Let's do science! Experiment A: SPROUTING

I just ordered some test equipment. Petri dishes with media, stains, and microscope slides. I have a couple very good microscopes on hand. I like having first hand knowledge, and doing science. So let's do sprout science! My plan is to get some materials to sprout, and sprout them under various conditions typically used at home, and measure the growth of any organisms other than sprouts. I need input as to the proper things to test. I tentatively plan to culture the dry seeds, then start the sprouting process and take cultures at the end of day 1, day 2, have no idea how long it takes but every day, then the finished washed sprouts, then maybe after a day in the fridge, after two hours in the bowl, etc. I think there are 10 plates in the initial order.

1. What is the best seed/bean to begin with? Most commonly used?
2. What procedure should I use to sprout them? (Kind of water, paper towels or what, temperature etc.)
3. How long are the finished sprouts kept, and how are they fed?

This way, I can check the bacterial growth during the sprouting process and see what it is.

I'm not a biologist, so I will have to go research what stains to use for what organisms. Etc. If anyone here has a background in microbiology or any related experience, would you please chime in? I ordered a soy based plate because that medium is supposed to support a wider range of organisms - including listeria - that don't grow on agar agar. I don't have an incubator so I'll have to figure out how to rig something up. Advice?

If the results are interesting, I'll order more agar and do more tests. It would be interesting to look at mouth bacteria in the bird, bacteria on cage and toys, on food dishes, on other food items, on commercial feeds, in water bowls ... The experiment is open to all! Collaboration is the way. If other people want to do the same tests at home, I can give the link to the stuff I ordered. Perhaps someone here knows more about the correct test equipment?

Addendum: testing of pre-sprouted sprouts, purchased from store?

I do not have the depth in this area to be of much help. That said, what to sprout would seem the first target, since there is likely a wide base in what is both provided in dry form and therefore what is sprouted.

We provide a very wide dry base, covering seeds, grains and nuts. We do not provide beans. From a sprouted base, we only sprout grains. This as a result of long held family fears of sprouted product that reaches back to my Great Grandmothers on both sides, passed down to Grandmothers, mother and finally me. That history in place, if tomorrow, Science came forward with a statement that sprouted (you name it) will provide extended life to everyone (not possible) I wouldn't touch the stuff. That likely based on what my Great Grandparents warned against having proven so very true of the years.

Regarding seed, we do not provide Sun Flower or any of the Sun Flower Seed like seeds, nor do we provide peanuts either in their shell or not. The exception is the rare roasted peanut (not shelled).

The few grains that the Mrs' sprouts are during the late Spring, Summer with them on a moist paper towel set in the Sun light. Not the best method for an experiment.

It would seem to me that two different groups would make sense. The first group would be those 'spouts' that have been tied to serious outbreaks resulting in illness and/or death. The second group would be those with no reported illness and/or death.

Just my two cents!

You will need a control to make the experiment valid. Since your aim is to see what's there under normal conditions, and you're not using a clean lab, you will want to see what's there just if you do nothing. Additionally, you'll want to do a dishes with swabs of your hands, your prep surfaces (before you do any prep), and utensils used. Don't forget the under side of your faucet and your sink. Swab any item you use before you use it. If you don't you can't positively say any bacterial growth actually comes from the sprouts.

At a rudimentary level stains are used to identify bacteria.

I caution you here, you are not setting yourself up for anything more than testing for the presence/absence of bacteria. Unto itself, it is not an informative endeavor. There's E. coli everywhere.

The question that is more helpful is HOW MUCH bacteria is present, is it an infectious dose? E. Coli has a very high infectious dose value, depending on your reference ranging 10^5-10^8 (100,000 to 100,000,000 individuals). This means that in order to get sick you need a LOT of individual bacteria at once to get sick. Just because you are able to culture some (assuming you are able to isolate it from only the sprouts) doesn't mean you started with enough to cause illness.

Also, identifying the species of bacteria you have cultured could be nearly impossible without DNA analysis. Many species can share the same colony growth patterns. The media you choose to use will also limit the bacteria you will be able to culture. More pathogenic bacteria, for example, are likely to grow on blood agar as opposed to general agar.

chris-md said:

At a rudimentary level stains are used to identify bacteria.

I caution you here, you are not setting yourself up for anything more than testing for the presence/absence of bacteria. Unto itself, it is not an informative endeavor. There's E. coli everywhere.

The question that is more helpful is HOW MUCH bacteria is present, is it an infectious dose? E. Coli has a very high infectious dose value, depending on your reference ranging 10^5-10^8 (100,000 to 100,000,000 individuals). This means that in order to get sick you need a LOT of individual bacteria at once to get sick. Just because you are able to culture some (assuming you are able to isolate it from only the sprouts) doesn't mean you started with enough to cause illness.

Also, identifying the species of bacteria you have cultured could be nearly impossible without DNA analysis. Many species can share the same colony growth patterns. The media you choose to use will also limit the bacteria you will be able to culture. More pathogenic bacteria, for example, are likely to grow on blood agar as opposed to general agar.

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I was afraid of something like this. I am hoping to at least get some size measurement of the colonies, if any. Maybe I won't be able to identify the specific organisms, though. It may be that the results don't provide any useful information, but what the heck ... it will be fun to try, just to see what happens. Depending on the outcome, maybe repeat with different parameters or different media. That's one thing about science, you can't control the outcome, you just have to experiment and revise and repeat.

Parrot Forums love science!
There was recently a big to-do in national news (new study on food safety; I forget the source) that condemned sprouts and strawberries as the dirtiest produce of all. Pop science or legit? Dunno.

Kentuckienne said:

chris-md said:

At a rudimentary level stains are used to identify bacteria.

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I was afraid of something like this. I am hoping to at least get some size measurement of the colonies, if any. Maybe I won't be able to identify the specific organisms, though. It may be that the results don't provide any useful information, but what the heck ... it will be fun to try, just to see what happens. Depending on the outcome, maybe repeat with different parameters or different media. That's one thing about science, you can't control the outcome, you just have to experiment and revise and repeat.

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GaleriaGila said:

Parrot Forums love science!
There was recently a big to-do in national news (new study on food safety; I forget the source) that condemned sprouts and strawberries as the dirtiest produce of all. Pop science or legit? Dunno.

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This is the sad state of Science Today! Resulting from the huge wave of 'Junk Science' (Pop Science) that has floored all parts of our existence and knowledge pools. Well Founded and Structured Science has to compete with on going mudded trash that only serves to confuse and steal funding from legit Science. Venting done!

I think you'll want 2 or 4 test. 1 being the control of sprouting in just water; another sprouting in grapeseed extract treated water; 2 other possible tests could include vinegar and then lemon juice.

It doesnt hurt to test to see the most effective method of keeping sprouts from culturing bacteria. You might show there is little or all the difference between the methods if nothing else?

Some good suggestions. Find the best growing medium for e.coli and just focus on that. One way to measure quantities is to count individual colonies which might be informative or might be too close. I also saw a reference to an e.coli test kit that looke quantitative. I am open to using GSE but that's a whole another thing...a couple of paper I saw found artificial preservatives in most of the commercial GSE available, even though the manufacturers claimed not to put any in. Maybe the first test should be just to test washing methods (frequency etc.) or sprouting temperature vs time? Get an idea of how the bacteria grow, then try to see how to minimize them.

Count the colonies? No. That tells you how many times you released bacteria from your swab as you moved it along the plate. On agar a single bacteria can ultimately grow into a colony given time.

You really don't have the equipment, technique, or background to draw the conclusions you're trying to draw. Its a fun exercise, but amounts to children's kitchen science (watch, I can grown bacteria!!). Just bear in mind you aren't going to be able to drawn any conclusions at the end of this.

chris-md said:

Count the colonies? No. That tells you how many times you released bacteria from your swab as you moved it along the plate. On agar a single bacteria can ultimately grow into a colony given time.

You really don't have the equipment, technique, or background to draw the conclusions you're trying to draw. Its a fun exercise, but amounts to children's kitchen science (watch, I can grown bacteria!!). Just bear in mind you aren't going to be able to drawn any conclusions at the end of this.

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I have an extensive background in laboratory science, albeit in organic and petroleum chemistry and not in biochemistry. I do expect to be able to generate useful data, so it's important to design the experiments properly. That's why I've put posts in this thread, to see if anyone else has input. Equipment is not a problem, as I have several high end microscopes on hand and can order the correct supplies, and as far as technique and background, I think that's covered too.

I don't usually talk about my nerdy history, since it puts people to sleep. A literature search is the best way to turn up correct information from peer-reviewed journals, but participative science is not beyond the grasp of the average person, and I thought people might be interested in the process. It sounds like you do have some biotech or medical experience, so any contributions would be appreciated!

The scientific method is a beautiful thing. I consider experimental design and statistical analysis to be fascinating. I am loving this thread... enjoying the science and seeing how we interact with it.

Kentuckienne, I am a biologist by training, so I have experience in biological experimental design.

Not to belabor the point (understand I'm not at all trying to be nasty or come at you, just adding the actual biological experience here), but my contribution here is to temper your expectations - not necessarily dissuade you from doing it. You aren't able to correctly identify the bacteria you are culturing - last checked you don't have a several thousand dollar DNA sequencer. If you can't even identify what you are culturing you cannot draw any valid conclusions.

Microbiology is fickle that way. As you somewhat implied yourself when you acknowledged no biological experience above, biology is not chemistry.

I found that news stuff I was talking about.
High bacteria levels in bean sprouts: CBC probe - Manitoba - CBC News
Growing Sprouts in Retail Food Establishment - CFP Issues 02-III-01 and 04-III-012
Not safe to eat: Three foods to avoid - CNN.com
and about home-growing...
https://www.foodsafety.gov/keep/types/fruits/sprouts.html
Fortunately, we don't LIKE them, so it's no skin off our proverbial buttfeathers.

chris-md said:

Kentuckienne, I am a biologist by training, so I have experience in biological experimental design.

Not to belabor the point (understand I'm not at all trying to be nasty or come at you, just adding the actual biological experience here), but my contribution here is to temper your expectations - not necessarily dissuade you from doing it. You aren't able to correctly identify the bacteria you are culturing - last checked you don't have a several thousand dollar DNA sequencer. If you can't even identify what you are culturing you cannot draw any valid conclusions.

Microbiology is fickle that way. As you somewhat implied yourself when you acknowledged no biological experience above, biology is not chemistry.

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I came across a reference to "E. Coli test kit" online so I started looking for those. I know, it's not DNA accurate, but perhaps accurate enough for our purposes? Here's a sample of what I found - would appreciate advice from anyone.

Coliscan EasyGel, test kit for 10 tests is about $30, uses pre-treated dishes and media that inhibits other bacteria to quantify (I assume roughly) E. Coli:
https://www.micrologylabs.com/page/93/Coliscan-Easygel

General presence/absence tests for coliform and e.coli, simply indicate present or not, without quantifying how much is in each sample. These are fairly inexpensive and available from lots of places, such as "Industrial Test Systems 487197 WaterWorks EZ Cult Bacteria Test" from Amazon

Interesting and simple semi-quantitative test from Aquagenx, haven't found the price yet:
https://www.aquagenx.com/wp-content/uploads/2015/11/-v4-CBT-Instructions-_DrinkingWater_Sobsey1.pdf

So far, Coliscan EasyGel looks like the best option, but I'll keep looking.

The CDC says:
Reduce your risk of getting sick from raw sprouts.
Sprouts are a known source of foodborne illness.
Children, older adults, pregnant women, and people with weakened immune systems should avoid eating raw sprouts of any kind (including alfalfa, clover, radish, and mung bean sprouts).
Cook sprouts thoroughly to reduce the risk of illness. Cooking sprouts thoroughly kills the harmful bacteria.
Request that raw sprouts not be added to your food. If you purchase a sandwich or salad at a restaurant or deli, check to make sure that raw sprouts have not been added.

GaleriaGila said:

I found that news stuff I was talking about.
High bacteria levels in bean sprouts: CBC probe - Manitoba - CBC News
Growing Sprouts in Retail Food Establishment - CFP Issues 02-III-01 and 04-III-012
Not safe to eat: Three foods to avoid - CNN.com
and about home-growing...
https://www.foodsafety.gov/keep/types/fruits/sprouts.html
Fortunately, we don't LIKE them, so it's no skin off our proverbial buttfeathers.

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With respect I fail to see the relevance of most of these links as they go back to 2011, we are in 2017 are we not? If you want to put up links to be taken seriously try and find something that is recent and can be taken seriously or it is just scare mongering. You will of course be adhering to the recommendation on eggs, meat, seafood, milk products, fruit juices? The one on pet food was interesting though. I am happy to read any info but make it timely and relevant please?

Edit: Digging deeper myself I have found this https://www.cdc.gov/salmonella/reading-08-16/advice.html This relates to 2016 so is worth reading IMO. So in the interests of safety all those in the US that buy their sprouts for their own consumption or their birds need to exercise caution.
I am happy with the source of sprouts I am able to purchase in the UK/EU.

Kudos Kentuckienne for doing the science. I look forward to hearing about the experiment!

Hey, what's the worst that can happen, I learn nothing. More likely I'll learn something on a macro scale that provides some useful information. I probably learn more from doing literature searches and relying on trusted sources, but it's so much FUN to play with science! Just the mental effort to design an experiment, carry it out and interpret the results is worthwhile even if the results aren't useful. It's good thinking practice. That's why I opened the project to everyone who wants to be involvement or have input. I sure don't know what I'm doing, but that's never stopped me yet!

Kentuckienne said:

Hey, what's the worst that can happen, I learn nothing. More likely I'll learn something on a macro scale that provides some useful information. I probably learn more from doing literature searches and relying on trusted sources, but it's so much FUN to play with science! Just the mental effort to design an experiment, carry it out and interpret the results is worthwhile even if the results aren't useful. It's good thinking practice. That's why I opened the project to everyone who wants to be involvement or have input. I sure don't know what I'm doing, but that's never stopped me yet!

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I'm dying here! Have we got any results yet?