A scalable culturing system for the marine annelid Platynereis dumerilii

Abstract

40Platynereis dumerilii is a marine segmented worm (annelid) with externally fertilized embryos 41 and it can be cultured for the full life cycle in the laboratory. The accessibility of embryos and 42 larvae combined with the breadth of the established molecular and functional techniques has made 43 P. dumerilii an attractive model for studying development, cell lineages, cell type evolution, 44reproduction, regeneration, the nervous system, and behavior. Traditionally, these worms have 45 been kept in rooms dedicated for their culture. This allows for the regulation of temperature and 46 light cycles, which is critical to synchronizing sexual maturation. However, regulating the 47 conditions of a whole room present limitations, especially if experiments require being able to 48 change culturing conditions. Here we present scalable and flexible culture methods that provide 49 ability to control the environmental conditions, and have a multi-purpose culture space. We 50 provide a closed setup shelving design with proper light conditions necessary for P. dumerilii to 51 mature. We also implemented a standardized method of feeding P. dumerilii cultures with 52 powdered spirulina which relieves the ambiguity associated with using frozen spinach, and helps 53 standardize nutrition conditions across experiments and across different labs. By using these 54 methods, we were able to raise mature P. dumerilii, capable of spawning and producing viable 55 embryos for experimentation and replenishing culture populations. These methods will allow for 56 the further accessibility of P. dumerilii as a model system, and they can be adapted for other aquatic 57 organisms. 58 59 Nereidid worms such as Platynereis have been popular in studies of development and 87 fertilization because of transparent, abundant, and comparatively large eggs and embryos [1,2]. As 88 researchers like Edmund Beecher Wilson did in the late 19th century, many labs today benefit 89 from Platynereis as a model organism for addressing a wide range of biological questions such as 90 cell type evolution, nervous system evo-devo and activity, reproductive periodicity, circalunar 91 cycling, endocrinology, regeneration, post-embryonic segment addition, stem cell biology, 92 fertilization, oocyte maturation, embryonic and larval development [3-16]. The sexual worms 93 broadcast spawn, producing thousands of externally-developing embryos. The embryos are large 94 enough to inject but small (about 160 µm in diameter) and transparent enough to image live and 95 fixed samples. Platynereis has a relatively quick and highly synchronized embryogenesis: it takes 96 only 18 hours from fertilization to hatching as a planktonic trochophore larva in P. dumerilii 97 [2,17]. This allows researchers to study embryonic development over the course of just one day. 98 A number of tools and techniques are already established in P. dumerilii [3] including 99 microinjection [5,12,18], transgenesis and genetic tools [18-21], single cell RNA sequencing [22], 100 behavioral tracking [8,...

Small-molecule mimicry hunting strategy in the imperial cone snail, Conus imperialis

“…S6). Thus, the feeding biology of C. imperialis is more complex than previously recorded (24)(25)(26).…”

“…Thus, a model system was required. P. dumerilii is one of the few polychaete model systems used in biological research (24), and it is consumed by worm-hunting cones (25,26). To determine whether P. dumerilii would provide a suitable model, we reinvestigated this question using a validated polymerase chain reaction (PCR) approach that has been recently applied to worm-hunting cones (26).…”

An automated aquatic rack system for rearing marine invertebrates

“…List of parts and suppliers for components used in the aquatic rack system. Note: Additional files 3,4,5,6,7,8,9,10,11,12,13,14,15,16 are available at Github [41].…”

“…f View of the IR sensor that detects the presence and absence of food in the Teflon feeding line (see Additional files 9,10,11,12). g View of the DC motor controller for the protein skimmer and a small microcontroller unit that regulates power to the skimmer (see Additional files 15,16,17,18). h High magnification view of the Tunze DC motor control board showing the points where 30-gauge wire connections are made to the microcontroller (see Additional file 15).…”

“…The major challenge for working with any marine organism, including slipper snails, is the time and effort required to maintain and rear enough animals to support an active research program. While some groups are beginning to address issues of standardizing laboratory culture for marine organisms, for example, the annelid Platynereis dumerilii [15] and the hydrozoan Clytia hemispherica [16], these systems have limited automation and require manual feeding. To address these challenges, we have developed an automated marine aquatic system that monitors and regulates seawater quality and incorporates automated feeding.…”