Bowman ducklow grc_pms_2017

Page 1

Patterns in microbial gene expression during the spring transition along thewestern Antarctic Peninsula

1Je� S Bowman, 2Hugh W Ducklow1Scripps Institution of Oceanography at UCSD, 2Lamont-Doherty Earth Observatory at Columbia University

Palmer Station

Ice Station

Station B

Station E

Arthur Harbor - 15-meter Landsat 8November 19, 2015

1 km

Introduction

In the Arctic and Antarctic sea ice is an important regulator of ecologi-cal processes in the water column. Sea ice stabilizes the water column by bu�ering wind stress, while limiting the transmission of light necessary for photosynthesis. To better understand the role of sea ice on ecological pro-cesses in the Antarctic we took advantage of unusual ice conditions in Arthur Harbor on the western Antarctic Peninsula during the 2015 spring seasonal transition. Rare, persistent land-fast sea ice allowed us to estab-lish a sea ice station approximately 200 m o�shore of Palmer Station. Water samples were collected from the sea ice station and incubated under simu-lated 10 m depth light conditions. Repeated sampling of the incubations and at the ice station allowed a direct comparison of ice-covered and ice-free conditions. When conditions allowed sam-ples were collected from Palmer LTER stations B and E, located in the marginal ice zone, providing a fur-ther comparison with the water column under partial sea ice cover.

Ice Sta�on Sta�on B Sta�on E Incuba�ons Time (days)4-Nov X -711-Nov X X 013-Nov X 215-Nov X X 417-Nov X 618-Nov X 725-Nov X 1426-Nov X X 1530-Nov X 192-Dec X 227-Dec X 2714-Dec X 34

Methods

Sample were collected with a peristaltic pump directly onto sterile 0.2 µm �lters and immediately �xed with 2-mercaptoethanol. Total RNA was extracted with the MoBio Powerwater RNA kit, treated with DNase, and rRNA depleted. Libraries were prepped from QC’d mRNA and sequenced at 2 x 100 on the Illumina HiSeq at a density of nine samples per lane. Sequence reads were analyzed with a new extension of the paprica pipeline for metabolic inference. In brief, overlapping reads were assem-bled with PEAR and mapped with BWA to a database of unique, taxonomi-cally referenced coding sequences extracted from all the completed ge-nomes in Genbank (Archaea, Bacteria, and virus), and from the Marine Mi-crobial Eukaryotic Transcriptome Sequencing Project (MMETSP). Each mapped reference genome (or reference transcriptome) is viewed inde-pendently in our analysis, allowing the identi�cation of di�erentially ex-pressed genes even as the relative abundance of the target organism changes over time.

Above: Sample density for the ice station, stations B and E, and the incu-bations. Black cell indicates when a sample was taken. Below: Sample lo-cations at the south end of Anvers Island, o� the western Antarctic Penin-sula.

Results

0 (B

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72 (D

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k)

Hours (treatment)

MMETSP0449MMETSP1057MMETSP0974MMETSP1445MMETSP0503MMETSP0929MMETSP1357MMETSP1160MMETSP1114MMETSP1178MMETSP1374MMETSP0168MMETSP1080MMETSP0105MMETSP1465MMETSP0943MMETSP0420MMETSP1126MMETSP1061MMETSP1310

−144

(A)

0 (B

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120

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360

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480

(N)

Hours (treatment)

MMETSP0449MMETSP1100MMETSP1178MMETSP1126MMETSP1061MMETSP1310MMETSP0929MMETSP1160MMETSP1114MMETSP1465MMETSP0105MMETSP1080MMETSP1357MMETSP0168MMETSP1445MMETSP0420MMETSP0943MMETSP1057MMETSP0974MMETSP0503

168

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384

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552

(O)

670

(N)

840

(R)

Hours (treatment)

MMETSP1178MMETSP1445MMETSP0449MMETSP0420MMETSP1057MMETSP1067MMETSP0902MMETSP0737MMETSP1100MMETSP1465MMETSP1058MMETSP1429MMETSP1336MMETSP0878MMETSP0319MMETSP1160MMETSP0974MMETSP1310MMETSP1126MMETSP1061

0.09

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Dominant eukaryotic genera according to relative abundance ofmapped transcripts

A) MMETSP0449, Strombidium: Mixotrophic ciliate, http://forum.mikroscopia.comB) MMETSP1445, Pyramimonas: Green algae (chlorophyte), nordicmicroalgae.keyC) MMETSP1178, Phaeocystis: Mixotrophic �agellate, http://cfb.unh.edu/phycokeyD) MMETSP1057, Cyclotella: Centric diatom, http://cfb.unh.edu/phycokeyE) MMETSP1067, Thalassiosira: Centric diatom, http://oceandatacenter.ucsc.edu

1e−01 1e+01 1e+03 1e+05 1e+07

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MMETSP0449 B C

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1e−01 1e+01 1e+03 1e+05 1e+07

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1e−01 1e+01 1e+03 1e+05 1e+07

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MMETSP0449 D G

Mean of normalized counts

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1e−01 1e+01 1e+03 1e+05

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12

34

MMETSP1057 B C

Mean of normalized counts

Log2

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1e−01 1e+01 1e+03 1e+05

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Mean of normalized counts

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4.1.1.211.1.1.421.3.7.22.3.1.2254.3.1.121.15.1.13.4.24.842.7.11.17|2.7.11.181.1.1.441.3.1.331.1.1.18|1.1.1.3702.2.1.13.1.13.44.2.99.181.1.1.401.1.1.215|1.1.1.262.3.2.264.3.1.16|4.3.1.17|4.3.1.18|5.1.1.184.2.1.226.1.1.13.6.1.15.3.1.13.3.1.12.5.1.543.4.25.13.6.1.5|3.6.1.62.1.1.3101.5.1.8|1.5.1.95.2.1.81.11.1.15|1.11.1.176.1.1.102.4.1.216.1.1.63.6.3.441.11.1.64.1.2.131.3.1.38|1.3.1.931.1.1.866.1.1.52.7.3.36.2.1.4|6.2.1.53.2.1.1236.6.1.12.7.1.21.5.1.201.7.1.1|1.7.1.26.6.1.1|6.6.1.22.7.1.25|2.7.7.44.3.2.11.23.5.12.6.1.100|2.6.1.101

1e−01 1e+01 1e+03 1e+05

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1e−01 1e+01 1e+03 1e+05

−0.4

−0.2

0.0

0.2

0.4

MMETSP1445 E I

Mean of normalized counts

Log2

(fold

cha

nge)

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1e−01 1e+01 1e+03

−10

12

MMETSP1445 I N

Mean of normalized counts

Log2

(fold

cha

nge)

●

●

●

●

●

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2.3.2.233.4.19.122.7.12.16.2.1.33.4.23.52.7.11.1|2.7.11.112.7.3.33.3.2.61.1.1.34|1.1.1.883.4.22.153.6.3.11.4.1.31.3.1.38|1.3.1.934.2.1.33.6.3.142.4.99.183.4.11.14|3.4.11.26.2.1.4|6.2.1.51.1.1.865.3.4.16.1.1.53.4.14.2|3.4.16.23.1.1.233.5.1.101.2.1.704.2.1.752.7.7.69|2.7.7.781.8.4.11|1.8.4.122.7.1.173.4.22.162.4.1.2281.8.4.111.3.5.22.6.1.96.3.4.10|6.3.4.11|6.3.4.15|6.3.4.92.7.11.1|2.7.11.17|2.7.11.193.1.30.13.1.11.2|4.2.99.182.4.1.1421.16.5.11.3.1.832.8.1.7|4.4.1.161.11.1.12|1.11.1.93.1.1.312.7.8.412.7.2.33.6.1.13|3.6.1.581.6.5.23.5.2.22.3.1.20|2.3.1.22|2.3.1.765.4.99.261.6.5.91.13.11.52.4.1.133|2.4.1.275|2.4.1.901.13.11.272.7.7.601.6.1.23.5.4.25|4.1.99.122.8.1.62.7.6.5|3.1.7.21.1.1.1003.3.1.12.7.11.11.10.2.2|3.4.24.642.7.11.22|2.7.11.233.1.4.35|3.1.4.531.2.1.134.2.1.331.14.19.11.1.1.35|4.2.1.17|5.1.2.3|5.3.3.82.5.1.48|4.4.1.1|4.4.1.81.1.1.188|1.1.1.2833.6.3.251.8.1.93.4.23.25|3.4.23.53.1.4.532.1.1.432.7.7.271.8.4.8|1.8.4.94.2.3.13.6.1.13|3.6.1.221.1.1.284|1.2.1.46|1.2.98.12.7.1.192.7.1.23|2.7.1.865.1.99.61.16.3.12.2.1.62.7.11.126.3.2.51.1.1.941.4.1.23.1.4.22.7.11.243.6.3.83.6.3.92.7.1.503.1.3.57|3.1.3.75.3.1.26|5.3.1.62.7.3.2|2.7.3.32.4.2.93.1.2.222.7.11.17|2.7.11.182.7.11.1|4.6.1.16.1.1.171.14.13.205|1.14.14.1|1.14.99.452.6.1.133.1.1.972.1.1.143|2.1.1.412.7.9.16.1.1.15|6.1.1.171.1.1.423.1.3.32.5.1.104|2.5.1.126|2.5.1.16|2.5.1.792.4.1.251.4.1.13|1.4.1.141.1.1.225.4.99.91.1.1.951.7.7.12.6.1.163.4.21.26|3.4.21.834.1.1.351.1.1.2|1.1.1.212.1.1.134.1.1.171.1.1.856.3.1.142.1.1.9|2.1.1.n43.2.1.1136.3.5.56.6.1.11.1.1.371.1.1.442.4.1.211.4.7.14.2.1.91.7.1.1|1.7.1.23.6.4.12|3.6.4.134.1.1.372.6.1.44|2.6.1.512.6.1.11.1.1.330|1.1.1.621.3.1.332.1.2.101.14.19.31.5.3.1|1.5.3.76.3.4.22.1.1.2871.14.19.3|1.14.19.306.1.1.112.5.1.622.1.1.2032.4.2.141.3.1.936.3.5.31.7.1.12.4.2.291.2.1.84.1.3.276.3.2.132.1.1.3193.4.22.15|3.4.22.30|3.4.22.672.4.1.181.3.1.6|1.3.5.43.6.3.543.1.3.123.13.1.11.14.19.203.1.1.n23.5.4.2|3.5.4.4|3.5.4.401.13.11.243.4.21.1023.4.25.12.7.11.113.2.1.244.2.1.23.4.14.41.2.1.3|1.2.1.31|1.2.1.83.6.4.126.2.1.54.1.2.133.1.3.163.4.22.342.3.3.161.11.1.62.7.1.683.7.1.21.2.1.36.1.1.36.1.1.63.2.1.1231.11.1.15|1.11.1.171.8.4.121.2.4.15.2.1.83.4.11.186.1.1.92.1.1.2051.1.1.402.3.2.261.10.3.91.2.1.122.6.1.522.7.1.403.4.22.434.2.99.186.1.1.44.2.1.223.6.1.11.1.1.35|1.1.1.n12|4.2.1.107|4.2.1.1195.3.1.92.7.7.61.1.1.215|1.1.1.261.2.4.44.3.1.16|4.3.1.17|4.3.1.18|5.1.1.181.11.1.154.2.1.246.3.2.32.7.7.73.1.3.16|3.1.3.483.6.4.62.1.1.625.4.2.11|5.4.2.43.2.1.15.5.1.41.17.4.13.1.1.33.6.3.42|3.6.3.441.1.2.4|1.1.99.393.1.3.11|3.1.3.371.4.4.22.7.1.205.3.1.12.1.2.16.2.1.452.4.1.13.4.22.25|3.4.22.30|3.4.22.6|3.4.22.672.5.1.63.1.4.17|3.1.4.35|3.1.4.531.15.1.1

1e−01 1e+01 1e+03 1e+05

−2−1

01

2

MMETSP0449 F L

Mean of normalized counts

Log2

(fold

cha

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1e−01 1e+01 1e+03 1e+05

−10

12

MMETSP0449 L O

Mean of normalized counts

Log2

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1e−01 1e+01 1e+03 1e+05

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MMETSP0449 O R

Mean of normalized counts

Log2

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1e−01 1e+01 1e+03 1e+05

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1.5

MMETSP1445 F L

Mean of normalized counts

Log2

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1e−01 1e+01 1e+03 1e+05

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1.5

MMETSP1445 L O

Mean of normalized counts

Log2

(fold

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2.7.1.683.7.1.21.2.1.36.1.1.36.1.1.63.2.1.1232.3.2.233.4.19.122.7.12.16.2.1.33.4.23.52.7.11.1|2.7.11.112.7.3.33.3.2.61.1.1.34|1.1.1.883.4.22.153.6.3.11.4.1.31.3.1.38|1.3.1.934.2.1.33.6.3.142.4.99.183.4.11.14|3.4.11.26.2.1.4|6.2.1.51.1.1.865.3.4.16.1.1.53.4.14.2|3.4.16.23.1.1.233.5.1.101.2.1.704.2.1.752.7.7.69|2.7.7.781.8.4.11|1.8.4.122.7.1.173.4.22.162.4.1.2281.8.4.111.3.5.22.6.1.96.3.4.10|6.3.4.11|6.3.4.15|6.3.4.92.7.11.1|2.7.11.17|2.7.11.193.1.30.11.16.5.11.3.1.832.4.1.1422.8.1.7|4.4.1.161.11.1.12|1.11.1.93.1.1.312.7.8.412.7.2.33.6.1.13|3.6.1.581.6.5.23.5.2.22.3.1.20|2.3.1.22|2.3.1.765.4.99.261.6.5.91.13.11.52.4.1.133|2.4.1.275|2.4.1.901.13.11.272.7.7.603.5.4.25|4.1.99.123.1.11.2|4.2.99.183.1.4.35|3.1.4.531.2.1.134.2.1.331.14.19.11.1.1.35|4.2.1.17|5.1.2.3|5.3.3.82.5.1.48|4.4.1.1|4.4.1.81.1.1.188|1.1.1.2833.6.3.251.8.1.93.4.23.25|3.4.23.53.1.4.532.1.1.432.7.7.271.8.4.8|1.8.4.94.2.3.13.6.1.13|3.6.1.221.1.1.284|1.2.1.46|1.2.98.12.7.1.192.7.1.23|2.7.1.865.1.99.61.16.3.12.2.1.62.7.11.122.8.1.62.7.6.5|3.1.7.21.1.1.1001.5.1.202.1.1.3104.1.99.171.14.99.452.7.13.3|2.7.4.62.2.1.73.4.11.21.23.5.13.4.22.15|3.4.22.30|3.4.22.672.7.1.503.1.3.57|3.1.3.75.3.1.26|5.3.1.62.7.3.2|2.7.3.32.4.2.93.1.2.222.7.11.17|2.7.11.182.7.11.1|4.6.1.16.1.1.171.14.13.205|1.14.14.1|1.14.99.452.6.1.133.1.1.972.1.1.143|2.1.1.413.6.3.92.7.11.243.6.3.83.1.4.26.6.1.1|6.6.1.22.7.1.24.1.1.396.3.2.51.1.1.941.4.1.24.3.2.12.6.1.836.1.1.202.7.7.42.6.1.100|2.6.1.1011.14.19.32.7.9.16.1.1.15|6.1.1.176.3.5.56.6.1.11.1.1.371.4.7.14.2.1.91.1.1.442.4.1.213.6.4.12|3.6.4.134.1.1.372.6.1.44|2.6.1.512.1.2.102.6.1.11.1.1.330|1.1.1.623.1.3.32.5.1.104|2.5.1.126|2.5.1.16|2.5.1.792.4.1.251.4.1.13|1.4.1.141.1.1.225.4.99.91.1.1.951.7.7.12.6.1.161.1.1.423.4.21.26|3.4.21.834.1.1.351.1.1.2|1.1.1.212.1.1.134.1.1.171.1.1.856.3.1.142.1.1.9|2.1.1.n43.2.1.1131.5.3.1|1.5.3.76.3.4.22.1.1.2871.14.19.3|1.14.19.306.1.1.112.5.1.622.1.1.2032.4.2.141.3.1.936.3.5.31.7.1.12.4.2.291.2.1.84.1.3.276.3.2.132.1.1.3191.14.19.305.1.3.12|5.1.3.62.6.1.42|2.6.1.882.1.1.951.6.1.21.8.4.123.3.1.12.7.11.11.10.2.2|3.4.24.642.7.11.22|2.7.11.231.3.1.331.7.1.1|1.7.1.22.5.1.104|2.5.1.162.4.1.181.3.1.6|1.3.5.43.6.3.543.1.3.123.13.1.11.14.19.203.1.1.n23.5.4.2|3.5.4.4|3.5.4.401.13.11.243.4.21.1023.4.25.12.7.11.113.2.1.244.2.1.23.4.14.41.2.1.3|1.2.1.31|1.2.1.83.6.4.126.2.1.54.1.2.133.1.3.163.4.22.342.3.3.161.11.1.63.1.3.11|3.1.3.372.7.7.73.1.3.16|3.1.3.483.6.4.62.1.1.625.4.2.11|5.4.2.43.2.1.15.5.1.41.17.4.13.1.1.33.6.3.42|3.6.3.441.1.2.4|1.1.99.391.11.1.15|1.11.1.175.2.1.81.10.3.91.2.4.11.2.1.123.6.1.12.6.1.522.7.1.403.4.22.434.2.1.224.2.99.186.1.1.43.4.11.186.1.1.92.1.1.2051.1.1.402.3.2.262.7.7.61.1.1.35|1.1.1.n12|4.2.1.107|4.2.1.1195.3.1.91.1.1.215|1.1.1.261.2.4.44.3.1.16|4.3.1.17|4.3.1.18|5.1.1.181.11.1.154.2.1.246.3.2.31.4.4.22.7.1.205.3.1.12.1.2.16.2.1.452.4.1.13.4.22.25|3.4.22.30|3.4.22.6|3.4.22.672.5.1.63.1.4.17|3.1.4.35|3.1.4.531.15.1.1

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MMETSP1057MMETSP0449

MMETSP1445MMETSP0449

MMETSP1445MMETSP0449MMETSP1057

Genes coding for enzyme involved in antibiot-ic biosynthesis and pigment production are upregulated at 192 hours in Cyclotella.

Genes coding for enzyme involved in extracel-lular DOM acquisition are upregulated at 24 hours in Cyclotella.

Cluster includes genes coding for bio-synthesis and phosphate metabolism, upregulated in Strombidium at 72 hours.

Cluster includes genes coding for bio-synthesis and phosphate metabolism, upregulated in Strombidium at 72 hours.

Relative abundance of most abundant genomesAs fraction of total mapped reads

Incubations

Ice Station

Station B

Di�erential expression for select genomesFor genomes and treatments outlined in grey in the heatmaps

A B C

D E

Preliminary observations of expression patternsBlue is signi�cantly downregulated, red is signi�cantly upregulated

Conclusions and Next Steps Strombidium spp. (MMETSP0449; mixotropic ciliate) dominated in the early spring below sea ice. Within 72 hours of exposure to 10 m light conditions the assem-blage had shifted to centric diatoms. By contrast Phaeocystis spp. (MMETSP1178; a mixotrophic �agellate) and chlorophytes were most abundant in the marginal ice zone, although mixotrophic ciliates were also observed. Diatoms did not dominate the marginal ice zone phytoplankton assemblage until nearly 35 days after the start of the experiment. These results highlight the role of reduced light and vertical mixing in suppressing the spring diatom bloom. There are clear di�erences in the gene expression patterns among the dominant protists. We are now exploring these patterns to identify the di�erent physiological responses to the onset of spring and summer conditions.

Acknowledgements

We would like to thank Jamie Collins, Nicole Couto, Ribanna Dietrich, Chelsea Farischon, Ashely Gonzalez, Colleen Hansel, and Conor Sullivan for their help with �eld sampling and their intellectual contributions. This work is funded by NSF PLR 1656344 to JSB and NSF PLR 1440435 to HWD.

The scatterplots show the results of the analysis for di�erentially expressed genes with DESeq2. The magnitude of log2(fold change) (y-axis) indicates how much expression has increased or decreased between timepoints. The color of the points gives the level of signi�cance according to the binomial test (red: p < 0.05, black: p >= 0.05). The products of di�erentially expressed genes are given in the heatmaps at right.

Alkaline phosphatase, secretion, DNA polymerase. upregulated in Strombidi-um at 480 hours.

Genes coding for peptidases, hydro-lases, and biosynthesis genes are all downregulated in Strombidium at 24 hours

Link to poster on the web

Email presenting author

Can this gene expression dataset help your research? We are happy to share the data as gene abundance tables (suitable for non-experts), all accompanying meta-data, and collaborate on any analysis.

2017 GRC Polar Marine Science

A) Chlorophyll a concentration. B) Bacterial production. T0 is 11 November 2015. The last Station B sampling point displayed corresponds to 552 hours on the heat-maps at right.