Accelerated sulfur cycle in coastal marine sediment beneath areas of intensive shellfish aquaculture

Abstract

Prokaryotes in marine sediments taken from two neighboring semi-enclosed bays (the Yamada and Kamaishi bays) at the Sanriku coast in Japan were investigated by the culture-independent molecular phylogenetic approach coupled with chemical and activity analyses. These two bays were chosen in terms of their similar hydrogeological and chemical characteristics but different usage modes; the Yamada bay has been used for intensive shellfish aquaculture, while the Kamaishi bay has a commercial port and is not used for aquaculture. Substantial differences were found in the phylogenetic composition of 16S rRNA gene clone libraries constructed for the Yamada and Kamaishi sediments. In the Yamada library, phylotypes affiliated with delta-Proteobacteria were the most abundant, and those affiliated with gamma-Proteobacteria were the second-most abundant. In contrast, the Kamaishi library was occupied by phylotypes affiliated with Planctomycetes, gamma-Proteobacteria, delta-Proteobacteria, and Crenarchaeota. In the gamma-Proteobacteria, many Yamada phylotypes were related to free-living and symbiotic sulfur oxidizers, whereas the Kamaishi phylotype was related to the genus Pseudomonas. These results allowed us to hypothesize that sulfate-reducing and sulfur-oxidizing bacteria have become abundant in the Yamada sediment. This hypothesis was supported by quantitative competitive PCR (qcPCR) with group-specific primers. The qcPCR also suggested that organisms closely related to Desulfotalea in the Desulfobulbaceae were the major sulfate-reducing bacteria in these sediments. In addition, potential sulfate reduction and sulfur oxidation rates in the sediment samples were determined, indicating that the sulfur cycle has become active in the Yamada sediment beneath the areas of intensive shellfish aquaculture.

FIG. 1.

Map of study sites.

FIG. 2.

Neighbor-joining trees showing phylogenetic relationships among phylotypes obtained in this study and reference 16S rRNA gene sequences retrieved from the nucleotide sequence databases for γ-Proteobacteria (A) and δ-Proteobacteria (B). The bold vertical bar to the right of the tree in panel A represents the range of Thiotrichales, while the one to the right of the tree in panel B represents Desulfobulbaceae. The numbers at the branch nodes are bootstrap values per 100 trials; only values of >50 are shown. Accession numbers for the reference sequences are shown in parentheses.

FIG. 3.

Phylogenetic distribution of clones in each library. Y3, the Yamada March library; Y8, the Yamada August library; KM, the Kamaishi library (in August); Antarctic sediment, a 16S clone library for Antarctic continental shelf sediment (4).

FIG. 4.

qcPCR assays for quantifying rRNA gene copies of phylotypes related to SOB and SRB. Ratios of specific rRNA gene copies to the total bacterial rRNA gene copies are shown. The names of qcPCR assays refer to Table 3. Data are means (n = 3 samples for each sediment sample and n = 9 samples for K-mean and Y-mean), and error bars represent standard deviations.

FIG. 5.

SOR and SRR of marine sediment samples. Data are means (n = 3 samples for each sediment sample and n = 9 samples for K-mean and Y-mean), and error bars represent standard deviations.