Determination of the Optimal Concentration of Medium Supplements Promoting the Recovery of Dormant Mycobateria in Culture
Location
CSU 253/4/5
Start Date
23-4-2007 1:00 PM
End Date
23-4-2007 3:00 PM
Student's Major
Biological Sciences
Student's College
Science, Engineering and Technology
Mentor's Name
Timothy E. Secott
Mentor's Department
Biological Sciences
Mentor's College
Science, Engineering and Technology
Description
Johne's disease is a chronic intestinal disease of cattle that is caused by Mycobacterium avium subsp. paratuberculosis (Mpt). Diagnostic testing is typically unrewarding until years after infection, consequently leading to massive spread of Mpt and substantial economic loss. Previous experimentation showed that reducing oxygen tension through the use of a reducing agent, Oxyrase, or supplementing the medium with Mpt-conditioned medium improved the recovery of Mpt from dormancy. However, while the concentration of conditioned medium that improved recovery was relatively constant, that of Oxyrase varied widely from experiment to experiment. In order to determine whether this was a consequence of the percent of viable cells in dormant cultures, we used a two-color fluorescence assay to estimate cell viability and compare these data with the concentration of conditioned medium or Oxyrase. A standard curve was prepared for Mpt using a BacLight Live:Dead fluorescence kit. The standard curve was used to calculate the ratio of viable to nonviable organisms in dormant Mpt cultures. Two-fold serial dilutions of Mpt conditioned medium or Oxyrase was added to cultures of dormant Mpt in rich medium and growth was monitored by measuring optical density for up to 21 days. A determination of the relationship between cell viability and the effectiveness of recovery promotants is in progress and will be reported.
Determination of the Optimal Concentration of Medium Supplements Promoting the Recovery of Dormant Mycobateria in Culture
CSU 253/4/5
Johne's disease is a chronic intestinal disease of cattle that is caused by Mycobacterium avium subsp. paratuberculosis (Mpt). Diagnostic testing is typically unrewarding until years after infection, consequently leading to massive spread of Mpt and substantial economic loss. Previous experimentation showed that reducing oxygen tension through the use of a reducing agent, Oxyrase, or supplementing the medium with Mpt-conditioned medium improved the recovery of Mpt from dormancy. However, while the concentration of conditioned medium that improved recovery was relatively constant, that of Oxyrase varied widely from experiment to experiment. In order to determine whether this was a consequence of the percent of viable cells in dormant cultures, we used a two-color fluorescence assay to estimate cell viability and compare these data with the concentration of conditioned medium or Oxyrase. A standard curve was prepared for Mpt using a BacLight Live:Dead fluorescence kit. The standard curve was used to calculate the ratio of viable to nonviable organisms in dormant Mpt cultures. Two-fold serial dilutions of Mpt conditioned medium or Oxyrase was added to cultures of dormant Mpt in rich medium and growth was monitored by measuring optical density for up to 21 days. A determination of the relationship between cell viability and the effectiveness of recovery promotants is in progress and will be reported.
Recommended Citation
Rock, Kelly E.. "Determination of the Optimal Concentration of Medium Supplements Promoting the Recovery of Dormant Mycobateria in Culture." Undergraduate Research Symposium, Mankato, MN, April 23, 2007.
https://cornerstone.lib.mnsu.edu/urs/2007/poster-session-A/5
