Collecting Soil Biofilms by the Buried Slide Technique

Instructor Version (go to Student Version)

Subject Area(s)	microbiology
Intended Audience	high school biology, independent study/science fair, introductory undergraduate microbiology, advanced college level
Type	laboratory exercise
Revision Date	April 8, 2008

CONTENT

This exercise describes a method for collecting soil biofilms on glass slides through direct contact between soil and microscope slide, in the field or in the laboratory. This technique is described in “Methods for Studying the Ecology of Soil Micro-organisms” (Parkinson et al.1971). In this exercise a microscope slide is placed in contact with soil. Organisms representative of the soil population will form a biofilm on the slide. Parkinson (1971) cautions not to assume that the microbial population observed on the glass slide is similar to the undisturbed soil. The glass slide alters the dynamics of the soil ecosystem by providing a new and novel substratum. 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.

PREREQUISITES

Students should be able to define a biofilm, describe the differences between biofilm (surface-attached) and planktonic (free-floating) bacteria, and describe why bacteria tend to grow on surfaces. Some knowledge of soil structure is also useful.

INSTRUCTIONAL OBJECTIVE

Given readily accessible materials, detailed instructions and appropriate drawings, a student will be able to collect biofilms containing soil organisms on standard glass microscope slides.

INSTRUCTIONAL PROCEDURES

1. Students will be provided with a complete set of materials required for collecting soil biofilms along with detailed instructions and diagrams for carrying out this procedure.
2. To enumerate cells collected from soil, there are preprinted slides such as those available from Erie Scientific Co. These slides have circular areas of known surface area (πr²). Using these as coupons, the student can know precisely the area from which cells are being harvested. See: http://www.eriesci.com/index.aspx.
3. After 1-3 weeks, students can retrieve the buried slides and stain them for observation under a microscope.
4. Classical simple and differential stains such as Crystal violet, Gram stain and spore stain may be used on these slides.
   
   Note: Since biofilms are 98% water, staining without fixation is preferred. Heat fixation dehydrates the slide so that the structure of the biofilm is greatly altered.

Students can use the “Flow Through Gram Stain” described elsewhere in this collection or any of a number of stains for visualizing the soil biofilms. Ruthenium red and Alcian blue are recommended for staining extracellular polysaccharides.

MATERIALS AND EQUIPMENT

Quantity	Description
1	1 x 3 inch glass microscope slides, cleaned and sterile
1	sterile forceps
1	long bladed spatula or knife
1	staining kits
1	microscope
1	clay or plastic flowerpot or a plastic cup

ASSESSMENT / EVALUATION

Assessment may be made by the instructor through a visual evaluation of each student's stained slides.

FOLLOW-UP ACTIVITIES

The biofilms prepared in this exercise may be used in a variety of other ways. Students may enumerate the cells on the slide or measure the thickness of the biofilm.

Companion exercises:

Drop Plate Method for Counting Biofilm Cells
Harvesting and Dispersing of Cells from Biofilms (Standard Method)
Harvesting and Dispersing of Cells from Biofilms (Alternative Method)
Measurement of Biofilm Thickness
Isolation of Biofilm Populations from Soil Crumbs by Flotation
Flow Through Gram Stain

REFERENCES

Microorganisms in surface films from soil crumbs
Harris PJ
Soil Biol Biochem 1972; 4:105-106

Methods for Studying the Ecology of Soil Micro-organisms
in Parkinson D, Gray TRG, Williams ST (eds.)
Blackwell Scientific Publications, Oxford, 1971, pp 30-31

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Educational Program Curricula and Teaching Resources

Supported in part by the Waksman Foundation for Microbiology
Developed in collaboration with Dr. John Lennox, Penn State University-Altoona
© 1999-2008 Center for Biofilm Engineering, http://www.biofilm.montana.edu