Buried Slide Technique

This technique for the direct microscopic observation of soil microorganisms is described in Methods for Studying the Ecology of Soil Micro-organisms (Parkinson et al. 1971).
  1. Clean the microscope slides in acid alcohol and heat sterilized them.
  2. Slits are made in a soil sample with a sterile knife (dip knife blade in alcohol and flame).
  3.  

  4. Insert the slides into slits made with a sterile knife in the soil to be sampled.
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  6. Once the slides are inserted, the soil should be firmed up around the slide to bring it into close contact.
  7. The soil is then covered (a petri dish cover or plastic wrap may be used) and labeled.
  8. The slides should remain is in place for 1 - 3 weeks after which the slide is removed from the soil. This is done by removing the soil plug from its container and carefully breaking the soil away from the slide
  9.  

  10. One side of the slide is wiped clean with a paper towel.
  11. The slide is then heat fixed and stained with an appropriate stain (phenolic aniline blue is recommended.
  12. Alternatively the slide can be gram stained by the traditional technique or by the flow through technique shown elsewhere in this collection.
  13.  

In this exercise the slide comes in contact with the soil moisture and organisms representative of the soil population will form a biofilm on the slide. Parkinson (1971), however, notes that one should not assume that the microbial population observed on the glass slide is similar to that in the undisturbed soil. The glass slide alters the dynamics of the soil ecosystem by providing a new and novel substrate. The organisms that colonize the slide can be assumed to be soil microorganisms but the proportion, distribution and relationships of the organisms may be drastically altered.

 This image shows a mixed bacterial population that has developed on the surface of a soil crumb or aggregate. The bacteria are stained with acridine orange and are seen fluorescing under UV light. The bacteria exist in large groups of similar colonies or sheets of bacteria. The colonies are usually one cell thick. Their exact location may be related to food substrates in the soil material. Some isolated bacteria either have been split off from larger colonies or may have been deposited on the soil crumb surface from the soil water.

The film containing the bacterial colonies is removed from the surface of a fresh soil aggregate by placing the aggregate onto a sintered glass filter funnel and very gently wetting the aggregate from below. As the water film moves up the soil aggregate, the surface film is removed and transferred onto the water surface. The water level is raised further until it is accessible at the top. If the filter and film are transferred onto a very clean microscope slide by touching the slide carefully to the water surface, the film containing the bacteria adheres very firmly to the slide and can be gently rinsed and stained. The spatial relationships of colonies are well preserved if care is taken (from the ASM Biofilms collection).

References:

Harris, P.J. 1972. Micro-organisms in surface films from soil crumbs. Soil
Biol. Biochem. 4:105-106.

Parkinson, D., T.R.G. Gray and S.T. Williams, Eds., 1971, Methods for Studying the Ecology of Soil Micro-organisms, Blackwell Scientific Publications, Oxford p 30-31.