GoKawiilHOV observations
We conducted this investigation during the TS29-3 cruise (7 February to 18 March 2023) aboard the R/V Tan Suo Yi Hao, using the full-ocean-depth HOV Fendouzhe.
Thirty-two HOV surveys were conducted on the sea floor along the trench axis, spanning more than 1,200 km and distributing relatively evenly from the western to the central and southeastern regions. The estimated trench-floor area is about 14,400 km2, calculated in Global Mapper 26.1 by delineating the areal extent of the trench bottom. The HOV video survey had a field of view about 5 m in width, calibrated using 10-cm laser scale points. The surveyed area was estimated from the view width and the sea floor transect length recorded by the in situ footage. Each transect was 2.4–5.5 km long (4.3 km on average), with a cumulative length of 127.72 km for all 32 dives, yielding a total surveyed area of about 0.64 km2. The diving map (Fig. 1) was generated using Global Mapper 26.1, with basemap data from Global Multi-Resolution Topography (GMRT) Synthesis (http://www.gmrt.org) under a CC BY 4.0 license51.
Collecting and processing of whale-fall fauna samples
Whale-fall fossil samples were collected using the submersible’s two hydraulically powered manipulator arms, operated by the submersible’s pilots, and stored in geological baskets. The whale-fall bones in the sulfophilic stage were collected using the arms and kept in the biobox. Some associated free-living whale-fall fauna species, including gastropods, squat lobsters and brittle stars, were collected using the slurp sampler mounted on the submersible. On retrieval of the submersible, the whale bones and associated specimens were immediately sorted, fixed and registered in the ship’s laboratory. For taxonomic purposes, specimens were preserved using either a 10% formalin or a 75% ethanol solution. Specimens for molecular analysis were preserved directly in −80 °C freezers.
Examination of the whale-fall fauna and bivalve chemosymbionts
Morphological identification was based on published literatures of deep-sea macrobenthos faunas, especially that reported from whale falls, cold seeps and hydrothermal vents.
For further molecular examination, whale faunal tissues (up to 0.5 cm3) were subjected to DNA extraction, library preparation and metagenomic sequencing at Novogene Co., Ltd. Metagenomic sequencing was conducted using DNBSEQ-T10 (MGI) at Novogene to generate 2 bp × 150 bp pair-ended reads of about 50 Gb. Raw sequencing reads were qualified and assembled into contigs routinely. Assembled contigs were searched against the MetaCOXI database using the BLASTN program to extract mitochondrial 16S ribosomal RNA (rRNA) and cytochrome c oxidase I sequences52. The 18S and 28S rRNA gene sequences were predicted using rRNA_HMM53.
To detect potential chemosynthetic symbionts in three bivalves, microbiome analyses were conducted to obtain metagenome-assembled genomes (MAGs) of associated microbials from the investigated bivalve gill tissues. The assembled metagenomes were subjected to genome binning, duplicate removal and quality evaluation and finally were annotated using GTDB-tk (v2.4.0)54 against the GTDB database R220 (ref. 55). The dominate symbiont MAGs are listed in Extended Data Fig. 3d according to relative abundances of all microbial reads from each host bivalve species.
Density assessments
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