Chemical composition of single microbial particles quantified by x-ray
microanalysis
Kjell Magne Fagerbakke
Department of Microbilogy, University of Bergen, Norway. November 1995.
ISBN 82-7761-003-3
Contents part I:
Acknowledgements
Introduction
X-ray microanalysis
Bacterial volume
Bacterial macromolecules and elemental composition
Bacterial growt
Heterogeneity of bacteria
Extracellular structures
Cytoplasmic ions
Concluding remarks
References
Part II: List of papers
Paper I: Norland S, Fagerbakke K M & Heldal M (1995) Light
element analysis of individual bacteria by X-ray microanalysis (XRMA).
Applied and Environmental Microbiology 61: 13 5 7-13 62. Abstract
Paper II: Fagerbakke KM, Heldal M & Norland S (1996) Content
of carbon, nitrogen, oxygen, sulfur and phosphorus in native aquatic and
cultured bacteria. Accepted - Aquatic Microbial Ecology. Abstract
Paper III: Tuomi P, Fagerbakke K M, Bratbak G & Heldal M (1995)
Nutritional enrichment of a microbial community: The effects on activity,
elemental composition, community structure and virus production. -
FEMS Microbiology Ecology 79: 123 -13 4. Abstract
Paper IV: Fagerbakke K M, Heldal M & Norland S (1991) Variation
in elemental content among and within trichomes in Nostoc calcicola
79WA01 measured by X-ray microanalysis. - FEMS Microbiology Letters
81: 227-232.
Paper V: Fagerbakke K M, Heldal M, Norland S, Heimdal BR & Båtvik
H (1994) Emiliania huxleyi: Chemical composition and size of
coccoliths from enclosures and a Norwegian fjord. - Sarsia 79:
349355. Abstract
Paper I
Abstract A method based on X-ray microanalysis (XRMA) with the
transmission electron microscope for measure ment of total amounts of elements
in single microbial cells has been developed. All major elements in cells
except hydrogen can be measured simultaneously, XRMA provided N/C ratios
(means +/- standard errors of the mean) for stationary-phase and growing
Escherichia coli of 0.23 +/- 0.01 and 0.30 +/- 0.01, respectively,
while CHN analysis gave values of 0.276 and 0.307, respectively, for samples
from the same cultures. Analyses of free coccoliths from Emiliana huxleyi
provided weight fractions close to those of CaCO3: 0.35 +/-
0.01, 0.15 +/- 0.01, and 0.47 +/- 0.01 for calcium, carbon, and oxygen,
respectively. Calibration is based on monodisperse latex beads and on microdrops
of defined compounds. Elements in particles in the size range from 5 fg
to 500 pg are measured with a relative precision between 500 and 5,000
ppm, depending on size. As a single-cell method, XRMA avoids the shortcomings
of commonly used fractionation techniques associated with bulk methods,
which are based on centrifugation or filtration, On the basis of morphology
and XRMA, particles may be classified more precisely into groups (e.g.,
biotic versus abiotic) than is possible by bulk methods. Single-cell elemental
analysis may provide insight into topics like nutritional and energetic
status, macromolecular composition, and (by multivariate statistics) community
structure.
Keyword : dry-matter , electron-microscopy , fresh-water , biomass
, marine , phosphorus , biovolume , growth , ratio , bacterioplankton
Paper II
Abstract The content of carbon, nitrogen, oxygen, phosphorus
and sulfur was measured in individual cells from 6 native aquatic samples
and 4 samples of cultured bacteria by X-ray microanalysis using a transmission
electron microscope (TEM). The molar C:N:P ratio for the pooled sample
was 50:10:1. From length and width measurements of unfixed air-dried cells
we estimated cell volumes over a total range of 0.0026 to 15.8 
m(3),
and mean C:volume ratios of 30 to 162 fg m(-3)
for the samples included. For the marine samples we found mean N:C ratios
of 0.25 to 0.28, while cells from fresh or brackish waters had mean N:C
ratios of 0.17 to 0.20, indicating differences in nutrient availability.
The P:C ratios for the samples analyzed varied from 0.040 to 0.090, with
a pooled mean of 0.052, which is approximately twice that of the Redfield
ratio for P:C. For O:C ratios we estimated a pooled mean of 0.37 and a
range of 0.22 to 0.77 for all samples. We may conclude that slow-growing
or non-growing cells have low O:C ratios. The mean S:C ratio for all samples
was 0.031, with a range of 0.016 to 0.084 for the sample means. A general
conclusion is that single-cell analyses of elemental composition give important
information on the physiological conditions of cells and on possible nutrient
limitations. The rationale for this is the assumption that changes in macromolecular
composition are due to nutrient availability.
Keyword : x-ray microanalysis , carbon , nitrogen , oxygen ,
sulfur , phosphorus , volume , ratios
Paper III
Abstract Viruses are active members of the microbial community
in natural waters but little is known about the factors that regulate their
activity and production. In this study we have investigated the effects
of increased availability of organic nutrients and inorganic phosphate
on activity, elemental composition, community structure and virus production
in a natural bacterial community. The fraction of active cells in the community
as estimated from microautoradiography of cells assimilating 3H-labeled
thymidine ranged from 0-22%, but changes in the elemental composition of
the cells indicated that more than 90% of the cells were active. The increase
in carbon and energy availability stimulated virus production more than
bacterial biomass production, while the increase in phosphate availability
stimulated biomass production rather than virus production. A decrease
in morphological diversity of the bacterial community was paralleled by
a reduction in the virus-to-bacteria ratio (VER) but the relationship between
bacterial diversity and viral activity is uncertain. Our general conclusion
is that nutrient availability, in addition to the bacterial activity, also
affects the viral activity, and that both of these may affect the structure
and diversity of the bacterial community.
Keyword : bacteria , phage , virus , virus-like-particle , vlp
, nutrient , x-ray microanalysis , activity
Paper V
Abstract Single Emiliania huxleyi (Lohmann) Hay et Mohler,
type A, coccoliths harvested from mesocosm enclosures (June 1991) with
different nutrient regimes, and the Norwegian fjord, Samnangerfjorden (October
1992), were analyzed with X-ray microanalysis in a Transmission Electron
Microscope (TEM). The average molar Ca : C ratios of the free coccoliths
varied within a range of 0.65-1.0, dependent on growth condition of E.
huxleyi. Free coccoliths sampled from enclosures with high production
or density of E. huxleyi, had Ca : C ratios near 1. Coccoliths sampled
from Samnangerfjorden, and enclosures with low production or cell density
the Ca : C ratios were close to 0.7. The excess of carbon in the coccoliths
compared to CaCO3 is probably due to organic matter, mainly
carbohydrates. It is concluded that the amount of organic matter associated
with coccoliths of E. huxleyi, is dependent on the growth conditions,
and carbohydrate may protect the coccoliths from dissolution. A higher
Ca : C ratio (0.8 versus 0.7) was determined in the central parts of free
coccoliths from slow growing algae. Average content of carbon, oxygen and
calcium in single coccoliths was 0.28, 0.87, and 0.67 pg (1.4 : 3.2 : 1
molar ratio) respectively, with an estimated average total weight of 1.8
pg.
Keywords : transmission electron-microscopy , inorganic carbon
, huxleyi lohmann , prymnesiophyceae , photosynthesis , calcification ,
haptophyceae , deposition , bacteria , bloom
Dissertation Database
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