LAB 1: INTRODUCTION TO THE MICROSCOPE AND COMPARISON OF SIZES AND SHAPES OF MICROORGANISMS

 

This assignment must be completed prior to attending this lab!

Before attending this lab you must:

1. View the following YouTube videos

2. Complete the Lab 1 Assignment Quiz found on your Blackboard site.

This quiz will be part of your core points for this lab. Quizzes must be completed at least 2 hours prior to the start of your lab. The purpose of the videos and the quiz is to prepare you for performing this lab. Therefore, late quizzes are not accepted.

Video 1: Preliminary Tips for Using a Microscope

Video 2: Focusing Using Oil Immersion (1000X) Microscopy

Video 3: Focusing Using Low Magnifications: Using the 10X Objective

 

DISCUSSION

In this lab, you will become familiar with the use of the microscope (particularly oil immersion microscopy) and will compare the relative size and shape of various microorganisms.

A. BACTERIAL SHAPES, ARRANGEMENTS, AND FORMS

Bacteria are unicellular prokaryotic microorganisms that divide by binary fission, a process by which one bacterium splits into two.

YouTube movie showing binary fission in bacteria.
YouTube movie showing fluorescing imaging of binary fission in bacteria.

 

Review of Prokaryotic and Eukaryotic Cells from Unit 1.

There are three common shapes of bacteria:

The cocci come in 5 different arrangements; the bacilli in 3 different arrangements; and the spirals in 3 different forms.

1. Coccus

A coccus-shaped bacterium is usually spherical, although some appear oval, elongated, or flattened on one side. Most cocci are approximately 0.5 - 1.0 micrometer (µm) in diameter and may be seen, based on their planes of division and tendency to remain attached after replication, in one of the following arrangements (see Fig. 1A):

a. Division in one plane produces either a diplococcus (see Fig. 1A and Fig. 1B) or streptococcus (see Fig. 1A and Fig. 1C) arrangement.

2. streptococcus: a chain of cocci
- Photomicrograph of a streptococcus
- Scanning electron micrograph of Streptococcus pyogenes
-Scanning Electron Micrograph of Enterococcus

b. Division in two planes produces a tetrad arrangement (see Fig. 1A and Fig. 1D).

c. Division in three planes produces a sarcina arrangement (see Fig. 1A).

    sarcina: a cube of 8 cocci
    - Photomicrograph of a sarcina

It is difficult with a conventional light microscope to tell a tetrad arrangement (square of four cocci) from a sarcina arrangement (cube of eight) so in our lab, anytime you see ba square of four cocci, say it is either a tetrad or a sarcina arrangement.

d. Division in random planes produces a staphylococcus arrangement (see Fig. 1A and Fig. 1E).

staphylococcus: cocci in irregular, often grape-like clusters
- Photomicrograph of a staphylococcus, negative image
-
Scanning electron micrograph of Staphylococcus aureus
- Scanning electron micrograph of methicillin-resistant Staphylococcus aureus (MRSA)

As you observe these different cocci, keep in mind that the procedures used in slide preparation may cause some arrangements to break apart or clump together (see Figs. 1D and 1E). The correct form, however, should predominate. Also remember that each coccus in an arrangement represents a complete, individual, one-celled organism.

2. Bacillus (rod)

A bacillus or rod is a hotdog-shaped bacterium having one of the following arrangements (see Fig 2A):

a. bacillus: a single bacillus (see Fig 2A and Fig 2B)
- Photomicrograph of a bacillus
- Scanning electron micrograph of a Pseudomonas aeruginosa, a bacillus

- Scanning electron micrograph of Escherichia coli O157H7, a bacillus

b. streptobacillus: bacilli in chains (see Fig 2A and Fig 2C)
- Photomicrograph of a streptobacillus

c. coccobacillus: oval and similar to a coccus (see Fig 2A, 2D, and 2E)

A single bacillus is typically 0.5-1.0 µm wide and from 1- 4 µm long. Small bacilli or bacilli that are dividing or have just divided by binary fission may at first glance be confused for diplococci or cocci (see Fig. 2A) so they must be observed carefully. You will, however, be able to see bacilli that have not divided and are definitely rod-shaped as well as bacilli in the process of dividing.

3. Spiral

Spiral-shaped bacteria occur in one of three forms (see Fig. 3A):

a. vibrio: an incomplete spiral or comma-shaped (see Fig. 3A and Fig. 3B)
- Photomicrograph of a vibrio
-
Scanning electron micrograph of Vibrio cholerae

b. spirillum: a thick, rigid spiral (see Fig. 3A and Fig. 3C)
- Photomicrograph of a spirillum

c. spirochete: a thin, flexible spiral (see Fig. 3A and Fig. 3D)
-
Photomicrograph of a spirochete
- Scanning electron micrograph of the spirochete Leptospira
; courtesy of CDC
- scanning electron micrograph of the spirochete Treponema pallidum; courtesy of CDC

The spirals you will observe range from 5-40 µm long but some are over 100 µm in length. The spirochetes are the thinnest of the bacteria, often having a width of only 0.25-0.5 µm.

To view a nice interactive illustration comparing size of cells and microbes, see the Cell Size and Scale Resource at the University of Utah. (Genetic Science Learning Center. (2010, September 2) Cell Size and Scale. Retrieved October 19, 2017, from http://learn.genetics.utah.edu/content/cells/scale/ )

YouTube movie illustrating Size Comparison of Microorganisms created by Gracia Alvaro Montoya, MetaBallsStudios (MBS), United Kingdom, Nov., 2017

 

Concept map for Lab 1

B. YEASTS

Yeasts, such as the common baker's yeast Saccharomyces cerevisiae (see Fig. 4), are unicellular fungi. They usually appear spherical and have a diameter of 3 - 5 µm. Yeasts commonly reproduce asexually by a process called budding. Unlike bacteria, which are prokaryotic, yeasts are eukaryotic.

- Scanning electron micrograph of Saccharomyces cerevisiae

To view a nice interactive illustration comparing size of cells and microbes, see the Cell Size and Scale Resource at the University of Utah. (Genetic Science Learning Center. (2010, September 2) Cell Size and Scale. Retrieved October 19, 2017, from http://learn.genetics.utah.edu/content/cells/scale/ )

YouTube movie illustrating Size Comparison of Microorganisms created by Gracia Alvaro Montoya, MetaBallsStudios (MBS), United Kingdom, Nov., 2017

 

C. MEASUREMENT OF MICROORGANISMS

The approximate size of a microorganism can be determined using an ocular micrometer (see Fig. 5) , an eyepiece that contains a scale that will appear superimposed upon the focused specimen.

To view a nice interactive illustration comparing size of cells and microbes, see the Cell Size and Scale Resource at the University of Utah.

D. FOCUSING

Focusing With The 1000X Oil Immersion Objective - Olympus CX31 Microscope (see Fig. 7)

 

Videos reviewing techniques used in this lab:
Preliminary Tips for Using a Microscope
Focusing Using Oil Immersion (1000X) Microscopy
Focusing Using Low Magnifications: Using the 10X Objective

 

Focusing With The 10X Objective - Olympus CX31 Microscope (see Fig. 7)

 

Concept map for Lab 1

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SPECIMENS

Prepared slides of the following bacteria:

On-line demonstration slides of the following bacteria:

Broth culture of Saccharomyces cerevisiae

Human hair

 

PROCEDURE

TIPS FOR MICROSCOPIC OBSERVATIONS

Move the slide around until you see an area representing the true arrangement of each organism. Also remember:

1. Using oil immersion microscopy (1000X), observe and measure the bacteria listed below.

a. Staphylococcus aureus: Staphylococcus species, as the genus name implies, are cocci possessing a staphylococcus arrangement (cocci in irregular, often grape-like clusters). Measure the diameter of a single coccus.

b. Escherichia coli: Escherichia coli is a small bacillus. Estimate the length and width of a typical rod.

c. Treponema pallidum: Treponema pallidum is a spirochete - a thin, flexible spiral. On this slide you are examining a direct stain ofTreponema pallidum, the bacterium that causes syphilis. Measure the length and width of a typical spirochete.

d. Spirillum: Spirillum species appear as thick, rigid spirals. Measure the length and width of a typical spirillum.

When finished, remove the oil from the prepared slides using either a paper towel or a Kim wipe and return them to their proper tray.

2. Observe the on-line demonstration slides of the following bacteria:

a. Micrococcus luteus: Micrococcus luteus can appear as tetrads or cubes of 8. This strain is a coccus usually exhibiting a tetrad or a sarcina arrangement. Note the shape and arrangement.

b. Neisseria gonorrhoeae: Neisseria species are cocci usually having a diplococcus arrangement. Note the shape and arrangement.

c. Streptococcus pyogenes: Streptococcus species, as the genus name implies, are cocci that usually possess a streptococcus arrangement (cocci in chains). Note the shape and arrangement.

d. Bacillus megaterium: Bacillus megaterium appears as large bacilli in chains (a streptobacillus). Note the shape and arrangement.

3. Prepare a wet mount of baker's yeast (Saccharomyces cerevisiae).

a. Using a pipette, put a small drop of the yeast culture on a microscope slide and place a cover slip over the drop.

b. Using your iris diaphragm lever, reduce the light for improved contrast by moving the lever almost all the way to the right.

c. Observe using oil immersion microscopy. Measure the diameter of a typical yeast cell.

d. When finished, discard the coverslip in the broken glass disposal container, wash the slide with water and dry with a paper towel, and use the same slide again for step 4.

4. Prepare a wet mount of your hair.

a. Remove a small piece of a hair from your head and place it in a small drop of water on a slide.

b. Place a cover slip over the drop and observe using oil immersion microscopy.

c. Measure the diameter of your hair and compare this with the size of each of the bacteria and the yeast observed in steps 1-3.

d. Discard the slide and coverslip in the broken glass disposal container.

5. At the completion of the lab, remove the oil from the oil immersion objective using lens paper and put your microscope away.

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RESULTS

1. Make drawings of several of the bacteria from each of the four prepared slides and indicate their approximate size in micrometers.


Diameter = _____ micrometers

Shape =

Arrangement =

 

Length = _____ micrometers

Width = _____ micrometers

Shape =

 

Length = _____ micrometers

Width = _____ micrometers

Shape =

Form =

Length = _____ micrometers

Width = _____ micrometers

Shape =

Form =



2. Make drawings of several of the bacteria from each of the four demonstration slides and indicate their approximate size in micrometers.

Shape =

Arrangement =

 

Shape =

Arrangement =

 

Shape =

Arrangement =

Shape =

Arrangement =

 



3. Make a drawing of several yeast cells and indicate their size in micrometers.

 

Diameter = _____ micrometers

 

4. Make a drawing indicating the size of the bacteria and yeast observed above relative to the diameter of your hair.

 


hair

 

Diameter = _____ micrometers

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PERFORMANCE OBJECTIVES FOR LAB 1

After completing this lab, the student will be able to perform the following objectives:

DISCUSSION

RESULTS

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SELF-QUIZ

Self-quiz

Answers

 

Practice Slides: Shapes, Arrangements, and Forms

Slide 1 (ans) Slide 9 (ans)
Slide 2 (ans) Slide 10 (ans)
Slide 3 (ans) Slide 11 (ans)
Slide 4 (ans) Slide 12 (ans)
Slide 5 (ans) Slide 13 (ans)
Slide 6 (ans) Slide 14 (ans)
Slide 7 (ans) Slide 15 (ans)
Slide 8 (ans)    

 

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Lab Manual Table of Contents


Creative Commons License
Microbiology Laboratory Manual by Gary E. Kaiser, PhD, Professor of Microbiology
is licensed under a Creative Commons Attribution 4.0 International License.

Last updated: August, 2017
Please send comments and inquiries to Dr. Gary Kaiser