Insights into the unique torpor of Botrylloides leachi, a colonial urochordate

Abstract

Rough environmental conditions make the survival of many multi-cellular organisms almost impossible, enforcing behavioral, morphological, physiological and reproductive rejoinders that can cope with harsh times and hostile environments, frequently through down-regulation of metabolism into basal states of dormancy, or torpor. This study examines one of the most unique torpor strategies seen within the phylum Chordata, exhibited by the colonial urochordate Botrylloides leachi, which enters a state of hibernation or aestivation in response to thermal stress, during which all of its functional colonial units (zooids) are entirely absorbed and the colony survives as small remnants of the vasculature, lacking both feeding and reproduction organs. Tissue vestiges then regenerate fully functional colony when re-exposed to milder environmental conditions. The whole metamorphic cycle of hibernation and arousal was studied here and divided into seven major stages, during which the anatomical characteristics of the zooids, the blood cell populations and the expression patterns of some "stem cell" markers were monitored. The first two phases are associated with the shortening of the blastogenic cycles from the typical 7-day cycle to 3-5day long cycles and with the significant diminution of zooids, leaving a carpet of vasculature. During hibernation this colonial carpet is made of a twisted, opaque and condensed mass of vasculature, loaded with condensed masses of blood cells that possess two types of multicellular structures, the 20-50µm "morula-like" opaque balls of cells, and small single-layer epithelial spheres, "blastula-like" structures (50-80µm). Arousal from hibernation starts with the emergence of several clear tunic areas among the vasculature lacunae, which then turn into transparent buds that become progressively larger and opaque. This is followed by sluggish, newfangled cell movement within the vasculature, which increases in intensity and rate over time. A closer examination of the vasculature revealed dramatic vicissitudes in the blood cell constituency as hibernation progressed, which is manifested by the appearance of two novel cell types not recorded in regular colonies, the multinucleate cells (MNC) and storage cells, each with 2-3 distinct cell morphs. Using mixtures of pre-labeled where half stained with a florescent marker for membranes and half stained for DNA we recorded within 2-3 days from onset new MNC stained by both staining, attesting for the de novo formation of MNC through cells fusion. At the outset of hibernation we documented high expression levels of PIWI, PL-10 and PCNA in cells residing in cell islands (CIs), which are the specific stem cell niches found along the endostyle at the ventral side of the zooids. During hibernation, most of the PIWI/ PL-10/PCNA cells were the MNCs, now located in the newly shaped and dilated vasculature, where they increased in numbers. Also, most of the PCNA cells were identified as MNCs. We further documented that the Bl-PIWI RNA (in situ hybridization) an...

Artificial seawater based long-term culture of colonial ascidians

“…We repeatedly observed that colonies isolated in smaller tanks will present signs of stress and enter into hibernation within a few days, as observed following environmental stresses like low temperatures (Hyams et al, 2017). Based on the communicated crucial role of inoculating artificial seawater with a suitable microbiota (Oliveri, personal communication, unpublished), we decided to investigate the role of bacteria for the survival of Botrylloides in ASW.…”

Colonial Life-history: A Major Evolutionary Transition, Involving Modularization of Multicellular Individuals and Heterochrony (Miniaturization and Adultation)

“…This system is essential during tissue repair, as it transports factors and cells required for regeneration (Brown et al, 2009a. Additionally, in case of adverse climatic conditions, colonies enter a dormancy stage through the resorption of all zooids, leaving only blood vessels (Hyams et al, 2017). The blood vessel system in Symplegma + Botryllids colonies maintains physiological cohesion and allows communication between zooids.…”

Evolution of colonial life history in styelids tunicates involves changes in complexity patterns

“…Blood vessels systems maintain cohesion and homeostasis of the colony. Vessels have the capacity to rearrange in case of external disturbance (Gutierrez & Brown, 2017) ; or by the dormancy at cold season, by the resorption of all zooids maintaining only the blood vessel system (Hyams et., 2017). The plasticity of the blood vessels system is decisive in the capacity of self-organization of the colonies.…”