Laboratory 11 - Lab Report PDF

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Lydia Rankin BMS 504.07 March 5th, 2019 Laboratory 11: Anaerobic Microbial Growth Purpose: The purpose of this lab was to examine the effect of oxygen on microbacteria. Materials/Methods: Exercise one was testing oxygen requirements of 5 different bacteria on thioglycolate broth. First, 5 tubes containing thioglycolate broth were obtained and labeled correctly. The 5 bacteria being examined, Escherichia coli, Clostridium sporogenes, Micrococcus luteus, Bacillus subtilis, and Bacillus cereus were aseptically transferred into the corresponding thioglycolate tube by inserting an inoculating loop into the media two-thirds of the way down being cautious not to touch the bottom of the tube. It was important to hold the tubes carefully to avoid shaking. The caps were loosely placed on the tubes which were incubated at 37 ∘ C for 1-2 days. Exercise two utilized nutrient agar media plates to test oxygen requirements for all 5 microorganisms. Two nutrient agar plates were obtained and divided into 5 equal sections as shown in diagram 1. Using aseptic technique and fishtailing technique, each section of the 2 plates were inoculated with the corresponding bacterial species. One nutrient agar plate was incubated at 37 ∘ C under aerobic conditions and the other was incubated at 37 ∘ C under anaerobic conditions by placing the plate in an anaerobic chamber with a GasPak. Diagram 1.

B c

E c

C s

M l B s

Results: Results from exercise 1 are displayed in Table 1 below. According to the observations E. coli, C. sporogenes, and B. cereus are facultative anaerobes, and M. luteus and B. subtilis are obligate aerobes. Results from exercise 2 are displayed in Table 2 below.

Table 1. Exercise 1- Testing oxygen requirements on thioglycolate broth

Ec

Cs

Ml

Bs

Bc

Observation

Cloudy

Cloudy

Clear

Clear

Cloudy

Results

Facultative Anaerobe

Facultative Anaerobe

Obligate Aerobe

Obligate Aerobe

Facultative Anaerobe

Picture

Table 2. Exercise 2- Testing oxygen requirements on nutrient agar media plates Aerobic Conditions

Anaerobic Conditions

Ec

Decent growth

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Cs

Small dots of growth

Small growth

Ml

Yellow streak

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Bs

Thick growth

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Bc

LOTS of growth, completely cloudy

Tiny growth

Discussion: On nutrient agar plates E. coli can be grown in aerobic conditions only indicating it is most likely an obligate aerobe. Thioglycollate tubes show that E. coli is a facultative anaerobe because growth is best at the top of the tube where oxygen is most present but still is present throughout the tube. These results do not correspond. E. coli is a facultative anaerobe meaning there is an error in the nutrient agar growth. On nutrient agar plates C. sporogenes can be grown in aerobic and anaerobic conditions indicating it is most likely a facultative anaerobe. Thioglycollate tubes show that C. sporogenes is a facultative anaerobe because growth is best at the top of the tube where oxygen is most present but still is present throughout the tube. These results do correspond with each other but are incorrect since C. sporogenes is an obligate anaerobe. On nutrient agar plates B. cereus can be grown in aerobic and anaerobic conditions indicating it

is most likely a facultative anaerobe. Thioglycollate tubes show that B. cereus is a facultative anaerobe because growth is best at the top of the tube where oxygen is most present but still is present throughout the tube. These results do correspond with each other and are correct. On nutrient agar plates M. luteus can be grown in aerobic conditions only indicating it is most likely an obligate aerobe. Thioglycollate tubes show that M. luteus is an obligate aerobe because growth only occurs at the top of the tube where oxygen is present. These results do correspond with each other and are correct. On nutrient agar plates B. subtilis can be grown in aerobic conditions only indicating it is most likely an obligate aerobe. Thioglycollate tubes show that B. subtilis is an obligate aerobe because growth only occurs at the top of the tube where oxygen is present. These results do correspond with each other and are correct. Lab Questions: 1. Biologically an anaerobic microorganism is one that forms ATP without oxygen. The respiratory electron transport chain is utilized but the final electron acceptors are molecules such as sulfate, nitrate, or sulfur instead of oxygen. Anaerobic microorganisms lack oxygen detoxifying enzymes superoxide dismutase and catalase resulting in growth inhibition or even death when in the presence of oxygen. In contrast, aerobic microorganisms utilize oxygen as the final electron acceptor in the respiratory electron transport chain and do have oxygen detoxifying enzymes catalase which converts H 2 O 2 to H 2 O and O 2 and superoxide dismutase which converts superoxide radicals to H 2 O 2 or O 2 ....