Advances in microbial screening | Luca Potenza | TEDxUniversityofWarsaw
URL: https://www.youtube.com/watch?v=QRpxrspf4Gw Video ID: QRpxrspf4Gw ============================================================ good evening everybody it's a big pleasure to have you here to have you still here I know I know it's a bit late isn't it but I will try my best to wake you up with the microscopic topic of today's micro talk we're having together here so speaking of microscopic things microorganisms or microbes or germs if you prefer the term they're everywhere aren't they have you noticed that they're everywhere there are in the soil there are in the water there are inside outside of our bodies as well and it's not a surprising fact but Humanity started to work to collaborate with them since the dawn of time like really before Christ long time ago and perhaps raise your hand if there is anybody here that would like to guess what were the first biotech products invented by humans yes no it's a no okay I'm going for it then beer beer alcool in other words all right so yes great answers and correct I would say Bread and Wine but still uh at the end of the day uh it's really it's selling has we need more than just food in life as as humans don't we and J side yes we were able to work with them with microbes without even seeing them because notoriously they're quite tiny it wasn't in fact since the the the 17th century that we've started to to visualize these tiny little bacterial cells with the Advent of microscope more or less uh in the in the way we're intending it today cool but that was just about it so we were able to perhaps classify microorganisms based on shapes or motility but really nothing more fast forward then to the 20th century where we started to manipulate to uh cultivate and even more importantly to select microbes at our own advantage in general and what you can see here is a very oldfashioned technique also known as Petri dishes for microbiologists in other words we are having a container can be plastic can be glass we fill that with medium cultivation medium so in other words nutrients and space for microbes to grow and we let them grow so what you seen here all these little dots are actual colonies originating from one single cell and they leave they they proliferate on top of this medium on this solid medium okay so among all these dots you can see some of those got actually a h around them a transparent Hollow around them which indicates visually to has the the the researcher the microbiologist that that specific colony is doing something very important for our experiments for instance I think that everybody in here at least once got tested for some sort of bacterial infection so what happening you're going to your doctor they got a swap of your saliva or blood and then they send this sample this unknown and let's say complex sample to the lab for what for screening so for instance in case of a bacterian infection the answer you would like to get out of these methods is what kind of drugs should I take to get rid of this infection so quite important answers we are in searching for unfortunately such whole fashion techniques are still used today and I'm saying unfortunately because the reality is that these methods we are going to call them conventional traditional methods today they they they come with with with disadvantages like one they are solid and this little detail actually is very important when you want to screen um a microbial population uh F fact most of the microbes don't like this scenario this cultivation media on on solid cultivation medium for different reason uh there are data out there actually showing us that if you're lucky by doing this um yeah if you're lucky you can get the 5% out of the total life of a sample so thumbs down and fortunately we moved on okay because in this case also it's worth mentioning that microbes compete for life really they compete for space they compete for for nutrients and as a result you may Overlook uh rare strains slow growing strains um because the fast growing will be the first one to show up bias representation but on top of that there kind of cost and time consuming procedure but over all the question I got for you is like how can we depict the the complexity of Life uh with such um bias techniques right because we clearly have some results but can we do it better did we already answer is yes we did we're still executing complex experiments in the lab but the laboratory uh became very very small so small that it can be compared with a coin for instance so it's a Neo it's not a poly swaty I apologize but for the next next time you get the reference anyway and you can also see inside of this lab on a chip device which is very tiny device as big as a as a as a inch square inch you can see some corridors you can see some channels you can see some rooms and all these designs allowed us to execute experiments multi-step complex experiments and on top of that in the process of Designing and fabricate such Laboratories we do have high degrees of of customization we can design our lab uh in the way we want to use it in other words so we're talking already about miniaturization keep this in mind for for for the next slides and I told you we can execute experiments inside of these devices and these experiments come in a droplet format so let's talk about droplets in this very very linear and simple workflow we can see uh three fundamental steps where we are using the droplets that we generate has um individual experiments so the droplet is sort of bioreactor is a sort of of of private Universe for the cell we encapsulate inside of the droplet in fact we do encapsulate one up a droplet since we are going for single step studies in this case first off we're making droplets we put and sell up a droplet and then without droplets we incubate our bacteria so from one cell you will have two cells four cells eight cells etc etc etc exponential growth you will obtain a colony inside confined in the droplet and even more importantly the third step you see what's going on inside droplets as we did with the microscope and you select what's growing inside of drop droplets so specifically uh these yellow ones that have been selected from the pool of droplets are the equivalent of this one Colony with transparent Hollow you've seen at the beginning so we really really really want to find the needle in high St this very tiny fraction of microbes that for some reasons are useful to us and before we dive into these three fundamental steps I think it's more mentioning that at the first step of this protocol we have already eliminated big big disadvantages associated with conventional methods because for instance number one in this case droplets are liquid which is thumbs up for most microbes they do prefer liquid medium and also via single set encapsulation we um prevent basically competition among strains so there is a quite High chance there are quite higher chances to to detect every every single strain even the slow growing ones this this train that are very rare in the in the original sample overall better results and we said number one let's make our experiment what you're seeing here is uh first of a series of Labon chip device that you would see today it's a droplet generator pretty straightforward we're generating droplets and we're doing it by mixing oil with a cultivation medium which is technically speaking gain um water with some nutrients for the microbes for allow for for growth and in a way that's somehow personally at least it resembles a lot mayonnaise I think we're all familiar with mayonnaise um I'm not a chef but as long as I remember you have to mix thoroughly a few components like vegetable oil and uh lemon juice and some egg okay so you mix them thoroughly and you get some very tiny tiny Aquos droplets made out of lemon juice of egg bits in an oily background let's say and this is what's going on here we're generating microscopic droplets at a very fast rate so 2,000 droplets per second namely which picture yourself it's like 2,000 experiments per second being generated unmatched numbers for conventional techniques and the CH on top is that all these droplets with have equal volume and this equal volume is is still there after the second step so the incubation where we let our bacteria grow you can see here uh life of some sort inside droplets because again the droplet is the universe for this for this microbes so they can proliferate independently from all the others without competition they're leaving happily there and here you can see also empty droplets and the secret here is that we statistically we prefer to have one single Dr single trer droplet and Screen some empty ones um instead of having maybe the chance of two or three set a droplet in in the in the experiment so we just dilute the sample a bit to achieve 99% single set encapsulation and I know that picol lers micrometers are not like everyone's cap of te but perhaps you want to guess just raise your hand again it's not a test how many droplets and therefore how many experiments you could squeeze in a volume of a vodka shot dear polish audience I see some faces are waking up with the magic word right 10,000 what's your name sir nice to meet you th me with me no you're quite far off uh from from the the actual number the actual number is more like 400 million droplets so again picture in case of Petri dishes the one we've seen at the beginning they're like this big and this tall you probably need this entire theater to put that into perspective so again miniaturization and miniaturization also mean means uh cost savings most of the time and this is especially the case so okay we have our droplets we have life in it we got move on so we're going towards the culmination towards this this third step the selection process of our experiments once again here we have uh yet another cheap device and this chip is designed in a way that we are having optic fibers inside of this laboratory capable or reading through the droplets these that look like massive pillars are actually micro optic fibers in the chip they read the colors of the droplets in a very fast Manner and one positive one pass in the channel we can detect that but also trigger the Sorting event in a way that we can pull the positive experience we can really retrieve that for whatever Downstream applications you you you got in your mind and let's picture this for some reason your droplets you got milon droplets to screen and you're interested in the 0.1% positive droplet so very tiny fraction the needle in a Hast stack in a form of of of experiment all your droplets are red so every time a droplet is red is passing you just say no I don't care no I don't care no I don't care but when a pink one which is your positive pass in the channel you will detect the pink color and you will say I want that I want to to retrieve that and we're doing this in a very fast manner once more in this case we're talking about a thousand of droplets per second thousand of experiment per second pretty big numbers again and the equivalent of you doing this for instance is like let's picture this your grandma gave you a bag of candies and they like thousand of candies they're all red for some reason you don't like this red ones but you like the tiny fraction the 10 pink uh candies in this bag and the equivalent of you doing this is like you got your bag you open and in one second you have on one side the red candies you don't like and on the other side you have your 10 uh Pink candies ready to be to be eaten so with that said we we we had a look on a very very powerful tool for single cell studies and to study what well live in other words on a very microscopic level um we have very big advantages coming from such protocols so we talked about minorization cost savings um we're going very fast which is usually good but also we can have the great chance to have less biased results which is definitely the point number one of such approach and such approach these tiny droplets were actually born within the academ environment not a long time ago like two decades ago more or less and they already brought us quite big discoveries in certain aspects within the Academia uh there were actually some they were Game Changer especially for single cell studies genomic transomics and go and so on but they also found uh application within like the industrial sector in pharmaceutical cosmetic preparation and what I do really hope personally with all the premises we got here today is I like to see such big discoveries being um implemented even more in the real world scenario for instance in hospitals where it can definitely and ultimately help ourselves the people