X-RAY TOMOGRAPHIC MICROSCOPY AS A MEANS TO SYSTEMATICALLY TRACK EXPERIMENTAL DECAY AND FOSSILIZATION

ABSTRACT Laboratory-based decay experiments have become commonly used to supplement our understanding of how organisms enter the fossil record. Differences in how these experiments are designed and evaluated, however, including dissimilarities in qualitative decay-scoring indices superimposed on variability in model organisms, renders any semblance of comparison between studies unreliable. Here, we introduce the utility of X-ray tomographic microscopy (μCT) as a means for reliable and repeatable analysis of soft-tissue decay experiment products. As proof-of-concept, we used a relatively simple experimental design with classic studies as comparators, and present our analytical protocol using μCT for capturing the entire volume of the decay subject. Segmentation software then allows for 3D volume analysis and high-resolution internal and external character identification. We describe the workflow from sample preparation, contrast-staining, and data collection to processing and analysis of the resulting data, using peppermint shrimp (Lysmata wurdemanni) as model organisms, and compare our results to previous taphonomic studies. These methods allow for improved visualization and quantification of decay and internal volume analysis with minimal handling as compared to traditional qualitative scoring methods. Using the same scoring criteria as previous studies, this study revealed similar decay results for certain features, while we were additionally able to detect other feature loss or alteration earlier—importantly without need for potentially distortive sample handling. We conclude that μCT is a more effective, straightforward, and exact means for extracting quantitative data on the progression of decay and should be adopted in future studies, where available, to streamline and standardize comparisons.

Uridine-5’-tri-phosphate is a candidate component of the soluble sex pheromone bouquet in a marine shrimp, Lysmata wurdemanni

Characterisation of distance sex pheromones in decapods is challenging, although great efforts have been made in this field in the past 50 yr. In a previous study, we identified a component of the distance (soluble) sex pheromone bouquet of the peppermint shrimp Lysmata wurdemanni as being a uridine-5’-di-phosphate (UDP)-like chemical. However, UDP itself does not elicit the full pre-copulatory ‘approach and follow’ behaviour in peppermint shrimp. Here, we tested the hypothesis that the UDP-like chemical is uridine-5’-tri-phosphate (UTP), a metabolic product of chitin synthesis, and that this is the component of the distance sex pheromone of L. wurdemanni. We ran a series of bioassays to examine whether UDP, UTP or mixtures of the 2 compounds elicit male mating behaviour. Our results showed that male L. wurdemanni responded to UTP by displaying stereotyped courtship behaviour—the same behaviour elicited from males when exposed to water that previously contained moulting females. Combining UTP and UDP did not enhance the intensity of this courtship behaviour. The minimum effective concentration of UTP found to elicit courtship behaviour in the male shrimp was between 10-6 and 10-7 M. HPLC analysis showed the presence of UTP in the moulting water of female shrimp and also the partial conversion of UTP to UDP during the sample preparation procedure. Both the bioassay and chemical analysis results presented in this study suggest that UTP is a component of the distance sex pheromone in L. wurdemanni, and that the major peak in the chromatogram of a L. wurdemanni pheromone identified in previous studies might be a breakdown product of UTP.