Bacterial Exposure Mediates Developmental Plasticity and Resistance to Lethal Vibrio lentus Infection in Purple Sea Urchin (Strongylocentrotus purpuratus) Larvae

Abstract

Exposure to and colonization by bacteria during development have wide-ranging beneficial effects on animal biology but can also inhibit growth or cause disease. The immune system is the prime mediator of these microbial interactions and is itself shaped by them. Studies using diverse animal taxa have begun to elucidate the mechanisms underlying the acquisition and transmission of bacterial symbionts and their interactions with developing immune systems. Moreover, the contexts of these associations are often confounded by stark differences between "wild type" microbiota and the bacterial communities associated with animals raised in conventional or germ-free laboratories. In this study, we investigate the spatio-temporal kinetics of bacterial colonization and associated effects on growth and immune function in larvae of the purple sea urchin (Strongylocentrotus purpuratus) as a model for host-microbe interactions and immune system development. We also compare the host-associated microbiota of developing embryos and larvae raised in natural seawater or exposed to adult-associated bacteria in the laboratory. Bacteria associated with zygotes, embryos, and early larvae are detectable with 16S amplicon sequencing, but 16S-FISH indicates that the vast majority of larval bacterial load is acquired after feeding begins and is localized to the gut lumen. The bacterial communities of laboratory-cultured embryos are significantly less diverse than the natural microbiota but recapitulate its major components (Alphaproteobacteria, Gammaproteobacteria, and Bacteroidetes), suggesting that biologically relevant host-microbe interactions can be studied in the laboratory. We also demonstrate that bacterial exposure in early development induces changes in morphology and in the immune system. In the absence of bacteria, larvae grow larger at the 4-arm stage. Additionally, Schuh et al.Bacteria-Mediated Developmental Plasticity bacteria-exposed larvae are significantly more resistant to lethal infection with the larva-associated pathogen Vibrio lentus suggesting that early exposure to high levels of microbes, as would be expected in natural conditions, affects the immune state in later larvae. These results expand our knowledge of microbial influences on early sea urchin development and establish a model in which to study the interactions between the developing larval immune system and the acquisition of larval microbiota.

Symbiotic Life of Echinoderm Larvae

“…Shortly thereafter and on multiple occasions since these original observations, studies have identified subcuticle bacteria in the developmental stages of echinoderms. These symbionts, thus far, have been found in three asteroids (Cameron and Holland, 1983;Bosch, 1992;Cerra et al, 1997), one ophiuroid (Walker and Lesser, 1989), and one echinoid (Heyland et al, 2018;Schuh et al, 2019) (Figure 1; Table 1). In these five species, subcuticle bacteria have been observed within the mouth and gut lumen, out-pockets of the extracellular matrix that surrounds the larval body, embedded in the inner layer of the secondary cuticle of the rudiment epidermis, and are engulfed and, in some cases, digested by epidermal cells (Cameron and Holland, 1983;Walker and Lesser, 1989;Bosch, 1992;Cerra et al, 1997;Heyland et al, 2018;Schuh et al, 2019).…”

“…Like many other studies of animal and plant microbes, the bacterial communities associated with echinoderm larvae are species-specific and taxonomically distinct from the environmental microbiota (Galac et al, 2016;Carrier and Reitzel, 2018;Carrier et al, 2018b), suggesting that these communities are, at least in part, selected by the host. This host-specificity, however, appears to be lost when larvae are cultured under traditional laboratory settings for rearing the developmental stages of marine invertebrates (i.e., fine-filtered or artificial saltwater) (Schuh et al, 2019). Specifically, Strongylocentrotus purpuratus larvae cultured under traditional laboratory settings associate with bacterial communities that are less diverse in total taxa and the phylogenetic breadth of those taxa, and retain ∼40% of the OTUs harbored by "wild-type" counterparts (Schuh et al, 2019).…”

“…This host-specificity, however, appears to be lost when larvae are cultured under traditional laboratory settings for rearing the developmental stages of marine invertebrates (i.e., fine-filtered or artificial saltwater) (Schuh et al, 2019). Specifically, Strongylocentrotus purpuratus larvae cultured under traditional laboratory settings associate with bacterial communities that are less diverse in total taxa and the phylogenetic breadth of those taxa, and retain ∼40% of the OTUs harbored by "wild-type" counterparts (Schuh et al, 2019). This implies that studying larval-associated bacterial communities is most accurately performed at near-natural conditions, such as by filtering ambient seawater to 5-µm to remove most debris and planktonic predators while retaining the environmental microbiota (Carrier and Reitzel, 2018;Hodin et al, 2019).…”

“…Larval-associated bacterial communities are variable in community membership and composition but exhibit nonrandom shifts that correlate with multiple components of larval biology and ecology. These communities, for example, are established on unfertilized eggs but not the sperm of sea urchins (Carrier and Reitzel, 2019b;Schuh et al, 2019). When cultured using coarsely (5-µm) filtered seawater, echinoderm larvae exhibit a development-based succession in symbiont composition and, using fluorescent in situ hybridization, these bacteria localize in the mouth and gut lumen (Carrier and Reitzel, 2019b;Schuh et al, 2019).…”

“…These communities, for example, are established on unfertilized eggs but not the sperm of sea urchins (Carrier and Reitzel, 2019b;Schuh et al, 2019). When cultured using coarsely (5-µm) filtered seawater, echinoderm larvae exhibit a development-based succession in symbiont composition and, using fluorescent in situ hybridization, these bacteria localize in the mouth and gut lumen (Carrier and Reitzel, 2019b;Schuh et al, 2019). Following fertilization, the diversity of these communities increases by ∼20% during the early embryonic stages and decreased by nearly ∼85% following hatching and through metamorphosis (Carrier and Reitzel, 2019b).…”