Transcriptomic Analysis and Time to Hatch Visual Prediction of Embryo Development in the Ornate Spiny Lobster (Panulirus ornatus)

Abstract

The ability to conduct closed life-cycle culture of tropical spiny lobsters, Panulirus ornatus, at the Institute for Marine and Antarctic Studies (IMAS) provides a unique opportunity to investigate specific developmental stages during embryogenesis. The production of closed life-cycle lobsters provides access to embryos at defined time points, yet physical staging is challenging due to their small size and environmental factors impacting their development. A transcriptome comprising 11 distinct stages across the 30-day P. ornatus embryonal development period allowed the establishment of the most detailed transcriptomic library of embryogenesis across decapods. A series of key genes across the 11 stages of embryonal development were characterized. The expression of neuropeptides was reported across P. ornatus embryos, suggesting they are primarily active during the later stages of embryogenesis as the nervous system develops and the animal prepares to hatch. Gastrula-specific genes, nanos and brachyury, presented an expression profile indicating gastrulation occurs early in embryogenesis. In addition to the molecular tools used to characterize embryo development, certain developmental characteristics, such as eye spot development, provide a measurable indicator that can be visualised. Hatch prediction models based on visual characteristics were shown to be an accurate method to predict the timing of the hatch for P. ornatus embryos kept at 26°C. The combination of morphological and visual measures with transcriptomics can be used to further define and establish the groundwork for future characterisation and staging of P. ornatus embryogenesis.

Ontogeny of the Cytochrome P450 Superfamily in the Ornate Spiny Lobster (Panulirus ornatus)

“…The CYP450 transcripts' gene expression was mapped across the extensive transcriptomic dataset established for P. ornatus at the University of the Sunshine Coast, and available on CrustyBase [48], encompassing 11 stages of embryogenesis [32], 12 stages of larval development [33], the hepatopancreas across 6 moult stages in juveniles (generated in Chapter 4), and tissues from adult P. ornatus [34]. All 25 CYP450s detected in embryo development show distinct differential expression, with minimal overlap in the genes expressed on each day of embryogenesis.…”

“…Ten of the CYP450s were expressed only in day 0 embryos, including CYP302A1d (a Disembodied paralog), Spook, and CYP3A2-like. It is possible that transcripts with expression specific to day 0 of embryogenesis are linked with initialising the embryonic development and cell division as the day 0 embryos were collected at the 4-8 cell stages [32]. Then there is a group of eight different CYP450s with expression from day 3 to day 9, including Shed B and CYP302A1a.…”

“…Then there is a group of eight different CYP450s with expression from day 3 to day 9, including Shed B and CYP302A1a. The genes with high expression on these days are likely to be implicated in processes such as gastrulation, where the germ layers appear, which occurs on day three [32], followed by segmentation and formation of the nervous system and organs [49]. This is followed by a set of eight different CYP450s expressed from day 12 to day 24, which include CYP2L1-like, CYP18A1, and Shed C. Both CYP18A1 and Shed C are essential for the conversion of 20HE [41], and their expression across the mid-stages of embryogenesis suggests 20HE may be implicated in the significant structural changes taking place, following the segmentation and establishing the body structures.…”

“…There are several established transcriptomic libraries currently available for P. ornatus, detailing the complex life stages and tissues: an embryo development transcriptome of eleven stages during the 30-day embryo development [32]; a twelve-stage transcriptome describing the metamorphic life stages of phyllosoma, puerulus and juvenile [33]; hepatopancreas from juvenile P. ornatus across six stages of a complete moult cycle (Chapter 4); and multiple tissues derived from immature and adult P. ornatus from males and females [34]. The tissues used include eyestalk, brain, thoracic ganglia, antennal gland, testis, sperm duct, ovary, oviduct, hepatopancreas, gills, heart, stomach, intestine, muscle, epidermis, fat and hemocytes.…”

“…A comprehensive transcriptome is available for P. ornatus, detailing samples obtained from eleven embryo stages [32], twelve distinct metamorphic stages ranging across the lobster life cycle from actively predating phyllosoma, to the non-feeding puerulus, to the predating juvenile [33], and 52 additional tissue samples obtained for 17 tissues from adult and juvenile P. ornatus [34]. This is a valuable resource that can be used for the identification of cytochrome P450s within the species across the life stages and particularly how they change in expression across metamorphosis and the tissues.…”