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Abstract

Objectives 

The overall research objective of this study is to assess the influence water availability has on structural diversification, community composition, production, and carbon sequestration in microbial mats. The specific goals for this observatory are to: 

www.SanSalMO.net

Tim SteppeHans PaerlLou Anne CheshireMelissa Leonard

Alan Decho

Jay Pinckney

Participants Collaborators

Virginia Tech

University of NC-Wilmington

University of Miami

tim_steppe@unc.eduhans_paerl@unc.edu

awdecho@gwm.sc.edu

pinckney@ocean.tamu.edu

UNC-CH Institute ofMarine Sciences

USC-Columbia Dept. of Environmental HealthSciencesTexas A&M Dept. of Oceanography

Anhydrophilic, Halotolerant Microbial Mats of San Salvador, Bahamas

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180

0

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100

150

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250

300

0

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20

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50

60

Salt PondSeawater

n.d.

n.d.b.d.b.d.

Tem

pera

ture

(o C)

Salin

ity

(psu

)N

H 4+

(N)

(mg

L-1 )

NO x

- (N

)(

g L-1

)

Mar.1999

Mar.2001

Mar.2000

Oct.2001

b.d.

b.d.

Mar.2002

Oct.2002

Date

Mar.2003

 Salt Pond salinity exhibits both inter- and intra-annual variation. Salinity and temperature measurements contributed by Elyse Voegeli.

Abundances of Extracellular Polymeric Secretions (EPS) in three different layers of the Salt Pond Microbial Mat: (1) An “orange” surface “ layer (L1); a “green” cyanobacterial layer (L2); and a “purple” Chromatium sp. Layer (L3). Significantly higher abundances of EPS occur in the surface L1 layer, and at sites where water-cover occurs most often.

L1(U)

L2(M)

L3(L)

Surface of MatX-Section of Mat X-Section of Mat

The surface layer microbial communities of Salt Pond mats form crenulated “polymer towers” that extend upward during water cover (see X-section). When examined using confocal scanning laser microscopy (CSLM), these polymer towers contain dense arrays of cyanobacteria and heterotrophic bacteria enveloped in a dense gel matrix of extracellular polymers (EPS). Dense colonies of cells suggest chemical signaling may occur in these towers. Also, clusters of cells contained within amphiphilic (hydrophobic/hydrophilic) EPS.

Con NH4 NO3 P04 NH4/P04 NO3/P040

0.5

1

1.5

2

2.5

3 Sea WaterSalt Pond

Con NH4 NO3 P04 NH4/P04 NO3/P040123456789

Treatment

H14C

0 3- U

ptak

e (n

mol

C c

m-2

h-1 )

Nitr

ogen

ase

Act

ivity

(nm

ol C

2H4

cm-2

h-1

)

0

500

1000

1500

2000

2500

3000

3500

4000

0 200 400 600 800 1000 1200 1400

DarkLight

Dep

th in

Sed

imen

t (µm

)

O2 Concentration (µM)

Light and dark profiles of dissolved oxygen concentration in hypersaline microbial mats. Oxygen gradients change from anoxic under dark conditions to ca. 10 times O2 saturation under sunlight. EPS may provide a buffering mechanism to prevent oxidative damage to photosynthetic enzymes.

Combined results of short-term nutrient bioassays from March 2002 and 2003. Mat pieces were collected and incubted in Salt Pond water or seawater ammended with nutrients (NH4

+ 20 μM; NO3

- 20 μM; and/or PO42- 5 μM). We observed no

significant stimulation of photosynthesis or nitrogenase activity (N2 Fixation) due to nutrient additions. Both forms of nitrogen repressed nitrogenase activity, while phosphorus appeared to ameliorate any N repression. Salinity appeared to affect 14CO2 upatke more than it did NA. These observations suggest water availability and salinity, in particular, have the largest impact on production and cycling in the mats.

0.1 subs/site

T10717T10011

T10012T10010

T10710T10008

T10016

T12313T12304

Desulfosarcina variablisT10017

T12319T12302

T12310T12317

T10006

T12309T10707

T12315T12316

T10715

Desulfovibrio longusDesulfovibrio africanus

T10014T10019

Desulfomonas pigraT10716T12305

T12306T10709

T10013

T12312T12303

Desulfotomaculumacetoxidans

T10713T10719

T10001T12318

Desulfoarculus baarsiiT10005

T10020T10002

T10003Desulfobulbus rhabdoformis

T10702T12314

T12301T10701

Archaeoglobus fulgidis

86100

100

94

70

67

8868

100

100

51100

59

61100

100

96100

100

73

93100

59

86

0.1 subs/site

T10717T10011

T10012T10010

T10710T10008

T10016

T12313T12304

Desulfosarcina variablisT10017

T12319T12302

T12310T12317

T10006

T12309T10707

T12315T12316

T10715

Desulfovibrio longusDesulfovibrio africanus

T10014T10019

Desulfomonas pigraT10716T12305

T12306T10709

T10013

T12312T12303

Desulfotomaculumacetoxidans

T10713T10719

T10001T12318

Desulfoarculus baarsiiT10005

T10020T10002

T10003Desulfobulbus rhabdoformis

T10702T12314

T12301T10701

Archaeoglobus fulgidis

86100

100

94

70

67

8868

100

100

51100

59

61100

100

96100

100

73

93100

59

86

T38L22T30U11

T30U10Phormidium sp.

T12307Pseudoanabaena sp.

NC mat cyanoT30U9

T30U02T323U03

T12306T38M06

Lyngbya lagerhaemiiDermocarpa sp.

Plectonema sp.T323L03T12304

Myxosarcina sp.Xenococcus sp.

Cyanothece sp.Aphanazomenon sp.

Anabaerna oscillaroidesNostoc commune

Lyngbya sp. SG1Synechococcus sp.

Synechocystis sp.Gloeothece sp.

Trichodesmium sp.Trichodemium thiebautii

T38L05T38M14

T10719T38U19

T323U09T38U23

T10711T10722

T38M12

Azotobacter chromatiumVibrio diazotrophicus

Azospirillum brasilenseRhodobacter rubrum

T10020T38M15

T10023NC Mat 0729 D10NC Mat 0729 D12

Desulfomicrobium baculatusDesulfovibrio vulgaris

Desulfovibrio salexigensT38M01

T10712

T323M18T323L05

T10002T10014

T10715T323L03

T10713Desulfovibrio gigas

T38U01NC Mat 0729 D11

T30U15T38U12

T323L12NC Mat 0909 D09

T323L07T323L21

NC Mat 0729 D09T38L08

T323L14T30L02

T323U23T323LU21T323L09

T38L18Clostridium pasteurianum

T30L13T30L06

T30M16T30L20

T10718Desulfobacter curvatus

Desulfonema limicolaDesulfosporosinus orientis

Clostridium cellobioparum Meth.voltae

0.1 subs/site

97

9964

100

90

99

52100

71

7278

64

100

86

8351

5192

60

67

89

68

10072

79

100

80

100

10062

100

10055

cyanobacteria

heterocystous

bet a/ga mm

aalpha

anaerobesdelta SR

B, gram

+s, etc

T38L22T30U11

T30U10Phormidium sp.

T12307Pseudoanabaena sp.

NC mat cyanoT30U9

T30U02T323U03

T12306T38M06

Lyngbya lagerhaemiiDermocarpa sp.

Plectonema sp.T323L03T12304

Myxosarcina sp.Xenococcus sp.

Cyanothece sp.Aphanazomenon sp.

Anabaerna oscillaroidesNostoc commune

Lyngbya sp. SG1Synechococcus sp.

Synechocystis sp.Gloeothece sp.

Trichodesmium sp.Trichodemium thiebautii

T38L05T38M14

T10719T38U19

T323U09T38U23

T10711T10722

T38M12

Azotobacter chromatiumVibrio diazotrophicus

Azospirillum brasilenseRhodobacter rubrum

T10020T38M15

T10023NC Mat 0729 D10NC Mat 0729 D12

Desulfomicrobium baculatusDesulfovibrio vulgaris

Desulfovibrio salexigensT38M01

T10712

T323M18T323L05

T10002T10014

T10715T323L03

T10713Desulfovibrio gigas

T38U01NC Mat 0729 D11

T30U15T38U12

T323L12NC Mat 0909 D09

T323L07T323L21

NC Mat 0729 D09T38L08

T323L14T30L02

T323U23T323LU21T323L09

T38L18Clostridium pasteurianum

T30L13T30L06

T30M16T30L20

T10718Desulfobacter curvatus

Desulfonema limicolaDesulfosporosinus orientis

Clostridium cellobioparum Meth.voltae

0.1 subs/site

97

9964

100

90

99

52100

71

7278

64

100

86

8351

5192

60

67

89

68

10072

79

100

80

100

10062

100

10055

cyanobacteria

heterocystous

bet a/ga mm

aalpha

anaerobesdelta SR

B, gram

+s, etc

T38M14T30L16

T30U10T30L23T30U04

T323U09T38M21

T38M13T38U23T38U08

T38M10T323M16T30M14T38M17

T38M07T38L07

T30M12T30U18

T323U01T38U13

T30M18T30U15T38L09

T30L13T30L06

Oscillatoria sp. OH25T30M24

T323M01T30M06

T323U21T30M11

T30M07T38M15

T38U18T30M19

Nodularia sp. PCC9350Anabaena flos-aquae

Anabaenopsis sp. PCC9215Symploca semiplena

Trichdesmium thiebautiiLyngbya aestuarii

Halospirulina sp. BAJA95T30U16

Halothece sp.

Halomicronema sp. TFEP2

Leptolyngbya sp. PCC9221T38L14

T38U21Cyanothece sp. PCC7418

Aphanothece sp. ATCC43922T38L03

T38U01T323U22

Lyngbya sp. PCC7419T323L01

T30M05T38U12

T38L01T323U19

T30L03

T323U15T38UL11

T38U22T323L04

T323L02T38L04

Leptolyngbya sp. PCC7104CY38L08

Escherichia coli0.1 subs/site

100

100

88

62

86

85

5370

52

9683

6077

T38M14T30L16

T30U10T30L23T30U04

T323U09T38M21

T38M13T38U23T38U08

T38M10T323M16T30M14T38M17

T38M07T38L07

T30M12T30U18

T323U01T38U13

T30M18T30U15T38L09

T30L13T30L06

Oscillatoria sp. OH25T30M24

T323M01T30M06

T323U21T30M11

T30M07T38M15

T38U18T30M19

Nodularia sp. PCC9350Anabaena flos-aquae

Anabaenopsis sp. PCC9215Symploca semiplena

Trichdesmium thiebautiiLyngbya aestuarii

Halospirulina sp. BAJA95T30U16

Halothece sp.

Halomicronema sp. TFEP2

Leptolyngbya sp. PCC9221T38L14

T38U21Cyanothece sp. PCC7418

Aphanothece sp. ATCC43922T38L03

T38U01T323U22

Lyngbya sp. PCC7419T323L01

T30M05T38U12

T38L01T323U19

T30L03

T323U15T38UL11

T38U22T323L04

T323L02T38L04

Leptolyngbya sp. PCC7104CY38L08

Escherichia coli0.1 subs/site

100

100

88

62

86

85

5370

52

9683

6077

Sites and Design Photosynthesis and Nitrogenase Activity Mat & Water Chemistry (Salt Pond)

EPS Characterization

Diversity of Key Biogeochemical Functional Groups

23m 11m 7m 3m 0m

Site

dsrAsulfate-reducers

Like many Bahamian Islands, San Salvador Island (24o05' N, 74o30' W) contains numerous shallow, hypersaline (45 to 322 ‰) lakes. The lakes are subjected to intense irradiance (> 2100 μE m-2 s-1), high temperatures (> 35o C) and chronic nutrient depletion. Highly productive microbial mats blanket the shallow sediments in many of the lakes. The overall research objective of this study is to assess the influence water availability has on structural diversification, community composition, production, and carbon sequestration in microbial mats. Three transects, 26 meters in length, have been established along a natural desiccation gradient in one of the hypersaline lakes, Salt Pond. Samples for community composition, extracellular polymeric substances (EPS) content, C & N content, and microscopic documentation are collected during each site visit (two to three times a year). Rates of key C, O, and N cycling processes (photosynthesis and N2 fixation) are obtained. In cooperation with the staff from the Gerace Research Center, Salt Pond’s salinity and temperature are being measured every 10-21 days. From March to July, Salt Pond’s salinity increased from ~ 110‰ to over 320‰. Light and dark vertical O2 distribution profiles of the mat’s upper 5 mm indicate that, under dark conditions, anoxia reaches the mat surface. When exposed to light (1,500 µmol m-2 s-1, 10 min), O2 was detected as deep as 5 mm with concentrations (ca. 800% O2 saturation) peaking at 1 mm depth. Light and dark cycles create a dynamic chemical environment that changes from anoxic to hyperoxic conditions within minutes. How EPS may buffer against drastic changes in redox conditions is being examined. Nutrient addition bioassays (e.g., NH4

+, NO3

-, and PO42-) indicate salinity levels and not nutrient availability has the greatest impact on

these crucial biogeochemical processes. Sequencing surveys of cyanobacterial 16S (primary producers), dsr (sulfate reducers/carbon mineralizers), and nifH (diazotrophs) genes show that diverse assemblages comprise the key functional groups of microorganisms. We are currently analyzing the sequence distributions to determine if there are any differences along the gradient. Carbohydrate analyses have led to the discovery of “amadori products" (APs) in the Salt Pond mats. APs are unique protein-carbohydrate linkages that form when basic amino acids cross-link with carbohydrate carboxyl groups. This is the first report of APs being found in natural systems. The potential for amadori products to act as a further defense (e.g., scytonemins, mycosporine amino acids, etc) against UV is being investigated.

NifH

3-D Reconstruction of TowerPolymer Towers

EPS

NifHdiazotrophs

Cyanobacterial 16Sprimary producers

T323U08

transect

meter markupper, middle, or lower portion of mat

clone number

10-Mar-03 19-Apr-03 29-May-03 8-Jul-030

50100150200250300350

051015202530354045

Date

Temperature

Salin

ity (P

SU)

10 20 300 10 20 30

4

9

14

19

24

10 20 30

Carbon:Nitrogen Ratio

Tran

sect

Pos

ition

Mar. 2002 Mar. 2003Oct. 2002

1) Describe the structural and microbial diversity of the mat communities in relation to water availability.

2) Assess the influence water availability has on primary production extracellular polymeric substances (EPS) production, and EPS degradation.

3) Isolate and characterize desiccation tolerant organisms4) Develop a conceptual model linking climate and water budget data, water availability, and

primary production.