earlier in this chapter I mentioned that bacteria can undergo horizontal Gene transfer so two bacteria in the same culture can actually exchange genetic material this requires that the two cells are in close contact this is not going to happen with one cell and then with another cell that's on the other side of the plate or the culture they would have to be in direct contact this is what we call conjugation so they would actually mate where the bridge would form and genetic material would flow from one bacteria to the other I often use the analogy of conjugal visits in prison so you know there has to be direct contact so that's why I often use as an analogy so in order for bacteria to transfer genetic material there has to be a passage way and this passage way is a conjugation pilus or conjugation Bridge there's a couple of different names for this this is an F pilus sex pilus or pillis however it is that you'd like to say it and the directions to making this conjugation pillis reside in a fertility plasmid or an F plasmid so if you remember from the beginning of this chapter I mentioned that plasmas have nice to have genes these are not NE necessary genes for the survival this is like extra DNA not all bacteria have plasmids if a bacteria has an F plasmid a fertility plasmid it has the genes to direct the synthesis of this conjugation bridge and that's how genetic information can flow from one bacteria to another so if we have a bacteria and it has an F plasmid we would call that the f plus cell that's the donor cell which will donate DNA the recipient is going to be the F minus cell now another term they'll use is male and female I mostly use f+ and donor and F minus and recipient so in a culture if we have a bacteria that has an F plasmid and it's an f+ it's a donor cell eventually all of those cells are going to become F plasmid cells because one bacteria is going to send genetic information to the next then that cell will become an f+ cell and then that cell will donate genetic material to the F minus cell and that cell will become an f plus cell and so forth so if we wait long enough all of those bacteria in culture will actually become those f+ cells so I mentioned that the genes to construct this conjugation Bridge are going to be found on plasmids so remember not all bacteria will have plasmids and these fertility plasmids have the genes the instructions to make this conjugation pilli without that genetic information is not going to be able to pass from one bacteria to the next I also mentioned that plasmids can contain nice to have information some of this information could actually be resistant genes and so we're going to spend a little bit of time talking about resistance resistance means that an antibiotic that once killed a bacteria now can no longer kill that bacteria that bacteria is now resistant to that antibiotic so if you take a look at this picture this is a picture of bacterial conjugation so on the left hand side we've got our donor cell our f plus cell it has a chromosome and you notice that it has an F plasmid that F plasmid has the instructions to make that conjugation pillis so now that conjugation pillis forms remember that's something unique to bacteria and it can actually exchange genetic information with that F minus cell that recipient cell so once that conjugation Bridge forms now some DNA can actually transfer in the case of this bacteria the plasmid a copy of the plasmid is actually being transmitted from that donor cell to the recipient cell and now at the end of conjugation when you look at that bottom picture those two cells are identical so now that recipient cell has now become an f+ or a donor cell so I keep mentioning this resistance so I want to talk a little bit about resistance and what this means we can actually test for antibiotic resistance in the lab in fact we're going to be doing that um coming up in an upcoming lab if a bacteria is resistant to an antibiotic that means that it has these mechanisms that work around that antibiotic killing it we're going to get into the specifics of those mechanisms coming up but if we had a bacteria and it had a resistant Gene and we grew it in the presence of that antibiotic that it had a resistant Gene for it would actually live there's nothing that would kill it that antibiotic is not going to work on it so it would live it would grow and we would probably assign a little plus to say yes there's growth if that bacteria is suscep ible now it doesn't have a workaround from the antibiotic killing it so it's susceptible it would actually die we would not see growth on that plate if we plated this organism in the presence of that antibiotic and it would die we would put a minus sign so to find out if a bacteria has a resistant Gene we could actually plate it in the presence of that antibiotic if it has that resistance Gene it will live if it does not have that antibiotic resistant Gene then it will die just a little note that I want to make sure the vocabulary is correct it's not that the antibiotic is resistant it's that the bacteria is resistant to the antibiotic sometimes we end up confusing those but it really is the bacteria that has a work around that antibiotic killing it again later on we're going to get into those specific mechanisms so a little example here so I have two strains of eoli so if we were to look at their biochemistry they would type out as eoli but you notice that they differ a little bit genetically this is why they're strains so strain one has a chromosome obviously it has to have that to survive and it has a streptomycin resistant gene on that chromosome if you take a look at strain two it has a chromosomal Gene of course and it also has a plasma you notice that strain one does not have a plasm so on the chromosomal genes of strain 2 it has a now resistant Gene meaning that it would be resistant to that particular chemical on its plasmid it has an ampicillin resistant Gene so there's a media that if we're testing for resistant genes and conjugation we actually use an lb augur and this is a special augur that allows uh the bacteria to kind of open up and remain healthy so that's a type of media that we would use so if we were to actually plate these two bacteria in a variety of situations this is actually what we would see so if we were to Plate our Str one in plain augur it would actually grow there's nothing in that media to kill it so I'm going to put a little plus sign there if we were to Plate strain 2 in that plain lb media there's nothing in it to kill it so it would grow so we would see growth we would note that with a positive so this is just to show we want to do a control just to show that this bacteria is growing it's able to use the nutrients in that media now let's take a look at a situation that second row where we have the media but we've added ampicillin to the media so strain number one would strain number one grow if you notice it has a streptomycin resistant Gene it does not have an ampicillin resistant Gene no ampicillin resistant Gene so that ampicillin and the media would actually kill it so we would not see growth I'm going to put a little minus sign there for strain number two would that grow if you notice that strin number two has an ampicillin resistant gene on the plasmid that ampicillin resistant Gene is going to give it a mechanism around that ampicillin killing it so we would expect growth so I'm going to put a little Plus in that column if we look at the third situation we've got the media with a streptomycin added to the media if you look at strain number one strain number one has a streptomycin resistant Gene so it's got to work around from that antibiotic killing it so we would actually see growth if we look at strain number two strain number two does not have a streptomycin resistant Gene so it would die we would not see growth so I'm going to put a minus there now let's kind of combine antibiotics and see what happens so if we pl at strain one in the media with both streptomycin and ampicillin in the media it would have to have both of those genes to survive so strain number one has a streptomycin resistant Gene it does not have an ampicillin resistant Gene the streptomycin in the media is not going to kill it but the ampicillin will so we would not see growth because that ampicillin would kill it if we look at strain number two there's an ampicillin resistant Gene but no streptomycin resistant Gene ampicillin in the media is not going to kill it the streptomycin will so again we're not going to see growth if we look at the case of n strain number one does not have a Nal resistant Gene so we would not see growth if we look at strain number two it does have an L resistant gene on its chromosome so so it would have that workaround so we would see growth so we would put a positive so this is what we would call a confirmation just to find out what genes and what genes we suspected our strains of eoli had so if we take a look at a picture and how those genes those resistant genes can be transferred if we have our donor cell our f+ cell it has that fertility plasma it's next to that recipient cell it has that F plasma so it can have the directions to make that conjugation Bridge conjugation Bridge forms and now if you notice that f+ cell is actually transmitting a copy of its plasmid so now at the end of this conjugation and procedure that F minus cell looks exactly like that f+ cell now it's become the donor cell now if let's say that that f+ cell has an ampicillin resistant gene on that plasmid now that cell next to it that was a recipient now has that ampicillin resistant Gene so when we talk about resistance and what an issue resistance is and how resistance can spread among a population this is exactly how it can occur and why it can occur so quickly because conjugation can happen these donor cells can then transmit a copy of their antibiotic resistant Gene so this is a big issue so here is an experiment that we actually did in lab and on the right hand side I actually have like what our strain would look like and so if we grew a stra strain and lab and we wanted to find out if it was resistant to certain antibiotics certain chemicals we would grow it in the presence if we see growth we know it has a resistant Gene if we don't see growth then we know it does not have that resistant Gene so this is actually a case where mating happened conjugation happened so I gave you that example of strain one and strain two so strain one and strain two made it and so what happened is that a brand new organism was made it has a different genetic map so it's a different strain so if we take a look at what strain three will actually grow on this can give us an idea of what its chromosomal map looks like so if you take a look strain number three grew in the presence of ampicillin so does that mean that it has that ampicillin resistance Gene yes it does it has that osym resistant Gene so I'm just going to put a little a right there for our ampicillin resistant Gene now we grew it in the presence of streptomycin does strain three have a resistant Gene for streptomycin yes it does because we see growth so we know that it has a streptomycin resistant Gene and an ampicillin resistant Gene now does strain number three have a nail resistant Gene if you take a look at that plate where there's no growth it was actually grown in the presence of Na there's no growth it does not have that na resistant Gene our last scenario we plated our conjugate our strain three in streptomycin and ampicillin remember in order for it to grow in the presence of both of those antibiotics it has to have both of those resistant genes and it does if you look at our chromosomal map it has the illum resistant Gene and the strep resistant Gene we found that out by plating them in those separate situations but when we combine them you can see that it's got to work around for both of those antibiotics so our strain three has an ampicillin resistant Gene and a streptomycin resistant Gene and this is how we can tell if a bacteria has a antibiotic resistant Gene so another thing that can happen is that the plasmid can actually incorporate itself into the chromosomal DNA this is what we call a high frequency recombination or HFR recombination means that we're combining DNA so that plasmid is going to meld into to the chromosomal DNA so if you take a look at this picture we have our f+ cell at the top with an F plasmid and in that next picture that F plasmid becomes incorporated into the chromosome so now it becomes an HFR cell this HFR cell is still the donor cell so now it can make with a recipient cell and some of those chromosomal and plasma genes can actually move so I know we've been talking about plasmids um so sometimes we get that idea in our head that oh only plasma genes can move no chromosomal genes can move plasma genes can move as well if we see a case of an HFR you can see where the combination of those plasmids and chromosomal genes can actually be transmitted to that recipient cell this is often what we call jumping genes now sometimes that plased actually comes back out of the DNA and it might actually take some of that chromosomal DNA with it because it looks a little different we go from calling it um an F plasmid to becoming an F Prime plasma just because it looks a little different so we have to give it a little different designation so here's my drawing of our HFR cell so it's got that circular chromos it has that red plasmid and then that plasmid becomes incorporated into the DNA and now we have an HFR cell if that plasmid comes back out it's not going to be a perfect excision it's going to end up taking some of the chromosomal genes with it and now it becomes an f-prime plasmid if you look at that it's got the red f plasmid genes but it also has a little bit of the blue the chromosomal genes so because it looks a little different we have to give it a little different designation which is why we call it an fime so again this is just part of that scrambling and how genes can kind of be mixed up so here is our HFR cell and then you can see where that plasmid comes back out and now it's an fime plasmid so now we've got our F Prime donor cell and it donates a copy of its plasma to a re recipient cell now that recipient cell looks the same as that donor cell but now it has the benefit of whatever chromosomal genes were there so if let's say there was an ampicillin resistant gene on the plasmid and by that high frequency it actually pulled out a streptomycin resistant Gene now we're actually donating two resistant genes instead of one so this is what we often call jump jumping genes and jumping genes is the ability of these genes to move these genes can move from plasma to plasma they can also move from plasma to the bacterial chromosome the ability to move from one location to another is what we call transposition and this is made possible by these transposable elements this is often what we call jumping genes it can jump from one thing to the other these transposable elements actually have a genetic sequence on the DNA but then there's also the codes for an enzyme transposes on the ends kind of like bookends of these transposable elements that allow it to actually be excised out so that it can actually move from one place to the other so if you take a look at this plasmid it has this transposable element so on the middle we've got our transpose on genes and then then we've got those inverted repeats that are kind of like bookends of these transposable elements there could be an antic resistant Gene of this transposable element if it moves from plasmid to plasmid or plasmid to chromosome then it's going to take those antibiotic resistant genes with it so this is a good way to see how resistance can arise so quickly with bacteria we're going to be talking a lot more about resistance um coming up in the upcoming chapters and it also kind of gives you an idea of how these bacteria can increase their genetic diversity and kind of do that scrambling of genes