Culture medium for specific bacteria

[iPeace]

Hi all,
My dog has chronic kidney disease, and takes a probiotic that is meant to help reduce uremic toxins. In the summer, I tried adding the contents of a capsule to a sample of my dog's urine. It turned cloudy within a half hour, and after an hour or so, the smell was greatly reduced, and the microscopic view was full of active bacteria. Very unscientific and uncontrolled experiment, but interesting all the same. Around 10/17 I started using a new lot of the probiotic. My dog is not doing as well, and the options are: progression of the kidney disease, kidney support food that is not what it claims to be, or a probiotic that is not biologically active.

I repeated the prior experiment using the new lot of probiotic. The sample did not get cloudy and the smell did not change. I did not bother with the microscopic view. Possibilities here are: 10 - 15 °F cooler temperature, different growth medium (urine sample), or inactive probiotic.

Seems the next step is to do a proper culture to see what grows. "Experimenting with the Microscope" (Krauter) suggests meat broth in agar-agar as a generally good growth medium, but also says different bacteria need different growth medium. Can anyone suggest a good growth medium for the bacteria listed for this probiotic?
S. thermophilus (KB19)
L. acidophilus (KB27)
B. longum (KB31)

[Alan Smith]

I have been out of the Bacteriology business for a long time, but McConkey Agar comes to mind If you look at a fresh sample you might also see flagellates like E.Coli and Proteus Mirababilis. But it sounds like a case for antbiotics.

https://www.sigmaaldrich.com/GB/en/prod ... gIxzPD_BwE

[Alan Smith]

Sorry- I misread your post. McConkey agar is for urinary bacterial culture. As were my comments about flagellate bacteria If you want to grow typical probiotic bacteria then full-cream milk is a start - providing you have a source of the bacteria you want. I am sure you can find YT videos about making live yoghurt for example. Making milk jelly -with gelatin would enable you to pick out strains of bacteria you want (they grow as spots on the plate) but doing all that requires a reasonable amount of practice and know-how. And an incubator.

[apochronaut]

Here is a systematic study of literature related to probiotic use in humans for renal disease. Probiotics are used to lower levels of certain intestinal compounds, thus taking the pressure off of the kidneys.
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6533949/

One of the characteristics of various bacterial species is that their required temp. for optimal growth varies widely, with published ranges of the 3 species mentioned s.thermophilis 35-42° c.., l.acidopholis 37° c. and b.longum around 42-45 ° c. being close enough but also different enough to require considerably different incubation periods in order for equal populations to grow at various temperatures . You can easily see that although there is some overlap between the three species, it isn't necessarily great and your original test might have favoured one over the other and your subsequent test , if the temp. was significantly lower, none of them .
Cultured or bioengineered strains can also vary widely from their wild cousins in many ways, since cultured strains often emerge out of industrial convenience, so even though b.longum for instance is known to heavily populate the intestines of human babies, b.longum kb31 might not be capable of competing well enough to do so.

All three bacteria are used in the fernentation of or are known to ferment dairy, so I would use re-pasteurized skim milk or skim milk made from powder, then pasteurized. Picking a number out of a hat, I would think that around 40° c. would be a happy medium or if you wanted to be more precise, do each separately at the median of it's optimal range. A good growth should occur after about 8 hours even clabbering at about 40°c. but don't expect too much until after 4 hours.
Small quantities will culture faster than larger ones but also be more susceptible to possible temp. fluctuations.

[iPeace]

Just to clarify, I am trying to prove that the probiotics I am currently giving my dog are, or are not, biologically active, ie, that they are capable of forming bacterial colonies. If they are capable, then I can look elsewhere for what might be effecting my dog, if they are not capable, then I need to replace them with a different lot of the same, or a different probiotic. The probiotics I use are shipped on ice, overnight. How they were handled before shipping can effect their potency.

The difference in temperature between my two crude experiments was due to the difference in ambient temperature between summer and fall.

I will need to incubate the cultures at dog body temperature, about 39°C.

My plan is to sprinkle a small amount of a capsule's contents on one half of a petri dish containing sterilized growth medium, the blank half serving as a control. Probably 2 petri dishes since it is not really more work.

[apochronaut]

Yes, I understood that. Firstly those 3 probiotic strains do not necessarily grow in urine. They might but that isn't how they are supposed to work. If the capsules are time release and capable of bypassing the extreme ph of a dog's stomach , they are supposed to grow in the intestine and assist in breaking down various compounds that might overload a weakly functioning kidney.
They will all grow in dairy, so pasteurized or sterilized skim milk at 40°c for about 8 hours will show if they are active or not.

[SuiGenerisBrewing]

MRS is the standard bacteriological medium used to grow lactic acid bacteria, and will nicely support the grow all three species in your probiotic capsules. However, Bifidobacterium longum is a strict anaerobe and is unlikely to grow under the conditions that you would have in a home lab. Both Streptococcus thermophilus and Lactobacillus acidophilus are aerotolerant anaerobes and will grow fine when exposed to air.

[iPeace]

I put enough milk to cover the bottom in each of two petri dishes. I put the probiotic powder in one dish, left the other as a control. My incubator is a cooler with a 7 W incandescent lamp for heat. It gets up to 87 °F. Within 10 hours, the control was still fluid, while the innoculated dish was congealed. Seems good enough to prove activity.