This Week in Microbiology
With Vincent Racaniello, Elio Schaechter, Michael Schmidt, and Michele Swanson
Episode 165: Pumping copper
Aired December 1, 2017
Vincent: This Week in Microbiology is brought to you by the American Society for Microbiology at asm.org/twim.
Vincent: This is TWIM, This Week in Microbiology, episode 165, recorded on November 16, 2017. I’m Vincent Racaniello and you are listening to the podcast that explores unseen life on Earth. Joining me today from Small Things Considered, Elio Schaechter.
Elio: Hello there.
Vincent: How are you, Elio?
Elio: I’m doing fine, I’m really doing very well.
Vincent: Good to have you back.
Elio: Considering my advanced age (laughs)
Vincent: Also joining us from Ann Arbor, Michigan, Michele Swanson.
Vincent: How are you?
Michele: I’m okay, I’m a little chillier than the last time we talked.
Vincent: The temperature is dropping, isn’t it.
Michele: Yeah, but it’s supposed to be 50 here on Saturday, so still a little crazy.
Vincent: Are all your leaves down or do you still have some up?
Michele: Some. Some are still up on the trees but a lot are down.
Vincent: Also joining us from Charleston, South Carolina, Michael Schmidt.
Michael: Hello, everyone!
Vincent: I bet it’s 80 degrees there, right?
Michael: No, we have the same cold that is up in Michigan, but it’s gonna be about 60s today.
Vincent: Do your leaves fall off the trees?
Michael: They do but they never get color because we never have frost. They just fall.
Vincent: They just drop? That must be interesting.
Michael: It is interesting.
Vincent: So basically no one comes to Charleston to see fall colors.
Michael: No, they come to Charleston, the only color we have is after you boil the shrimp.
Vincent: Actually, they come to Charleston to see Michael Schmidt.
Michael: We’re gonna have a virology, one of ASM’s last conferences is gonna be here at the end of the year in December on virology. I got the email yesterday about this virology contest that ASM is sponsoring here in Charleston.
Vincent: They really should have a TWIV in association with that.
Michael: I would think so!
Vincent: I’ve heard nothing of it. Okay, Michael, you are up first with a copper snippet.
Michael: A copper snippet! Today’s paper, or today’s snippet is from the American Journal of Infection Control and is by a group from Grinnell College which is located in Grinnell, Iowa, with the work being led by Dr. Shannon Hinsa-Leasure and being principally conducted by her undergraduate students, Zina Ibrahim, Alexandra Petrusan and Patrick Hooke. Grinnell is a small liberal arts college–
Elio: A good one, too.
Michael: It’s a fantastic one. The undergraduates have a strong tradition of conducting research, and here are three undergraduates under the guidance of Dr. Hinsa-Leasure who conducted an outstanding piece of work, and the paper is entitled “Reduction of bacterial burden by copper alloys on high touch athletic centers.” So as next week is Thanksgiving–
Michael: Surfaces, I’m sorry. As next week is Thanksgiving, I think Vincent picked this paper to encourage us all to hit the gym the day when the majority of Americans wear loose fitting clothes so that they can enjoy the bounty of their labors. So today we are going to go to the gym, and I know what you might be saying. There is no safe place from these damn microbes. But consider that athletic centers have been locations for the transmission of community acquired infections for quite some time.
For those of you who follow the NFL before concussive injuries took the spotlight in NFL medicine, the NFL had seriously investigated the issue of moving drug resistant microbes among their players. They were very much concerned about this with perhaps the most famous being Brandon Noble of the Redskins who was featured in a number of Washington Post stories and subsequent to his knee surgery contracted an especially recalcitrant variant of methicillin resistant Staphylococcus aureus or MRSA. Then in the early part of this decade the issue reached a tipping point in the NFL where teams were actually going to forfeit because one team had a MRSA infestation and they didn’t want to use the locker rooms, they didn’t want to play.
So that forced the NFL to work with the Duke Center for Antimicrobial Stewardship and Infection Prevention, where they now provide infection prevention education to all 32 teams. This effort is supported by both the NFL and the National Football Players Association. The only reason I am bringing this up is because what is good for the NFL is good for the general population.
Vincent: Wait, does that mean we have to bang our heads together, Michael?
Michael: Well, you know, those of us who submit grants on a routine basis feel that we’re already doing that.
Vincent: You’re right, you’re right.
Michael: So as you can imagine, the program that the Duke group set up focuses on several key aspects of infection prevention including, and this is the reason I put this little link in, is cleaning and disinfection of facilities and equipment as well as infection prevention education for both the staff, the players, and the coaches. The paper here today introduces us to the use of a continuously active antimicrobial surface where this intervention as introduced here is an integral piece of common equipment used in most gyms, CrossFit centers, rehab centers, and so it should offer a way because what copper is doing is it is continuously inactivating the microbes that fall on them or are transferred from touch, so you simply are reducing the bio burden on these surfaces so you can limit your exposure to the microbes and if you have any micro abrasions or cuts or you have a tendency to rub your eyes, you know, just prevent moving microbes.
So the study assessed the capacity of these copper alloys to reduce bacterial burden associated with these high touch athletic center equipment. So what they did is they designed a first rate study where copper alloy weights and grips were rotated with rubber coated and stainless steel controls in an undergraduate college athletic center over a 16 month period.
Elio: Remind us, how do they add the copper, actually? It’s not clear from the paper. Did they coat the surfaces?
Michael: No, so this is actually a tube, just like the plumbing fixtures in large buildings, they actually use either a copper sleeve over it because it had to support the kilos of weight. This was not a coating, this was actually a solid copper.
Elio: Oh I see, that’s what it was, okay.
Michael: So why this is important is there is a literature out there that not only does this affect NFL teams, namely community acquired microbes being transferred in these facilities, but surfaces in high schools are also at risk. High school wrestling teams have tested positive for community acquired MRSA, which represents approximately 25% of all the MRSA infections that the medical community saw in the United States. And if you think about it, in the US, 57 million people are members of at least one athletic facility. So their hypothesis was pretty straightforward. Would there be a decrease found in the bacterial burden found on the copper alloy surfaces tested compared with the bacterial concentrations associated with the standard non copper equipment?
So Dr. Hinsa-Leasure and her team determined the bacterial burden following the protocol that has been out in the literature for quite some time of sampling high touch surfaces and they did this in the control and they used 5 types of commonly used equipment. They used control grips, they looked at dumbbells, and they had a whole set of ranges, and it’s interesting because in the US, we measure weight training in pounds, and Dr. Hinsa-Leasure and her team reports the values in kilos because that’s how the rest of the world measures weight.
Elio: Good for them!
Michael: So the dumbbells were 6.8 kilos, 9.1, and it comes into play as they report their data. The US EPA recognizes five families of copper alloys as antimicrobial with their sole criteria for this designation being that they will reduce the bacterial concentration on that surface by 99.9% within 2 hours of an in vitro bacterium being placed on the alloy. The alloy family that was used here was a copper alloy called C706, which is 90% copper and 10% nickel by weight, and they used this over pure copper because the 90/10 version of this alloy doesn’t tarnish from exposure to human sweat. The silver coins that you all carry around in your pockets in the US are actually copper. They are clad in two layers of cupron nickel which is 75% copper and 25% nickel and they’re on a core of pure copper. So US coinage is actually antimicrobial.
Elio: This is true for dimes, not for the rest.
Michael: Dimes too, no, dimes, nickels, and quarters.
Elio: Oh really, all of them? Wow.
Michael: All of them. We got rid of silver in 1965, so if you’re out there–
Vincent: I remember, I remember that. Because the new coins have the little copper rim around the edge.
Michael: Yeah. You can actually visually see it as the platting begins to wear off. So their first, the bulk of their data are associated with the bacterial burden was indeed significantly decreased on these high touched alloy surfaces, and the copper grips had significantly lower bacterial loads than the control items with the average decrease being 94%. They measured a lot of these, they looked at 543 copper objects and 536 for the control, and for every type of grip test that the reductions in bacterial counts decreased from 85 to 97% on these surfaces. The thing that harbored the most microbes were the control dumbbells. And they carried the highest loads with around ten thousand colony forming units per hundred centimeters squared, which is similar to the concentration of bacteria that we see associated with the rails of hospital beds, the most highly soiled surface in a hospital.
Vincent: Michael, I’m just gonna take this as a good reason not to go to a gym (laughter)
Michael: You’re saying that, Vincent, is, break out your pants with the wider waist after next week. Winston Churchill had his fat suits, he said. So Dr. Hinsa-Leasure broke these things down by item and found that the lightest dumbbell also had the highest bacterial load. So the rule of thumb is that if you’re gonna go to the gym, use the heavy weights because they are likely cleaner and lesser clad in copper. And they do a beautiful job of presenting both the mean and median and they have a clever way of doing this. The means are the bars on their graph and the median numbers are the triangles. And remember, the median sort of throws out the outliers and that gets into the other discussion of where do the bacteria come from? And they are associated with your skin, they are associated with hair follicles that are falling out, and they are just everywhere in the gym.
So that is why they were reporting at both mean and median, because the means, if you have someone who has a little bit of stool on their hands as they were pulling on their gym clothes, they may have contaminated themselves with some resident material as the infectious disease people refer to it, our fecal patina. We are all covered in a fecal patina that we routinely wash off each day but sometimes we don’t wash too well and so you may pull some along and it will come along. They also characterize the most common species and isolates that they found and not surprisingly they were staphylococci and micrococcus which are normal Gram positive organisms that we see on people, and the staphylococcus was the most common genus.
Antibiotic susceptibility testing, they looked at three drugs, vancomycin, erythromycin, and linezolid, and these were conducted on 17 isolates from copper surfaces and 18 isolates from the control. They found that erythromycin resistance was found in 6 copper isolates and 8 controls. That’s not surprising because the staphylococci that are on us are exposed to the antibiotics that we routinely encounter in our everyday lives. Vancomycin resistance is quite common, VRE is actually moved into hospitals from the community, as is MRSA and the others and they all carry drug markers, and so it is not surprising that these things have persisted. The reason this is all significant is because community acquired infections have persisted and continue to persist at the same rate while hospital infections because of the interventions on the behalf of the infection control community have been really going after HAIs in the hospital. Because of their interventions, those infections have dropped. But the community acquired data have not dropped as much as we would have liked to have seen. So that’s the paper in a nutshell.
Elio: I’d like to add that in the field of copper as an antimicrobial, Michael Schmidt has made major contributions! He hasn’t said that because of his innate modesty. (laughter)
Michael: Well, thank you for that.
Elio: Michael has made important contributions.
Vincent: Michael, the obvious question is, that everyone is thinking, would this decrease in the number of bacteria on these surfaces make any impact on transmission?
Michael: That’s a hard thing to test for and people are beginning to do that set of experiments in the hospital with C. diff, and Curtis Donskey was here last week and he is one of the well established researchers in the C. diff community, and what he has people do is he has them conduct an exam on a patient, whether it be gloves or hands, and then he has them place their hands on a petri plate and he can actually then recover C. diff spores. The trick he has to not pull your normal flora off your skin is he wipes you down with an alcohol wipe or wipes your gloves down with an alcohol wipe and of course the alcohol has no effect on the C. diff spores.
You can literally see how easy it is to transfer, in the case of C. diff, from patient to health care workers, hands or gloves, to another surface or something else. So many hospitals are now beginning to try to understand the movement of C. diff. And the NFL was certainly very much concerned about this that the players association as well as the owners have funded Duke to run this continuous infection control program for the NFL to limit the spread of these organisms in the locker rooms. It is a significant issue and it has indeed worked.
Vincent: Shouldn’t we just replace weights with copper cladding in some facility and then monitor the occurrence of infections?
Michael: That’s the best way to do it, but it’s hard because the number of interactions that have to take place, because for the most part, people don’t get sick. They’ll just become colonized. And so it’s a challenging study to do because most of us are healthy. The thing that really grosses me out in the gyms are the squishy grips because they’re sort of like sponges, and while you and I might put our sponge in our dishwasher to knock the load down, when was the last time you think those squishy grips on the treadmill were put in a dishwasher?
Michele: So wash your hands.
Michael: I do! And that’s why I’m healthy!
Vincent: So Michele, that’s what your solution would be?
Michele: Yeah, I mean, the NFL is especially vulnerable because of all the contact, especially the linemen, they end up with open wounds and then if they get MRSA in the open wound then it’s a real problem. But if you’re just lifting weights and you wash your hands after, I think there is less risk of having disease.
Vincent: Maybe the solution, Michele, is to make football clothing with embedded copper fibers (laughter)
Michael: Then you have problems with, because that’s a leeching technology where the copper is actually displaced by the sweat, and so then you are, it gets washed out because what’s the active ingredient in soap? It’s sodium laurel sulfate, so it’s the same difference as sweat. The sodium displaces the copper from the fabric and you end up with copper in your wash load.
Vincent: Oh, okay. What do you think, I mean in your studies, Michael, you looked at the rate of hospital acquired infections when you put copper in strategic places in the hospital room. Is that going to happen here or as you say it is too hard to do because most people don’t get sick?
Michael: It’s whether or not it’s affordable, and I just came back from the healthcare design show, and there are a number of new technologies entering the marketplace that are specifically beginning to figure out, as Elio tumbled to, is there an inexpensive way of placing an antimicrobial concentration of copper on very complex geometries? And that technology is just coming to market now, and so we are hoping that will lower the cost of getting some of these surfaces in the hospitals to help control infections. Because that’s what we found, and this is no great leap of faith to most microbiologists, if you lower the concentration of bacteria on critical high touch surfaces, you are going to lower the risk of acquiring a microbe from the built environment. And that’s what we showed, we cut the infection rate by almost 60%. That was a significant amount and it’s the hope for controlling some of these nasty things like carbapenemase resistant enterobacteria which seem to be much more harder to control than MRSA and VRE, and even C. diff.
Vincent: So Elio, next time you go to the gym, just lift the heavy weights, okay?
Elio: That’s it. That’s it. 100 pounds and up.
Michael: 100 pounds and up! Or at least 50 kilos.
Vincent: Thank you, Michael, that is great, and I like that these small colleges do this sort of thing.
Elio: It’s lovely.
Vincent: I was at a conference here in the New York area, it’s called Metropolitan Area Conference for Undergraduate Biologists. And it was students and professors from local small colleges that are mainly teaching schools but they like to do some research. And it was just so, I was lucky to be the keynote speaker, and these students were so engaged. They had a poster session, they were so excited about their work. It was just so great to see because we’re all impressed by what can be done at large universities but these small ones play a big role. So this is from one of those small colleges, so that’s great.
Michael: And this is a tradition of Grinnell. Grinnell has always encouraged its undergraduate community to conduct cutting edge research, and they, and this is just another example of them being successful. So, well done, Grinnell.
Vincent: Alright. For our paper we have an offering from Michele.
Michele: Yes. This is from the new issue, the November issue of MBio, it is entitled “Mechanisms of Surface Antigenic Variation in the Human Pathogenic Fungus Pneumocystis jirovecii.” And it is from Emanuel Schmid-Siegert, Sophie Richard, Amanda Luraschi, Konrad Muehlethaler, Marco Pagni, and Phillipe Hauser. And they are from the Swiss Institute of Bioinformatics and the Institute of Microbiology, both in Lausanne, Switzerland, and the University of Bern in Bern, Switzerland.
So they are looking at a fungal pathogen that causes a serious pneumonia, and you may have heard of it as PCP, pneumocystis pneumonia. This is a serious infection that especially affects people who are immunocompromised. So we see it in the elderly or people who have transplants because they are on immuno suppressive therapy, or it really came to the fore during the HIV epidemic in the early 80s, so by the late 1980s some 75% of people who were living with AIDs developed this pneumonia due to pneumocystis. So it can be treated, there are antibiotics that work but you require about a three week course to clear the infection and some of these drugs can cause side effects like nausea and rash but it’s really important that people be treated because the mortality rate can be anywhere from 5 to 30%.
So worldwide we are seeing about 400,000 cases of this PCP each year. Here in the US we don’t have a surveillance system so we don’t have good numbers. So despite this heavy burden of disease, we don’t know a whole lot yet about the fungus that causes this pneumonia, and that’s in large part because we have not yet figured out how to culture it. So a lot of the knowledge that we have comes from studying related fungi and drawing parallels, and then also more recently from doing whole genome sequencing. But what we do know is that the fungus undergoes a bi-phasic life cycle so we can look in the microscope and see that it can alternate between round cysts that have a rigid, thick cell wall, and that is thought to be the transmissible form, and then there is the replicative form, whose cell wall is thin and flexible and actually lacks chitin, which is a structural component of most fungal cell walls.
Elio: That’s really unusual, isn’t it.
Michele: Yeah, it looks more like an amoeba in shape. In fact, it was originally thought to be a protozoa but later they named it Pneumocystis carninia, which is–
Michele: Carnini? Which is a–
Michele: Say it again? Say it again, Elio?
Elio: It’s called Carinii.
Michele: Carinii, ah. So that’s the rat adapted but now it is known to be a separate species, jirovecii. So there also was some genomics work that set the stage for this study. Ling Ma and his colleagues published in 2016 Nature Communications a paper, the whole genome sequence, and also RNAseq profile data for a rat, a mouse, and a human pneumocystis species, and they recognize that a large amount of the genome, up to 5% of the genome encoded a repeated family of genes that encode surface glycoproteins. And we recognize that this large reservoir of surface glycoproteins falls in a category of genes we call contingency genes, which means that the organism has them to ready them for a future that is uncertain and unpredictable but possible, and we will see how that plays out in the infection.
Elio: Money in the bank (laughter)
Michael: Money in the bank.
Michele: That’s right. So what Schmid-Siegert and her colleagues establish here is that this Pneumocystis jirovecii is a quick change artist. It has built in its genome mechanisms to change its coat at a high frequency, and we call this antigenic variation. So this would alter how the fungus interacts with host cells but also alter how it is seen by the immune system. So that’s probably the selective pressure for this mechanism that we are going to learn about. So for this study, they obtained a bronchial lavage from a patient who happened to be HIV positive that was colonized with Pneumocystis jirovecii and they determined through a series of genetic and statistical tests to be less that 7% probability that this patient was co infected with a second strain. That was important because they wanted to be able to interpret their data in terms of what happens as a population expands and diversifies.
So they extracted the total DNA from this bronchial lavage and then used a trick to enrich for the DNA from the pneumocystis based on its known lack of methylation and then they amplified randomly by PCR, generated a library, and then used whole genome sequencing. In particular they relied on PacBio sequencing technology. This is important because with this technology you can get long reads of DNA in each run in the kilobase range. That is important because if the DNA contains a lot of repeated sequences it gets very difficult to correctly assemble. So with this PacBbio sequencing they were able to with confidence assemble the contigs into the chromosomal segments and learn not only about the sequence of each gene but also how they were arrayed on the chromosome.
So the other key point before I get into the data is we know from the literature that the pneumocystis have adapted to a particular host range, so Pneumocystis jirovecii looks to be a human specific lung pathogen. We don’t have any evidence that it grows elsewhere, and then other species in this genus are adapted to the rat or to the mouse. That’s relevant because it means that the pathogen, to persist, must be able to avoid clearance in the human lung, and we have also known from looking at the whole genome size from earlier studies that it is quite reduced relative to other fungal pathogens or fungi, rather. In other words it has thrown away a lot of its metabolic pathways and has a more restricted genome and that is a hallmark of a microbe that has adapted to a particular niche. You can deduce that it is probably scavenging nutrients from that niche and it is specialized for that site. So do any of you want to add any more background details about pneumocystis before I get into anything else?
Elio: No but you may want to compare or contrast it to other organisms that use antigenic variation as a survival strategy.
Michele: Yeah, so we will talk about that. A number of, once we talk about the mechanism, this is, as Elio points out, a very common virulence strategy in the microbial world to alter the surface and then the molecular details of how the microbe carries that out differs from species to species. How’s that for now?
Michael: That’s good.
Michele: So they have now collected whole genome sequence from the population of fungi that they isolated form the lungs of this one patient and then they used a whole suite of bioinformatic and statistical tools to reconstruct the genome and deduce probable mechanisms of this antigenic variation.
Elio: By the way, this is as good an example as I know of the marvels of modern technology because none of this would have been possible without recent developments in genomics and bioinformatics. This was a bug you could not cultivate, you couldn’t do anything. And here you are doing very advanced studies.
Michael: I should also point out that they did manual curation of the MSE genes that led to their classification of full length, partial, and pseudo genes, so there was, they’re not cheating with all of the computers, there was a heavy lift on behalf of the authors.
Elio: That’s true.
Michele: And also using the literature to make predictions.
Michael: That’s true.
Michele: So they had some precedence to guide their design. So when they had the full genome sequence, they identified in this population genome pool 113 different genes that encode MSG genes or major surface glycoprotein genes. Remarkably, only 2 of the 113 were identical. So there was tremendous variation in this pool of 113 surface glycoprotein genes.
Elio: That’s quite remarkable, isn’t it?
Michele: Yeah. So how does this work? What they were able to do by doing alignments and statistical tests, they were able to group these 113 genes into 6 different families, and they could compare each family and look within the family at diversity and also between families. In table 1 they have some of the characteristics that they identified. What they found in general was that the DNA sequence identity varied between 45% for say, family 6, to 83% identity for families 2 and 3. And at the protein level, the sequences varied between 21% for some families up to 73% identity for the protein sequence. They also found that the N terminus of most but not all of the families had a signal peptide which predicts then that the glycoprotein would be secreted out to the cell surface.
Elio: It’s a very important point, by the way, because it’s an important prediction that it comes through.
Michele: Right. Most of the families also have at their C terminus a signal for modification by a GPI, glycosylphosphatidylinositol modification. So this is a greasy moiety that would be added and is known to mediate attachment of proteins to membranes. So we have the picture now that the family of proteins being secreted and then stuck to the outside surface, so they are decorating the outer surface of the fungus. They also have between 1 and 14 glycosylation sites, so they are heavily glycosylated, and a number of the proteins have coiled coil domains and also leucine zipper motifs, and we can deduce from that that they have the potential to interact as dimers or multimers and create even more antigenic variation.
So if we look at figure 2 it shows the features of the proteins and lays out how each family differs. Here I want to emphasize that one of the families, MSG1, is an outlier in that it lacks a promoter sequence. Instead, at the 5’ end of the MSG1 family genes, there is a short segment that is predicted to mediate recombination. So it is called a conserved recombination junction element. It only has one but elsewhere in the genome there is another element called an upstream conserved sequence that has a strong promoter and an ATG start codon and at the 3’ end it has another one of these recombination sites.
Michele: We can deduce then that you can have recombination between these many different MSG1 family genes with the strong promoter and activate expression of the MSG1 family.
Elio: And is this mechanism, this is not entirely new, right? This finding that you can activate the gene by the combination.
Michele: That’s right. So that’s used throughout the microbial world, different mechanisms of recombination to turn genes on or off, or to change which cassette is expressed at what time, that’s right.
Elio: But in these, in fungi where this has been known before, probably.
Michele: Well the mating type loci for Saccharomyces cerevisiae, a and alpha, use a recombination based mechanism to turn on or off.
Elio: Yes, of course, of course.
Michele: So what we have then if we stand back and look is that the genome is loaded with these six different families of genes that each one with quite a bit of variation within the family and five of the six families can be expressed on their own and then the other family, the MSG1, can be activated by a recombination mechanism that links it directly to a very strong promoter. So there is data from similar systems that tell us that the MSG1 family protein would be expressed at high levels and probably be one of the dominant antigens, and then the other MSG2,3,4 whatever family members could also be expressed and sprinkled among a surface as well.
So they then want to verify their in silico analysis where they used sophisticated predictions about what genes are possible. They wanted to go in and ask in their population of genomes do they actually have evidence for this recombination event between an MSG1 cassette and the UCS promoter sequence. So they are able by PCR to interrogate their whole genome sequence, and indeed they identified 18 different combinations of MSG1 genes that had been recombined into the USC strong promoter. So they have evidence that the cell population is in fact undergoing recombination to turn on a particular MSG1 gene.
So the next set of data is especially interesting. They were able to, by sequencing the adjacent DNA and using that to align the contigs, they were able to map each of these MSG loci to the telomere of the yeast chromosomes. So the telomere is the far end of the chromosomes, all eukaryotes have them.
Elio: Being linear. Being linear, I should say, unlike most bacteria.
Michele: That’s true, that’s true, because eukaryotes have linear chromosomes, by and large. What’s true about chromosomes and telomeres is that there is great variation from species to species but there are some general rules that hold true, and one is that the telomeres have very funky structures. And also weird biochemistry. So they have different rules for how they manage to replicate their DNA, how they protect those free ends from nucleases and prevent the loose ends from being recombigenic and causing mutations and also in a number of species it has been established that some regions of the telomere are silenced. There is a specialized mechanism that can prevent gene expression or de-repress expression of genes in that locus. So it is set up for some really special DNA replication, DNA transcription, etc.
So we can’t know what tricks this particular pneumocystis is using, but we can expect that there will be some interesting biology. But there are two features of telomeres that are shared by many different species that promote gene diversification of loci that are present at telomeres. The first quality is that most telomeres are tethered at the periphery of the nuclear envelope which means the telomeres are spatially adjacent to one another, or close together, so you can have some interesting dedicated biochemistry happening at that site. They also have a lot of repeated sequences, telomeres do, which would promote then recombination by homologous recombination at those repeats.
So what would be the consequence of a crossover event at the telomere? Imagine that if you get a single crossover, two chromosomes could swap their ends and if you had a gene encoding a surface glycoprotein mapped down at the telomere and it is present in many different copies, then you could imagine a high rate of homologous recombination between these different genes that encode these surface glycoproteins and therefore promote gene diversification. Is that clear? Is that the general connection between the structure at the end of the chromosome and promoting high levels of recombination?
Vincent: Yep, I got it.
Michele: So we’ll see that that is actually, there is a precedent for that in a wide variety of organisms. So in particular we know that another fungus, Candida glabrata, also has surface proteins encoded at the telomeres. Again, it’s a large family and there is a dedicated mechanism that generates a lot of antigenic variation to promote colonization by that yeast. And in a different family we have Trypanosome brucei and also Plasmodium falciparum, the agent of malaria. They too are experts at evading host immune responses and specifically antibody dependent responses by altering their surface chemistry at a high frequency and again, they do so by having repeated genes at telomeres and specialized mechanisms to promote recombination based diversification of these families.
So the idea is that when a person is infected with a particular Pneumocystis jirovecii, as the bug replicates we are getting a high level of recombination at the telomeres, so in that population we are getting sub-populations that have a different coat, the quick change artist idea, and therefore the immune system would not see a heavy load of one particular antigen type and every time we raised an antibody against one type, there would be a sub population that no longer had that antigen and it would continue to grow while the other ones might be cleared. So we have a lot of potential mechanisms that can account for the ability for this pathogen to survive and colonize at least transiently in the lungs of humans and most likely similar mechanisms are working in the rat and the mouse as well because they have many of the same genetic features.
Vincent: What is it about the telomere, is it because they are anchored adjacent to each other and that promotes recombination, is that why it’s found in so many different organisms, this mechanism?
Michele: I think that’s one thing and the fact that you can have a single crossover and not be deleterious.
Vincent: Yeah, because it’s at the end, right.
Michael: Or maybe simply solution chemistry, it may be exposed to the solution because it’s not all knotted up in the chromatin.
Michele: That’s true.
Elio: That’s also true.
Michele: And then they also are known to have a lot of repeats but that’s kind of a chicken and egg, I don’t know if that’s intentional or why that is but that also promotes a lot of recombination.
Elio: So there are probably quite a few systems that take advantage of the unique properties of telomeres to conduct their, transact their business.
Elio: It’s just a busy place.
Michele: (laughs) So as Elio said, this is really impressive that even though we don’t yet know how to culture this organism, we can come up with some pretty specific molecular mechanisms to account for antigenic variation which could then drive hopefully one day new therapies to treat these nasty infections.
Michael: The scary thing is vaccines because that’s what the HIV community desperately needs is a vaccine against this horrible fungus and with this rearrangement it really limits that possibility unless they can find a common antigen that can elicit sufficient protection.
Vincent: If you can have immunity that is sterilizing and prevents replication, because it’s during the replication as Michele pointed out, that’s when you generate the diversity. If you can prevent that, that would work, and that’s hard, it’s hard to get sterilizing immunity with vaccines. But that would be the key.
Michele: The other feature of this life cycle that is relevant here is that the yeast can undergo, they can mate and fuse their nuclei, so that is another opportunity for the telomeres during meiosis to undergo this recombination.
Elio: Oh my.
Michele: So this cell is really built for a high level of recombination that would grant it antigenic variation or quick change artistry.
Vincent: That’s really cool.
Elio: In the most general terms, I think this goes a ways to explain why it is such a common pathogen. It’s not always a pathogen but it’s a common inhabitant of the human lung and rat lung and so forth because it has all these clever techniques at its disposal.
Vincent: That’s very cool. Are you finished there, Michele?
Michele: I am.
Vincent: You were able to get some information from the first author, Emmanuel Schmid-Siegert who says, well he is a computational scientist currently at the Swiss Institute of Bioinformatics, previously he was a post doc at the same place and he did his PhD in molecular plant physiology. He worked on arabidopsis, and then for his diploma thesis in biology which was at the University of Erlangen in Germany, he worked also on arabidopsis. So he’s had a bit of a change. He says he started his career as I said at Erlangen in plants and then decided to move to the University of Laussanne, again working on plants. He says that after a few months in this lab and completing his diploma thesis, the PI Edward Farmer offered him the possibility to work on this project as a full PhD thesis, and he did that, he published a couple of papers, and then he eventually moved on to Lausanne to see whether he would, he said, “warm myself up to a complete non wet lab work.” (laughter) “And after trying that out I accepted happily found that data analysis is very demanding but rewarding and fulfilling.”
He is very fond of the possibility to have a large variety of different research projects with various means of analysis and different organisms which means, for example, for each project you make a small team out of different colleagues to obtain the best expertise for that particular project. The project that Michele told use about today, he says was interesting and demanding as we tried to describe and analyze chromosomal regions which are extremely hard to resolve, they are located in sub-telomeric regions, they are very repeat rich, and after initial technical difficulties we are very proud and happy when we managed for the first time to hold in our hands assembled regions of MSGs.
During the further characterization of their families I benefited immensely from the long experience of our senior scientist, Marco Pagni, which was matched by the knowledge and manual curation from Phillipe Hauser. Manual curated. So he has some advice for younger colleagues. “As I switched completely my career path from a wet lab plant scientist to a more general computational scientist/bioinformatician I started to realize that one tends to develop a more and more narrowing field of expertise or research interest. I would recommend any young colleague to reconsider from time to time their options and opportunities, open their horizon and not to shy away from challenges, whether it might be another topic of research or even a new discipline.” There you go.
Michele: That’s so true. It also means you can then converse with people in a range of disciplines if you’ve had first hand experience and bring different ideas to your field.
Vincent: So he is married and his wife has a PhD and they have two kids.
Elio: And they’ll have a PhD some day (laughter)
Vincent: Pretty heavily in debt, that’s what we’re gonna call it here in the US, soon, if they take away the tax exempt status of tuition for graduate students.
Michael: Oh, that will be a disaster.
Vincent: Yep, I don’t think that should happen because it is hard enough to get people to go into science because it is one of the most poorly paid professions out there, at least professional professions. But let’s, we have a few minutes, let me read a few emails. I have one here from Anthony who sends a link to an article which I think Michele would like. It’s on CNN, was published on November 13, “Disneyland shuts down cooling towers over Legionnaires’ cases.”
Michele: Oh my.
Vincent: This happened in Southern California, they had an outbreak of Legionnaires’ disease, they had 9 people who visited the park in September who developed the disease and 3 other people who were in Anaheim but didn’t go to Disneyland, they also got sick. Listen to this, Michele, the ages were between 52 and 94. That’s typical, right?
Michele: It is. I was afraid you were gonna say they were 12, that would scare me. But I mean, it’s, I’m sad that elderly people got sick.
Vincent: So they shut down the cooling towers which is typically how these infections begin, right, Michele?
Michele: Yes. Cooling towers are a great way to spread it. They’re basically ponds that are open to the air and if wind blows across bacteria can travel long distances or they can be sucked into an institutional air conditioning system and dispersed through a building.
Vincent: So this is a cooling tower which runs air conditioning basically, right?
Vincent: So what do they have to do, they have to shut it down and clean it up?
Michele: Clean it out, yeah.
Vincent: But then it could get repopulated from the environment, right?
Michele: They usually do, or some of the legionella in the population will be in a dormant state and they are resistant to the biocides and so they’ll grow back once that selective pressure is lifted.
Vincent: So they did look and they found elevated levels of legionella in the towers, in the waters. And so they have to remained shut down until they can be verified to be free of contamination.
Michael: This happened in New York City, whatever happened to that? It sort of was in the news and it faded away. They go in with the biocides, clean up the water towers and the cooling towers and things got better?
Vincent: Do you know anything about that, Michele?
Michele: So for in the Bronx, that is true. They got better, but within two months they had another case and it was the same strains from the same cooling tower. So again, it’s hard to kill every bug. Legionella forms biofilms like many other bacteria and they are hard to clear. There are some incidents of hospital systems being colonized for decades despite remediation efforts.
Vincent: I’m surprised.
Michael: Often times they just get rid of the cooling tower and put in a new one.
Michael: Or they put it in a different spot.
Michele: Maybe they need copper, Michael, maybe they need copper up there.
Michael: Yeah, it’s probably an aluminum based system.
Vincent: They typically are because copper would be really expensive, but that might do it, right.
Michael: Actually, copper is not that much more expensive. Your biggest cost is in the fabrication and everybody who thinks copper is expensive, it is only for the electrical grade which is the stuff that literally comes from ingots that are recently mined. Most of the stuff that is used out for these antimicrobial applications are recycled because they are alloys and so you don’t have to worry about having pure metal. The pure metal is the expensive stuff. The recycled stuff is fairly inexpensive because there are very few applications other than wire where copper is in high demand, other than in China where copper is used as collateral for building things, they buy copper ingots and they use it lieu of gold as collateral.
Vincent: Michele, here in New York there are lots of water tanks on top of buildings that are made of wood. They can supply your showers and so forth. Are they ever a source of legionella or are they closed so that’s not a problem?
Michele: I think they are closed, yeah. Those water tanks are different from cooling towers.
Vincent: Cooling towers are a relatively recent invention and so is human infection by legionella, right, 1976, I think, right?
Michele: That’s when it was first discovered, yeah, because of the large outbreak, but then retrospectives still identify earlier cases. But you’re right, it’s a disease of technological progress.
Vincent: Air conditioning, huh.
Michele: And anything that generates water aerosols.
Vincent: I have one across the street, the building across from here, you may remember, there’s the Neurological Institute, they have a cooling tower on the roof which I can see in the summer, the water is pouring down the sides. The fans are whirring, and there’s huge aerosols, you know, up there.
Michele: So you no longer stand under there to get cool?
Vincent: No, I don’t go up there any more, yeah.
Michele: Now that you’re of a certain age.
Vincent: (laughs) I’m within the 52 to 94 range, I have to admit, yeah. 1965, which is when I remember what happened earlier, that we said this is reflective of my age. I don’t remember what we said. Anyway, I was 12 years old. I remember, when we switched, what were we talking about, Michael, at the beginning, where I said I remembered–
Vincent: Pennies, yeah.
Michael: And quarters.
Vincent: I was 12 years old. Alright, we have an email from Bailey, who writes:
I have listened to your podcast for over 2 years now but haven’t felt the need to write in until now. Having heard the great advice you’ve given to past listeners, I now found myself in need of your help. First, here is some background info on me. I’m currently an undergraduate at Penn State, majoring in Biotechnology with a minor in Chemistry. My choice of biotechnology is reflective of my preference to develop new processes derived from biological systems, over the purely inquisitive study of said systems. That is to say, I would like to do stuff with what we learn from microbes rather than just learn about them. I’ve been told by many that I will need to pursue grad school after I get my bachelors this December, if I am to ever become an attractive candidate for potential employers. This is where my dilemma arises.
On one hand, the opportunity of grad school presents a new and exciting range of biological fields to branch out from. On the other, I am conflicted on which field to pursue and fearful my choice will pigeonhole me for the rest of my life. For example, biomedical engineering promises an exciting job developing biotech systems but demands an engineering background I fear I lack. Bioinformatics and pharmacogenomics would allow me to capitalize on my programming skills but would force me to abandon the hands-on wet lab work I thoroughly enjoy. Pursuing a masters in biotechnology seems obvious given my major but the program seems so limited in scope now given my newfound options.
I feel like this is my last chance to choose a path, which in turn will define the rest of my career and by extension life. I feel trapped and don’t know where to turn for advice besides you four. Maybe you can help me, maybe you can’t… I understand it’s not a lot to go off on. Maybe just typing this out helps. Regardless, thanks for reading and thanks for the great content.
P.S if you are giving out free books still I’d love one
TL;DR can’t decide on what graduate program to pursue :/
And TL;DR means too long, didn’t read.
Vincent: Alright, who has advice?
Michael: I have some advice. Since they are into biotechnology and they have a minor in chemistry and they also have programming skills, fermentation is the answer, and there are a lot of microbreweries out there that are in desperate need of good microbiologists that understand how to cultivate strains, how to do the background on that fermentation, and if you can learn a little bit about active fermentation, you can effectively sign your own ticket, and I’m sure Penn State has many microbrews in town.
So if you go and chat with the folks running the fermenters, making the beer, and there’s a lot of stuff out there that is relying on fermentation, and the process control is often very critical in making batch to batch consistency, which often these microbrews have struggles with in the beginning. The way they are currently controlling it is they are buying commercially made inocula which are often very expensive. So if you could think about how to make, to effectively develop the inocula for them so they can limit that cost component in their production process, you’ll be worth your weight in gold. And you don’t necessarily need an advanced degree because you have the requisite skills and I’m sure Penn State’s program has rigorously prepared you to do those sorts of things.
Vincent: How about you, Michele?
Michele: Yeah, I would say, Bailey, I totally understand why you’re kind of in a panic and you feel like this next decision is gonna set the course for your life, but let me tell you that’s not true. Number one, yeah, as an undergraduate, you haven’t got enough experience to make such a big decision and the truth is that fields are changing all the time and so the work landscape 10 years from now is going to be different from what it is now. So give yourself, take some pressure off yourself, what I would do is just find a job and work for a couple of years in a field that interests you. You’ll meet some people that are in that field, they’ll be a great source of advice, you can see whether you are interested in the life that people are 5 or 10 years ahead of you are leading, and then you may or may not find that you need more graduate school, and you’ll be a better consumer, you’ll make a wiser choice about what kind of training you want to get. Even then, once you get a PhD, you’re not locked in to one career, even though I’ve stayed on the academic track I feel like every 5 or 10 years my career really looks quite different from what it did the previous period. So try not to put too much pressure on yourself, just go out and find a job that uses some skills that you have and see how you like it. Get the best training you can at every stage and keep your eyes open.
Vincent: Good advice. Elio, you got anything to add?
Elio: No, I think that Michele put it very well, I agree with everything that she said. Life experience is important and is the best teacher.
Vincent: And our author, the first author on the paper, wrote don’t be afraid to change completely, right. He went from wet lab to bioinformatics, so. Alright, good luck Bailey. Let me read one more from Adam who is a sophomore in high school.
I’m a sophomore in high school. Though I have been fascinated by microorganisms for several years now, I have only been subscribed to TWiM for a few months. I have future aspirations of being a microbiologist, but for now, I look up as much as I can about microbes. Your show discusses microbial functions in depth, describing cells and proteins and chemicals and molecules I’ve never heard of and can barely pronounce, and I don’t know what the heck you guys are talking about half the time, but I learn a little bit more every episode. This became evident to me while watching episode four of Star Trek: Discovery with my family.
Vincent: When the skeletal structure of the captain’s very large, unknown creature named “Ripper” is shown, I without even thinking said “Tardigrade.” A few seconds later, one of the characters said it was comparable to the “tardigrade of Earth, a microorganism.” Later in that episode, it’s revealed that the giant tardigrade is sensitive to what the trans-warp spores are doing. I again without thinking said “Chemocommunication.” I don’t think that’s a real word, but it suited my purposes. Later in the episode, it was revealed that that was exactly what was happening between the giant tardigrade and the trans-warp spores. I know these are probably the ramblings of sweaty high school nerd and this might not ever get read, but my point is, thank you. Thank you for cultivating my knowledge in microbiology and teaching me more than I ever realized.
Well, this is a good time to wrap up TWIM 165 and by the way, Adam, thank you, and keep on listening. That’s the way you learn, just immerse yourself, as Michele said, find something you’re really excited about and immerse yourself in it.
Michele: I could also direct Adam to the publications by the American Academy of Microbiology, there is an FAQ series that is written for the general public, and they have some on yeast and making beer, they have some on cheese, they also have some on flu and MRSA, all kinds of really cool topics, and they are meant for a general audience and you can certainly get a lot out of them. So just do a search for Academy FAQ ASM and you’ll come across the whole catalog.
Vincent: Alright, TWIM 165, you can find it at asm.org/twim. Apple Podcasts, send us your questions and comments, email@example.com, if you’d like to support us financially you can go to microbe.tv/contribute to find the variety of ways that we have. Thanks to everyone today, Michele Swanson from the University of Michigan, thank you Michele.
Michele: Thank you.
Vincent: Elio Schaechter at Small Things Considered, thank you, Elio.
Elio: Hey, my pleasure, thank you.
Vincent: Michael Schmidt is at the Medical University of South Carolina, thank you Michael.
Michael: Thank you, I’ll see you at the gym.
Vincent: (laughs) No no, I don’t go to the gym. I’ll see you, yeah, okay. I’m Vincent Racaniello, you can find me at virology.ws. I’d like to thank the American Society for Microbiology for their support of TWIM and Ray Ortega for post production. The music on TWIM is composed and performed by Ronald Jenkees, you can hear his work at ronaldjenkees.com. Thanks for listening, everyone, we’ll see you next time on This Week in Microbiology.
Content on This Week in Microbiology (microbe.tv/twim) is licensed under a Creative Commons Attribution 3.0 License.
Transcribed by Sarah Morgan.