Microbiology Method Forum

Danger in probiotic boosted home made yogurt ?

Mon, 20 Feb 2012 16:22:11 +0000

Hello dear members,

I hope I am not intruding into your field of study with this question coming from a microbiology neophyte. I'm actually a photographer/photo journalist wanna be.

That being said, I'll go ahead with my question anyway, in all its non-scientific glory.

I have been making fermented products (for myself, family and occasionnaly friends) for a while now : miso, shoyu, sauerkraut, yogurt, pickles, etc. Recently, I've been wondering if i could not "boost" my yogurt with more beneficial cultures that you find in the probiotic pills and drinks available in pharmacy and grocery stores. I've been wondering first, if they would survive (i've done the research and i think 6/7 strains would), if they would grow happily during my 8h incubation at 42C, and, mostly, if they would over grow and maybe be harmful to my regular yogurt strains and mostly to myself. Is there a point where too much of those probiotic strains could be harmful to me ? how much can those trains actually multiply during a 8h fermentation at 40-42C ?

To be more specific, i would be using Probaclac, Adults. It has the following strains : Bifidobacterium bifidum, Bifidobacterium longum, Lactobacillus acidophilus, Lactobacillus bulgaricus, Lactobacillus casei, Lactobacillus rhamnosus, Streptococcus thermophilus.

I was thinking of mixing one pill with my 1 tablespoon of yogurt to innoculate my 1 quart of milk (and some added powdered milk protein).

Does anyone expect a problem ? should i not even try it ? should i try and see if it screws with my digestive tracks ? Can i get my yogurt tested somewhere to see whats in my yogurt after the incubation period ? Should I not post here anymore ?

Thank you very much for your time.

Simon

HIV modification - replication independent reporter integration?

Fri, 17 Feb 2012 21:38:21 +0000

I'm currently trying to figure out if there is a way to put a reporter into the HIV genome that is not dependent on the HIV genome being replicating. Basically the effect should be the same no matter if the genome is transcribed or not.
I think if I just add some promoter into the construct the expression will go up once the LTR is active correct?
This is what I would like to avoid.

Edit: Before I forget, no, placing it antisense is no solution, affects the HIV expression.

How to..? Fungistatic & fungicidal evaluation.

Tue, 14 Feb 2012 12:35:43 +0000

How to evaluate fungicidal and fungistatic effects of a plant extract? Im using the pourplate method with SDA. Organism is Microsporum canis and Candida albicans. 10 ml Leafextract will be added with 10ml SDA in plate and when it solidifies, fungal incoulum will be stabbed at the center. Help!

some help needed for fluorescent focus assay for virus quantitation

Mon, 13 Feb 2012 17:48:08 +0000

I'm trying to set up a virus quantitation system based on fluorescence. I've read about this assay (FFA) first on Wikipedia, but since it wasn't very detailed, I looked up Flint's Principles of Virology (it was given as a reference for the assay).
It wasn't very informative, either. I would like to know how to calculate the FFU using this method.
There are two main options, really: one is to treat the fluorescent signal the same way as in the plaque assay. In this case I'm not really sure how to survey a whole plate for fluorescent signal under a microscope.
The second is to count the number of foci under low mag objective in a few fields, and interpolate. In this case I have the following issues: do you normalize the # of foci to the total # of cells? (After all you can actually count the number of infected cells, and don't need to worry about plaques. In a way it seems like a more accurate way to quantify viruses than the plaque assay.)
Should I count the cells and simply calculate TCID50 based on the number of infected vs total number?

I'm not really sure what to do. Please advise.

Turbidometric tables

Mon, 13 Feb 2012 13:32:18 +0000

Anyone come across a table for a turbidometer that tells you what %light transmittance equates to population density of microbes in suspension?

ask for a strain for the suicide vector called E. coli S17-1 pir

Mon, 13 Feb 2012 11:40:34 +0000

hi, there.

[/left]

recently,our lab focus on a project,in which a strain called E. coli S17-1 pir needed to have suicide plasmid preserved,but in the reference articles, no clue is mentioned for us to get it,so i come to ask for you guys' help.

[/left]

if you guys have it,please have a contact with the E-mails

[/left]

huangclong@yahoo.com.cn

[/left]

thank you.

[/left]

USP 51 and Microbial Recovery Incubation Time

Fri, 10 Feb 2012 16:46:46 +0000

Well, I'm looking to base a test method on USP 51 Antimicrobial Effectiveness Testing and I'm having trouble understanding what the USP is intending. Anyone done this before? I'll copy the relevant section:

PREPARATION OF INOCULUM

Preparatory to the test, inoculate the surface of a suitable volume of solid agar medium from a recently revived stock culture of each of the specified microorganisms. The culture conditions for the inoculum culture are described in Table 2 in which the suitable media are Soybean–Casein Digest or Sabouraud Dextrose Agar Medium .
To harvest the bacterial and C. albicans cultures, use sterile saline TS, washing the surface growth, collecting it in a suitable vessel, and adding sufficient sterile saline TS to obtain a microbial count of about 1 × 10⁸ colony-forming units (cfu) per mL. To harvest the cells of A. niger, use sterile saline TS containing 0.05% of polysorbate 80, and add sufficient sterile saline TS to obtain a count of about 1 × 10⁸ cfu per mL.
Alternatively, the stock culture organisms may be grown in a suitable liquid medium (i.e., Soybean–Casein Digest Broth or Sabouraud Dextrose Broth) and the cells harvested by centrifugation, then washed and resuspended in sterile saline TS to obtain a microbial count of about 1 × 10⁸ cfu per mL. [Note—The estimate of inoculum concentration may be performed by turbidimetric measurements for the challenge microorganisms. Refrigerate the suspension if it is not used within 2 hours. ]
Determine the number of cfu per mL in each suspension, using the conditions of media and microbial recovery incubation times listed in Table 2 to confirm the initial cfu per mL estimate. This value serves to calibrate the size of inoculum used in the test. The bacterial and yeast suspensions are to be used within 24 hours of harvest, but the fungal preparation may be stored under refrigeration for up to 7 days.

The table 2 they reference has a column for incubation temp, inoculum incubation time and microbial recovery time. This is what's throwing me off.
Way I see it, you get a starting population of 10^8 CFU's based on turbidometric measurements (and if anyone knows of a good chart to show this it'd be appreciated) and then you...what? Where do the incubation and recovery times come into play? They say to use the suspensions within 24 hours. How do you use these times to determine the population? I'd just take a sample and do a membrane filtration test on it to get the population, run concurrently with the actual inoculated samples.
Anyone run this before or understand what they are talking about with these "inoculum incubation time" and "microbial recovery times?" I suspect they are making it more complicated than it is.
First time post, hope to scan the archives soon to glean some good info

antibiotic assay

Fri, 10 Feb 2012 02:12:17 +0000

Anyone who is familiar with Gramicidin (antibiotic) assay? I have been performing this test for quite a while, but I could not come up with a valid result. I am using USP as my reference for the procedure (Spectrophotometry). The problem is, the turbidity of my assay do not have a trend even if I increase the concentration of the antibiotic used for the Standard. I am working with a pure culture of Enterococcus hirae, and all the materials used was sterilized, so I am confident that my problem with turbidity is not caused by contamination. Any insight will be highly appreciated. Thanks in advance.

problem in making a proper smear for nile blue staining of an unknown yeast

Thu, 02 Feb 2012 11:33:50 +0000

i have isolated an organism which looks like an yeast under the microscope and i want to find out if it is capable of producing polyhydroxybutyrate. the problem im facing is that the smear is loaded with weird looking things (not cells as i have performed gram staining and they wont ever stain). due to such interfering background im unable to confirm if the fluorescence is from cells or surroundings.

E.Coli trypsin resistance

Sat, 28 Jan 2012 23:19:19 +0000

Hi,

Does anyone have an idea what is the maximum viable concentration of trypsin that can be added to an E.coli (TG1) culture overnight ?

Thanks