Bacteria abundance and thymidine/leucine incorporation from R/V Thomas G. Thompson TT043, TT045, TT049, TT054 cruises in the Arabian Sea in 1995 (U.S. JGOFS Arabian Sea project)

Website: https://www.bco-dmo.org/dataset/2518
Version: final
Version Date: 2002-02-27

Project
» U.S. JGOFS Arabian Sea (Arabian Sea)

Program
» U.S. Joint Global Ocean Flux Study (U.S. JGOFS)
ContributorsAffiliationRole
Azam, FarooqUniversity of California-San Diego (UCSD-SIO)Principal Investigator
Ducklow, Hugh W.Marine Biological Laboratory Ecosystems Center (MBL - Ecosystems)Principal Investigator
Smith, David C.University of California-San Diego (UCSD-SIO)Co-Principal Investigator
Chandler, Cynthia L.Woods Hole Oceanographic Institution (WHOI BCO-DMO)BCO-DMO Data Manager


Dataset Description

Bacteria abundance and thymidine/leucine incorporation


Acquisition Description

See Platform deployments for cruise specific documentation


Processing Description

Methods for the bacterial component of US JGOFS Arabian Sea Process 1 and Process 7 Cruises

Bacterial Abundance

Bacterial abundance was determined by epifluorescence microscopy of
acridine orange- stained bacteria on Nuclepore filters ( Hobbie et al.
1977 ). Samples of between 50 to 100 ml of seawater for bacteria counts
were fixed with 0.2 m-filtered, EM-grade glutaraldehyde (1.25% final
concentration; Ted Pella).  Subsamples of 2 to 50 ml were vacuum
filtered (< 150 mm Hg) onto 0.2 m pore size, black, polycarbonate
membrane filters (Nuclepore).  During filtration, when a sample volume
had been reduced to 2 ml, 200 l of an 0.2 m-filtered acridine orange
working solution (0.05%) was added to the sample in the tower for a
final concentration of 0.005%.  Filtration rate was kept low enough to
allow at least 5 minutes of contact with the stain before filtration
was complete.  Blanks were 0.2 m-filtered seawater or deionized,
Milli-Q-treated water fixed and processed in the same manner as the
samples.  After filtration, filters were transferred quickly (while
still moist) onto fogged glass slides.  Before the filters could dry, a
coverslip with a drop of paraffin oil on the underside was placed over
the filter.  Slides were stored at approximately 4 degrees C for less
than 1 day, then transferred to a -20 degrees C freezer for longer term
storage.  Bacteria were counted at 1250' magnification by
epifluorescence microscopy with an oil immersion objective.  Each
sample was counted until either 20 fields or 200 cells had been
enumerated.


Bacterial Production

Bacterial production was estimated using both the 3H-thymidine (
Fuhrman and Azam 1980; Fuhrman and Azam 1982  ) and the 3H-leucine
(Kirchman et al. 1985 ) incorporation methods as modified for
processing by microcentrifugation by  Smith and Azam (1992) .  Aliquots
of seawater (1.7 ml) were incubated with 20 nM 3H-leucine or
3H-thymidine in the dark.  The length of incubation ranged from 1 to 6
hours depending on the depth from which the sample was collected.  The
incubations were terminated with the addition of trichloroacetic acid
(TCA; 5% v/v final).  Samples which received the addition of TCA prior
to the radioisotope served as blanks.  The samples were centrifuged at
16000 x g for 7 min and the supernatant was removed.  The pellet was
washed once with 5% TCA and once with 80% ethanol.  Liquid
scintillation cocktail was added directly to the centrifuge tube which
were then radioassayed.

References

 Fuhrman, J. A. and F. Azam 1980. Bacterioplankton secondary production
     estimates for coastal waters of British Columbia, Antarctica and
     California. Appl. Environ.  Microbiol. 39: 1085-1095.

 Fuhrman, J. A. and F. Azam 1982. Thymidine incorporation as a measure
     of heterotrophic bacterioplankton production in marine surface waters:
     evaluation and field results. Mar.  Biol. 66: 109-120.

 Hobbie, J. E., R. J. Daley and S. Jasper 1977. Use of nuclepore
     filters for counting bacteria by epifluorescence microscopy. Appl.
     Environ. Microbiol. 33: 1225-1228.

 Kirchman, D., E. K'Nees and R. Hodson 1985. Leucine incorporation and
     its potential as a measure of protein synthesis by bacteria in natural
     aquatic systems. Appl. Environ.  Microbiol. 49: 599-607.

 Smith, D. C. and F. Azam 1992. A simple, economical method for
     measuring bacterial protein synthesis rates in seawater using
     3H-leucine. Mar. Microb. Food Webs. 6: 107- 114.

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Parameters

ParameterDescriptionUnits
eventevent number, from event log YYYYMMDD
stastation number, from event log dimensionless
sta_stdArabian Sea standard station identifier dimensionless
castCTD cast number, from event log consecutive within station dimensionless
botCTD rosette bottle number dimensionless
presssample depth decibars
bact_het_origheterotrophic bacteria abundance, original units; microscopy cells/liter *10^9
bact_het_sestandard error for heterotrophic bacteria cell counts; original units cells/liter *10^9
bact_het_micheterotrophic bacteria abundance; DMO converted units, microscopy cells/milliliter
thy_incorpthymidine incorporation picomoles/liter/hr
thy_sdstandard deviation thymidine incorporation picomoles/liter/hr
leuc_incorpleucine incorporation picomoles/liter/hr
leuc_sdstandard deviation leucine incorporation picomoles/liter/hr


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Instruments

Dataset-specific Instrument Name
Niskin Bottle
Generic Instrument Name
Niskin bottle
Dataset-specific Description
Niskin bottles were mounted on the CTD rosette.
Generic Instrument Description
A Niskin bottle (a next generation water sampler based on the Nansen bottle) is a cylindrical, non-metallic water collection device with stoppers at both ends. The bottles can be attached individually on a hydrowire or deployed in 12, 24, or 36 bottle Rosette systems mounted on a frame and combined with a CTD. Niskin bottles are used to collect discrete water samples for a range of measurements including pigments, nutrients, plankton, etc.


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Deployments

TT043

Website
Platform
R/V Thomas G. Thompson
Report
Start Date
1995-01-08
End Date
1995-02-05
Description
Purpose: Process Cruise #1 (Late NE Monsoon)

Acquisition Description
PI: Farooq Azam and David Smith of: Scripps Institute of Oceanography dataset: Bacteria abundance and thymidine/leucine incorporation dates: January 09, 1995 to January 31, 1995 location: N: 22.483 S: 10 W: 57.2999 E: 68.75 project/cruise: Arabian Sea/TTN-043 - Process Cruise 1 (Late NE Monsoon) ship: Thomas Thompson Sampling Seawater samples (250 ml) were taken from the Niskin bottles mounted on the CTD rosette and transferred directly into brown high-density polyethylene bottles. The bottles were maintained within 2 degrees C of the in situ temperature until the start of the incubations or fixing.

Processing Description
Methods for the bacterial component of US JGOFS Arabian Sea Process 1 and Process 7 Cruises Bacterial Abundance Bacterial abundance was determined by epifluorescence microscopy of acridine orange- stained bacteria on Nuclepore filters ( Hobbie et al. 1977 ). Samples of between 50 to 100 ml of seawater for bacteria counts were fixed with 0.2 m-filtered, EM-grade glutaraldehyde (1.25% final concentration; Ted Pella). Subsamples of 2 to 50 ml were vacuum filtered (< 150 mm Hg) onto 0.2 m pore size, black, polycarbonate membrane filters (Nuclepore). During filtration, when a sample volume had been reduced to 2 ml, 200 l of an 0.2 m-filtered acridine orange working solution (0.05%) was added to the sample in the tower for a final concentration of 0.005%. Filtration rate was kept low enough to allow at least 5 minutes of contact with the stain before filtration was complete. Blanks were 0.2 m-filtered seawater or deionized, Milli-Q-treated water fixed and processed in the same manner as the samples. After filtration, filters were transferred quickly (while still moist) onto fogged glass slides. Before the filters could dry, a coverslip with a drop of paraffin oil on the underside was placed over the filter. Slides were stored at approximately 4 degrees C for less than 1 day, then transferred to a -20 degrees C freezer for longer term storage. Bacteria were counted at 1250' magnification by epifluorescence microscopy with an oil immersion objective. Each sample was counted until either 20 fields or 200 cells had been enumerated. Bacterial Production Bacterial production was estimated using both the 3H-thymidine ( Fuhrman and Azam 1980; Fuhrman and Azam 1982 ) and the 3H-leucine (Kirchman et al. 1985 ) incorporation methods as modified for processing by microcentrifugation by Smith and Azam (1992) . Aliquots of seawater (1.7 ml) were incubated with 20 nM 3H-leucine or 3H-thymidine in the dark. The length of incubation ranged from 1 to 6 hours depending on the depth from which the sample was collected. The incubations were terminated with the addition of trichloroacetic acid (TCA; 5% v/v final). Samples which received the addition of TCA prior to the radioisotope served as blanks. The samples were centrifuged at 16000 x g for 7 min and the supernatant was removed. The pellet was washed once with 5% TCA and once with 80% ethanol. Liquid scintillation cocktail was added directly to the centrifuge tube which were then radioassayed. References Fuhrman, J. A. and F. Azam 1980. Bacterioplankton secondary production estimates for coastal waters of British Columbia, Antarctica and California. Appl. Environ. Microbiol. 39: 1085-1095. Fuhrman, J. A. and F. Azam 1982. Thymidine incorporation as a measure of heterotrophic bacterioplankton production in marine surface waters: evaluation and field results. Mar. Biol. 66: 109-120. Hobbie, J. E., R. J. Daley and S. Jasper 1977. Use of nuclepore filters for counting bacteria by epifluorescence microscopy. Appl. Environ. Microbiol. 33: 1225-1228. Kirchman, D., E. K'Nees and R. Hodson 1985. Leucine incorporation and its potential as a measure of protein synthesis by bacteria in natural aquatic systems. Appl. Environ. Microbiol. 49: 599-607. Smith, D. C. and F. Azam 1992. A simple, economical method for measuring bacterial protein synthesis rates in seawater using 3H-leucine. Mar. Microb. Food Webs. 6: 107- 114.

TT045

Website
Platform
R/V Thomas G. Thompson
Start Date
1995-03-14
End Date
1995-04-10
Description
Acquisition Description
PI-provided images: Surface leucine incorporation Surface thymidine incorporation Surface bacterial abundance Surface mean cell volume PI: Hugh Ducklow of: Virginia Institute of Marine Science dataset: Bacterial abundance, cell volume, thymidine and leucine incorporation dates: March 15, 1995 to April 07, 1995 location: N: 22.4858 S: 9.9993 W: 57.3007 E: 68.7532 project/cruise: Arabian Sea/TTN-045 Process Cruise 2 (Spring Intermonsoon) ship: R/V Thomas Thompson

Processing Description
Methods: Bacteria were enumerated essentially as described in our EqPac data documentation and the article in the EQPAC I DSR volume. Multiple images of all samples were acquired, processed and analyzed with Zeiss software. We used a new algorithm which removed most, but possibly not all of the prochlorophytes, so these data represent heterotrophic bacterial counts only. We will be comparing samples with Rob Olson and David Caron to constrain the bacteria counts. We also include estimates of the mean volume of cells in each sample. Bacterial abundance times cell volume yields total biovolume per liter, an index of bacterial biomass, to which carbon per unit volume factors can (and will) be applied. All raw images have been archived and can be made available by separate arrangement. Bacterial production was estimated by the incorporation of 3H-thymidine and 3H-leucine into cold, 5% TCA-insoluble extracts, essentially as described in our earlier JGOFS work, although using the new method of Smith & Azam (see the Azam data documentation for Process-1). Production estimates are forthcoming. Analytical precision for bacterial abundance and cell volume. We do not routinely perform replicate determinations of bacterial counts. However, reanalysis of 92 slides from the March series gave the following coefficients of variation (sd/mean) for bacterial abundance and cell volume: property mean st.dev. CV (%) n - - - - - - - - - - - - - abundance* 0.53 0.061 11.7 92 cell volume 0.037 0.006 16.4 92 - - - - - - - - - - - - - *units for abundance are x 10^9 cells/liter. volumes are um3/cell; these are mean values for 92 replicate determinations of each property. Hugh Ducklow tel 804-642-7180 Virginia Institute of Marine Sciences fax 804-642-7293 Box 1346 email duck@vims.edu Gloucester Point, VA 23062-1346

TT049

Website
Platform
R/V Thomas G. Thompson
Start Date
1995-07-17
End Date
1995-08-15
Description
Acquisition Description
PI: Hugh Ducklow of: Virginia Institute of Marine Science dataset: Bacterial abundance, cell volume, thymidine & leucine incorporation dates: July 18, 1995 to August 13, 1995 location: N: 22.5268 S: 9.911 W: 57.2997 E: 68.7507 project/cruise: Arabian Sea /TTN-049 Process cruise 4 (Middle SW Monsoon) ship: R/V Thomas Thompson

Processing Description
Hugh Ducklow's notes on methodology Comments: Incorporation data are only available for stations 13 - 30 as Omani customs would not allow our isotopes into the country. Methods: Bacteria were enumerated essentially as described in our EqPac data documentation and the article in the EQPAC I DSR volume. Multiple images of all samples were acquired, processed and analyzed with Zeiss software. We used a new algorithm which removed most, but possibly not all of the prochlorophytes, so these data represent heterotrophic bacterial counts only. We will be comparing samples with Rob Olson and David Caron to constrain the bacteria counts. We also include estimates of the mean volume of cells in each sample. Bacterial abundance times cell volume yields total biovolume per liter, an index of bacterial biomass, to which carbon per unit volume factors can (and will) be applied. All raw images have been archived and can be made available by separate arrangement. Bacterial production was estimated by the incorporation of 3H-thymidine and 3H-leucine into cold, 5% TCA-insoluble extracts, essentially as described in our earlier JGOFS work, although using the new method of Smith & Azam (see the Azam data documentation for Process-1). Production estimates are forthcoming. Analytical precision for bacterial abundance and cell volume. We do not routinely perform replicate determinations of bacterial counts. However, reanalysis of 92 slides from the March series gave the following coefficients of variation (sd/mean) for bacterial abundance and cell volume: property mean st.dev. CV (%) n - - - - - - - - - - - - - abundance* 0.53 0.061 11.7 92 cell volume 0.037 0.006 16.4 92 - - - - - - - - - - - - - *units for abundance are x 10^9 cells/liter. volumes are um3/cell; these are mean values for 92 replicate determinations of each property. Hugh Ducklow tel 804-642-7180 Virginia Institute of Marine Sciences fax 804-642-7293 Box 1346 email duck@vims.edu Gloucester Point, VA 23062-1346

TT054

Website
Platform
R/V Thomas G. Thompson
Start Date
1995-11-30
End Date
1995-12-28
Description
Acquisition Description
PI: Farooq Azam and David Smith of: Scripps Institute of Oceanography dataset: Bacteria abundance and thymidine/leucine incorporation dates: December 01, 1995 to December 26, 1995 location: N: 22.5011 S: 9.9789 W: 57.302 E: 68.7849 project/cruise: Arabian Sea/TTN-054 - Process Cruise 7 (Early NE Monsoon) ship: Thomas Thompson Sampling Seawater samples (250 ml) were taken from the Niskin bottles mounted on the CTD rosette and transferred directly into brown high-density polyethylene bottles. The bottles were maintained within 2 degrees C of the in situ temperature until the start of the incubations or fixing.

Processing Description
Methods for the bacterial component of US JGOFS Arabian Sea Process 1 and Process 7 Cruises Bacterial Abundance Bacterial abundance was determined by epifluorescence microscopy of acridine orange- stained bacteria on Nuclepore filters ( Hobbie et al. 1977 ). Samples of between 50 to 100 ml of seawater for bacteria counts were fixed with 0.2 m-filtered, EM-grade glutaraldehyde (1.25% final concentration; Ted Pella). Subsamples of 2 to 50 ml were vacuum filtered (< 150 mm Hg) onto 0.2 m pore size, black, polycarbonate membrane filters (Nuclepore). During filtration, when a sample volume had been reduced to 2 ml, 200 l of an 0.2 m-filtered acridine orange working solution (0.05%) was added to the sample in the tower for a final concentration of 0.005%. Filtration rate was kept low enough to allow at least 5 minutes of contact with the stain before filtration was complete. Blanks were 0.2 m-filtered seawater or deionized, Milli-Q-treated water fixed and processed in the same manner as the samples. After filtration, filters were transferred quickly (while still moist) onto fogged glass slides. Before the filters could dry, a coverslip with a drop of paraffin oil on the underside was placed over the filter. Slides were stored at approximately 4 degrees C for less than 1 day, then transferred to a -20 degrees C freezer for longer term storage. Bacteria were counted at 1250' magnification by epifluorescence microscopy with an oil immersion objective. Each sample was counted until either 20 fields or 200 cells had been enumerated. Bacterial Production Bacterial production was estimated using both the 3H-thymidine ( Fuhrman and Azam 1980; Fuhrman and Azam 1982 ) and the 3H-leucine (Kirchman et al. 1985 ) incorporation methods as modified for processing by microcentrifugation by Smith and Azam (1992) . Aliquots of seawater (1.7 ml) were incubated with 20 nM 3H-leucine or 3H-thymidine in the dark. The length of incubation ranged from 1 to 6 hours depending on the depth from which the sample was collected. The incubations were terminated with the addition of trichloroacetic acid (TCA; 5% v/v final). Samples which received the addition of TCA prior to the radioisotope served as blanks. The samples were centrifuged at 16000 x g for 7 min and the supernatant was removed. The pellet was washed once with 5% TCA and once with 80% ethanol. Liquid scintillation cocktail was added directly to the centrifuge tube which were then radioassayed. References Fuhrman, J. A. and F. Azam 1980. Bacterioplankton secondary production estimates for coastal waters of British Columbia, Antarctica and California. Appl. Environ. Microbiol. 39: 1085-1095. Fuhrman, J. A. and F. Azam 1982. Thymidine incorporation as a measure of heterotrophic bacterioplankton production in marine surface waters: evaluation and field results. Mar. Biol. 66: 109-120. Hobbie, J. E., R. J. Daley and S. Jasper 1977. Use of nuclepore filters for counting bacteria by epifluorescence microscopy. Appl. Environ. Microbiol. 33: 1225-1228. Kirchman, D., E. K'Nees and R. Hodson 1985. Leucine incorporation and its potential as a measure of protein synthesis by bacteria in natural aquatic systems. Appl. Environ. Microbiol. 49: 599-607. Smith, D. C. and F. Azam 1992. A simple, economical method for measuring bacterial protein synthesis rates in seawater using 3H-leucine. Mar. Microb. Food Webs. 6: 107- 114.


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Project Information

U.S. JGOFS Arabian Sea (Arabian Sea)


Coverage: Arabian Sea


The U.S. Arabian Sea Expedition which began in September 1994 and ended in January 1996, had three major components: a U.S. JGOFS Process Study, supported by the National Science Foundation (NSF); Forced Upper Ocean Dynamics, an Office of Naval Research (ONR) initiative; and shipboard and aircraft measurements supported by the National Aeronautics and Space Administration (NASA). The Expedition consisted of 17 cruises aboard the R/V Thomas Thompson, year-long moored deployments of five instrumented surface buoys and five sediment-trap arrays, aircraft overflights and satellite observations. Of the seventeen ship cruises, six were allocated to repeat process survey cruises, four to SeaSoar mapping cruises, six to mooring and benthic work, and a single calibration cruise which was essentially conducted in transit to the Arabian Sea.



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Program Information

U.S. Joint Global Ocean Flux Study (U.S. JGOFS)


Coverage: Global


The United States Joint Global Ocean Flux Study was a national component of international JGOFS and an integral part of global climate change research.

The U.S. launched the Joint Global Ocean Flux Study (JGOFS) in the late 1980s to study the ocean carbon cycle. An ambitious goal was set to understand the controls on the concentrations and fluxes of carbon and associated nutrients in the ocean. A new field of ocean biogeochemistry emerged with an emphasis on quality measurements of carbon system parameters and interdisciplinary field studies of the biological, chemical and physical process which control the ocean carbon cycle. As we studied ocean biogeochemistry, we learned that our simple views of carbon uptake and transport were severely limited, and a new "wave" of ocean science was born. U.S. JGOFS has been supported primarily by the U.S. National Science Foundation in collaboration with the National Oceanic and Atmospheric Administration, the National Aeronautics and Space Administration, the Department of Energy and the Office of Naval Research. U.S. JGOFS, ended in 2005 with the conclusion of the Synthesis and Modeling Project (SMP).



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Funding

Funding SourceAward
National Science Foundation (NSF)

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