Pelagic fish camouflage in shallow waters from the humboldt current system through intracellular structures and reflectance mechanisms

Pelagic fish have evolved specialized biogenic multilayer reflectors composed of stacks of intracellular anhydrous guanine crystals separated by cytoplasm, giving notorious silvery appearance to their skin. While the reflective properties of guanine crystals and their utility for fish camouflage have been shown in other fish species, this is the first evaluation on fish species from the southern hemisphere, and from the Humboldt current system. This is one of the most productive systems on earth, having particular oceanographic conditions such as upwelling, and thus under strong selection pressures. In this study, we conducted a comparative analysis of four pelagic species, Sardine, Anchovy, and Snoek, known for their silvery characteristics, and Mote sculpin, which lacks silvery features. We aimed to explore the biological mechanisms underlying light reflectivity in fish species and to understand how fish skin microstructures affect whole fish light reflectance and intensity in the visible spectrum. We measured the reflectance of individual fish using hyperspectral imaging and characterized the guanine crystal/cytoplasm layers within the skin of each fish using high-resolution scanning electron microscopy. These Scanning Electron Microscopy (SEM) images were analyzed using the 2D discrete Fourier transform to extract the spatial patterns that govern the light interaction with the guanine crystal structures. A novel spatial frequency analysis approach applied to SEM images explained reflectance differences between species with similar spectral behavior. Furthermore, this study presents the first fish classifiers based on the analysis of spatial frequency features, achieving up to 92.14% accuracy using a K-Nearest Neighbors classifier, highlighting the functional and taxonomic relevance of guanine microstructure organization. Our findings confirm, on pelagic fish species from the Humboldt current system, that silvery species have a chaotic distribution/arrangement of guanine crystals, whereas non-silvery species have a more organized arrangement. Accordingly, Fourier analysis indicated that silvery fish are capable of scattering light uniformly across the visible spectrum. In contrast, the Mote sculpin shows a stronger scattering of red light, distinguishing it from silvery fish.