Freshwater Sponge Tubella variabilis Presents Richer Microbiota Than Marine Sponge Species

Abstract

Sponges can host diverse and abundant communities of microorganisms, which constitute an interesting source of bioactive compounds. Thus, to broaden our knowledge about the diversity of the microbiota that is found in freshwater sponges, the microbial community of Tubella variabilis was analyzed using culture-independent and culture-dependent approaches. Additionally, sponge-associated bacteria were compared with those living in the surrounding waters. Bacteria were also tested for antimicrobial production. Overall, the microbial composition identified comprises at least 44 phyla belonging mainly to Proteobacteria and low percentages of Bacteroidetes, Acidobacteria, and Verrucomicrobia. Alphaproteobacteria was the dominant class in T. variabilis while Betaproteobacteria was dominant in freshwater. Our data also revealed a high richness of bacteria in comparison to another freshwater sponge and 32 marine sponges. A global comparison of the structure of microbiota of different sponges showed that the main structuring factor may be the sponge environment, with T. variabilis and all freshwater sponges clustering together, and far away from the marine sponges. Bacterial strains from sponges and from freshwater were isolated and 163 morphotypes were phylogenetically identified. These belong to 26 genera, of which 12 were exclusively found in sponge samples and three only in freshwater. Inhibitory activities were also detected among 20-25% of the isolates from sponges and freshwater, respectively. This study presents new information on the composition of the microbial community found in freshwater sponges, which is diverse, abundant and distinct from some marine sponges. Moreover, the antimicrobial activity observed from the bacterial strains might play an important role in shaping microbial communities of the environment.

Keywords: Spongillida; bioactive compounds; culture-dependent approach; global sponge-microbiome; sponge-associated bacteria.

FIGURE 1

Relative abundance of bacterial groups found in Tubella variabilis and surrounding freshwater, obtained by 16S rRNA gene sequencing and classified using RDP database. (A) Relative abundance of bacterial phyla. (B) Relative abundance of Proteobacteria classes. (C) Relative abundance of Bacteroidetes classes.

FIGURE 2

Relative abundance of the top 30 most abundant OTUs (3% dissimilarity cutoff) found in the surrounding freshwater and T. variabilis. Each bar represents the average relative abundance of a given OTU. The dark blue mark represents the difference between the abundance of both environments. ^∗p < 0.05, ^∗∗p < 0.01 (t-test).

FIGURE 3

Global richness comparison of the microbial community of T. variabilis with two freshwater sponges and 32 marine sponges. The bars represent the average number of OTUs (n = 3 or 2 per sponge) with the standard deviation. Bright blue bars are freshwater sponges and in dark blue bars are marine sponges.

FIGURE 4

Non-metric multidimensional scaling based on OTU distribution of T. variabilis with two freshwater sponges and 32 marine sponges. Freshwater sponges are represented in blue dots and marine sponges in black dots. Ordination stress was 0.18 in the scale of 0–1.

FIGURE 5

Distribution of bacteria isolated from T. variabilis and surrounding freshwater samples. (A) Venn diagram shows the numbers of common and unique bacterial strains isolated from sponges and from freshwater. (B) Phylogenetic composition of the bacteria culturable community from the T. variabilis and freshwater samples. Stacked column bar graph depicting the absolute abundances of bacterial genera and family isolated. Bacterial isolates belonging to Actinobacteria (^∗), Bacteroidetes (¥), Firmicutes (¤), and Proteobacteria (§) are indicated.

FIGURE 6

Venn diagram that illustrates the relationship between OTUs that were detected in the T. variabilis samples by culture-independent and culture-dependent analysis. A comparison of the overlap in terms OTUs in the three different sponge samples (T. variabilis A, B, and C) and their cultivable fractions (isolates). The numbers of OTUs from and shared by each sample are represented inside the oval shapes and the main bacterial genera are indicated. Venn diagram of OTUs at distance 0.03.