence of animals tested before and after out of water transport (Figure 1A) .
As both physiological parameters such as stress hormones and tools available for empirically documenting physiological changes in crabs remain less advanced compared to higher order aquaculture species, it is difficult to ascertain whether this change in fluorescence can be categorized as an indicator for specific levels of stress. A recent study exposing adult RKC to low oxygen levels documented anoxic stress in hemolymph through an increased concentration of lactic acid (Mota et. al., 2021). Imagery-based estimates of changing fluorescence , like the one demonstrated in this study, should be linked with such empirical approaches in future studies to further explore the utility of non-invasive techniques for assessing RKC welfare. These findings indicate that biofluorescence in RKC is dynamic and further investigation may reveal whether such fluorescence could be used to monitor the welfare of decapods in captivity.
Lorentzen, G., Voldnes, G., Whitaker, R. D., Kvalvik, I., Vang, B., Gjerp Solstad, R., ... & Siikavuopio , S. I. (2018). Current status of the red king crab (Paralithodes camtchaticus) and snow crab (Chionoecetes opilio) industries in Norway. Reviews in Fisheries Science & Aquaculture, 26(1), 42-54.
Mota , V. C., Siikavuopio , S. I., & James, P. (2021). Physiological responses to live air transport of red king crab (Paralithodes camtschaticus). Fisheries Research, 237, 105882.
Funding: DeepVision project (Nofima, Norway), EATFISH (H2020 #956697).