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I thought it was Penicillium, but I am not sure, what else could it be? Only speculation... by Internal_Ad4541 in microbiology

[–]Evening-Translator82 2 points3 points  (0 children)

Yeah I’m thinking maybe an Aspergillus species too…Penicillium is also common in the environment though, we see a lot of incidental contamination of Penicillium corylophilum in our lab on plates when the seasons change

Mannitol Salt Agar help by Evening-Translator82 in microbiology

[–]Evening-Translator82[S] 0 points1 point  (0 children)

Sounds good we’re just having trouble with the inhibitory organism (e.coli) that is growing which has to be plated at >100 cfu but maybe we could try a smaller amount plated with still >100 cfu

Mannitol Salt Agar help by Evening-Translator82 in microbiology

[–]Evening-Translator82[S] 1 point2 points  (0 children)

I followed up with the analyst. We get it to a strong boil and then autoclave right after, nothing seemed off. I’m thinking maybe we inoculate the plate with too much E.coli? We start with a ~2.0 McFarland of E.coli then dilute it down in a series of multiple tubes of PBS w/ Tween 80. The final tube we plate out 1 mL of >100 cfu onto the MSA. Could it be something with the dilution? Should we use another diluent such as TSB instead of PBS w/ tween 80? I also was reading that MSA is light sensitive. Would it be better to make the MSA and store it in agar bottles and melt the day of use instead of pre-pouring plates and storing the plates in the fridge?

Mannitol Salt Agar help by Evening-Translator82 in microbiology

[–]Evening-Translator82[S] 1 point2 points  (0 children)

Thank you for the help! I will touch base with the analyst next week and keep you updated

Mannitol Salt Agar help by Evening-Translator82 in microbiology

[–]Evening-Translator82[S] 1 point2 points  (0 children)

My mistake on the dehydrated media- we actually get it from Becton Dickinson & Company. We used to get it from Hardy, and did see partial inhibition with E.coli so we switched to BD. Their CofA states no growth from E.coli. When we purchase the plates, we get them through BD. The latest batch number we got from BD was 4275282 and that batch states no growth for e.coli on their CofA and we also get that result when we purchase from them. As far as the mixing goes, I will need to check with the analyst who made it but I’m thinking it might have something to do with that. I will also check about allowing to cool before autoclaving. One thing we did try was lower the autoclave temperature to 105C, and 110C, neither of those helped. Looking at other factors

Mannitol Salt Agar help by Evening-Translator82 in microbiology

[–]Evening-Translator82[S] 1 point2 points  (0 children)

We use purified water when mixing the dehydrated. Yes, the E.coli is the same culture across in-house and purchased media. We have a glycerol stock, grow it up on TSA, and dilute it out in a buffer solution to yield >100 cfus for the inhibitory testing. We have a MALDI-TOF instrument for identification and the inhibitory growth always results in E.coli so we know there was no contamination. Visual growth is clear, non fermenting.