Scientist Stories: Jean-Laurent Casanova, Are Infectious Diseases Only Infectious?

We're going to start with a simple question. Can one of you tell me what's a Ukronia? It's obvious? Everybody knows? So who knows? Is there a student who can define a eukarya? None, or at least one? Yeah, one here. Are you raising your hand, or are you on the contrary asking me not to designate you? So nobody knows what's a eukarya among the students? Interesting. So eukarya is— a narrative that tells, imagines what would be the present day if something that did not happen in the past had happened. For example, what would be New York if Christopher Columbus had not discovered America? That's a Uchronia. It's a very interesting exercise. For example, what if the French had won the War of Seven Years against Britain in the 18th century? Probably you'd be all speaking French. It would be much easier for me, frankly. And we lost that war just because we didn't care, which was, of course, stupid. Anyway, so the Uchronia here is to give you an idea of what we're going to discuss now, that is the notion that infectious diseases are not only infectious but also genetic. The Uchronia is what if Darwin in 1865 had read Mendel. So Mendel, as you know, was a monk in Brno, or Brno for the Slavic pronunciation, in Moravia in the south of the Czech Republic, what is currently the Czech Republic and was Moravia, part of the Austro-Hungarian Empire in these days. And he discovered working with peas the laws of inheritance. I suppose you've learned the laws of inheritance, what is a recessive, a dominant trait. Well, these were discoveries made by this genius working in, in Bruin. So he published his paper in 1865, and nobody cared. Nobody understood the paper. His laws were rediscovered in 1900. And the Uchronia here is what if Darwin had read and understood Mendel's paper in 1865? In these years, Darwin had already lost 3 children of fever, like Pasteur. And unlike Pasteur, he married his first cousin. In Europe until the end of the 19th century, and it currently still is the case in, you know, maybe a third of the human population, he married his first cousin. It was very common to marry someone from the same village and very common to marry a direct relative. So he married his first cousin, had 9 or 10 kids, lost 3 of fever. And the uchronia is that if he had read Mendel and understood the laws of inheritance, he would have deduced from the observation of his own family that death of fever is autosomal recessive. That is, is transmitted as a genetic trait. Because when you marry your cousin, your children are going to be 6.25% homozygous in their genome. And therefore, the likelihood to have an autosomal recessive disease, whether predisposing to infection or otherwise, is much higher. If you marry someone who comes from the other end of the globe, the likelihood of having children with an autosomal recessive disorder, it's much smaller because you marry someone who's much more distant. You understand me, right? We're all cousins. I mean, we're all cousins. But your more or less close or distant cousins. So if you marry your first cousin, your children are more homozygous than if you marry a very, very distant cousin. But that did not happen. It's a Uchronia. But it gives you a flavor of what I'm trying to tell you now and what we're going to discuss, this notion that childhood fever was discovered to be infectious, but it could have been discovered to be a genetic trait based on the observation in this family and other families where childhood fever runs seemingly like a genetic disorder because it affects multiple children and sometimes born to parents who are consanguineous. Now, this idea that infectious diseases can be genetic is actually old. I told you earlier that the idea predates the germ theory. Throughout the 16th, 17th, 18th century, the debate was whether fever was intrinsic or extrinsic. And Pasteur himself, when he discovered the infectious cause of the diseases of silkworms, he noted that one of these two diseases, the flasherie— there are two diseases of silkworms. Febrine and flesh-eat. And he noted that one of them, the flesh-eat, is not only infectious but also inherited. That's a quote that I've translated from Pasteur's book. In the course of hereditary flesh-eat, it is not the microbe that is transmitted from the parents to the offspring, but the predisposition to disease. So Pasteur could have gone in that direction. His life took him elsewhere. He discovered microbes. Then he started to try to vaccinate against microbes. But he was aware that there was more than the microbe. And that's a very good quote from this extraordinary book. So the idea that infectious diseases are also genetic is ancient. At the turn of the 20th century, when the problem was posed, the problem that we were discussing before the break, interindividual variability in the course of infection, some children developing, dying of infection when others infected by the same microbe remain OK. When that problem was posed, the idea, of course, that there could be a genetic influence, a genetic determinism of severe infections was put forward. And several lines of evidence quickly suggested that this idea was to be taken seriously. The first one shown here is a paper from 1905 that was published in the Journal of Agriculture. In 1905, this investigator a botanist in Cambridge University, UK, discovered that wheat— the wheat with which you make bread— that there was a disease of wheat called yellow rust caused by a fungus. It's an infectious disease of that particular plant. Well, he discovered that this disease is genetic. And that's a quote from the paper. You know, if you look at the second half of this text, the extent of infection there increased steadily, and a second count reduced the number of relatively immune plants to 64, whilst the remaining 195 were infected for the most part badly. Now the ratio 64 to 195 seems to be too close an approximation to the ratio 1 to 3 to be a mere accident. And taken in conjunction with the fact that the F1 generation was so badly attacked, I quote, "it is fair proof that susceptibility and immunity are definite Mendelian characters, the former being the dominant one." So in 1905, plant biologists studying a fungal infection of wheat convincingly established that this infection was not only caused by a fungus, but also by a Mendelian, that is, a monogenic inheritance. So from these years onward, as you know, the laws of inheritance had been discovered in plants. So plant biologists and plant geneticists were ahead of the game when compared with human geneticists and animal geneticists. In 1940, it was proposed that infectious diseases of plants in general, not only this fungal infection of wheat, but infections of tomato, cucumber, radish, you can name it, that these infections are Mendelian traits, that they follow an autosomal recessive or dominant pattern of inheritance. And these data were then confirmed from the '90s onwards with the identification of the genetic— molecular genetic basis of resistance or susceptibility to infection in plants. So 1905, this idea was documented at least in one type of plant. From the '20s onward, the idea that the germline genetic background may influence the outcome of infection was also documented in animals, in particular by Leslie Webster, who worked here at Rockefeller. And who in a series of beautiful papers established that different types, different strains of mice, rats, or guinea pigs were differently vulnerable or resistant to one bacterium or another. And in a beautiful review that he wrote in the Journal of Heredity, he reviewed these studies that he had published in the Journal of Experimental Medicine, a journal that is that has been, since the turn of the 20th century, edited and published here at Rockefeller. He came up with this beautiful title that is explicit, Heredity in Infectious Disease. It's a beautiful paper that establishes the rules by which animals experimentally challenged with bacteria mostly, but also a few viruses, and fungi were vulnerable or resistant depending on the genetic background. So both in plants and in wild plants or in cultivated plants and in inbred animals, experimental or natural infections were shown to be genetic in a number of instances between 1905 and 1940. How about humans? For humans, a number of studies were conducted from the '20s onward, and I'm going to review quickly a few of these studies. Like the studies in plants and the studies in animals, these findings established that infectious diseases had a strong germline genetic component, but they did not identify genes, that is molecules or cells responsible for susceptibility or resistance. As you know, the identification of molecules and cells involved in physiological or pathological processes, that's a story that begins after World War II. And we're still before World War II. I'm still reviewing the evidence in plants, animals, and humans that between 1910 and 1950 supported this notion that infectious diseases were not only infectious but also genetic disorders. And I'm going to show you one of these studies. This is a beautiful paper, again published in the Journal of Heredity by Sister Flavia, who obviously was like Mendel, not only religious but also a scientist. And she published a family in which appendicitis segregates like a genetic trait, as though it's Huntington's disease or Parkinson's disease could segregate. Now what is interesting with appendicitis is that it is an infectious disease. Of the appendix. It's a little tissue at the end of your colon, at the beginning of your colon to be more precise. And it is caused by bacteria that are natural commensals, that is bacteria that live within your gastrointestinal tract. And about 5% to 10% of people develop appendicitis. I don't know if I'm authorized to ask you who has appendicitis, but it's probably 1% to 10% here. If I were in France, I would ask here. I'm being very careful here. All these questions of privacy, all these things. Anyway, I don't want to know who had appendicitis. But I can tell you it's probably 1% to 10%. So before surgery, people died of appendicitis. It was simple. And then thanks to surgery, people recover. I had appendicitis, for example, and I recovered after surgery. So what's interesting with appendicitis from the point of view of the genetic theory of infectious diseases is that it's infectious, but it's not contagious. Let me repeat. It's infectious, but it's not contagious because the infection of the appendix is caused by bacteria that normally reside within your gastrointestinal tract. And you can shake hands. You can kiss on the cheeks. It's not going to transmit appendicitis. So the segregation and even the clustering of cases of appendicitis in a given family cannot be ascribed to contagion. If you have a family where half of the members have had tuberculosis, well, you can say, as I alluded to earlier, you can say, well, they have TB because they're sharing food. When one is coughing, he's contaminating the others. It's difficult to disentangle the contagion from the predisposition within a household for an infectious disease that is contagious. But appendicitis is an infectious disease that is not contagious because it is caused by normally commensal microorganisms, bacteria that reside within your intestine and your colon. So that was very smart, very clever of Sister Flavia to try to collect and describe what we call multiplex families, that is families in which there are multiple cases patients affected by the same condition. And in that beautiful family, she really noted that, well, there was more, clearly more cases of appendicitis than predicted based on the frequency of appendicitis in the general population. It's not 1% to 10% here. It's like 25% or 30%. And it really segregates like a genetic disorder. Anyway, when I read this paper, I couldn't refrain from calling the College of St. Francis to ask whether they knew more about Sister Flavia. But I got a picture. That's what they had. So that's a picture of her in the '40s when she published that paper. So other examples followed. This is a beautiful study of 39 cases of appendicitis in a single family pedigree. That's remarkable. I mean, even for real genetic disorders—cystic fibrosis, sickle cell disease, and others—you very rarely see families with 39 patients. So this is a family with 39 cases of appendicitis. So these data suggested the mere clustering of this infectious disease that is not contagious in itself suggested that there was more than the bacterium and that it may in particular have something to do with the germline genetic background and the transmission of variations in the genome of these patients in these families with appendicitis. OK? So that's one type of study. It's not compelling. It's suggestive. A second type of study was much more compelling. These are called the twin studies, and in particular, the concordance twin studies. So there are two types of twins. You know that. You know you have the identical twins and the fraternal twins. That's the common denomination. A more scientific denomination are the monozygotic twins, versus the dizygotic twins. The monozygotic twins are indistinguishable. They're the identical twins. And the fraternal or dizygotic twins, it can be a boy and a girl. So the identical twins are identical. They have the same germline genetic background. Same genes. They're clones, so to speak. OK? The fraternal twins only share a proportion of the genome. They have the same parents. They can resemble each other. If both parents are red hair, well, maybe they're going to be red hair. There's a good likelihood. So they're going to resemble each other for a number of traits. But still they're going to be different. One may be very tall and the other shorter. One may be a genius in mathematics and the other may be great at basketball. There is diversity within the sibship. But still they do share some genetic background. That makes a big difference between the monozygotic and the dizygotic twins. Now, how about their environment? Well, they all live— the sibs, the twins, live in a given household. That is, the monozygotic twins are exposed to the same environment, in particular, the same microbes. They share meals. They live in the same apartment or the same house. They go to the same school, typically, and so on and so forth. The fraternal twins, too. So the environment is shared whether you deal with monozygotic twins or fraternal twins. The only difference is the germline, the genetic background. It's identical for the monozygotic twins, whereas it's only in part shared for the fraternal twins, the dizygotic twins. OK? So early on in the 20th century, geneticists came up with a good idea. And that applies to infectious diseases or any condition of interest, actually any variation of interest. It may be not necessarily a disease. It can be the height or the color of the eye or anything you like. They assess, they measure the concordance rate for any phenotype comparing the monozygotic pairs and the dizygotic pairs. Let's take the example of tuberculosis. This is a study that was conducted in Germany. And what you see is— if you can— yes, that works. You see here that the fraternal cotwins —their concordance of TB is around 20%, 25%. That is, if one of the two sibs has TB, if you study, for example, 100 pairs, in 25 of the 100 pairs, the second twin also has TB for the fraternal twins. OK? Now, for the monozygotic twins, it's 87%. That is, clearly the difference between the fraternal twins, the concordance of 25% for the fraternal twins and the concordance of 80% for the identical twins, this difference indicates, given that the environment is constant and shared for any given pair of twins, it indicates that the difference between 25% and 85% is due to the germline genetic background. So this indicated that TB was not only infectious, but also genetic. And the result here is as strong as can be. If you take severe genetic disorders that have what we call a complete penetrance, the concordance is going to be 100%. But for conditions that have relatively high penetrance, 80% or 85% concordance, is already very high. So these data strongly suggested, even more so than the clustering of cases of appendicitis in some families, that TB and other diseases that I won't review here had a strong human genetic basis. The third type of study that was conducted in these years were the adoptee studies. Now, the graph here does not come from a study in the '30s or '40s. It comes from the most beautiful study of the type that was conducted in Scandinavia in the '80s. Now, what is an adoptee study? When a child is adopted, you can compare the cause of death when this child dies as a child, an adolescent, an adult, or later in life, was the cause of death of his biological parents with whom he or she never lived, and the cause of death of the foster parents with whom the child shares no genetic similarity but shares the environment because he or she is living with the foster parents. Therefore, the comparison of the cause of death of adopted children with biological versus foster parents is a remarkable way to disentangle the influences of the genetic background, the biological parents, and the environment, the foster parents. So a large study was conducted in Scandinavia in the '80s. And it was agnostic in the sense that it analyzed the causes of death, premature death in particular, before the age of 40 or 50 in adopted children. Agnostic in the sense that it did not focus particularly on infectious diseases or a particular type of infection. It simply registered all adopted children in Scandinavia. And ask, OK, what do they do— what do they die from? And what was the cause of death of the foster parents and the biological parents? And does the cause of death of the adopted children correlate with the cause of death of the foster parents, in which case the environment is at work, or of the biological parents? In which case the genes are at work. Now, if you consider the infectious diseases, that is the adopted children who die of infection, they die of infection not when the foster parents die of infection, but when the biological parents with whom they never lived died of infection. So that was amazing. It was extraordinary results because of all human diseases tested, including cancer, including atherosclerosis, vascular problems, the most genetic disease of all, that is the group of diseases that showed correlation between death of the adopted children and death of the biological parents, not the foster parents, was infection. That is, even if the foster parents died of infection, it didn't lead to death of infection of the adopted children. Conversely, if the biological parents had died of TB or pertussis or else, the kids, the adopted kids, were much more likely to die of infection. So that indicated strongly that infectious diseases were actually genetically determined. So that took us to roughly 1950, and this is where we should have posed in the most pessimistic scenario a couple of hours ago. That was to indicate that I wanted to have a break, and we're not going to have a break now. We're going to go on. Now that I've set the stage, we're moving to the early '50s. So by the end of the '40s and the early '50s, there is evidence in plants, in animals, and in humans that infectious diseases are, A, infectious, and B, genetic. The infectious agents are increasingly well-known. But the genes that make some children or some adults sick of an infection or another are not known. What I've reviewed so far are sets of data that provide suggestive, sometimes compelling evidence that human genes are at work, but I haven't told you anything about the actual genes, the actual mechanisms by which certain microbes can kill one person while being harmless in another. So the cellular, the molecular era, the modern era, that starts in the '50s, in the 1950s. And it starts with the identification of the first primary immunodeficiencies. The first primary immunodeficiencies are conditions that are Mendelian trait, typically recessive, and that are associated with the occurrence of multiple recurrent severe, often lethal infectious diseases. Now, I want to ask you, who here knows or who doesn't know what's a Mendelian trait? Who knows what's a Mendelian trait? All right. That's pretty good. So for the others, and perhaps for some who think they know what it is but might benefit from a little more information, a Mendelian trait is a phenotype that segregates with a single gene lesion, with a genotype characterized by a single gene lesion, and that displays complete penetrance. That is, all individuals carrying the at-risk genotype display the phenotype. For example, an autosomal recessive trait, 25% of the children in a sibship are going to display blue eyes or red hair or whatever. OK, that's an autosomal recessive trait. That's one of the different types of Mendelian transmission. So in the early '50s, several primary immunodeficiencies were described. And the term primary immunodeficiency refers to Mendelian inborn errors of immunity. That is, anomalies of white blood cells or of components of the blood in the plasma molecules that are missing and the absence of which segregates like a Mendelian trait. Two examples are agammaglobulinemia. Some children were born without the antibodies that you referred to earlier. Another example is neutropenia. Some children are born without neutrophils, the granulocytes. That's a type of leukocyte that is needed to engulf invading microorganisms. Now, these two hematological and immunological anomalies segregate as Mendelian traits, and they're invariably associated with multiple recurrent severe infectious diseases. That was discovered in the '50s. And these examples provided conclusive evidence that some infections, at least, were caused by genetic disorders. And for the first time, there was more than an indirect genetic evidence. There was a molecule or a cell that was missing in the patient. Some patients lacked neutrophils. Others lacked immunoglobulins. So the field of primary immunodeficiency was in part started here, not by Bob Good, who worked then in Minnesota, but by Henry Kunkel. The field has expanded. There are now probably 300, maybe close to 400 Mendelian primary immunodeficiencies. The field has evolved in multiple directions that I will not review. I just want to emphasize, to stress that there are Mendelian primary immunodeficiencies that do not underlie infectious diseases, but a variety of conditions that can be inflammatory, autoimmune, or even malignancies, and the famous allergies that one of you earlier was interested in. Regardless, what I'm showing on the slide here is a typical Mendelian transmission of an immunological anomaly that segregates with an infectious phenotype. The left lower pedigree— this is a family in which there's an X-linked recessive transmission. Of a lack of gamma globulins, a lack of immunoglobulins due to a lack of B lymphocytes. The B lymphocytes are the cells that make the antibodies. These two patients in black are maternally related, and the gene that is mutated in this patient is on the X chromosome. As you know, boys have only one copy of the X chromosome, so a single mutation is sufficient for a boy to be sick. Whereas women in the family having two X chromosomes are carriers of the mutation, but they're not sick. They transmit the mutation to boys. So this is one example. On the right panel, you have two other examples, three typical transmissions, three typical Mendelian transmissions. The upper right pedigree, it's an autosomal recessive transmission. You have two of five children born to first cousin parents. And these two patients were born without T cells. Because they didn't have T cells, they had a variety of infections different from the infections seen in the patients without B cells. Pneumocystis is a fungus. BCG is a live vaccine for TB that can cause disease in patients without T cells. These patients do not have T cells because they are mutated in RAG1 or RAG2. Genes that are necessary for the development of T cells and also of B cells. But it's the lack of T cells that's the primary driver of these severe multiple recurrent opportunistic infections in these patients. And the third pedigree in the bottom right panel is a family in which not only two sisters, but also the father suffered from birth onward from a lack of neutrophils, granulocytes, the phagocytes in the blood, the most common phagocytes in the blood. And because of the lack of these phagocytes, these 3 patients in 2 generations develop one after another a variety of fungal and bacterial infections. So these are 3 examples of primary immunodeficiencies. That are characterized by a Mendelian pattern of inheritance, the occurrence of immunological or hematological anomalies in the patients, and the occurrence of multiple recurrent severe infectious diseases. So these data from the '50s onward, and still each week there's a new primary immunodeficiency that is discovered, These data provided compelling evidence that infectious diseases were not vaguely genetic, but that they could be genetic due to the specific absence of a particular molecule or a particular cell. These immunological and hematological phenotypes connected the genotype and the phenotype, the infectious phenotype and the mutations. So these patients with primary immunodeficiency is, however, rare. They're actually very rare. Most of you have had infectious diseases, but I would say based on the number here that probably none of you had 2, 3, or more severe infectious diseases. Patients, children with such an unusual presentation, they are probably, I would say, 1 in 20,000, but not more than 1 in 10,000 or 1 in 20,000. And if you recall what we discussed earlier, infectious diseases until the end of the 19th century killed half of the patients. Not 1 in 10,000, not 1 in 20,000, but half of them. So that clearly indicated that genes were at work and that mutations in genes could underlie infectious diseases. But the observation of these primary immunodeficiencies did not, by itself, crack the problem of infectious diseases in the general population. Of the half among you who, without vaccine, without hygiene, without antibiotics, would have died of one infectious disease or another. But these data suggested that the field was on the right track and that there was some sense to try to to discover genetic lesions underlying infectious diseases in children without immunological or hematological anomaly, at least without detectable or overt immunological phenotypes, and in children or adolescents or young adults who would not develop one infection after another, but who would suffer in their lifetime from one, from a single infectious disease. And you have here on the left the criteria of the primary immunodeficiencies as defined in the '50s. I alluded to them— multiple infections, recurrent infections. There's an immunological anomaly. The infections are likely to be opportunistic, that is, to be seen only or preferentially in patients with immunological anomalies. These infections are often rare, like the syndromes themselves. And they're familial because they're Mendelian traits. Now, the idea that we have tested over the last two decades is that children, adolescents, and young adults who suffer from a single infectious disease, whether viral, bacterial, fungal, or parasitic, a disease that does not necessarily recur, that is idiopathic in the sense that it is not associated with a detectable immunological anomaly, a disease that is not opportunistic, that is a disease known by the general public, not necessarily by highly specialized physician. It could be influenza. It could be tuberculosis. It could be diseases, infectious diseases that you've heard of. That is relatively common and that is sporadic, that is, that does not cluster in families. Well, we've tested the hypothesis that such conditions, such infectious diseases, could be genetic. And we were encouraged to do so first by all the data that I've reviewed, the evidence in plants, animals, and humans garnered between 1910 and 1950. And B, the discovery of the primary immunodeficiencies, which although very rare, clearly indicated that some infectious diseases in children with unusual phenotypes could be genetic. The third reason that suggested we were on a good track was the discovery from the '70s onwards of mutations in a pathway known as the complement. And I will not get into the details. Suffice it to say that there were in the literature case reports of individuals with mutations in this mysterious complement pathway. And some of these patients would be completely fine until the age of 5, 10, or 15 years when they would suddenly develop one type of bacterial infection. And that suggested that perhaps there was some value to test this idea at a broader scale. So we proceeded in two steps. In the first step, we analyzed what I refer to as Mendelian infections. That is pedigrees, as shown here, in which there is no immunological or hematological anomaly. These patients do not lack the antibodies or the granulocytes or anything else. They look OK, like you and me. If you analyze their blood, if you look at their lymphoid tissues, they have tonsils, they have a spleen, they have a thymus. They look OK. And they are vulnerable to a single infection that is unlike the patients with the primary immunodeficiencies that I've described before, the patients in the 3 families represented here suffer from a single type of infection. However, it runs in the family. That is, you have an X-linked recessive transmission in the bottom left. You have an autosomal recessive transmission in the upper right panel and an autosomal dominant transmission in the lower right panel. So in these 3 families, you have a Mendelian transmission of an infectious disease without an immunological anomaly. So the 2 differences with the primary immunodeficiencies that we've described before is, A, that there's no immunological anomaly, and B, that there's a single infection that runs in the family. So this is rare, but it served as a transition between the primary immunodeficiencies and the sporadic infections that we'll discuss in 10 minutes. So infections that run in families, such as these three kindreds, to my knowledge, there are only 5 of them. And they're listed here. There are 5 Mendelian infectious diseases. There are families with mycobacterial disease caused by environmental mycobacteria or the BCG vaccine or TB. There are families where you see TB in an uncle, two brothers. So mycobacterial disease can segregate like Mendelian traits. You also have families in which Epstein-Barr virus-driven these diseases segregate as Mendelian traits. For example, a B-cell lymphoma in one generation, hemophagocytosis in the next generation, and then in the third generation, you're going to have an EBV-driven agammaglobulinemia in two patients. So Epstein-Barr virus can cause a variety of diseases, but they can segregate as Mendelian traits. A third type of infection that can segregate as Mendelian trait is caused by papillomaviruses. They're the viruses that cause warts and that cause skin cancer, the non-melanoma skin cancers. Well, in some families, you have an autosomal recessive transmission of warts and non-melanoma skin cancer. The fourth infection that can segregate as a Mendelian trait is chronic mucocutaneous candidiasis. There are patients who can't control this commensal fungus. So they have oral thrush, white deposits on the tongue. They also have the fungi that gradually invade their nails or the space between the toes or the fingers. It's called intertrigo. So that also can run in families as a Mendelian trait. And the fifth infection that can be Mendelian is called invasive dermatophytic disease. You've heard of athlete's foot. It's caused by another fungus, a group of fungi called dermatophytes. So athlete's foot is benign. It's just some irritation and deposits between your toes. But in rare patients, the dermatophytes not only attack the skin, but they go through the skin. They can attack the bones. And from the bones, they can disseminate through the body, reach the liver, the spleen, and the brain. So this is invasive. And that infection can also follow a mode of inheritance that is typically autosomal recessive. So there were 5 infections. That were known from the '40s onward to segregate as Mendelian traits in otherwise healthy individuals without any immunological anomaly. So we decided to go ahead and try to test whether these seemingly genetic infections were truly genetic. And we attempted to discover the underlying genes and their mutations and the cellular and molecular basis underlying the determinism of infectious diseases. So very briefly, for the first group of infections, mycobacterial diseases, over the last 20 years we've discovered a group of about 20 genetic disorders. There are mutations in at least 10 different genes, and for several genes, different types of mutations that collectively define— as I said, about 20 genetic disorders. There is genetic heterogeneity, that is mutations in different genes, each patient being mutated in a particular gene, each family. There is also allelic heterogeneity because depending on the family, the type of mutation in any given gene may vary, and the impact of these mutations may vary depending on the type of mutation. But there is physiological and immunological homogeneity. That is, all these gene products converge for the production of gamma interferon, which is a cytokine, a molecule secreted by lymphocytes to activate macrophages, or the response to gamma interferon in macrophages and other similar types of phagocytes. So if you're born with a mutation that diminishes your production of this cytokine, gamma interferon, or if your cells do not respond well to gamma interferon, well, that's going to run in families. And the individuals carrying this mutation will be prone to develop mycobacterial disease, including in particular TB. So the study of these families added a layer of understanding for the pathogenesis of TB. You remember what we discussed this morning? I showed you that Robert Koch had discovered that Mycobacterium tuberculosis is the microbial agent of TB. I then told you that Tobal Smith I said that this was a big surprise because everybody thought that TB was genetic because it ran in families. And I also told you that the discovery that M. tuberculosis can be latent in tissues provided one of the first evidence that microbes are necessary but not sufficient for development of infectious diseases. Well, the study— of the Mendelian forms of TB and the identification of the underlying genetic lesions, the fact in particular that mutations that impair gamma interferon secretion or response to underlie TB really provided proof that TB, at least in some families, is not merely just an infectious disease, but also a genetic disorder. Now, the second type of Mendelian infection is caused by the Epstein-Barr virus. And I'm not going to go into the details because that is a group of infections that my laboratory has not studied. Suffice it to say that there are families in which disease caused by Epstein-Barr virus can kill. It can be a lymphoma. It can be what we call a macrophage activation syndrome, agammaglobulinemia, and other manifestations. But it has been shown that mutations in various genes that affect the control of EBV-infected B cells by cytotoxic T cells— when this control is disrupted by mutations, then the EBV transforms the B cells or alters the B cells, and that results in a variety of phenotypes that can kill patients. So this is a viral disease, a group of viral diseases that is not only viral but also genetic. Another type of viral disease that is also genetic is epidermiodysplasia verruciformis. These are patients who unfortunately are covered with warts. If you go to the swimming pool, you know, you can have wart. Probably many of you have had warts as children, but rarely— that is really rare— but some individuals unfortunately develop warts one after another. And warts are caused by papillomaviruses, a group of viruses that infect keratinocytes. The keratinocytes are the cells that form the epidermis. The skin external layer. And these patients develop warts again and again. They're disseminated, and in the second or third decade of life, they develop skin cancer. The warts transform and develop skin cancer, non-melanoma skin cancer. So Gerard Ott discovered the first genetic etiologies, and that is a project that we've taken over in the lab, where we've discovered new genetic etiologies of epidermiodysplasia variciformis. And the take-home message here is that there are mutations that impair the ability of keratinocytes, the cells in your skin, to control papillomaviruses. You're doing otherwise well. Your leukocytes in the blood do well. You control other viruses. Other pathogens in general well. But with mutations in these genes, your keratinocytes cannot control papillomaviruses. And as a result, you develop warts and skin cancer. So this is a third example of a Mendelian infection. The fourth example are mutations in another cytokine or in pathways that control the production of another cytokine or the response to that cytokine. It is not gamma interferon that, as we've seen, is mutated in patients with mycobacterial disease. This is interleukin-17. If you're born with a mutation that diminishes the production of IL-17 or the response to it, well then, you're going to develop your nails, your tongue, your oropharynx, your toes, your fingers, will suffer from invasion by a commensal that we all carry and that is typically innocuous, Candida albicans. And these patients develop, therefore, mucosal, mucocutaneous candidiasis. And it is Mendelian. It runs in families because the infectious disease, the clinical manifestation, follows the gene the mutation in the gene that controls IL-17 immunity. And then the fifth infection that is also Mendelian is invasive dermatophytic disease. So you have here some pictures of patients with invasive dermatophytic disease. And we've discovered that these patients are mutated in CARD9. We have no idea of the mechanism, of the immunological mechanism. This is why I'm not showing you any pictures. Graph with cells. I'm just showing you the genotype, the mutations in the gene, and the phenotype, pictures of the patients. But we do not have an understanding of the molecular and cellular basis. We don't know what connects these mutations in this gene known as CARD9 with the infectious phenotype. Now, what matters is that these 5 infectious diseases that ran in families as Mendelian traits in patients otherwise healthy, normally resistant to other microorganisms, without any detectable immunological or hematological anomaly. Well, these Mendelian infections were proven to be caused by actual mutations in specific genes. And the identification of these mutations in specific genes, with the exception of invasive dermatophytic disease, led to the identification of a mechanism of disease, of a cellular and molecular basis of disease, which— the identification of which goes well beyond these 5 rare types of infection. Let me explain to you what I mean here. If you take patients with AIDS, that is, patients who are infected with HIV, they develop mycobacterial disease, and they also develop candidiasis. These are two of the most common infectious diseases in patients with AIDS. What we knew from AIDS was that when their T cells, when their CD4 T cells would drop, that would increase the likelihood that they develop both mycobacterial disease and candidiasis. But the fine molecular and cellular basis of these two infections was elusive. What we've learned from the study of these Mendelian infections is that protective immunity against mycobacteria is controlled by gamma interferon, and protective immunity against peripheral Candida fungi is controlled by IL-17. Therefore, by inference, candidiasis in AIDS patients and mycobacterial disease in AIDS patients are caused by diminished IL-17 and diminished gamma interferon immunity respectively. Now that takes us a bit far from the genetic theory, but I wanted to illustrate and to stress the importance of these studies beyond the analysis of the genetic basis of infectious diseases. So at that stage, we had 5 infections that were Mendelian and that we showed to be— we and others showed to be genetic. But again, like with primary immunodeficiencies, these 5 Mendelian infections do not account for half of a generation to die of infection. Nevertheless, like primary immunodeficiencies, these data suggested that we were on the right track. So the fundamental problem is shown here. And I've taken here 3 infections, but I could have taken 3 others. As you know, there are hundreds of infectious diseases— viral, bacterial, fungal, or parasitic. But let's take this example. Why would a child suffer from influenza without any family history? Why would another child suffer from herpes simplex virus encephalitis? Why would a child suffer from TB? In these three families, there's no more Mendelian segregation. These are sporadic cases. The cases are sporadic. There is no immunological or hematological anomaly. There's no family history. This is the typical infectious disease that would kill until the end of the 19th century. And that still hurts not half of a generation, but still a significant number of kids in the US and even more so in the developing world. That's the fundamental problem that we've been interested in. Why would a child previously healthy in a family that has never suffered from infectious disease, why would that child end up in the intensive care unit with influenza or TB or herpes simplex encephalitis or anything else? The core, the key problem. But based on everything that I've discussed with you so far, we've decided to tackle this problem. But we haven't tackled it the way it was tackled before. I'm not going to go too much into the details of the history of genetics. But let's say that if you have children with a variety of sporadic infectious diseases, and you're interested in testing the idea that these infections are genetic, the first thing you need to come up with— if you remember my second or third slide, let's say that the problem is good and let's say that the hypothesis is good— you need to come up with an approach that is also good. And to come up with a good approach, you need to come up with a specific hypothesis that is correct. And in genetics, there have been, briefly, schematically, two schools of thought— the classical geneticist, Mendelian qualitative geneticists, and the population geneticists. And suffice it to say that the people who've tested the genetic theory of infectious diseases over the last 30, 40 years, they've searched for common variations in big populations of patients. For example, 5,000 African children with TB, and they search for common variations that would enhance the likelihood to discover— to cause TB. In short, these studies did not really work. The geneticists behind that were— had made important contributions from the '40s onward, in particular, in theoretical genetics. But this approach, to cut a long story short, essentially did not lead to the discovery of genetic variations underlying severe infections. The best paper in the field was published in 1954. That gives you an idea. It's like if the best car were still the car that was made in 1954. It would suggest that the car industry has not done too well. So when the best paper of a field is from 1954, you wonder whether the field is really that strong. So that gives you an idea briefly of the field of population genetics applied to infectious diseases. The field that conversely worked better is the field that followed the study of primary immunodeficiencies and Mendelian infections that I discussed with you earlier. What you see now are the same 3 families. The only difference is that there's a vertical bar in one healthy individual in each of these three families. Now, does somebody here— a student, not a teacher— does somebody here know what it means when you put a vertical bar in a pedigree? I mean, in a square or a circle? Yes? Is it plate carrying of that? Or not like asymptomatic? Exactly. You deserve something. Pretty good. So yes, this is someone who carries the at-risk genotype but is healthy. So the thing here is that with a pencil, just by adding these 3 bars, one in each family, you come up with a hypothesis that is plausible and testable. The hypothesis is that these sporadic common unexplained infectious diseases may be caused by single gene lesions that do not display complete penetrance. Now, who can define the term penetrance? A student again. Yes.

What if, like, if the genes in a family is the amount that a person carries it, in a sense? So, like—

You're not too far, but—

So say there's like PCOS in a family, like one— like say one sister has complete polycystic ovarian syndrome, whereas like her sister just has like irregular menstrual cycles. So it's sort of like you have like aspects of a gene but not necessarily the full— Well, this is related.

It's the notion of variable expressivity. That you're discussing. But the penetrance itself is the probability for an individual who carries the genotype at risk to be sick. So in other words, the penetrance corresponds to a calculation based on the proportion of black circles and black squares versus squares or circles that have a vertical bar. For example, here you would say it's 50%. Half of them are sick and half of them are not sick. So in that case, roughly speaking, you would say the penetrance is about half, about 50%. OK? Now the type of manifestations, that's variable expressivity. One may have a full-blown disease, the other may have a mild form of disease. So here's the answer. So penetrance is a term that was coined in 1926 by Oscar Vogt, who was a brilliant neuroanatomist working in Germany. And he said the penetrance is the probability of an individual carrying the at-risk genotype to be sick. That's the clinical penetrance. So the model that we've tested is very simple. We've said there are 5 infections that are Mendelian. That is, they are monogenic with complete penetrance. Could it be that the other infectious diseases are monogenic but with incomplete penetrance, a penetrance that could be 50% or 20% or 60%? And the model that we've proposed is shown here. It's a model in which regardless of the viral, bacterial, fungal, or parasitic cause of infection, the occurrence of a life-threatening infection in the course of primary infection, typically in childhood but also in adolescence and young adults, the determinism of severe infection is caused by a single gene lesion with incomplete penetrance. Now, if this model is correct, it has implications because we can understand infectious diseases. We can prevent them. We can offer genetic counseling, molecular diagnosis. And we can also treat patients, prevent or treat infection with a molecule that is missing. I'm going to give you an example. We recently identified in Kuwait a child who is mutated in gamma interferon. I showed you earlier that mutations that prevent the production of gamma interferon or the response to gamma interferon underlie mycobacterial disease, including TB. But of all the genes we had identified, none of them were the gamma interferon itself. Recently, we've identified two cousins in Kuwait. They have mycobacterial disease because they lack gamma interferon. They have mutations on both copies of the gamma interferon gene. So we're going to treat them with recombinant gamma interferon because it's been a drug— you know, you can buy it in the pharmacy next door. It's been on sale for 30 years. There was no reason to use it in these patients until we discovered that their mycobacterial disease was caused by mutations in the gamma interferon itself. Providing them with gamma interferon is like treating a diabetic patient with insulin. You just replace, if you like, the piece in the machine that doesn't work because of a mutation. So there are clinical implications, and there are also immunological implications. And you're interested in science, I know some of you may be interested in medicine, but your core interest in science. Well, what we can do by the identification of mutations that underlie infectious diseases in humans is with 20,000 genes, protein-coding genes, and maybe 40,000 or 50,000 isoforms— isoforms. Who knows what's an isoform? Who knows what are, let's say, two isoforms encoded by a given gene.

Yes? A form of the gene?

Say it again? A form of a gene? A form of the gene, not really. You're almost there. Does someone have a better definition of the term isoform? Yes, somebody there? I don't see. No? So let me help you. A gene comprises a variety of exons. And depending on which exons assemble to form a messenger RNA, a given gene may encode for different proteins. So one gene may code for two or more proteins. There are genes that code, for example, for 10 or 12 proteins. OK? So there are isoforms. So with 20,000 genes, we do more than 20,000 proteins. We do at least 40,000 or 60,000 proteins. So if infectious diseases are genetic as a rule, or if at least a sizable proportion of infectious diseases are genetic, caused by these single gene lesions that do not display complete penetrance, well then with 7 billion people, having babies every 25 or 30 years, given the mutation rate in the germline, which is pretty high, you know, 10 to the minus 8, 10 to the minus 9, well then we can hope in 20 or 30 years to identify mutations in most, if not all, genes that control host defense in natural conditions. And therefore, you know, there are immunological scientific implications of that work, if you like. Which brings me to a book that I encourage you to read, at least those of you who are really passionate about science. It's called Pasteur's Quadrant by Donald Stokes. It was published 20 years ago. And it's an interesting essay about the history and philosophy of science. And there's a beautiful figure in that paper that I've reproduced here. He classifies science depending on whether the research is inspired by considerations of use— medical use, for example— and a quest for fundamental understanding. Of course, if there is no consideration of use and no quest for fundamental understanding, there is no science. So that's why this square is empty in the lower left square. Now, there are scientists who pursue exclusively fundamental questions. And the example he takes here of pure basic research is Niels Bohr. There are also scientists who pursue— who do pure applied research, such as Edison, your compatriot. But he takes Pasteur as the prototypical example of a research that is use-inspired and basic at the same time. That is, the implications are clinical, medical, but also the implications are biological and scientific. And I like to think that the genetic dissection, the human genetic of infectious diseases, is both use-inspired and a quest for fundamental understanding. So with this idea in mind, we've discovered over the last 20 years mutations underlying a variety of sporadic infections shown here, including severe influenza, herpes simplex, and syphilis, and TB, the three examples that we took. And for TB, we've discovered mutations in the same genes that I alluded to and other genes. Again, these mutations underlie the circuits or disrupt the circuits that normally produce gamma interferon or respond to gamma interferon. What led us to the discovery of TB in some of these patients is that there is incomplete penetrance for some of these disorders for infectious diseases caused by poorly virulent mycobacteria. And that led us to the identification of mutations in patients and families with TB as the sole phenotype. We've also discovered mutations causing invasive pneumococcal disease. The picture you see here is taken from the cerebrospinal fluid. When you do a spinal tap, when you put a needle in the back of the spine in a child or an adult, you get some liquid, the liquid that surrounds the brain, the meninges. And if you have a meningitis, well, you have leukocytes as shown here. They shouldn't be here. They're the purple cells. And the little blue dots, they're the cocci. These are pneumococci that shouldn't be there either. So this is taken from a child with a pneumococcal meningitis in whom we've discovered mutations. We've discovered a variety of mutations, mutations that impair the development of the spleen and that makes these patients vulnerable to pneumococcus, also mutations in pathways called TLRs and IL-1 receptors. I'm not going to get into the details. But if you have inborn errors in this pathway, you're prone to develop pneumococcal disease. This is illustrated here. These are the mutations in the pathway in gray. But then the mutations in the pathway in green here that are related underlie a completely different infectious disease, which is herpes simplex encephalitis, shown here. What we've shown over the last 15 years is that herpes simplex virus, which is a benign virus— I bet given your age that probably 70%, 80% of you have already been infected by herpes simplex virus 1. And the only thing it does once you're infected, it establishes latency here in your trigeminal ganglion. And from there, it's going to reactivate from time to time. You're going to have a blister, herpes labialis, blister on the lips upon sun exposure or if you're stressed to come to Rockefeller, that sort of things. Then you get a blister. And that's OK. Then the virus reestablishes latency and sleeps for a while. And a few months later, it's going to pop up again. That's the classic life cycle of herpes simplex virus 1. However, unfortunately, in about 1 in 10,000 children, the virus doesn't stop here. It goes from the nostrils or the mouth directly to the brain. And it causes a terrible, dreadful encephalitis that used to kill until acyclovir was discovered. And with the advent of acyclovir 25 years ago, the children, if they're treated in time, they survive. But they suffer from terrible neurological sequelae, cognitive impairment, recurrent seizure. It's a terrible infectious disease. And what we've discovered is that this disease, which strikes once, strikes otherwise healthy children, is relatively common, is not opportunistic. These patients do not have detectable immunological or hematological anomalies. It's sporadic in nearly all cases. Well, we've discovered that this infectious disease is not only viral, it results from single gene inborn errors in a pathway that controls the production of antiviral molecules in the brain. And if you're born with a mutation in this pathway, you develop encephalitis. I see a question here. So yes, I mean, that can be one of the sequelae when you can have cognitive impairment and poor memory. Absolutely. So this is a viral disease that is also genetic. So you see mutations in related pathways can underlie different infectious diseases. Infectious diseases, which brings me to this idea. If you recall what I told you earlier, Archibald Garrod had proposed following his discovery of inborn errors of metabolism that infectious diseases could be caused by inborn errors of immunity. And if you compare neophenylketonuria, which is a typical inborn error of metabolism, with herpes simplex encephalitis, you see that in both cases, you have a disease of the brain. In both cases, there is a genetic lesion. There's a mutation in a gene. And in both cases, the development of disease, its occurrence, depends on the exposure to an environmental trigger— the virus, herpes simplex virus 1 in the case of herpes simplex encephalitis, and phenylalanine in the case of phenylketonuria. The reason why whenever you buy food in the grocery store or candies or anything, it gives you the content of phenylalanine is because people with phenylketonuria, if they don't eat phenylalanine, they're going to be OK. But if they eat phenylalanine, they are at risk to develop brain disease. So phenylketonuria and herpes simplex encephalitis, in a way, are both genetic disorders and disorders, the clinical manifestations of which are triggered by an environmental stimulus— phenylalanine in one case and herpes simplex virus in another. I see that time is running, and I'm going to close this talk with influenza virus because the first virus, the first evidence that influenza was viral was discovered here at Rockefeller by Richard Shope. And he published his paper in the early '30s. It was a swine influenza that was followed a few years later by the discovery in Britain of the first influenza viruses infecting humans. And from that period onward, all we know about influenza is that it is caused by influenza virus. And most people infected by influenza virus are OK. A little bit of fever, but they're fine. Some are going to spend 3, 4 days in bed. And rare patients infected by influenza are going to be admitted to the emergency room and then the intensive care unit. And some of them will die. It is 1 in 10,000, 1 in 100,000, depending on the virus. Some influenza viruses are more virulent than others. You hear about avian flu or swine flu. Yes, these pandemic viruses are more pathogenic in humans than the seasonal flu. But regardless whether avian flu, swine flu, or seasonal flu, it's only a relatively small proportion of infected individuals who will end up in the intensive care unit intubated and ventilated. So the question in the field, like for other infectious diseases, is why would influenza virus kill some teenagers and not others? So very little was known, in fact nothing. And we tested the hypothesis because the known primary immunodeficiencies, none of them underlie severe flu. We tested the hypothesis that severe flu might be caused by mutations in completely novel genes not mutated or not related to genes mutated in primary immunodeficiencies. We enrolled children with severe flu. We sequenced their genome. And we discovered mutations first in a patient from Western France, from Angers. That's the chest X-ray of that particular patient. And she was found to carry two mutations, one inherited from the mother, another inherited from the father. It's an autosomal recessive transmission, so far apparently with complete penetrance, but we don't know. There's only one child. And this patient developed life-threatening flu, not only because she was infected by the flu virus, but also more importantly, primarily because she has RF7 deficiency. It's a genetic disorder that we've shown prevents them from making interferon in the lungs where influenza virus first invades the human body. So I hope I've taken you from 1800 to RF7 through a long journey, and I hope you'll remember that infectious diseases are infectious, evidently, but that for some of them They're also genetic disorders. I want to— before you go, you're going to wait a few more seconds because, A, we have questions for you. Things will come. Don't run away. You see, I try not to make you suffer too much. But I want to thank the people in the lab. That's a picture of my lab. Yeah, that's a picture of my lab. I took them hiking. That picture was taken in the east of France at the Mont-Dore after I took them to visit their hometown and actually the house in which Pasteur was born and also the house in which Claude Bernard was born. Both houses were transformed into museums. I took my lab there, and then I took them for a hike. Thank you for your attention.

So at the beginning, you showed the data that showed that humans, about half of them die in infancy or before adulthood. Is it the same in other animals in the wild? That's a very good question.

That's okay? So that's a very good question. You know, what's the mortality rate of infection in other species? The answer, I don't know. It's a question that I've asked myself often. There's very little data. We know much more about humans in natural conditions than other species for obvious reasons. So, OK, I'm going to bring the box here. So, yeah, you handle that. That's right. She handled the— that's a great question too. So yes, can we fix the mutations with CRISPR-Cas9? We cannot do it now for technical ethical reasons, but is that going to happen in your lifetime, for example, while you're in college or grad school? Yes. That's coming soon. So I've heard in non-academic circles that the susceptibility of patients to severe sickness after they get the flu may have to do with whether they're very old or very young. Is that known, or how much has to do with genetics versus just just how feeble the patient is? Well, that brings us to one of the slides in which I discussed the 4 theories. So in a given age, there is inter-individual variability. So yes, the age has an impact. For example, herpes simplex encephalitis is a condition of young children. But in a given age, there is variability. So for flu, what we know is that it strikes preferentially people, whether young or adults, who have a preexisting pulmonary comorbidity. That is known. But the other risk factors for any group age were unknown. And that's in that context that we discovered RF7 deficiency.

Hi. So you said that getting appendices out is like passed on, right?

Yeah? That appendicitis?

Appendicitis, yeah, yeah, that, right? So is that— can that be— can that happen with gallbladders?

Can you what?

Can that happen with gallbladders?

With the gallbladder? Oh, cholecystitis, that's what you're saying. Is that— So that's another good example, but it's more a disease of people in their 40s or 50s, and it's more rare. Cholecystitis is, I would say, no more than 1 in 10,000 people, whereas appendicitis is more common. So it's easier to study its segregation. But it's a good question. It's also an infection that's not contagious, absolutely. Is atherosclerosis Is this a disease that people are genetically predisposed to? And if so, what is its inheritance pattern? So atherosclerosis is in part genetic, but it's not infectious, and we're not going to discuss it today. So the whole point of the lecture was, yeah, diseases can be— diseases and infections can be hereditary, but can the opposite be true, that despite not being passed down in an extreme environment, can you also contract diseases and infections? As I say, you know, English is not my native language. You have to be indulgent here. What are you exactly trying to say here? So you mentioned that there are rare cases where there's no history of a disease or an infection And people still contract certain diseases and infections. So in extreme environments, is that more likely? Do you understand what he's saying?

I think he's asking— the best way to get around it is he's asking if extreme environments can cause infection in extreme situations.

Are you saying that could there be infections that are not influenced by the genetic background? Yeah. You know, the fact is no, because the most virulent pathogens— there are examples that I did not discuss of resistant individuals. For example, even HIV. There are people who have 5, 10 unprotected relationships per day. And they do not get infected because they are mutated in CCR5. You said in the beginning that every living organism evolves, right? So I'm assuming that bacteria evolves too, or the pathogens. So what is stopping the pathogens of smallpox to coming back? All right. Now let me give you a trick. I'm going to tell you something that I tell my students every Monday, every Friday, we have a lab meeting. We're 25 in the lab. And what happens now reminds me of the lab meeting. And you know, they're big boys. They're graduate students and postdocs. And it's a small room. And nobody understands what they're talking about. So because they don't speak loudly enough. So you have the microphone. Please, what you're going to do is you're going to speak loudly so that so that we can all understand what you say. Speak to the microphone, don't be shy. Ask your question so that everybody can hear. Go ahead, repeat it. So you said in the beginning that every living organism evolved, right? Right. So I'm assuming that pathogens evolved too, right? Right. So for example, let's say the bacteria or the pathogen of smallpox which killed a lot of people early in history disappeared because we had vaccine. So assuming they evolved, what is stopping them from killing a lot of people in this time? Oh, it may come back in a different form. This is absolutely possible. There are emerging and re-emerging microbes, absolutely. And by the way, do you see when people speak loudly, you can hear questions much better. Good question. How long could an infectious virus stay in someone's gene strand in their— like, how long can an infectious strand stay in someone's hereditary genes? How long could it last before— For your lifetime. You can be infected with a virus at birth and keep it until the age of 90. So my great-grandmother could have an infection disease and it wouldn't come into effect until my third son? Exactly. Absolutely. Okay. All right. Thank you. Thank you. Okay. I think here they need the microphone here. There's a cluster. Bring it here.

You know, there's a cluster of questions I kind of have the mic in my hand already, so like—

Now speak to the microphone, please.

Can you please explain what are the first steps in creating a vaccine? The first? Like, what are the first steps you would take in creating a vaccine for a disease?

The first steps that people take to create a vaccine? Okay, well, first you need to know the microbe that causes disease. Then you have two options, two types of vaccines. You have a first possibility which consists in attenuating that particular microbe. That is, you generate a microbe that is a cousin of the causal microbe and that has lost its virulence but that has retained its immunogenicity. That's one possibility. The other possibility is you simply kill that microbe or synthesize part of that microbe and inoculate it. These are the two types of microbes— of vaccines.

So before, you were talking about the twin studies and everything. And I was thinking about nurture versus nature and how genes are expressed through different environments and stuff like that. So in this study, you were saying that a lot of— well, all of the people lived in the living in the same house or apartment or whatever the case may be. So like if this study was to be done with twins, or like identical twins or fraternal twins that lived in different houses, would that change this study completely or would it just be a different study?

No, prerequisite for the twin studies is that the twins live in the same household. You know, if it were not the case,— it would be impossible to conduct the twin study. So it's a prerequisite.

Thank you. First of all, good presentation, Dr. Kosanova. I liked it.

OK. What did she say? She said she liked your presentation.

Really? Thank you. I've been thinking about this question since the break for lunch. This topic was introduced before we went on break. You mentioned that certain— antibodies are made after acquiring an infection. And then sometimes these infection agents can kind of serve as a vaccine for other diseases. You mentioned that, correct? Right. And then I got to thinking, would an example be Edward Jenner with his discovery of the vaccine of smallpox for cowpox?

Brilliant. Excellent. Brilliant. Thank you.

Absolutely. That's an example?

Yeah, yeah, yeah, yeah. She deserves something.

So you were talking about the genetic theory of infectious diseases, how we can build an immunity up by one of the four ways. Is it possible to build an immunity from two or more ways? Like you can have the genetic factors plus the ecological factors to build an immunity?

Absolutely. I should have stressed that these four theories Theories are not mutually exclusive. Absolutely. I completely agree. Good point. They're good. They're good. Next. Yeah, circulate the microphone.

Go ahead. So I remember from my bio class that certain genes code for proteins that can prevent certain symptoms from from showing up. And I was wondering, can genes exist within the same family that provide immunity from symptoms of infections that are also inherited?

At one stage, you moved your microphone away from your mouth, and I missed a word. Are you asking if some mutations Prevent infection? Yeah. Yeah, well, as I said earlier to one of your colleagues, there are mutations in genes that prevent infection with HIV. There are mutations in genes that prevent infection with norovirus, which is a cause of diarrhea, and also Plasmodium vivax, which is a cause of malaria.

But I was wondering, could those genes exist within the same family?

Well, the genes— we all have the same genes. The mutations run in families, yes. Okay. Okay. More, yes, circulate, circulate the microphone.

Hi. Stand, stand. Okay. So the more common diseases that like infect the human population, are they because of like sharing drinks and stuff like that, or they're more just because they're like humans are easily susceptible? Susceptible. To them? Did you understand that question? Oh yeah, so like herpes simplex 1, is that because everybody shares drinks or is it just because it's kind of an easy disease to catch?

No, the virus, the herpes simplex virus 1 is transmitted when you shake hands or when you kiss on the cheeks. So that's the transmission of the virus. But the development of the encephalitis occurs in individuals with mutations. And the mutations are transmitted in families because kids, you know, are born to parents.

My question was, you said that the— oh, stop. The diseases that are hereditary that you were talking about, that I was wondering, is there a higher risk of contracting these diseases in a family in an urbanized area or rural area? Where you can't—

where you aren't as exposed to pathogens and other people. Right. So the area in which you live has an impact on the microbes you're going to encounter, but it does not have an impact on the genes that you will inherit and the mutations in these genes. OK?

So if the same disease is affecting a rural and urban area, How would the— what's the risk? Is there a higher risk of the urban or rural area contracting the disease?

Well, it depends on the infection. For example, TB is more likely to be caught in big cities where people, you know, get a ride on the bus or in the subway. And, you know, a person with TB can cough and transmit the virus, the bacterium, because because there's contagion. Conversely, other infections are more common in rural areas because you can catch them by, in a farm, you know, playing with animals. So it depends on the infection. Some infections are more common in rural areas and others in big cities.

All right. Thank you.

On your left. On the left. So I was just wondering the difference between the immunological phenotypes of a primary immunodeficiency versus a single gene inborn error, is that just that primary immunodeficiencies are less specific to diseases? Exactly. So in primary immunodeficiencies, the anomaly is so profound and so deep that there's vulnerability to a bunch of infections. Exactly. Thank you. Well, we alluded to that earlier. One of your colleagues spoke of the CRISPR-Cas9 dependent cure in these patients.

That would, in essence, be gene therapy, exactly. Hi. I was wondering, what was the defining moment where you knew you wanted to study infections?

When I was— that's a great question. When I was a resident in pediatrics. Good question. Does anyone here want to become a pediatrician? Oh, good. Very good. Well, then you have to come and see me. I'll give you advice. OK? It's the best job. That's the great thing. OK. Yeah, go ahead. We're out of gifts now. OK. Go ahead. Are these genes that increase susceptibility to infectious disease equally spread out on all the chromosomes, or are they concentrated on any particular ones? That's right. There's no particular clustering on a given chromosome, no. It's throughout the genome. If someone has a microphone, go ahead. Ask a question. Shoot.

In the recent breakthroughs in the CRISPR-Cas9 system, what are some of the holdbacks, and where do you see it in the future?

What are the drawbacks? Yeah. Frankly, I see mostly advantages. I don't see any potential problem here. Problems could be ethical, you know, if someone in his kitchen would start doing genetic engineering.

But otherwise, I see only advantages of this. Hi. OK. So I'm a twin. So you said that recently, that you said that identical twins have the same germline. So if a twin that lived elsewhere, like say my brother, lived not with me and he contracted tuberculosis, would I have a much higher chance, or would I have the same chance of getting that disease if he were to live with me every day?

If you're identical twins? Yes. Yes, if you live together, you would have a high likelihood to contract the disease. But if he— But if he lives elsewhere, no, because you need the infectious agent. During your, during your talk, you briefly touched on bacterial antibiotic resistance. So I kind of clicked in my mind a few weeks ago, I watched this short video about viral therapy since antibiotics becoming slowly obsolete because bacteria is becoming more and more resistant. What is your thoughts on using a virus to kill bacteria? I mean, viral, do you think it's gonna go anywhere? Do you think we're to be developing more antibiotics until— I don't know. I'm aware of this line of research. I don't know. Time will tell. It's always interesting to explore.

Hello? Yeah. Okay. So I have a question. As you described before that a virus can cause a loss of memory, is it a partial loss of memory or a total? And if it is, then how long is it for?

Lifetime, or is it for like a temporal loss? Well, it depends on the virus. It depends on what kind of virus caused encephalitis. It depends on the patient. It depends on how quickly the diagnosis was made and the treatment given. So there's no uniform answer to your question. Thank you. You're welcome. You mentioned also the whole thing about isoforms and how there can be multiple genes on the same section of a strand of DNA.

So how is it that—

or what is it that dictates which gene is going to be expressed if it's the same in order in the genome? Well, that's another good question. But this is controlled by what is called regulatory regions. These are the regions that determine the spatial and temporal expression of genes.

So you mentioned that as humans evolve, microbes evolve. And vice versa. So I was wondering, because this is a common occurrence with the flu, how can scientists stay updated with how fast it adapts?

How fast the virus adapts? Yeah. Well, because when there's an outbreak of flu, the virus is being analyzed, sequenced. OK. Hello? Hello? Yeah, go ahead. You first.

Yeah, you have to come. When a disease is asymptomatic, is the immune system constantly fighting it, or is it just like dormant and has no effect?

That's a great question, and we don't have the answer.

Hey, um, so like just in the case of like sickle cell disease, like, um, giving an immunity to malaria, and that's like genetically linked, like how exactly is that possible? Like are the genes like on the same chromosome, like closely linked, like how exactly?

Right, there may be mutations that underlie different infectious diseases, and also there may be mutations that are linked with other mutations, that is the two mutations underlying two different infections, that's possible, absolutely. Okay, thank you. Yeah, go ahead. I don't— oh, it's mic. So in cases of herpetic encephalitis, can it be caused by other strains other than just HSV-1, or is that exclusive? No, it's for herpes simplex, it's herpes simplex 1 for the childhood disease. There's another form which is the neonatal disease that's caused by herpes simplex 2. But it's a different type of disease.

OK. So can bacterial infections act as epimutagens? And if so, can those epimutagens affect the susceptibility to future bacterial infections?

There's no data suggesting that Candida infection can facilitate a superinfection.

My question also has to do with HSE. What are the consequences of herpes simplex— I forgot the last word.

What are the side effects or symptoms? The symptoms of herpes simplex encephalitis is fever, loss of consciousness, and seizure.

Go ahead. It's OK.

Go ahead.

You talked about identical twins and fraternal twins, but what about Siamese twins? Is it— The what? Siamese twins?

Conjoined twins. Conjoined twins. Oh. Well, that's unrelated. I mean, the question is whether they're identical or fraternal. That's going to make a difference to study the nature-nurture dichotomy that your colleague alluded to. I mean, whether they're connected or not, it doesn't have an impact. What matters is their genetic background.

Thank you. Hi, Dr. Casanova. During the first part of the—

Can you raise your hand? Oh, yeah, you're here.

Right here. The first section, there was a pedigree trace chart that actually showed— I don't remember who it was, but they had 5 children. 3 out of the children died from fever. But they were all female. So I was wondering if this infectious disease is sex— can it be sex-linked?

That's another great question. You guys are all asking good questions. So yeah, the X-linked recessive disorders affect boys only. I'm not aware of genetic disorders underlying infectious diseases in females. But it's not because I'm not aware of it, it has not been discovered that someday somebody is not going to discover it. That's an interesting idea. Thank you. Go ahead. Speak to the microphone. Closer. Since each living organism is unique, do they produce their own individual antibodies like for the same disease, would they differ? And if so, would there be different degrees of strength for antibodies depending on the— Very good. Very good question. So yes, each individual makes his or her antibodies. And if you compare, for example, all your antibodies against flu, there will be some public structures, that is structures shared by a number of you, and private structures, that is structures of antibodies that are unique to each of you.

Absolutely. I have a question about one of your earlier cases. You had a case with a woman named Martin, and then you said that it was in, I believe it was like the 1980s or something. And it was a woman that came in and she had skin lesions that were down to the bone and a lot of different viral-type symptoms, what would be the underlying reason for you to— I'm guessing the same. What would be the reason to think towards a hereditary reason for the disease being so severe instead of a pathogen making it so difficult?

Well, it's not or. And, you know, I don't want to give you the false impression that I've told you that infectious diseases are not infectious. They're infectious and genetic. So it's a balance, you know. It's a conflict between two organisms, a small one and a big one. It's a conflict between two genomes, the microbial genome and the host genome. OK, so the outcome of this conflict is determined by the respective virulence of the bug and vulnerability of the host. All right, no more questions? Thank you.