Analysis of bacterial composition in marine sponges reveals the influence of host phylogeny and environment

FEMS Microbiology Ecology, 93, 2017, fiw204 doi: 10.1093/femsec/fiw204 Advance Access Publication Date: 3 October 2016 Research Article RESEARCH ARTICLE Danilo T. Souza1,2 , Diego B. Genuário1 , Fabio Sérgio P. Silva1,2 , Camila C. Pansa1,2 , Vanessa N. Kavamura1 , Fernando C. Moraes3,4 , Rodrigo G. Taketani1 and Itamar S. Melo1,∗ 1 Environmental Microbiology Laboratory, Embrapa Environment, 13820-000, Jaguariúna, SP, Brazil, 2 College of Agriculture ‘Luiz de Queiroz’, University of São Paulo, 13418-900, Piracicaba, SP, Brazil, 3 Rio de Janeiro Botanical Garden Research Institute, 22460-030, Rio de Janeiro, RJ, Brazil and 4 National Museum, Federal University of Rio de Janeiro, 20940-040, Rio de Janeiro, RJ, Brazil ∗ Corresponding author: Environmental Microbiology Laboratory, Embrapa Environment, Rodovia SP 340-Km 127,5, 13820-000, Jaguariúna, SP, Brazil. Tel: +55 (19) 3311-2665; Fax: +55 (19) 3311-2640; E-mail: One sentence summary: Linking sponge phylogeny to associated bacteria. Editor: Julie Olson ABSTRACT Bacterial communities associated with sponges are influenced by environmental factors; however, some degree of genetic influence of the host on the microbiome is also expected. In this work, 16S rRNA gene amplicon sequencing revealed diverse bacterial phylotypes based on the phylogenies of three tropical sponges (Aplysina fulva, Aiolochroia crassa and Chondrosia collectrix). Despite their sympatric occurrence, the studied sponges presented different bacterial compositions that differed from those observed in seawater. However, lower dissimilarities in bacterial communities were observed within sponges from the same phylogenetic group. The relationships between operational taxonomic units (OTUs) recovered from the sponges and database sequences revealed associations among sequences from unrelated sponge species and sequences retrieved from diverse environmental samples. In addition, one Proteobacteria OTU retrieved from A. fulva was identical to sequences previously reported from A. fulva specimens collected along the Brazilian coast. Based on these results, we conclude that bacterial communities associated with marine sponges are shaped by host identity, while environmental conditions seem to be less important in shaping symbiont communities. This is the first study to assess bacterial communities associated with marine sponges in the remote St. Peter and St. Paul Archipelago using amplicon sequencing of the 16S rRNA gene. Keywords: Aplysinidae; Chondrosidae, Associated symbionts; Verongimorpha; Saint Peter and Saint Paul Archipelago; Brazil INTRODUCTION Sponges (phylum Porifera) are the oldest multicellular animals still extant on Earth. These animals evolved over 600 million years ago and currently encompass over 8600 species that inhabit different marine and freshwater ecosystems (Webster and Thomas 2016). The evolutionary success of sponges is closely linked to their capacity to maintain a rich community of Received: 14 April 2016; Accepted: 2 October 2016  C FEMS 2016. All rights reserved. For permissions, please e-mail: 1 Analysis of bacterial composition in marine sponges reveals the influence of host phylogeny and environment 2 FEMS Microbiology Ecology, 2017, Vol. 93, No. 1 verse and shaped by both host and environmental factors (Erwin et al. 2012; Schmitt et al. 2012; Pita et al. 2013b). However, few studies have compared sponge microbial communities in closely related species that coexist in close spatial proximity (Erwin et al. 2012; Hardoim et al. 2012; Webster et al. 2013a; Cuvelier et al. 2014; Naim et al. 2014). The major driving forces regulating microbial diversity and the evolutionary history of these communities remain unclear; therefore, understanding the microbial patterns that are present in closely related sponge species is of special interest (Hardoim et al. 2012; Webster and Thomas 2016). In the present survey, the phylogenies of three marine sponges (Aiolochroia crassa, Aplysina fulva and Chondrosia collectrix) that live in close spatial proximity were examined to determine the bacterial species that are specific and shared among phylogenetically related host sponges. To accomplish this, the compositions of the bacterial communities associated with 12 sponge specimens were characterised. Sponge samples were collected from the St. Peter and St. Paul Archipelago (SPSPA) in Brazil. To ensure the accurate identification of target sponges, host phylogenetic relatedness was assessed using sequences from the standard CO1 barcoding fragment. This is the first study to assess bacterial communities associated with marine sponges in the SPSPA using amplicon sequencing of the 16S rRNA gene. MATERIALS AND METHODS Study site The SPSPA is one of the smallest and most isolated archipelagos in the world. It is located ∼1000 km from the city of Natal, Rio Grande do Norte State, North-eastern Brazil (0◦ 55 02 N, 29◦ 20 44 W) and is a federal Environmental Protected Area maintained by the Brazilian Navy. The archipelago is composed of 10 small islands and islets distributed across a 1.5-ha area (Fig. 1). The rocky shores of the archipelago are mainly devoid of unconsolidated substrates, with the exception of one cove that forms gradually from a 3 to 20 m depth and includes a mosaic of rocky and gravel bottom. The unique marine fauna harbours high endemism of fishes and sponges and have attracted growing attention from researchers (Edwards and Lubbock 1983; Moraes 2011). Despite the high local fishing pressure, the SPSPA suffers relatively little anthropogenic disturbance, at least with respect to industrial and domestic pollutants, due to its isolation from the mainland and sparse human occupation. Sponge and seawater sampling Twelve sponge specimens (i.e. four replicates from three sponge species) located at a similar depth (14–15 m) and within a few meters of each other were sampled during a single dive within a protected area at the cove of SPSPA, all inhabited the same rocky substrate. The temperature of the water was 25◦ C, and the specimens were living in an area that was exposed to light. A small fragment from each specimen was collected and placed separately into a 50-mL Falcon tube containing seawater; the tubes were then stored on ice for transport to the laboratory. The sampled sponges were previously identified as belonging to the families Aplysinidae: Aiolochroia crassa (Hyatt 1875) and Aplysina fulva (Pallas 1766) and Chondrosidae: Chondrosia collectrix (Schmidt 1870), all from the subclass Verongimorpha (Moraes 2011; Erpenbeck et al. 2012). Phylogenetic inference methods utilizing sequences of the standard CO1 barcoding fragment were used to aid in the molecular identification of the sponge species (Fig. S1, microorganisms that can comprise up to 40% of their biomass (Hentschel et al. 2012). These microorganisms play key roles in nutrient cycling (Fan et al. 2012; Ribes et al. 2012) and are considered impor (...truncated)