Dataset Information

A comparative study of microbial diversity and community structure in marine sediments using poly(A) tailing and reverse transcription-PCR.

ABSTRACT: To obtain a better understanding of metabolically active microbial communities, we tested a molecular ecological approach using poly(A) tailing of environmental 16S rRNA, followed by full-length complementary DNA (cDNA) synthesis and sequencing to eliminate potential biases caused by mismatching of polymerase chain reaction (PCR) primer sequences. The RNA pool tested was extracted from marine sediments of the Yonaguni Knoll IV hydrothermal field in the southern Okinawa Trough. The sequences obtained using the poly(A) tailing method were compared statistically and phylogenetically with those obtained using conventional reverse transcription-PCR (RT-PCR) with published domain-specific primers. Both methods indicated that Deltaproteobacteria are predominant in sediment (>85% of the total sequence read). The poly(A) tailing method indicated that Desulfobacterales were the predominant Deltaproteobacteria, while most of the sequences in libraries constructed using RT-PCR were derived from Desulfuromonadales. This discrepancy may have been due to low coverage of Desulfobacterales by the primers used. A comparison of library diversity indices indicated that the poly(A) tailing method retrieves more phylogenetically diverse sequences from the environment. The four archaeal 16S rRNA sequences that were obtained using the poly(A) tailing method formed deeply branching lineages that were related to Candidatus "Parvarchaeum" and the ancient archaeal group. These results clearly demonstrate that poly(A) tailing followed by cDNA sequencing is a powerful and less biased molecular ecological approach for the study of metabolically active microbial communities.

SUBMITTER: Hoshino T

PROVIDER: S-EPMC3684792 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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A comparative study of microbial diversity and community structure in marine sediments using poly(A) tailing and reverse transcription-PCR.

Hoshino Tatsuhiko T Inagaki Fumio F

Frontiers in microbiology 20130618

To obtain a better understanding of metabolically active microbial communities, we tested a molecular ecological approach using poly(A) tailing of environmental 16S rRNA, followed by full-length complementary DNA (cDNA) synthesis and sequencing to eliminate potential biases caused by mismatching of polymerase chain reaction (PCR) primer sequences. The RNA pool tested was extracted from marine sediments of the Yonaguni Knoll IV hydrothermal field in the southern Okinawa Trough. The sequences obta ...[more]

PMID: 23785366

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Project description:ObjectivesThe marine benthic nitrogen cycle is affected by both the presence and activity of macrofauna and the diversity of N-cycling microbes. However, integrated research simultaneously investigating macrofauna, microbes and N-cycling is lacking. We investigated spatio-temporal patterns in microbial community composition and diversity, macrofaunal abundance and their sediment reworking activity, and N-cycling in seven subtidal stations in the Southern North Sea.Spatio-temporal patterns of the microbial communitiesOur results indicated that bacteria (total and β-AOB) showed more spatio-temporal variation than archaea (total and AOA) as sedimentation of organic matter and the subsequent changes in the environment had a stronger impact on their community composition and diversity indices in our study area. However, spatio-temporal patterns of total bacterial and β-AOB communities were different and related to the availability of ammonium for the autotrophic β-AOB. Highest bacterial richness and diversity were observed in June at the timing of the phytoplankton bloom deposition, while richness of β-AOB as well as AOA peaked in September. Total archaeal community showed no temporal variation in diversity indices.Macrofauna, microbes and the benthic n-cycleDistance based linear models revealed that, independent from the effect of grain size and the quality and quantity of sediment organic matter, nitrification and N-mineralization were affected by respectively the diversity of metabolically active β-AOB and AOA, and the total bacteria, near the sediment-water interface. Separate models demonstrated a significant and independent effect of macrofaunal activities on community composition and richness of total bacteria, and diversity indices of metabolically active AOA. Diversity of β-AOB was significantly affected by macrofaunal abundance. Our results support the link between microbial biodiversity and ecosystem functioning in marine sediments, and provided broad correlative support for the hypothesis that this relationship is modulated by macrofaunal activity. We hypothesized that the latter effect can be explained by their bioturbating and bio-irrigating activities, increasing the spatial complexity of the biogeochemical environment.

Project description:Little is known about the diversity and succession of indigenous microbial community during debromination of polybrominated diphenyl ethers (PBDEs). This study examined the diversity and dynamics of microbial community structure in eight saline (mangrove and marine) and freshwater sediment microcosms exhibiting different debrominating capabilities for hexa-BDE 153, a common congener in sediments, using terminal restriction fragment length polymorphism (T-RFLP) and clone library analyses. The results showed that microbial community structure greatly differed between the saline and freshwater microcosms, likely leading to distinct variations in their debrominating capabilities and pathways. Higher relative abundances of Chloroflexi and Deltaproteobacteria succeed by Alphaproteobacteria and Betaproteobacteria were detected in the two mangrove microcosms with the fastest debrominating capabilities mainly via para pathway, respectively; the dominance of Alphaproteobacteria resulted in less accumulation of tetra-BDEs and more complete debromination of lower brominated congeners (from di- to tetra-BDEs). Meanwhile, the shifts in both microbial community structure and PBDE profiles were relatively small in the less efficient freshwater microcosms, with relatively more ortho and meta brominated products of BDE-153 resulted. Coincidently, one of the freshwater microcosms showed sudden increases of Chloroflexi and Deltaproteobacteria by the end of incubation, which synchronized with the increase in the removal rate of BDE-153. The significant relationship between microbial community structure and PBDEs was confirmed by redundancy analysis (18.7% of total variance explained, P = 0.002). However, the relative abundance of the well-known dechlorinator Dehalococcoides showed no clear correlation with the debrominating capability across different microcosms. These findings shed light in the significance of microbial community network in different saline environments on enhancement of PBDE intrinsic debromination.

Dataset Information

A comparative study of microbial diversity and community structure in marine sediments using poly(A) tailing and reverse transcription-PCR.

Publications

A comparative study of microbial diversity and community structure in marine sediments using poly(A) tailing and reverse transcription-PCR.

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