Research ArticleMICROBIOLOGY

Methanotrophic bacterial symbionts fuel dense populations of deep-sea feather duster worms (Sabellida, Annelida) and extend the spatial influence of methane seepage

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Science Advances  03 Apr 2020:
Vol. 6, no. 14, eaay8562
DOI: 10.1126/sciadv.aay8562
  • Fig. 1 In situ images of the two annelid species featured in this study.

    A new species of serpulid annelid (Laminatubus) and a new species of sabellid (genus Bispira), at a site known as Jaco Scar at 1768 to 1887 m depth (9°7.1′N, 84°50.4′W), along the convergent margin off the west coast of Costa Rica. Insets show the worm species in life position. Photo credit: HOV Alvin, Woods Hole Oceanographic Institute.

  • Fig. 2 Carbon and nitrogen isotope data for native and experimental animals.

    Biplots of δ13C and δ15N values for the crown radioles of (A) the serpulid Laminatubus n. sp. and (B) sabellid, Bispira n. sp. from the Jaco Scar seep, sampled directly upon collection from the active seep (“Native”), after an in situ transplant for 16 months to an area of no methane venting (“Transplants”), or after shipboard incubation with either 12CH4 (“controls”) or 13CH4 (example “13CH4-incubated”). “Nonseep” sabellids were also collected from an offshore seamount but were not identified beyond family. The right-hand vertical shaded bar highlights the measured values of particulate organic material (“POM”) [collected from CTD (conductivity/temperature/depth) casts over the area of active seepage]. The left-hand vertical shaded bar indicates the δ13C for methane from the water column above Jaco Scar seeps (−62 to −50‰) reported by Mau et al. (21). (C) Changes in δ13C over time are shown for both species. Two 13CH4-incubated Laminatubus that were observed to have a decreased relative abundance of putative methanotrophs based on 16S rRNA analysis (0.3 and 14%; see also Fig. 5) are shown with an asterisk in the symbol.

  • Fig. 3 Microscopy of Laminatubus n. sp. crown radioles.

    Light microscopy (A and B), fluorescence (C and E) plus corresponding bright field (D), and transmission electron (F to H) microscopy of Laminatubus n. sp. radioles. A FISH probe (MTC851) was designed to be an exact match to the putative methanotrophic symbionts [related to Methylococcales MMG-2, shown in orange (C) or green (E)]. DAPI-stained nuclei of host cells are shown in blue. TEM images show the bacteria, with dense internal membranes, on the epidermal surface of the host, and also possibly in the process of being engulfed (arrows) and completely engulfed [arrowheads in (F) and (G)] by host cells. (H) A close-up of a single filament, attached to the worm epidermis. n, nucleus; bac, bacteria. Scale bars, 1 cm (A), 10 μm (C), 15 μm (D and E), and 1 μm (F to H). Photo credit: Greg Rouse, Scripps Institution of Oceanography, and Shana Goffredi, Occidental College.

  • Fig. 4 Microscopy of the Bispira n. sp. crown radioles.

    Light microscopy (A and B), fluorescence (C and E) plus corresponding bright field (D), and transmission electron (F and G) microscopy of Bispira n sp. radioles. A FISH probe (MTC851) was designed to be an exact match to the putative methanotrophic symbionts [related to Methylococcales MMG-2, shown in orange (C) or green (E)]. DAPI-stained nuclei of host cells are shown in blue. TEM images show the bacteria, with dense internal membranes, embedded in the cuticle of the host, and also possibly in the process of being engulfed (arrows) and completely engulfed [arrowheads in (F) and (G)] by host cells. n, nucleus; mv, microvilli; bac, bacteria. Scale bars, 0.5 cm (A), 10 μm (C), 15 μm (D and E), and 1 μm (F and G). Photo credit: Greg Rouse, Scripps Institution of Oceanography, and Shana Goffredi, Occidental College.

  • Fig. 5 Relative abundance of bacterial phylotypes, based on 16S rRNA.

    Bacterial community structure for crown radioles of (A) the serpulid Laminatubus n. sp. and (B) sabellid Bispira n. sp. from Jaco Scar seep, Costa Rica. Tissues were sampled directly upon worm collection from the active seep (native), after transplant for 16 months to areas of very little/no methane venting (“600 m” and “1400 m” away) or after shipboard incubation in either 12CH4 or 13CH4 (example “13CH4-incubated”). Bispira were not transplanted and thus do not have specimens for those categories (“N/A”). Each color on the graph represents a distinct genus-level phylotype or lowest level available. Phylotypes were grouped to 99% 16S rRNA sequence similarity. Dominant phylotypes within the Methylococcales Marine Group 2 are indicated by either purple or orange bars. Genera that were not putative aerobic methanotrophs are shown in light blue (Arenicella), black (Moritella) or gray (all others). The raw Illumina 16S rRNA barcode sequences and metadata collected in this study are available from the Dryad Digital Repository (https://doi.org/10.5061/dryad.wdbrv15jq) and the NCBI Small Read Archive (BioProject # PRJNA599018).

  • Fig. 6 Bathymetric map of the Jaco Scar seep region, as surveyed by the AUV Sentry.

    Location of the Jaco Scar seep study site, off the west coast of Costa Rica, and the 44-km track surveyed by the AUV Sentry, equipped with high-resolution mapping tools (sidescan sonar, multibeam echosounder, and digital still camera), traversed during four dives in 2017 and 2018. Moving at an average speed of 0.54 m/s, Sentry traveled at an average altitude of 6.9 m and took ~20 photos/min. Out of a total of 42,272 downward-facing photos, 28,962 photos were annotated with serpulids. Serpulid presence was categorized by the occurrence of other obligate seep fauna within the same 5-m grid cell (seep-associated) and the absence of other obligate seep fauna within the same grid cell (non–seep-associated). The distribution of seep fauna and carbonate rock without serpulids is also shown.

Supplementary Materials

  • Supplementary Materials

    Methanotrophic bacterial symbionts fuel dense populations of deep-sea feather duster worms (Sabellida, Annelida) and extend the spatial influence of methane seepage

    Shana K. Goffredi, Ekin Tilic, Sean W. Mullin, Katherine S. Dawson, Abigail Keller, Raymond W. Lee, Fabai Wu, Lisa A. Levin, Greg W. Rouse, Erik E. Cordes, Victoria J. Orphan

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