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Clade-Specific Quantitative Analysis of Photosynthetic Gene Expression in Prochlorococcus.

Fernández-Pinos MC, Casado M, Caballero G, Zinser ER, Dachs J, Piña B - PLoS ONE (2015)

Bottom Line: After optimizing sample collection methodology, we analyzed a total of 62 stations from the Malaspina 2010 circumnavigation (including Atlantic, Pacific and Indian Oceans) at three different depths.Sequence and quantitative analyses of the corresponding amplicons showed the presence of high-light (HL) and low-light (LL) Prochlorococcus clades in essentially all 182 samples, with a largely uniform stratification of LL and HL sequences.Synechococcus cross-amplifications were detected by the taxon-specific melting temperatures of the amplicons.

View Article: PubMed Central - PubMed

Affiliation: Department of Environmental Chemistry, IDAEA-CSIC, Barcelona, Catalonia, Spain.

ABSTRACT
Newly designed primers targeting rbcL (CO2 fixation), psbA (photosystem II) and rnpB (reference) genes were used in qRT-PCR assays to assess the photosynthetic capability of natural communities of Prochlorococcus, the most abundant photosynthetic organism on Earth and a major contributor to primary production in oligotrophic oceans. After optimizing sample collection methodology, we analyzed a total of 62 stations from the Malaspina 2010 circumnavigation (including Atlantic, Pacific and Indian Oceans) at three different depths. Sequence and quantitative analyses of the corresponding amplicons showed the presence of high-light (HL) and low-light (LL) Prochlorococcus clades in essentially all 182 samples, with a largely uniform stratification of LL and HL sequences. Synechococcus cross-amplifications were detected by the taxon-specific melting temperatures of the amplicons. Laboratory exposure of Prochlorococcus MED4 (HL) and MIT9313 (LL) strains to organic pollutants (PAHs and organochlorine compounds) showed a decrease of rbcL transcript abundances, and of the rbcL to psbA ratios for both strains. We propose this technique as a convenient assay to evaluate effects of environmental stressors, including pollution, on the oceanic Prochlorococcus photosynthetic function.

No MeSH data available.


Related in: MedlinePlus

Cladograms of sequences from amplicons derived from field and laboratory samples compared to the closest sequences identified by BLAST.Two qRT-PCR products were sequenced for each sample from Atlantic, Indian and Pacific 1 stations, and only one for Pacific 2 samples (more information in supplementary S4 Table). Amplicons from surface, DCM, and DCM+40 samples from the different stations are identified with "S", "D" and "D40" letters. Panels a-c correspond to HL amplicons, whereas panels d-f correspond to LL amplicons. For each ecotype, cladograms corresponding to rnpB (a,d), rbcL (b,e) and psbA (c,f) genes are presented. The trees also include sequences from the GenBank. LL Prochlorococcus, HL Prochlorococcus, and Synechococcus (Syn.) sequences are indicated by dark green, light green and red colors, respectively. Amplicons from MIT9313 and MED4 strains grown in the lab were also sequenced and included in the analysis as MIT9313-Lab and MED4-Lab, respectively. Sequences for uncultured microorganism isolates are marked as U.P. (identified as Prochlorococcus), U.Syn. (identified as Synechococcus) or U.O, (not identified).
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pone.0133207.g004: Cladograms of sequences from amplicons derived from field and laboratory samples compared to the closest sequences identified by BLAST.Two qRT-PCR products were sequenced for each sample from Atlantic, Indian and Pacific 1 stations, and only one for Pacific 2 samples (more information in supplementary S4 Table). Amplicons from surface, DCM, and DCM+40 samples from the different stations are identified with "S", "D" and "D40" letters. Panels a-c correspond to HL amplicons, whereas panels d-f correspond to LL amplicons. For each ecotype, cladograms corresponding to rnpB (a,d), rbcL (b,e) and psbA (c,f) genes are presented. The trees also include sequences from the GenBank. LL Prochlorococcus, HL Prochlorococcus, and Synechococcus (Syn.) sequences are indicated by dark green, light green and red colors, respectively. Amplicons from MIT9313 and MED4 strains grown in the lab were also sequenced and included in the analysis as MIT9313-Lab and MED4-Lab, respectively. Sequences for uncultured microorganism isolates are marked as U.P. (identified as Prochlorococcus), U.Syn. (identified as Synechococcus) or U.O, (not identified).

Mentions: Sequence analysis of amplicons from four different stations at the three sampled depths demonstrated that they all encoded the targeted genes (Fig 4 and supplementary S4 Table). Sequences from HL amplicons were very similar in all cases to reported sequences from HL Prochlorococcus strains belonging to the eMED4 and eMIT9312 ecotypes (S4 Table). Similarly, amplicons from LL primers showed strong sequence similarities to LL Prochlorococcus strains included into the eMIT9313, eMIT9211, eNATL2A, and eSS120 ecotypes [20]. However, ten out of the 21 sequenced LL amplicons from surface samples were identified as Synechococcus sequences (S4 Table). This is keeping with the predominance of Synechococcus strains in conditions of high solar irradiance [10–12, 16–18, 22], and corroborates our interpretation of the amplicon Tm variability observed in Table 5 and Fig 3.


Clade-Specific Quantitative Analysis of Photosynthetic Gene Expression in Prochlorococcus.

Fernández-Pinos MC, Casado M, Caballero G, Zinser ER, Dachs J, Piña B - PLoS ONE (2015)

Cladograms of sequences from amplicons derived from field and laboratory samples compared to the closest sequences identified by BLAST.Two qRT-PCR products were sequenced for each sample from Atlantic, Indian and Pacific 1 stations, and only one for Pacific 2 samples (more information in supplementary S4 Table). Amplicons from surface, DCM, and DCM+40 samples from the different stations are identified with "S", "D" and "D40" letters. Panels a-c correspond to HL amplicons, whereas panels d-f correspond to LL amplicons. For each ecotype, cladograms corresponding to rnpB (a,d), rbcL (b,e) and psbA (c,f) genes are presented. The trees also include sequences from the GenBank. LL Prochlorococcus, HL Prochlorococcus, and Synechococcus (Syn.) sequences are indicated by dark green, light green and red colors, respectively. Amplicons from MIT9313 and MED4 strains grown in the lab were also sequenced and included in the analysis as MIT9313-Lab and MED4-Lab, respectively. Sequences for uncultured microorganism isolates are marked as U.P. (identified as Prochlorococcus), U.Syn. (identified as Synechococcus) or U.O, (not identified).
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4526520&req=5

pone.0133207.g004: Cladograms of sequences from amplicons derived from field and laboratory samples compared to the closest sequences identified by BLAST.Two qRT-PCR products were sequenced for each sample from Atlantic, Indian and Pacific 1 stations, and only one for Pacific 2 samples (more information in supplementary S4 Table). Amplicons from surface, DCM, and DCM+40 samples from the different stations are identified with "S", "D" and "D40" letters. Panels a-c correspond to HL amplicons, whereas panels d-f correspond to LL amplicons. For each ecotype, cladograms corresponding to rnpB (a,d), rbcL (b,e) and psbA (c,f) genes are presented. The trees also include sequences from the GenBank. LL Prochlorococcus, HL Prochlorococcus, and Synechococcus (Syn.) sequences are indicated by dark green, light green and red colors, respectively. Amplicons from MIT9313 and MED4 strains grown in the lab were also sequenced and included in the analysis as MIT9313-Lab and MED4-Lab, respectively. Sequences for uncultured microorganism isolates are marked as U.P. (identified as Prochlorococcus), U.Syn. (identified as Synechococcus) or U.O, (not identified).
Mentions: Sequence analysis of amplicons from four different stations at the three sampled depths demonstrated that they all encoded the targeted genes (Fig 4 and supplementary S4 Table). Sequences from HL amplicons were very similar in all cases to reported sequences from HL Prochlorococcus strains belonging to the eMED4 and eMIT9312 ecotypes (S4 Table). Similarly, amplicons from LL primers showed strong sequence similarities to LL Prochlorococcus strains included into the eMIT9313, eMIT9211, eNATL2A, and eSS120 ecotypes [20]. However, ten out of the 21 sequenced LL amplicons from surface samples were identified as Synechococcus sequences (S4 Table). This is keeping with the predominance of Synechococcus strains in conditions of high solar irradiance [10–12, 16–18, 22], and corroborates our interpretation of the amplicon Tm variability observed in Table 5 and Fig 3.

Bottom Line: After optimizing sample collection methodology, we analyzed a total of 62 stations from the Malaspina 2010 circumnavigation (including Atlantic, Pacific and Indian Oceans) at three different depths.Sequence and quantitative analyses of the corresponding amplicons showed the presence of high-light (HL) and low-light (LL) Prochlorococcus clades in essentially all 182 samples, with a largely uniform stratification of LL and HL sequences.Synechococcus cross-amplifications were detected by the taxon-specific melting temperatures of the amplicons.

View Article: PubMed Central - PubMed

Affiliation: Department of Environmental Chemistry, IDAEA-CSIC, Barcelona, Catalonia, Spain.

ABSTRACT
Newly designed primers targeting rbcL (CO2 fixation), psbA (photosystem II) and rnpB (reference) genes were used in qRT-PCR assays to assess the photosynthetic capability of natural communities of Prochlorococcus, the most abundant photosynthetic organism on Earth and a major contributor to primary production in oligotrophic oceans. After optimizing sample collection methodology, we analyzed a total of 62 stations from the Malaspina 2010 circumnavigation (including Atlantic, Pacific and Indian Oceans) at three different depths. Sequence and quantitative analyses of the corresponding amplicons showed the presence of high-light (HL) and low-light (LL) Prochlorococcus clades in essentially all 182 samples, with a largely uniform stratification of LL and HL sequences. Synechococcus cross-amplifications were detected by the taxon-specific melting temperatures of the amplicons. Laboratory exposure of Prochlorococcus MED4 (HL) and MIT9313 (LL) strains to organic pollutants (PAHs and organochlorine compounds) showed a decrease of rbcL transcript abundances, and of the rbcL to psbA ratios for both strains. We propose this technique as a convenient assay to evaluate effects of environmental stressors, including pollution, on the oceanic Prochlorococcus photosynthetic function.

No MeSH data available.


Related in: MedlinePlus