Design an experiment for measuring the activity of sulfuroxidizing bacteria in soil. If only certain species of the sulfur oxidizers present were
To discuss:
Design an experiment to measure the activity of sulfur-oxidizing bacteria in the soil. How to detect, if only certain sulfur oxidizing species present were metabolically active. How to prove the activity measurement was because of biological activity.
Introduction:
Sulfur oxidizing bacteria (for example, Thiobacillus, Thermothrix,Sulfolobus, and Thiothrix) can convert hydrogen sulfide (H2S) into sulfate (SO42) or other forms of sulfur. They can grow in various environments, for example, soil and water. MAR-FISH is a fluorescence in situ hybridization (FISH) combined with microautoradiography (MAR). MAR-FISH can simultaneously identify microorganisms and their metabolic property.
Explanation of Solution
An experiment to measure sulfur-oxidizing bacteria from soil:
- Isolate Thiobacillus species (gram-negative bacteria) from sulfide rich soil (for example, paddy field) followed by serial dilution in water.
- Add 5 ml water containing sample (bacteria) to 50 ml thiosulfate mineral salts medium (thiosulfate MSM).
- Incubate the medium at 30°C for 7 days. The formation of turbidity and reduced pH of the thiosulfate MSM medium indicates the bacterial growth.
- Further streak plate method (onto the solid medium of thiosulfate MSM) can be used to isolate the pure culture of Thiobacilli.
- Incubate the petri plate at 30°C for 7 days.
- Screen the purified culture of Thiobacilli in the MSM consisting of Na2S but not of Na2S2O3.
- The sulfide medium used for the growth of bacteria is the sole source of energy.
- Further, Thiobacillus can be confirmed by gram-staining.
- Sulfur-oxidizing bacteria, such as Thiobacillus species can be used to study.
To measure the activity of sulfur oxidizing bacteria in the soil, the radioisotope H235S should be used. The soil should be incubated with radioisotope (H235S) to know whether the sulfide oxidation occurred or not. After incubation, the presence of 35SO42- should be analyzed to prove the activity of sulfur-oxidizing bacteria in the soil. The presence of 35SO42- proves that the sulfide oxidation has occurred in the test. To prove that the oxidation occurred due to biological activity, sterile the soil (medium) before the addition of labeled H2S. This is used as a control in this experiment. If the oxidation activity observed in the experiment is biological, it should be absent in the control. The MAR-FISH can be used to identify the presence of metabolically active bacteria in the soil.
MAR-FISH (microautoradiography- fluorescence in situ hybridization) technique is used to evaluate the microbial identification (phylogeny) with measurement of metabolic activities. MAR-FISH is used to assess the metabolism of a specific radiolabeled substance by microorganisms in the natural sample and also help to identify those particular microorganisms. Therefore, MAR-FISH can be used to identify the metabolism of radioisotope H235S by microorganisms in this experiment.
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