BLT Stages

Abstract bachelor project 2017-2018: Comparative evaluation of different growth media for mycobacteria from specimens of animal origin

Background: Bovine Tuberculosis (BTB) is a chronic, debilitating disease caused by Mycobacterium bovis (M. bovis) that mostly resides in cattle. The bacteria affects the productivity of the animals and can influence the international market trading. Therefore detection of it becomes of high economic relevance. The best way of detection needs to be found so comparing the solid and liquid media directly gives a better understanding on what method should be used.

Aim: At the moment the University of Pretoria uses LJ-pyruvate slants for detection purposes. This is a solid media giving clear results after ten weeks and is made in such a way that it promotes growth of M. bovis and suppresses other growth.
The media that it will be compared to is the liquid Mycobacteria growth indicator tube (MGIT). A promising method that should give quicker results and does the controlling of growth automatically. Comparing these two methods in a controlled environment can give more insight on how they perform in practise.

Methodology: The method that will be used consist of either tissue samples or milk samples being homogenated and decontaminated before being inoculated on the media. The inoculation is then either on the LJ-pyruvate slants or in the MGIT tubes. The LJ-slants go into the incubator and will be monitored every week to notice any growth and the MGIT tubes go into the BACTEC® MGIT 320 machine where they will be automatically monitored. The samples that are being used were either previously redeemed negative or positive giving an idea on what growth to expect. Also a few milk samples will be used to have a different form of sample as well. If an off white colony forms on the LJ-pyruvate slants with an acid-fast look after Ziehl-Neelsen staining, it can be confirmed for M. bovis because it can be traced back to those previous results. The MGIT results are a bit more difficult because no colony assessment can be made from the liquid media. That is why these positive acid-fast samples will be controlled with Polymerase Chain Reaction (PCR).

Results: LJ-pyruvate slants got results as expected. The previously positive samples were not all positive now due to the fact that they have been stored away for a long time in a freezer and most of the bacteria could be dead now. A few of the MGIT samples first turned out positive on the machine, but when further PCR analysis was done they did not turn out to be M. bovis or even from the Mycobacterium genus. In the end six samples were positive for M. bovis on just one LJ-pyruvate slant and ten samples on both of them. Only one sample was just positive on the MGIT tube and lastly one sample was found positive for M. bovis on the two LJ-slants and the MGIT tube.

Conclusion: It can be concluded that the LJ-pyruvate slants are still more superior over the MGIT tubes for finding M. bovis growth when the normal protocol is used. In the future an altered broth can be used in the MGIT tubes for encouraging more M. bovis growth. A different alternative could be using a different form of PCR that can detect the bacteria that were now not detected in this research.