The Life Cycle of Aurelia aurita Depends on the Presence of a Microbiome in Polyps Prior to Onset of Strobilation

Abstract

Aurelia aurita's intricate life cycle alternates between benthic polyp and pelagic medusa stages. The strobilation process, a critical asexual reproduction mechanism in this jellyfish, is severely compromised in the absence of the natural polyp microbiome, with limited production and release of ephyrae. Yet, the recolonization of sterile polyps with a native polyp microbiome can correct this defect. Here, we investigated the precise timing necessary for recolonization as well as the host-associated molecular processes involved. We deciphered that a natural microbiota had to be present in polyps prior to the onset of strobilation to ensure normal asexual reproduction and a successful polyp-to-medusa transition. Providing the native microbiota to sterile polyps after the onset of strobilation failed to restore the normal strobilation process. The absence of a microbiome was associated with decreased transcription of developmental and strobilation genes as monitored by reverse transcription-quantitative PCR. Transcription of these genes was exclusively observed for native polyps and sterile polyps that were recolonized before the initiation of strobilation. We further propose that direct cell contact between the host and its associated bacteria is required for the normal production of offspring. Overall, our findings indicate that the presence of a native microbiome at the polyp stage prior to the onset of strobilation is essential to ensure a normal polyp-to-medusa transition. IMPORTANCE All multicellular organisms are associated with microorganisms that play fundamental roles in the health and fitness of the host. Notably, the native microbiome of the Cnidarian Aurelia aurita is crucial for the asexual reproduction by strobilation. Sterile polyps display malformed strobilae and a halt of ephyrae release, which is restored by recolonizing sterile polyps with a native microbiota. Despite that, little is known about the microbial impact on the strobilation process's timing and molecular consequences. The present study shows that A. aurita's life cycle depends on the presence of the native microbiome at the polyp stage prior to the onset of strobilation to ensure the polyp-to-medusa transition. Moreover, sterile individuals correlate with reduced transcription levels of developmental and strobilation genes, evidencing the microbiome's impact on strobilation on the molecular level. Transcription of strobilation genes was exclusively detected in native polyps and those recolonized before initiating strobilation, suggesting microbiota-dependent gene regulation.

Keywords: Aurelia aurita; asexual reproduction; life cycle; native microbiota; recolonization; reproduction; strobilation.

FIG 1

Life cycle of Aurelia aurita and study design for recolonization at various developmental stages. (A) Summary of the life cycle of the moon jellyfish Aurelia aurita. Sexual reproduction of the matured medusa results in the release of planula larvae that develop into benthic polyps. Asexual reproduction includes the transition into early and then late strobila, forming bodily segments to release precursor medusa in the form of ephyrae. This offspring generation is impaired in the absence of a microbiota. Sterile animals showed malformations characterized by a pale color, slim body shape, and a lack of tentacles represented in a gray color of early and late strobila (11). (B) Experimental design to identify a permissible time point(s) of recolonization. Native polyps are induced with the chemical inducer for the onset of strobilation (native conditions [NC]), which was removed by washing after 72 h. Similarly, sterile polyps were induced, resulting in impaired strobilation (sterile conditions [SC]). Sterile polyps were recolonized prior to induction (R1). Developed sterile strobila stages (light gray images) were recolonized after washing off the inducer (R2, R3); in a second approach, the inducer was again added when recolonization was performed (constant presence of inducer; R2i, R3i).

FIG 2

Analysis of the microbial community of Aurelia aurita. (A) ASV abundance based on the V1-V2 region of the 16S rRNA gene of the bacterial communities derived from native polyps, the inoculum used for recolonization, obtained microbiota from recolonized sterile polyps, and a control of the inoculum incubated in ASW are presented at the genus level and normalized by the total number of reads per sample. Bar plots represent averages of six replicates per treatment. The threshold for the represented taxa is >1% relative abundance; consequently, taxa with a mean relative abundance of <1% across all samples are collectively reported as “Others.” In the electronic version, a scroll over the colored box and a right mouse click will show the genus. (B) Box plot of α-diversity (Shannon index). Significance of comparisons is shown: *, P < 0.05; **, P < 0.01.

FIG 3

Transcription of Aurelia-specific strobilation genes depends on presence of a microbiota. (A) Transcription patterns (reported as ΔC_T using EF1 for normalization) of Aurelia-specific strobilation genes CL112, CL355, CL390, and CL631 in the life stages of polyps, early strobila, late strobila, and ephyra of native (left) and sterile (right) animals. Averages of four biological replicates with three technical replicates each are reported. (B) Fold changes of transcription of selected conserved developmental genes and A. aurita-specific strobilation genes in sterile polyps compared to the corresponding native life stage.

FIG 4

Timing of recolonization is crucial to restore asexual reproduction. (A) Morphology of polyps undergoing strobilation under various conditions. Left to right: native conditions (NC), sterile conditions (SC), and recolonization of sterile polyps (R1) of sterile early strobila (R2) and of sterile late strobila (R3). Photographs show the phenotypical appearance of polyps, early strobilae, late strobilae, and ephyrae (from the top). One hundred percent of the animals of NC and R1 released ephyrae, whereas 54% of R2, 16% of R3, and 17% of SC released a reduced number of ephyrae. The number of days postinfection is specified. Scale bars, 1 mm. (B) Bar plot summarizing the number of segments per animal at late strobila stage (9 days postinduction) and of released ephyrae per animal (14 days postinduction). Significance of comparisons are shown: ****, P < 0.0001. (C) Phenotypes of sterile polyps undergoing strobilation when sterile early strobilae (R2esm) and sterile late strobilae (R3lsm) were recolonized with their life stage-specific microbiota.

FIG 5

Impact of the native microbiota on the transcription of A. aurita-specific strobilation genes during asexual reproduction. (A) Heat map of normalized C_T values (ΔC_T) of genes CL112, CL355, CL390, and CL631 using the housekeeping gene EF1 for normalization. ΔC_T values represent an average of four biological replicates with three technical replicates for each. (B) Expression profile of CL390 during development over time for the different treatments. A complete summary of all assessed genes is presented in Fig. S3 in the supplemental material.

FIG 6

Direct contact between polyp and bacteria is assumed for a normal strobilation process. Strobilation was induced with the chemical inducer that is able to pass the membrane (0.2-μm sterile filter). Scale bars, 1 mm. (A) A single native polyp was physically separated by the membrane from the strobilation inducer-ASW solution. (B) A single sterile polyp was separated by the membrane that hinders microbial cells from passing from a single polyp or a pool of 10 native polyps. Chemical inducer was present from the beginning. (C) A single sterile polyp was separated by the membrane that hinders microbial cells from passing from a single polyp or a pool of 10 native polyps. After 8 days of preincubation, strobilation was induced.