Study of the microbial biogeography of deep ocean habitats has gained attention in recent years (Hamdan et al., 2013; Wilkins et al., 2013; Salazar et al., 2016; Hanson et al., 2019) with focus on how naturally occurring habitat features shape biogeographic patterns of sediment and water column communities (Hanson et al., 2012). While the importance of habitat features on biogeographic patterns is recognized (Hanson et al., 2012; Hanson et al., 2019), until recently, the significance of built habitats on microbial biogeography on the seafloor has been unexplored (Hamdan et al., 2018; Hamdan et al., 2021). Historic constraints (e.g., dispersal limitation) and contemporary features (e.g., habitat specificity) are the main drivers on microbial biogeographic patterns (Martiny et al., 2006; Hanson et al., 2019) in environmental settings. Both constraints have been studied in naturally occurring habitat features in the marine environment including hydrothermal vents, methane seeps, seamounts, and organic falls (Huber et al., 2007; Dick et al., 2013; Anderson et al., 2015). Anthropogenic structures in the marine environment include shipwrecks and energy infrastructure and are encompassed in the term “built environment” (Bugnot et al., 2020). Wood of terrestrial origin on the seafloor presents a habitat feature that may be derived from natural (wood falls) and anthropogenic (historic shipwrecks) sources. Wood delivered in the form of shipwrecks could play a role in deep-sea microbial biogeography.

The Theory of Island Biogeography (MacArthur and Wilson, 1967) explained three main considerations for island habitats: island size, distance from a mainland, and immigration and extinction rates. The Theory of Island Biogeography introduces a biotic equilibrium model in which richness reaches steady state when immigration equals extinction. At equilibrium, species richness and diversity decrease with distance from an island and has been used as indicators of “island-like” seabed habitats (Smith and Baco, 2003; Goffredi and Orphan, 2010; Hamdan et al., 2021). Natural marine habitat features exhibit island-like properties, including whale falls which evidence a distance effect with species richness being higher in proximity to fall sites than in the surrounding seafloor (Smith and Baco, 2003; Goffredi and Orphan, 2010). Anthropogenic structures, specifically, historic shipwrecks, have been recently considered island-like systems, as sediment microbiome diversity and richness declined as a function of distance from a historic metal-hulled shipwreck in the Gulf of Mexico (Hamdan et al., 2021).

While the factors that shape microbial biogeography for sediment and pelagic microorganisms associated with naturally occurring habitat features have been investigated, such studies are generally lacking for marine biofilms. Biofilms form on all submerged surfaces, and settlement of early colonizers and production of extracellular polymers leads to recruitment of microorganisms on hard substrates (Mugge et al., 2019; Mugge et al., 2021). Biofilm formation promotes settlement of macro-fauna (Hadfield, 2011; Dobretsov and Rittschof, 2020) on both natural and built habitat features, and occurs in three steps: adhesion, accumulation, and dispersal of macromolecules and biofilm microfauna (Garrett et al., 2008). Ultimately, dispersal of biofilms defines their ability to reach other habitats and is controlled by both historic constraints and contemporary features.

Considering the estimated 3 million shipwrecks in the world’s oceans, with the majority constructed of wood, shipwrecks may be a more significant source of wood to the seabed than recognized (UNESCO, 2017). Naturally derived wood is delivered to the deep-sea through riverine transport or storm events. While wood falls are most common along coastlines, they have also been observed at deeper water depths (Wolff, 1979; Bienhold et al., 2013). Wooden shipwrecks are found at all explored depths where past shipping lanes occurred. Experimental studies show wood falls are colonized by microorganisms and create hotspots of diversity (Palacios et al., 2009; Fagervold et al., 2012; Bienhold et al., 2013; Kalenitchenko et al., 2015). These studies show geographic location and water depth are important factors in structuring bacterial communities on submerged wood (Palacios et al., 2009; Ristova et al., 2017). Different experimental constraints, including wood type and immersion time also dictate the composition of wood-colonizing microbiomes (Palacios et al., 2009; Fagervold et al., 2012; Kalenitchenko et al., 2015).

The convergence of biofilm formation, maturation, and dispersal on submerged wood provides opportunity to understand how wood habitat features influence biogeography processes on the seabed.

The goal of this work was to determine how proximity to historic, wooden shipwrecks influenced diversity, richness, and community composition of microbial biofilms on two types of wood. This information may support understanding of how submerged wooden habitats influence biofilm dispersal in the marine environment. PERMANOVA revealed distance from shipwreck sites was a key structuring factor on phylogenetic composition for bacteria and archaea, but not for fungi. Alpha diversity extinction plots did not provide uniform evidence that shipwrecks function as island-like systems for biofilms in the surrounding environment given that a significant relationship was not generally discovered for alpha diversity. Bacterial alpha diversity at both sites and archaeal alpha diversity at Site 15470, however, were highest in samples collected nearest to the shipwrecks ( Figure 2 ) with a statistically significant relationship for archaea at Site 15470. A related study of sediment surrounding these sites also revealed archaeal diversity and taxa richness declined as a function of distance from site 15470 but not 15711 (Hampel et al., 2022). In that work, local environmental conditions, specifically differing organic matter supply between the sites was cited as a reason that alpha diversity decreased as a function of distance from 15470 which rests in a deeper, more oligotrophic setting compared to 15711. The two sites in this study are separated by 111 km of seafloor and 1275 m of vertical depth. Previous studies have shown that location and water depth are important factors driving marine sediment bacterial and archaeal community composition (Kallmeyer et al., 2012; Hoshino et al., 2020), and these factors may also influence biofilm composition. Site 15711 rests in shallower water near the coast (76 km from the Mississippi River delta), and thus receives more nutrient and carbon deposition relative to Site 15470. Discharge from the Mississippi River is also known to influence microbial community composition (Mason et al., 2016; Sánchez-Soto Jiménez et al., 2018). It is, therefore, likely that along with being a more eutrophic setting, 15711 has higher rates of immigration of new taxa from terresterial sources. In the context of The Theory of Island Biogeography, the site has greater proximity to a “mainland” source for immigration, resulting in an unequal extinction rate at the two sites. The dispersal of biofilms from the shipwreck would thus not be as evident at 15711 compared to 15470, the latter further from this “mainland” source and potentially at equilibrium. While the diversity and richness curves do not provide clear evidence for the shipwrecks as island-like systems, there is evidence that island-like functions relating to geographic location and taxa and nutrient supply impact biofilm dispersal around a shipwreck.

Across the two sites, archaeal and bacterial diversity and richness were generally highest nearest the shipwrecks, with secondary peaks near the ends of the transects, with the exception of archaea at Site 15711. The secondary peaks may be indicative of an ecotone, defined as a convergence zone between two ecosystems connecting along an environmental gradient (Levin et al., 2016). Ecotones function as diversity hotspots providing for co-location of taxa from adjacent ecosystems. If the shipwreck ecosystem is converging with the soft sediment dominated ecosystem, an ecotone may appear in water, sediment, or biofilm communities in between and would contain higher alpha diversity on par with end-member environments along the gradient. A recent study presented evidence of an ecotone in sediment microbiomes surrounding a metal-hulled shipwreck (Hamdan et al., 2021). The presence of an ecotone in this study indicates wooden shipwrecks harbor unique communities different from the background habitat of the surrounding seafloor and may be functioning as an island-like system. As noted above, however, the extent to which a shipwreck may function as an island-like system is influenced by geographic location in proximity to a source of taxa and nutrients.

UniFrac PCoAs and PERMANOVA tests showed community composition is influenced by site, wood type, and proximity to a shipwreck. The differences in the environmental setting between sites may explain the statistically significant effect of site in structuring all three domains in this study. Geographic location is known to be important in structuring wood-associated bacterial communities. A study by Ristova et al. (2017) found sunken wood communities were more similar within one geographic location than between different seas. However, wood type has also been shown to be a strong influence on bacterial communities, and more important in bacterial community composition than geographic location or depth (Palacios et al., 2009; Fagervold et al., 2012; Kalenitchenko et al., 2015). The current study agrees with those prior studies where bacteria are concerned, and notably, wood type was also a selective force on phylogenetic composition for fungi, although secondary to site. Although not statistically significant for archaea, wood type could be a selective force to a lesser extent than site and distance. Proximity to historic wooden shipwrecks also shaped community composition of biofilms on wood for bacteria and archaea, while the relationship was not significant for fungi. This is somewhat in conflict with results for alpha diversity where distance was not consistently a significant factor structuring diversity. This may indicate different dispersal mechanisms or different sources of taxa for individual members of the biofilm microbiome.

Previous studies have shown specific phylotypes have an affinity for forming biofilms on different types of sunken wood (Palacios et al., 2009; Fagervold et al., 2012; Kalenitchenko et al., 2015). Myxococcales has been found in association with sunken pine submerged for 20 months in the South Pacific (Fagervold et al., 2012). In the current study, one ASV associated with Myxococcales (Blrii41) was abundant in pine samples, had lower abundances at 25 m from the site, increased in abundance in the middle of the array, and then decreased in abundance at 100-124 m for both sites. This pattern was also seen at 15470 for oak samples. These data provide additional evidence that proximity is a structuring factor on microbiomes but is especially important to wood-associated taxa utilizing wood on the seafloor. Rhodobacteraceae has been found associated with oak in other studies and changes in abundance across distance in this study, exhibiting the same pattern described above for Myxococcales at 15470 (Kalenitchenko et al., 2015). The main archaeal phylotypes observed in this study from Nitrosopumilus are common in the Gulf of Mexico but have also been found on submerged wood (Kalenitchenko et al., 2015; Mason et al., 2016). Nitrosopumilus exhibits changing abundances across distances, although the patterns are not as clear as described for Myxococcales. While information on fungal communities on submerged wood is limited, this study found members of the phylum Ascomycota, which contains wood degraders, in high abundance (Pedersen et al., 2020). This includes those members of Sordariomycetes that exhibited a similar pattern of increasing abundance mid-transects as described for Myxococcales. This is another indication that while proximity overall is not as strong a structuring factor for fungal communities, it may be important to specific wood-associated fungal taxa.

This study reveals that wood-associated biofilm composition is impacted by proximity to wooden shipwrecks. Wood type is a highly selective force on bacterial community composition in deep-sea biofilm communities. Proximity to a shipwreck also shapes bacterial and archaeal community composition although questions remain for fungal communities. This study shows that the presence of wooden shipwrecks and similar anthropogenic structures on the seafloor should be considered in studies of marine microbial biogeography to support understanding of microbial dispersal in the deep-sea.

The datasets presented in this study can be found in online repositories. The names of the repository/repositories and accession number(s) can be found below: https://www.ncbi.nlm.nih.gov/bioproject/PRJNA802802.

LH designed the study. RDM conducted the data analysis. RDM and LH wrote the manuscript. All authors conducted the laboratory analyses and contributed to the manuscript. All authors contributed to the article and approved the submitted version.

Funding for this study was provided by the National Oceanic and Atmospheric Administration’s (NOAA) Ocean Exploration and Research Program under Cooperative Agreement NA18OAR0110286.

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

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