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a) A photograph of a spectrophotometer; a machine with a keypad and a place to put a sample. b) A diagram explaining how the spectrophotometer measures turbidity. A light source passes through a prism and a single wavelength is selected. If this beam of light passes through a clear broth, most of the light reaches the detector. If this beam of light passes through a cloudy sample, much of the light is reflected and refracted and only a little bit of light reaches the detector.
(a) A spectrophotometer is commonly used to measure the turbidity of a bacterial cell suspension as an indirect measure of cell density. (b) A spectrophotometer works by splitting white light from a source into a spectrum. The spectrophotometer allows choice of the wavelength of light to use for the measurement. The optical density (turbidity) of the sample will depend on the wavelength, so once one wavelength is chosen, it must be used consistently. The filtered light passes through the sample (or a control with only medium) and the light intensity is measured by a detector. The light passing into a suspension of bacteria is scattered by the cells in such a way that some fraction of it never reaches the detector. This scattering happens to a far lesser degree in the control tube with only the medium. (credit a: modification of work by Hwang HS, Kim MS; credit b “test tube photos”: modification of work by Suzanne Wakim)

Measuring dry weight of a culture sample is another indirect method of evaluating culture density without directly measuring cell counts. The cell suspension used for weighing must be concentrated by filtration or centrifugation, washed, and then dried before the measurements are taken. The degree of drying must be standardized to account for residual water content. This method is especially useful for filamentous microorganisms, which are difficult to enumerate by direct or viable plate count.

As we have seen, methods to estimate viable cell numbers can be labor intensive and take time because cells must be grown. Recently, indirect ways of measuring live cells have been developed that are both fast and easy to implement. These methods measure cell activity by following the production of metabolic products or disappearance of reactants. Adenosine triphosphate (ATP) formation, biosynthesis of proteins and nucleic acids, and consumption of oxygen can all be monitored to estimate the number of cells.

  • What is the purpose of a calibration curve when estimating cell count from turbidity measurements?
  • What are the newer indirect methods of counting live cells?

Alternative patterns of cell division

Binary fission is the most common pattern of cell division in prokaryotes, but it is not the only one. Other mechanisms usually involve asymmetrical division (as in budding) or production of spores in aerial filaments.

In some cyanobacteria , many nucleoids may accumulate in an enlarged round cell or along a filament, leading to the generation of many new cells at once. The new cells often split from the parent filament and float away in a process called fragmentation ( [link] ). Fragmentation is commonly observed in the Actinomycetes , a group of gram-positive, anaerobic bacteria commonly found in soil. Another curious example of cell division in prokaryotes, reminiscent of live birth in animals, is exhibited by the giant bacterium Epulopiscium . Several daughter cells grow fully in the parent cell, which eventually disintegrates, releasing the new cells to the environment. Other species may form a long narrow extension at one pole in a process called budding . The tip of the extension swells and forms a smaller cell, the bud that eventually detaches from the parent cell. Budding is most common in yeast ( [link] ), but it is also observed in prosthecate bacteria and some cyanobacteria.

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Source:  OpenStax, Microbiology. OpenStax CNX. Nov 01, 2016 Download for free at http://cnx.org/content/col12087/1.4
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