Transient shifts in bacterial communities associated with the temperate gorgonian Paramuricea clavata in the Northwestern Mediterranean Sea

Abstract

Background: Bacterial communities that are associated with tropical reef-forming corals are being increasingly recognized for their role in host physiology and health. However, little is known about the microbial diversity of the communities associated with temperate gorgonian corals, even though these communities are key structural components of the ecosystem. In the Northwestern Mediterranean Sea, gorgonians undergo recurrent mass mortalities, but the potential relationship between these events and the structure of the associated bacterial communities remains unexplored. Because microbial assemblages may contribute to the overall health and disease resistance of their host, a detailed baseline of the associated bacterial diversity is required to better understand the functioning of the gorgonian holobiont.

Methodology/principal findings: The bacterial diversity associated with the gorgonian Paramuricea clavata was determined using denaturing gradient gel electrophoresis, terminal-restriction fragment length polymorphism and the construction of clone libraries of the bacterial 16S ribosomal DNA. Three study sites were monitored for 4 years to assess the variability of communities associated with healthy colonies. Bacterial assemblages were highly dominated by one Hahellaceae-related ribotype and exhibited low diversity. While this pattern was mostly conserved through space and time, in summer 2007, a deep shift in microbiota structure toward increased bacterial diversity and the transient disappearance of Hahellaceae was observed.

Conclusion/significance: This is the first spatiotemporal study to investigate the bacterial diversity associated with a temperate shallow gorgonian. Our data revealed an established relationship between P. clavata and a specific bacterial group within the Oceanospirillales. These results suggest a potential symbiotic role of Hahellaceae in the host-microbe association, as recently suggested for tropical corals. However, a transient imbalance in bacterial associations can be tolerated by the holobiont without apparent symptoms of disease. The subsequent restoration of the Hahellaceae-dominated community is indicative of the specificity and resilience of the bacteria associated with the gorgonian host.

Figure 1. Sampling sites.

Map of the Northwestern Mediterranean basin (A) and an enlargement showing the location of the study sites (B).

Figure 2. Representative denaturing gradient gel electrophoresis (DGGE) profiles of the bacterial community composition.

DGGE analysis of bacterial communities associated with triplicate samples (A, B and C) of 3 P. clavata colonies (Pcla1 to Pcla3) and the surrounding seawater sampled in Riou in winter 2007. 16S rDNA was amplified with different sets of primers: 27F-GC/536R (lanes 1 to 10), EUB f933-GC/EUB r1387 (lane 11), 341F-GC/907RA (lane 12) and 341F-GC/907R (lane 13). The sequenced DGGE bands are indicated with a star.

Figure 3. Relative abundances of CfoI TRFs in P. clavata samples analyzed by T-RFLP.

Each row represents a TRF generated by CfoI digestion of PCR amplicons from samples that were collected from the different sites (Riou (Ri), Medes (Me) and Scandola (Sc)) during the seasons indicated in columns. TRFs were designated by their size (in bp) after binning. The filling colors represent the relative fluorescence of the TRFs according to the range presented in the color key.

Figure 4. Relative abundances of MspI TRFs in P. clavata samples analyzed by T-RFLP.

As described in Figure 3, with the restriction enzyme MspI.

Figure 5. MDS ordination plot of P. clavata bacterial communities.

2-D scatter plots of T-RFLP profiles from P. clavata colonies sampled at 3 sites in winter and summer from 2007 to 2010 are based on the Dice similarity matrix for the T-RFLP data retrieved from the amplified bacterial 16S rDNA digested with CfoI (A) and MspI (B). Each symbol represents the bacterial community of an individual sample from Riou (square), Medes (triangle) or Scandola (circle), and the colors correspond to different sampling seasons. Sample clusters are based on coordinates determined with the k-means method. Dotted ellipses contain 50% (grey dots) or 95% (dark dots) of the points that contribute to the cluster. The centers of the confidence ellipses are identified with a cross.

Figure 6. Bacterial diversity of clone libraries derived from P. clavata colonies.

Pie charts illustrating the composition of bacterial communities in the winter 2007 and winter 2008 samples from Riou (A) and the summer 2007 samples from Riou (B), Medes (C) and Scandola (D). The large pies represent the clone library composition at the family level (A) and at the phyla or class levels (B–D). The smaller pies in B–D represent the affiliations of Actinobacteria and Firmicutes sequences at the genus level. The relative abundances of the bacterial groups retrieved from the libraries are indicated as percentages.