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We are trying to grow liquid cultures of Mycobacterium smegmatis as a host for bacteriophages, but we have been having issues with contamination. Our latest attempt went like this:

  • Tuesday 1 pm: Added 20 ml 7H9 broth, 2 ml Albumine-Dextrose solution, 0.02g CaCl2, one colony from master M. smegmatis culture was added to 100 ml Erlenmeyer flask. Incubated in a shaker at 35° C.
  • Wednesday 6pm: Added 20 ul of carbenicillin (should have been added from the top, but the carbenicillin arrived a day late)
  • Thursday 1pm: liquid culture is turbid white and clumpy as expected.
  • Friday 2pm: Culture is still turbid white and clumpy. Inoculated a streak on left half of L-agar plate.
  • Saturday 3pm: Streak plate shows streaks of yellow-white colonies (as expected), but also a blue diffuse contaminant which smells very bad. This is similar to what we've seen in our previous attempt at growing this culture

Now, the question is: What could this blue contaminant be? The original culture protocol used Cycloheximide to control for fungal growth, but ours has not arrived yet.

A related question is. Where could it have come from?

Master M. smegmatis culture

Master M.smegmatis culture

Liquid M. smegmatis culture

enter image description here

Contaminated streak plate

Contaminated streak plate

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Pseudomonas aeruginosa is a likely candidate. This is a common skin organism and cultures/colonies often have a blue/green sheen. Pure cultures generally have a fruity/ester smell.

The blue/green pigment is a combination of pyoverdine and pyocyanin.

Of note is that it is also commonly antibiotic resistant having intrinsic antibiotic resistance to a range of antibiotics, including the penicillin group

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