The freshwater sponge Ephydatia fluviatilis harbours diverse Pseudomonas species (Gammaproteobacteria, Pseudomonadales) with broad-spectrum antimicrobial activity.

Tina Keller-Costa,Leo Van Overbeek,Rodrigo Costa,Jan Dirk Van Elsas,Alexandre Jousset,Melanie R Mormile

doi:10.1371/journal.pone.0088429

Abstract

Bacteria are believed to play an important role in the fitness and biochemistry of sponges (Porifera). Pseudomonas species (Gammaproteobacteria, Pseudomonadales) are capable of colonizing a broad range of eukaryotic hosts, but knowledge of their diversity and function in freshwater invertebrates is rudimentary. We assessed the diversity, structure and antimicrobial activities of Pseudomonas spp. in the freshwater sponge Ephydatia fluviatilis. Polymerase Chain Reaction – Denaturing Gradient Gel Electrophoresis (PCR-DGGE) fingerprints of the global regulator gene gacA revealed distinct structures between sponge-associated and free-living Pseudomonas communities, unveiling previously unsuspected diversity of these assemblages in freshwater. Community structures varied across E. fluviatilis specimens, yet specific gacA phylotypes could be detected by PCR-DGGE in almost all sponge individuals sampled over two consecutive years. By means of whole-genome fingerprinting, 39 distinct genotypes were found within 90 fluorescent Pseudomonas isolates retrieved from E. fluviatilis. High frequency of in vitro antibacterial (49%), antiprotozoan (35%) and anti-oomycetal (32%) activities was found among these isolates, contrasting less-pronounced basidiomycetal (17%) and ascomycetal (8%) antagonism. Culture extracts of highly predation-resistant isolates rapidly caused complete immobility or lysis of cells of the protozoan Colpoda steinii. Isolates tentatively identified as P. jessenii, P. protegens and P. oryzihabitans showed conspicuous inhibitory traits and correspondence with dominant sponge-associated phylotypes registered by cultivation-independent analysis. Our findings suggest that E. fluviatilis hosts both transient and persistent Pseudomonas symbionts displaying antimicrobial activities of potential ecological and biotechnological value.

Highlights

Sponges (Porifera) are sessile filter-feeding organisms that primarily lack evasive or behavioural defence strategies [1]
Commensal bacterial communities may fulfil important services required for the survival of their freshwater sponge host
In contrast with 16S rRNA gene analyses, the relative abundance of some Pseudomonas gacA bands from bulk water was enhanced in E. fluviatilis while several other bulk water gacA bands could not be readily detected in the sponge samples, resulting in reduced gacA band diversity in the latter PCR-Denaturing gradient gel electrophoresis (DGGE) fingerprints in comparison with the former (Fig. 1c)

Summary

Introduction

Sponges (Porifera) are sessile filter-feeding organisms that primarily lack evasive or behavioural defence strategies [1]. There is increasing evidence that bacterial symbionts are the actual producers of many spongederived antagonistic metabolites [6,7,8,9,10], and this aspect has triggered much research interest in the diversity and bioactive potential of bacteria from marine sponges [9,11,12]. Knowledge of microbial communities in freshwater sponges remains limited. Their ubiquity in continental water bodies [13], coupled to recent molecular findings on highly selected communities and specific lineages of bacteria that inhabit them [14] make freshwater sponges valuable models in symbiosis research. Commensal bacterial communities may fulfil important services required for the survival of their freshwater sponge host

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Conclusion