Autofluorescence in embryos and larvae of the giant clam Tridacna noae: challenges and opportunities for epifluorescence microscopy

Abstract

Epifluorescence microscopy has numerous applications in molluscan research. Methods rely on distinguishing target fluorescent signals from natural fluorescence, termed autofluorescence, of a specimen. Limited research has been conducted on the autofluorescence of marine bivalves, which hinders development of epifluorescence microscopy techniques in this sector. We examined autofluorescence in larvae of the giant clam Tridacna noae to identify challenges and opportunities in applying epifluorescence microscopy to the study of giant clams. Under blue-light excitation (lambda(ex) = 460-490 nm) three sources of autofluorescence were identified in larvae of T. noae: (1) green autofluorescence of the gut region; (2) green autofluorescence of the 'dark bodies' anatomical structure; and (3) red autofluorescence of ingested zooxanthellae, Symbiodinium species. Each of these sources of autofluorescence varied in prevalence and relative intensity over the course of larval development. The multiple sources and varied wavelengths of the observed autofluorescence presents challenges for applying fluorescent labelling techniques. However, our study identifies epifluorescence microscopy as a novel means of examining larval ontogeny and the uptake of zooxanthellae in giant clams.