Plagues, populations & survival by Stephen J O'Brien

so good evening and welcome to the fifth of the 2014 Darwin College lecture series which is as you know is on the broad theme of Clare plagues and just in case you've forgotten here it is behind us and welcome to all of you here in the lady Mitchell hall and to all the people who watch these lectures later on because they're freely available on the web and as most of you know this is very important year for Darwin College as we're celebrating the fiftieth anniversary of the foundation of the college as the first exclusive College for graduate students in the University of Cambridge and this is the first mixed college so from a very small beginning we're now among wunst of the one of the largest colleges with alumni encompassing every subject and spread around the world so this evening I'm very pleased to welcome professor Stephen O'Brien Stephen is a molecular biologist with very wide research interests including a plague he uses genome sequencing to understand diseases others who are hiv/aids as well as to inform the conservation of endangered species his team identified the first restriction gene that imparts immunity to HIV he's worked on the genomes of many species including cheetahs and red pandas and and on and on he's a founder of the genome 10k project that is assembling a genomic zoo the genomes of 10,000 vertebrate species he's had a four decades long career at the National Cancer Institute in the US during which he served for 25 years as the founder and chief of the genomic diversity lab then two years ago he crossed to the side of the Atlantic to join us at Petersburg State University where he is the chief scientific officer at the Center for genome bioinformatics so it gives me great pleasure to welcome Stephen O'Brien to speak on plagues population and survival [Applause] [Music] good evening is the microphone on okay good it's a special pleasure to come here to Cambridge and Darwin College in order to talk to you about a area that I think also think is very fascinating the subject of the series which are plagues plagues pestilence disease mortality horror we've all heard and read imagined some have experienced them but today science is beginning to weigh in a little bit on some of the details however in one man or woman's life we're not so concerned day by day because it doesn't happen very often in fact one observer mentioned to me that world pandemic seemed to occur about once a generation every 30 years or so as is illustrated by the slide here Spanish flu in 1918 polio in 1950 hiv/aids 1982 SARS 2003 there were others but none as horrific as these and during that time it has been really interesting to look back and look forward but with the new tools that allow us to not only explore questions unanswerable in the past but also allow us to make some recommendations about and observations about the disease plagues which have afflicted humankind one thing we have discovered is that there's a delicate balance between the host and their pathogens which is a Biddy akin to a deadly arms race waged fiercely almost daily and multiplied over individuals populations across geography and even among the 4,000 mammal species that survive on earth today that struggle is frequently lost it's no really one simply temporarily circled with the survivors rising once again to compete for another day I'm old enough to recall back in the late 60s when the US Surgeon General was a man named William Stewart and he pronounced proudly that end to the horrors of infectious disease he was encouraged by the success of antibiotics vaccines against smallpox measles mumps and polio he predicted a swish yet shift and biomedical emphasis from infectious diseases to chronic diseases that he didn't really know how to cure then came AIDS hepatitis B hepatitis C papilloma hantavirus SARS West Nile mad cow disease modern medicine is far from conquered these infections in fact I don't think we're even close I'm going to tell you three stories here today to illustrate how scientists including members of our team have learned from taking a population evolutionary and molecular approach to understanding diseases I'm going to start about 12 years ago when a severe flu-like outbreak began to appear in the emergency rooms of hospital around Guangzhou none in Hong Kong a devastating syndrome that defied any and all treatments it was a new to medicine to science to people and it was called the severe acute respiratory syndrome or SARS within nine months it traversed to 29 countries and ected over 8,000 people and killed almost 800 the alarming speed and virulence of transmission was dazzling and was illustrated by a couple of examples in one of the hospitals in Hong Kong a patient arrived with this disease which no one had ever seen before and within a few days 112 people who had come in contact with him had come down with same disease there was an apartment complex in Hong Kong called the Mori gardens a young woman who contracted SARS went to her apartment and stayed there but the virus spread through the ventilation system in women in a few weeks three hundred and twenty-one people were infected residents who never had any contact with his patient in March 2003 several groups who were studying this disease which was terrifying and changing the culture in southern China and in northern China worked independently to describe the cause of this disease it was a virus one that was familiar to veterinary practitioners but not so much to humans it was in a group called corona viruses this is an evolutionary tree which shows the genetic distance between some of the corona viruses that we know about about one third common colds are caused by corona virus this is one of the corona viruses here and this is one that causes human diseases but these over here cause disease and chickens and in turkeys and this is a mouse virus this is a domestic cat and dog virus was listing over here and this is a porcine or pig virus the SARS virus was in this group but it's sat out here in a limb as if it was really a new isolate that hadn't been seen before now initially there was a rush to find out where this virus came from the sequence of the virus was determined which led to the tree you're looking at right here and a number of specimens were looked forward to me reservoirs one of the ones that was identified was farmed palm civets a relative of the cat which were there were hundreds of thousands of them in captivity in China that were basically being used in the in the as a farm market for food so the government of China which was a little very uncomfortable with this one of the things they did was the order of the extirpation of about 10,000 of these these palm civets however the palm civet was only the unlikely passenger a few years later it was the horseshoe bat that was shown to be the primary reservoir from which this came from in June of 2004 the epidemic stopped stopped it subsided in May excuse me of 2003 presumably consequence of a draconian quarantine measures the epidemic scared us by its ferb fervent economic catastrophe which has been estimated in the order of tens of billions of dollars now ten or twelve years later we still don't have a really clear understanding of the precise mode of transmission we have no laboratory based clinical diagnosis other than PCR we have no vaccine and there's really no treatment the mortality of this virus was about 10% an emerging disease that spread at rather lightning field but it stopped as I said in May of 2003 and really hasn't surfaced so much except for a small outbreak in in the in in the Middle East which is going on right now now it did change the culture in China where people would wear wear masks there were no handshakes kisses or any other kinds of unmentionable contact because of the fear of what this was going on now as I said it was familiar to the veterinary profession because many of these viruses are known to cause disease in cats and I remembered when I began reading about this an effect that very few other people did which was an epidemic of a similar corner of coronavirus which took place back in the early 1980s and it occurred in a wildlife park in Oregon and inflicted this species the African cheetah at that time we had begun our interest in wildlife species particularly trying to find out the consequences of of historical events on their fitness and their adaptation in their breeding in captivity and we had discovered that the cheetah was remarkable among other cat species principally because it had a highly reduced level of genetic diversity relative to other cats it was an outlier of several thousand species that had been looked at with a series of different measurements that quantified the amount of overall variation if each of you looks at the person next to you there's 10 million DNA differences between you and that person cheetahs have fewer than a few hundred thousand between each animal a hundred times to a thousand times less and this was validated in a number of different methodologies which are available across the years because it was so surprising and it was explaining in some ways the difficulty that zoo managers were having in breeding cheetahs in captivity while at the same time they were very easy to breed other cats like lions and tigers and many other species one correlate of inbreeding in this particular cheetah population was that it's sperm morphology had a lot of problems this is a series of sperm which are malformed found in cheetahs three heads two heads bent tails droplets large heads tiny heads this is a very rare chain of sperm about 10 percent of cheetah sperm are normal the rest 90% are abnormal and the reason for the Cheetahs genetic uniformity and the consequences were imputed by other methodologies to trace back to a event that occurred in North America about ten thousand years ago when the glaciers retreated for the last time during that evolutionary epic called the Late Pleistocene something like 40 species of large mammals disappeared as the glaciers retreated for the last time these included the mastodons and mammoths the giant Seibert saber-toothed cats the giant ground sloths the direwolf several predatory Birds and the American cheetah which is actually evolved in North America as a sort of a sister species with Poulin sand so forth that particular event X all these large mammals we don't know really why but they all went away and Pumas even went away from North America to be refounded by migration from South America but that event was kind of interesting and it caught the conservation community by surprise because it meant that endangered species which have dropped to low numbers can contain a hidden hidden curse or ax hanging over their head and the cheetah was a good example and that could display itself and it adds it has been shown in many other species in different ways including congenital abnormalities that are elevated including reproductive at abnormalities such as a cheetah displayed but one of the things that was kind of interesting was that the immune system was also homogenized and this led us to worry and speculate that perhaps the Cheetahs genetic situation would make it sort of more vulnerable for any infectious disease that might drop into the species because genetic diversity itself among the immune system is basically something that is adaptive and keeps individual populations alive when pathogens overcome the defenses of the first individual that sort of the traditional wisdom as it were well no sooner had we speculated it when we saw a realization of this horrific prediction at a wildlife park which up at that time in the early 1980s was perhaps the most successful cheetah breeding organization in North America they had about 60 cheetahs and they were producing a number of different individuals a couple of cheetahs were imported from the Sacramento Zoo their names were Tom ooh and Sabah and those two became patient zero in an epidemic what happened to Tama and Sabu is they came to the zoo or to the Wildlife Safari Park they were isolated in quarantine as was the the method at that town but they had a fever and they had little twitches and they were looking a little bit jaundice and eventually in spite of aggressive symptomatic therapy by the veterinary clinicians on-site at the place the animals would collapse dehydrated and pass away at the necropsy which is a veterinary term for autopsy their bellies were opened up and it was filled with this milky proteinaceous fluid and this was something that the clinicians recognized because they'd seen it before in other cats particularly domestic cats as a symptom of a disease called feline infectious peritonitis feline infectious peritonitis is caused by a corona virus the first cousin of the of the SARS virus but distant enough and what it does is it generates a hyper immune reaction which accumulates immunoglobulin complexes in the peritoneum this milky fluid and strangulate s-- the tissues and the animals die the clinicians and the managers of this park were rather upset because they were worried that the corona virus might the feline infectious peritonitis virus might spread to other cheetahs and of course it did now because of the fact that banked serum samples were held by the clinicians and the veterinarians it was possible to track the appearance of antibodies against the standard FIP virus by simply measuring individuals over a period of time before the arrival of patient zero these five animals in this family were negative but within six months of the arrival of the patient zeros and their death they all developed antibodies and five of the six of these animals were dead within a year of this disease this family group was not the only individuals in the wildlife safari that succumbed to this disease in fact they all did the next slide is a blow-up of a number of different slides like this which were put together and published by Jonathan Heaney when he was a postdoc in our laboratory 20 years ago and this particular slide illustrates that for each cheetah in the park before 1982 they were all negative but within the next year every animal in ceará converted and developed antibodies because the virus was so infected as a threat that as it is excreted in the and in in the feces and it is because the hygiene in these places cannot be perfect it spread to all of them rather quickly through the food and through the hands and to the keepers the arrows indicate the deaths of these animals as you can see nearly all of them died about within three years there was a hundred percent morbidity which means a hundred percent symptoms there wasn't a normal stool in the park they all had diarrhea they all had the jitters there was jaundice everywhere and most of the animals like sixty percent of them would die within three years of the original outbreak this was a horrifying example of the homogeneity of this particular result of this of this individual and what we what we concluded was that perhaps that the animals had gotten a virus which had gotten into the inn and into the Cheetahs and it spread through it quickly perhaps because of the genetic uniformity of the immune system of these animals well when the SARS epidemic took place we had in our freezers tissues from the cheetah epidemic which would took place in 1980s now for those scientists in the audience you'll know that in the 1980s there was no PCR there was no sequencing we didn't know very much about these viruses but when the SARS epidemic took place back in 2003 we were more capable so what happened was some of the youngsters in the laboratory took a look at the sequence of the SARS virus and the corona viruses and they develop primers to go back into the Cheetahs to see first of all are they related to the SARS virus and second of all what is the situation with respect to their getting into the epidemic and so we wondered if we looked at the sequence of these individuals of the Cheetahs where would find out well mercifully a Ju which is a Latin abbreviation for a cheetah came out on the limb very close to the domestic cat version of FIP now what I should mention to you is that FIP and domestic cats is a the disease that pretty much had told us the story I just told you however one thing is important and that is that the incidence of mortality and domestic cats even an outbreak is seldom over 5% and that more morbidity is on the order of 10 percent that is symptoms 90% of them out don't get sick so the truth is that we were surprised at the cheetah outbreak seeing being so uniformed furthermore when we examine a series of different isolates of the cheetah virus it basically and compared it to domestic cat versions they were nested within the genetic cluster in terms of their genetic identity a long way away from the canine virus or the or the turkey virus and so forth they weren't like the SARS virus percolating out somewhere this was a B flat normal house cat virus was jumped into cheetahs furthermore we actually didn't experiment with these animals with it with these viruses we had a few house cats that were naive and we isolated virus from the cheetahs and we put them into tent house cats to see if they would develop disease none of them did and this is kind of what we expected for a normal house cat virus because remember I said that normally in house cats the mortality is less than 10 percent so if you go back then let's talk about what I've just said the mortality differs Aladin cheetahs the Cheetahs had between 60% mortality a hundred percent morbidity House cats was one to two percent mortality the SARS as bad as it was is five ten percent and the reason that we believe this is so is that the genetics of the cheetah was identical one of the other pieces of evidence that we had to reaffirm this this conclusion was there were ten Lions at wildlife sapphire they all became infected they all cero converted they all develop antibodies none of them got sick so it wasn't just a virulent virus which was hopping into all these species it was a normal house cat virus but it overcame the genetics of the first individual of the cheetah it looked around they were all the genetic or immunological clothes and it went through it like a train crew station our conclusion and the sobering conclusion was that the Cheetahs outbreak was excessive and horrible because the genetic diversity had been depleted by that long time ago situation in the Late Pleistocene it was a sobering example in people in breeding breeding cheetahs then immediately started to screen for this virus every time because the cheetahs have this hyper susceptibility my second story has to do with an outbreak and an epidemic that really began in 1982 or so it began earlier but we didn't know about it until the early 1980s when a series of actively gay men came down with a rare pneumonia and a rare cancer Kaposi sarcoma that immediately was shown to be the disease we now know as AIDS within a few years later it was discovered that HIV was the virus that causes it a Lenti virus related to a sheep virus and some of the other viruses we've seen in other species in fact there are Lenti virus relatives in a number of different species there's one in in in in monkeys there's one in cattle there's one in sheep and there's one in horses and these are photographs of these different lentiviruses well around around 1988-89 a woman named Marla Browne who lived in Petaluma California shows up at the UC Davis Veterinary College and says my cats kind of sick I think my cat has AIDS and the veterinarians looked at it and said yeah right however Niels Peterson who was a talented veterinary clinician at UC Davis said well let me look at your cat he took some blood out and he called cultured them with different kinds of tissue culture cells and then he asked whether or not those tissue culture cells two weeks later made any particles or excreted anything that had reverse transcriptase enzyme which is an indicator of a retrovirus like HIV one of them did he wrote a paper he characterized it he took the pictures he says we have feline immunodeficiency virus Marla Brown was right her cat had AIDS but she didn't have it from HIV she had it from a cat version the symptoms of feline AIDS are very similar and there have been a number of studies where cats throughout the world have been studied by talented clinical veterinarians if you look at this list of definitions it's basically very similar to hiv/aids flu-like symptoms weight loss depletion of the cd4 lymphocytes skin lesions bacterial infections opportunists infections is exactly what kills HIV AIDS patients these cats getting the same disease a pretty good model my colleagues and I by this time I don't know longer traveling to Africa Asia South America collecting specimens tissues and serum from many members of the cat family and I was terrified that perhaps some of these animals would be infected with FIV FIV is a virus which acts like HIV HIV at that time was as deadly a diagnosis as a bullet to the head 95% of the people with HIV died so I was terrified that the cats would habit to make a long story short Bob Olmstead and Gentry mounted a screen for FIV in non domestic cats to see how many of these wildlife species actually have a FIV they develop a test called Western blot which is an electro phoretic method where they took Marla ground Browns cat virus and they basically put it on a gel electrophoresis separated an electric current and then stained it with a labeled antibody which recognizes the ant about which would recognize the antibodies in a lion or a cheetah or a Puma and what you can see here is that the Lions actually several them showed a nice black band which that they had antibodies as if they had been exposed to something like Marla Brown sick cat the same thing with cheetahs there was a few of them that had it in the same thing with American Pumas we were terrified of this and mounted a survey of over 3,000 samples we had in the laboratory at the time and we found FIV antibodies in a lot of individuals this is an illustration of several populations of American Pumas the pie charts give you an indication of the percent of those animals that had antibodies against the feline AIDS virus as you can see throughout the West and Central America and South America virtually every population had a few animals if not half the animals that were infected with FIV when we looked at other species in South America such as the Ocelot the Margay the Jeffries cat the pampas cat they were all positive - not every cat but many of them had it in Africa ten percent of the Cheetahs had antibodies to feline immunodeficiency virus and in Africa leopards had it in fact it was endemic in about 12 of the 37 species of cats so we were terrified that we were just gonna have to sit around and wait for them to get sick we did some sequencing of the virus from every one of these species and produced a tree which is completely unreadable which I'm going to show you next and this is a evolutionary tree which only thing I want to say is that it is possible to relate the relationship between the sequences of many viruses to each other in the following way and the conclusion that I want you to point out is that all of these are Lyon viruses and all of these are pluma viruses and these are leopard viruses and these is actually hyena viruses down here what this slide is meant to tell you is that every virus that we see always has its closest relative another virus from the same species evolutionary biologists call this monophyly just means that they're all close together if they're in a lion are all close together in a puma but how did we interpret that well first of all it means that the viruses are not jumping between species all the time they jumped once and then they evolved inside the species and that's the way these viruses move around they jump from one species to another as is illustrated here now let's talk a little bit about the lions the Lions in the Serengeti are an amazing tourist attraction there's about 3,000 of them all together and they wander around in the ecosystem which is about the size of the state of New Jersey tracking wildebeest 100% of the Lions in the Serengeti over three years old are infected with FIV if you drive around and you see an adult lion looking on top of your Land Rover through binoculars there in fact to do with that 5e this is a photograph of our team who drew blood from these animals these animals are not dead they're quite happy they're anesthetized they all got up and walked away but I want you to believe me when I tell you that there was a time when we could gather specimens from these individuals and and look at them and what we found was that these animals were all infected with with feline immunodeficiency virus we isolated the virus sequenced it and so forth but the ecologist said you know Steve these animals are not getting sick I think they've evolved a genetic defense against this virus I think they're healthy when I say the ecologist I mean craig packer I mean and I mean the other line biologists at the time so if they weren't developing HIV or they weren't developing AIDS they weren't dying like the house cats why was that okay so we wrote a paper and we said well we think it's alright we don't think they're not sick and then what happened now I'm going to shamelessly read you one page of a very good book that I wrote a few years ago okay it's about a couple who were on safari in the Serengeti their name was Connie and Harold Chandler and this was the year was 1994 I think they rose at 4 a.m. an eager anticipation of the high point of their African safari a hot air balloon ride across the vast Serengeti topped off with a champagne breakfast Connie and Harold Chandler eagerly donned their banana republic vestments then hurried to the balloon launch pad a short land rover dried from the luxurious serán era tourist Lodge it was January 1994 the perfect season to view the expansive Vista of East Africa and its marvellous wildlife now the evening before their tour drivers had regaled them with apocryphal yarns of obliging Robert Redford and Meryl Streep and their crew during the filming of Out of Africa so the Chandler's gathered their snacks Sunstream and their camcorder onto the balloon deck in anticipation of a splendid Serengeti adventure now as the Sun crept up over the sweeping Serengeti Plain the balloon lifted them above endless herds of wildebeest zebra giraffe who had grazed the region to a brown stubble you know those fortunate enough to have visited a nice African game park would spend countless hours in animated conversation reliving their you for a foria most would also agree that even the best photographs are woefully inadequate and expressing his grandeur so I apologize right now for the photographs yet you just have to experience it to understand extraordinary enthusiasm that the savanna inspires now the Chandler's were flying actually and spiritually when Connie spotted a trio of young male lions strolling below the balloon descended and Harold started the camcorder rolling but then it happened the male in the rear began to twitch his whiskers soon his conniptions became noticeable enough to spook his brothers who took a few swats at the trembling adolescent and ran off probably to avoid the descending balloon the shivers progressed the tremors and then to severe muscle spasms then the unfortunate beached lurched into a grandma neurological cease seizure extending his front legs forcefully flailing him into the air and crashing to the ground for thirty agonizing minutes the animal thrashed about of as if it was possessed by demons finally as the violent seizures and gyrations gradually lost intensity the miserable creatures breezed his final torturous breath before the horrified balloon errs nobody felt much like drinking champagne the shaken terrorists had witnessed something that few people even trained military clinicians had ever seen the acute neurological collapse of a large predator but what had killed the lion was it a poison was Ana nasty germ was it a hereditary ailment the veterinarian in charge is Serengeti at the time was a talented clinician named melody roki the balloon pilot went notified her she mentioned that she had been seeing weird things than the lion populations over the few months including skinny lions when there was a lot of prey in fact what happened in the next six months was a thousand of the 3,000 lions living in the Serengeti ecosystems succumbed to a mysterious contagion between December 1994 and June 1995 dr. roki set up the equivalent of an international health campaign to try to find out what was the cause it was originally thought was FIV because that 5e is a relative a of HIV and there were press releases out that the Lions were dying of HIV because HIV causes AIDS and one of them one of the one of the display phenotypes pathologies of age Ivy's neurological symptoms melody wasn't sure she wanted to prove or disprove that hypothesis so she went out from dawn till dusk every day and darted every lion she could find she basically gave them a a physical examiner and she took tissues when she caught the animals were healthy got up and walked away the carcasses she was able to get some from anything she sent the tissues to veterinary college in Zurich the University of Tennessee Veterinary College Cornell Medical veterinary school in our laboratory she says what is it a talented pathologist in Tennessee Linda Munson looked at the brain tissues of these individuals and said I have seen this before this pathology looks like something that happens not in cats but in dogs distemper caused by our morbillivirus but we all know the veteran textplus tell us well canine more Villa virus canine distemper virus only affects dogs it will infect cats but dub caused any disease max appellate Cornell had monoclonal antibodies against the canine distemper viruses he put him down in the tissues of these guys and discovered bingo they had these red deposits which meant that his antibody was binding to something in this lion tissue and a number of animals that had died of whatever it was there were six animals out of 40 who did not have HIV or FIV who also died so that led us to exclude the possibility that HIV was the principal and the morbillivirus became the next best candidate during the period of the collapse of the population one-third were lost the incidence of antibodies against CDV he went from zero in 1992 to 10% 93 to 80 percent by the end of 1994 postdoc in the laboratory designed pcr primers for a number of genes in the canine distemper genome and amplified DNA from many of the tissues in lions what she discovered Margaret carpenter was that the Serengeti Lions indeed had sequences which were the first cousins of the canine distemper strain which was used for vaccines were from South Africa and of course the more bili viruses are relatives of more bili viruses in seals dolphins porpoises and the human measles and rinderpest virus winter past of course of cattle bars okay so and all the animals had it in fact when she looked at a number of lions hyena domestic dogs bat-eared foxes around the Serengeti she found that they all had a virus which was very similar to each other and a little bit distant from the domestic dog vaccine strain the cheet originally said was a match this was a very Catholic virus it didn't care what species it entered he just went into each one and it caused the disease and all now where'd it come from well the Serengeti and the surrounding areas has an indigenous population of people called the Masai tribesman who are pastoral Wanderers who herd cattle and most of them have dogs they did a survey Craig and his colleagues did a survey Sarah gasps going to the survey of the dogs blood samples in the next year so and they discovered that the incidence of antibodies against canine distemper in these domestic dogs was on the order of 60 to 80 percent now you can't get dogs into the Serengeti legally however because it's against the law to bring domestic dogs in so the intermediate is this guy Enis hyenas get crosswise with everybody they could crosswise the dogs they get crosswise lions and it's always bloody they were endemic with the distemper morbillivirus and that began vaccine campaign not of the Lions but of the domestic dogs with a herd immunity idea of getting the immunity down this was 15 years ago there hasn't been a serious outbreak since FIV was not responsible for the lion plague did it play a supporting role well I didn't know and melody wasn't sure we like to listen to our ecologist friends but we weren't sure so they began after this of rather widespread study across many areas of the Serengeti to look at Lions that were infected with FIV and those that weren't they collected specimens from each of eight different areas and they brought the materials back it was a great expense I had to hire helicopters to fly these women veterinarians around and they basically did an amazing job of collecting tissues and monitoring them and they looked for clinical correlates of HIV also SIV in these Lions and what the ones they looked at was how about the cd4 or gingivitis or Co viral infections these are normal hiv/aids defining conditions what happens when you monitor these which all of which were seen occasionally in lions and FIV versus FIV negative the answer was quite striking the red is positive for the disease listed on the bottom for gingivitis it was almost 90 percent in the positives and maybe 25 or 30% in the negatives for dehydration the same is true was twice as high in the FIV positives papillomaviruses 60% in the FIV positives 20% here less quantifiable was the presence of wasting animals that were skinny that were underfed that we're not doing very well and they were the histopathology demonstrated a chronic debilitation in Kruger Park and also in the Serengeti these were observed before during and after the CDV outbreak wasting the clinical chorus associated with these were these and every one of them was affected statistically significantly affected this was not an innocuous virus in these lions it was basically contributing to certain things however why do we think that it was innocuous well I think it's a little bit like a matter of the lions tend to postpone the immune collapse of their system until they die as something else there's Akademi analogous call this a completing mortality it's a little bit suppose HIV was just discovered suppose it was discovered in nineteen or 1865 in West Texas well at that time as illustrated by all the Cowboys turns including Louis and of these people didn't live very long they had a lot of competing mortality Indians would kill him snake bites would kill him disease would kill him they'd get shot by each other they were all dying before the vibe the disease could get them at the loser their eyes were infected with HIV you wouldn't know it the same way with the Lions so we think that the Lions are told us a little bit of lesson the lessons from these cats stimulated me in my post at the NIH to start looking for human diseases or human genetic explanations and resistances for deadly diseases and we began actually in the mid-1980s before this FIV outbreak and we were looking for human genes that polymorphic variants that influence the outcome of HIV exposure infection we gathered thousands of patients from epidemiologists who had characterized patients at risk for HIV they were people in risk groups actively homosexual men recipients of contaminated blood products before 1983 when the blood test was introduced people who were IV drug injectors who were sharing needles and city slums where the HIV prevalence was approaching 90% these were the risk groups some of them were getting AIDS some of them were not some of them when they got AIDS they progressed at different rates there are whole schools of Public Health dedicated to epidemiology including here and what they do is define heterogeneity in epidemics perfect as a geneticist I said yeah maybe some of that genetic that difference has a genetic basis the fledgling Human Genome Project was getting started and we were able to use the tools of amplification and allele recognition to test many many genes that were thought to be involved with HIV we discovered a number of age restriction genes we're probably the one that is the best-known was the receptor by which HIV enter cells when HIV encounters a t-cell it binds to the cd4 receptor and then to a chemokine receptor which is called ccr5 once this discovery was made by five separate groups in 1996 published in science nature and cell within five days of each other my group went to the lab where they had thousands of patients and began looking for mutations in the ccr5 structural genes they discovered a interesting deletion in some people called ccr5 Delta 32 it removed 32 base pairs leading to a frameshift and the wrong amino ask them to stop codon it was present in 10% of the Americans that we had in our cohorts the allele frequency which mean about 1% were homos they had two copies of this deletion they're pretty healthy these people didn't know they had a problem with ccr5 however when we went to the cohorts what we discovered was that people were homozygous for Delta 32 people who had two copies one from each parent they didn't have any ccr5 on the surface and the reason was that the garbage disposal detailed in the endoplasmic reticulum of cells recognized this crappy protein as being no good and eats it up the fork gets to the surface so people who were homozygous have no ccr5 in the surface so oh when we went to the cohorts and we said what's the frequency of the wild-type or the normal or the heterozygous or the homozygous we discovered and this is a summary of now 70,000 people remember that in a few minutes this number okay and what we saw was that the homozygotes were of equally frequent and people who were negative or positive for HIV and the heterozygotes worse too also but the individuals were homozygous never became infected there were no positives they were completely resistible reason this the doorway ccr5 which HIV enter cells is gone so we had a recessive restriction of HIV infection heterozygotes do become infected people with one copy of the normal one copy and when they do when they progress to AIDS they actually delay the onset of AIDS very slightly about two years it's statistically significant not as much as the infection but nonetheless there's this modest delay now I want to say a few things about ccr5 Delta 32 it's led to a whole discipline of trying to use it to help cure AIDS it's to understand some background Natural History but I want to tell you about its background because this is about plagues HIV or Delta 32 which protects you against HIV has an allele frequency of about 10% in Europeans however that frequency is not identical in all countries it's actually pretty high up in Sweden it's about 10% in Germany and in France 5% in Greek in Italy 0% in Saudi Arabia and 0% in Africa and 0% in Asia it's not present in the Asian ethnicities and it's not present in pure native Africans it's only present in Europeans and this gene frequency gradient is called a Cline and it's indicative of maybe something favoring it perhaps some adaptive value up north and then spreading down to the south now why does a gene which knocks out a perfectly good function like ccr5 what's the good function ccr5 is a chemokine receptor a chemokine receptor is a protein we know a little bit about its it has to do with the fact that when you have an inflammation in a joint because it's bruised or it's infected or something the inflamed joint shoots out these peptides called chemokines into the lymphoid system and they're meant to attack lymphocytes they bind to the lymphocytes through the chemokine receptor and then they home in to the abrasion and try to fix it I don't know how they fix it I don't think anybody really does but the point is that's what they do it's sort of one of the armaments of the immune system okay the reason that ccr5 seems to be dispensable is that there are other chemokine receptors that bind to the same chemicals so okay that makes sense why does the gene knockout like ccr5 existed a high frequency in europeans and not in africans or Asian peoples when was ccr5 Delta 32 born what can we learn about AIDS it's from the history of this variant and now I will quote not Charles Darwin but theodosius dobzhansky who is the namesake of the center that I direct in st. Petersburg he says nothing in biology makes any sense except in light of evolution if the Europeans have this and the Africans and the Chinese don't what that means is that the mutation probably occurred along the lineage after Caucasian ancestors European ancestors diverged from others we know from a lot of data that about a hundred thousand years ago the African peoples migrated Out of Africa into Europe and then dispersed into Europe and then over to Asia and then they split apart maybe 40 50 thousand years later they also made it within the end of Falls around this time and there's a certain fraction of all your genes that come from the Neanderthals we don't know if it's a good part of the bad part but that did come from the adults okay so this is beginning to make us think okay it's a recent addition but how when it took place the populations in Europe was on the order of hundreds of thousands of individuals so the day it was born in some baby let's say they were hundred thousand Europeans so if it occurred in the one mutation and the baby was born then all of a sudden it had a frequency one and two hundred thousand how did it get up to one in ten today among you which means that one in five of you is a carrier I had a resigns carrier well one of the possibilities is that the mutation had some kind of breathtaking advantage resistance perhaps to a deadly plague or infectious disease maybe it just good the people that had it and that caused a selective jack-up or rise or advantage that favored the carrier's plausible not proven a guess if you look at other populations for ccr5 mutations there's about 20 of them that our team discovered but there was something interesting about them this is Delta 32 the others are all around here usually when you have mutations in a gene it's bad for you it's sort of like taking a hammer to a Mercedes been its engine one time in a thousand it might be good the most the time it isn't so what really happens here is the frequency of mutations that alter amino acids they actually do something is filtered out quickly so that when you look at all the genes the 20,000 genes across the human genome the amount of what we call nonsynonymous amino acid altering mutations is on the order of 20 25% the rest of them are all synonymous that is their mutations that occur but they don't change the amino acid because they don't matter that's not true of ccr5 90% of the mutations in the ccr5 are amino acid altered the only other place we've seen that is in the antigen recognition site of the major histocompatibility complex class one HLA genes what do you say what does that mean HLA is one of the principal immunological gene families that defends against deadly infectious diseases it has enormous variation enormous hundred times a thousand times more than other genes and the reason is it's good for you to be diverse at your immune system remember the Cheetahs okay so the MHC has a large amount of diversity and the diversity is all in the site that recognitions peptide so that individual can recognize viruses and pathogens they've seen in the past to the ones they haven't seen the more the better remember the Cheetahs ccr5 has more amino acid substitutions than HLA so everybody now agrees this has not argued my friends my enemies they all agree ccr5 was elevated by a recent breathtaking selective pressure a history of natural selection but what caused it mr. Darwin helped our thinking here we don't know what caused it but there's another piece of data I want to tell you about and by the way if you get too tired to listen to me there's a short movie at the end which is pretty good okay it turns out that we can estimate when this mutation took place by looking at the neighborhood of genetic variation around the ccr5 gene why is that well when a mutation takes place on a chromosome it occurs on a chromosome that has a whole bunch of other mutations maybe a hundred upstream 100 downstream over a short space and they'll have some form or other sort of like a signature so the first generation you just have this one form with all this all these different variants next to it however as mutation and breeding takes place recombination clips it away and randomizes randomizes the variants on either side and the further away the variant the quicker it goes away from being associated and the closer is the longer takes so the links of the non-random parts is actually a chronometer which you can relate to time so we took this next slides unreadable to but it illustrates for you that there are a bunch of variance around ccr5 that we looked at it when we measured linked around it the average length of the stretches in Caucasian European Americans and Europeans is about 20,000 base pairs ccr5 it's a million much longer we developed a simple algebraic equation when you can do on a calculator that you can buy from the local Walmart to basically compute how long it was since the last selective event that favored ccr5 Delta 32 when we did that that was 700 years ago 700 years ago so I'm not very good student history like all you I went to the library and said what was going on in Europe 700 years ago and what was going on in Europe 700 years ago was the Black Death the Black Death so immediately the cause of the Black Death came as a candidate well maybe that jacked up ccr5 Delta 32 then we went ahead and said okay well we have this indirect data we have the dating and everything like this but let me tell you for those of you don't know what I'm sure most of you do there is that interesting history about the Black Death it started out in the Burmese ghettos probably sometime in the 12th century and there were supposed to have been tens of millions of Southeast Asians that have died and then it came across the Silk Road to the Black Sea and into Messina Sicily in 1348 and people were dying of this disease it is a disease caused by a bacterium called Yersinia pestis and it infects the individual the bacteria has a plasmid which has a bunch of proteins called yaps Yersinia operative proteins that go up to macrophages which is your first line of defense against bacteria and that it injects these poison darts into the macrophages and destroys their primary defense then it goes to the lymph nodes that grows up that explodes them and it causes these pleating black things called buboes where the name bubonic plague comes from it basically within a five-year period spread throughout Europe up to up to UK France Germany around they're taking between 30 and 70 percent of the population including here in Cambridge quite a few it was huge horrific thing and it's carried on the backs of black rats and trans transmitted in marmot pelts which which were which were brought back and the migration that I just described is illustrated in this old photograph about coming from Sicily and then up and then round back around to the black sing something on the order of 50 million Europeans succumb to this disease and it was a horrific disease and at that time of course there was no medicine there was no there was no HMOs or pharmacies these people all I had was their theology I thought the evidence was pretty good that bubonic plague was a strong candidate but other people thought that there were other things that could have been the cause of plague some people thought maybe smallpox some people thought anthrax there was a paper came out of Britain a few years ago that said it was a bola type fires typhoid these are all guesses as to what may have caused the Black Death and what also may have been responsible for ccr5 Delta 32 elevation I wasn't sure about it all but I knew that the devastation was enormous about eight years ago I heard about a town near Manchester called ium this little town of very few people maybe a thousand people that most of the people there are descended from the survivors of the Black Death they have a little museum there this is up near Manchester's you go see it and the people in the town are very proud of the fact that they are descendants of the Black Death and nobody else moved in there there haven't been any immigration so I wondered if the average frequency around Manchester and London and in other places in England was on the order of ten to eleven percent what's the what would happen if I looked at the ancestors or the descendants rather of the survivors of the Black Death so we went to the town and we drew blood from a hundred and twenty people actually we didn't draw blood we took us cheek swab and extracted the DNA and I was thinking that if Delta 32 was protective against Yersinia pestis then it would be higher in ium than it would be in the surrounding neighborhoods and I wouldn't tell you the story if the results didn't turn out the way I wanted to but they weren't as good as I wanted I mean basically the allele frequency came out to be 15 percent but the incidence of the of the homozygotes was 20% of homozygous was twice as high three or four percent it was only a hundred and some people so it wasn't statistically significant but it was in the right direction and I was encouraged by that but it was amazing to walk through this town where 50 percent of the people died during the Great Plague in the 17th century which was caused by Yersinia pestis and this reminds me to tell you that the Black Death is horrific it as was and as much as it changed everything wasn't the first or the last plague there were one every generation up till the Great Plague and I think the last one was in the 18th century in Marseille where they had 40% mortality the first one the earliest one was the Justinian plague which was in between 550 and 750 BC and that was said to have taken a hundred million people in Roman Empire can you imagine counting a hundred million people in Roman numerals this is basically an amazing result I don't know if the numbers are right but there was a lot of deaths it certainly was enough to explain the elevation if your city could be connected now a few years ago there was a paper published from a group which hasn't been repeated but it was an interesting result these workers were wondering whether our hypothesis was full all wet or true so they took mice which are hyper susceptible to Yersinia and they made knockouts that is they eliminates the cr5 so they had the equivalent of a delta thirty-two homozygote Mouse strain and a wildlife strain and then they infected it with your cine pestis and they quantified how much infection they got with or without ccr5 and the answer was that your cine a' pestis requires ccr5 to get into these cells that's what they reported photographs taken right out of the nature letter said money slatkin a good friend of mine from Berkeley did some mathematical calculations and modeling and says it can't be your Senia there wasn't enough mortality I said are you kidding of course there was enough mortality says I think was smallpox smallpox how are we going to test that this was back at the time where smallpox was being extirpated completely completely by the World Health Organization's then along came 900 that time there were biological anthrax mailers that killed several people in the United States and all of a sudden people became a little bit more worried about bioterrorism and deadly diseases the two places in the world where smallpox was housed was in Russia at a center and at the CDC in Atlanta Georgia I was working the NIH was housed on Fort Detrick which is has a history of being the center of excellence for biological warfare well that stopped but some of those guys are still there so I found them and I said you know can we test smallpox to see whether or not it requires ccr5 if I get you blood samples from people with alternative genotypes can we infect them he says we couldn't done this before but he says I think I can get permission so he did so we did the experiment and we exposed human cells to your senior pet or to smallpox and here's what we thought basically we found that the difference between the wild-type the heterozygous and the homozygous was indistinguishable smallpox gets into cells it doesn't care what the Delta 32 genotype is pretty good again supported at least in my head that we had it so ccr5 Delta 32 is the knockout mutation it seems innocuous it was born 600 years ago when the population was greater than hundred thousand that shows dramatic evidence for breathtaking selective elevation and it seems to be required for your sin iya and HIV to enter cells it has no effect on smallpox infection and the people in am poor survivors of the plague have at least an elevation if you believe the results and you can't defend them specifically but that's what we saw nonetheless so we need to do the same experiment with your Sania on human cells we're trying to arrange it I found some guys in Russia who worked in these bougie things and they're willing to do the experiments so we'll see what happens stay tuned one more piece of data ccr5 became interesting to many people because he was shown for example that the mix of mitosis virus used ccr5 and rabbits to enter mr. boo ptosis is a distant relative of smallpox it's one of the things that led the smallpox camp to get invigorated but many other viruses have been looked at for whether or not ccr5 is utilized by them especially given all those mutations that clearly had nothing to do with the black death but they were mutations probably to avert infections animals like or viruses like to use ccr5 one of my colleagues Phil Murphy at NIH and his postdocs did an experiment with knockout mice the same mice that were done for the use any experiments saying brain and they said does West Nile use it now West Nile of course is a flavivirus mosquito-borne had arrived in the United States now about 14 years ago it caused about 17,000 infections and about 678 were fatal they were monitored by the CDC in Colorado and in Atlanta 20 percent not all 20% of the inciteful infection progressed encephalitis an 80% or subclinical to date there's no vaccine and no effective treatments still wondered whether ccr5 was required for West Nile so he did a simple experiment he took a bunch of mice that were either ccr5 positive or ccr5 negative and exposed him to West Nile and mice are very sensitive to West Nile virus and what he expected was the Knockouts would be resistant but that's not what he found what he found was the wild-type were resistant and the scouts all died than 12 days ccr5 was the primary defense in these mice against West Nile did it work in humans he went and got a hold of as many patients as he could and he did Association tests and what he discovered was that when you looked at people infected with West Nile they either got this deadly encephalitis or didn't remember I said 20% did that the frequency of ccr5 Delta 32 and the negatives was low but the positives was really excuse me the Delta 32 homozygotes were elevated four to eight times and the people that got sick so yes Delta 32 is a risk factor a huge risk factor most of the people that died of of West Nile virus are Delta 32 homozygotes so a lot of my friends have come up to me before the state says hey I'm homozygous for Delta 32 isn't that nice because I'll never get HIV I said oh but now I tell all these people you're gonna have to avoid these cocktail parties in Florida in the summertime because they have mosquitoes they have West Nile and this is not a joke this is a serious risk factor for this and it's true whether you look at just coke Asians are all races together the difference is the Delta 32 genotype is what seems to be the major contributor to encephalitis when West Nile virus so it's a mixed blessing let's look back final couple of things in 1996 HIV entry receptors like ccr5 was discovered and the triple drug therapy was discovered but the translation of ccr5 has had some interesting variants one is that pharmaceutical companies began to develop antagonists to ccr5 binding HIV because they were generated by the resistance of the Delta 32 homozygotes so there's about ten of them and three of them are now FDA approved in the last few years and they're Salvage drugs that are being used to block ccr5 from binding HIV so this has been translated directly to the patients my last example is an anecdote this is Timothy Lee Brown Tim is an American he was living in Berlin he became HIV infected he's a gay man and he didn't he got on the powerful therapy didn't do very well he didn't like it and made him nausea so he didn't want to take it so he stopped it and then he developed acute myelogenous leukemia which is a death sentence for an AIDS patient his doctor was a talented clinician named Guerra hunter he said to him mr. Brown you can die however there is one option that we can do and that's we can do a bone marrow or stem cell transplantation we can use stem cells to make bone marrow cells to make lymphocytes and what that involves is a radiating year old lymphoid system getting a donor from a big bone marrow registry that matches your HLA type and then infusing that in there and then crossing our fingers but I gotta warn you if you don't take the drugs HIV will hide itself in reservoirs and will jump back so eventually it's gonna come back and you'll have to get back on the treatment he says okay I'll try it but then Guerra hunter did something very interesting he actually got two or three hundred people who were compatible with mr. Brown for donation he asked for pieces of blood from each one a gia typed them for ccr5 Delta 32 he was looking for a homozygote so that he could use Delta 32 as a homozygote donor for this bone marrow transplant number 77 was homozygous donor number 77 called up the guy got the blood already mr. Brown infused him with it he suspended the antiretroviral treatment he suspended the antiretroviral treatment because he wanted to give the graft a chance to grow back but he monitored mr. Brown for a week a month six months later a year later no antiretroviral drugs Delta 32 donor virus never came up he sent in an abstract to the world aids meeting he was given a poster I walked by the posters that's this this is really interesting I says this guy looks like he's suppressing HIV is the only guy in history everybody else has never nobody's ever done that and there's 40 million people infected with HIV he published an article in New England Journal two years later and everybody else really not a cure could be an anecdote we don't know exactly and then they set up a committee to study what happened mr. brown and they said you know he's probably still got HIV someplace that just hasn't had a chance to come out he's still I know drugs this is three years later no no HIV no about no rebound they said well we need to look and biopsy his tissues so if I give you a list of tissues you'd be horrified they wanted to look at his testes they wanted to look at his lymphoid cells his kidney his liver his his his his neurological tissues they wanted to look at everything and mr. Brown was so happy to be alive he agreed they went into it they use the most sensitive pcr methods available detect one half of a virus in a milliliter blood found nothing it's now I think seven or eight years later and everybody says mr. Brown was cured he has not recovered he goes to AIDS meetings now he gives talks and he's basically the one person in addition to that the problem the caveat is that this is exciting but it's not a treatment for everybody because one-third of people who get bone marrow transplant get a disease called graft-versus-host where the graft attacks the host and kills the patient one-third mortality is not what you want to hear when you're going into a procedure so what they've done is they've used zinc finger knockout technology this is a tenon and UCLA to knockout ccr5 in patients who have HIV who have a bad prognosis and then infuse these synthetic Delta 32s back into the patients and see whether or not they get the same good result as mr. Brown and the answer is that it's very promising but it's too early to say the truth is ccr5 has been a very interesting ride but HIV is not as horrible as it used to be I talked to you about the promising bone-marrow transplants some of the vaccines have partial there is something called pre exposure prophylaxis which is a combination of drugs given to people at high risk which will reduce infection and transmission by five or six volt there are vaginal Reiko besides that are important and interestingly enough the president's emergency program for AIDS relief was started by this gentleman President George Bush it's probably the one thing in his legacy that everybody agrees was good and it is basically to convince the Congress to put 15 billion dollars into treatment in the developing world to help stem this guy got the price down from ten thousand fifteen thousand to about $150 a month for treatment and millions of people are now on treatment he then added 48 billion dollars into 2009 for this is more than the people before George Bush and certainly more than the people after mr. Obama education is working to I gave you the story about the natural history finally this is the age quilt it was displayed in 1996 on Columbus Day and the Washington Washington Mall it's the last time it was displayed it's over 17,000 quilts each one the size of a coffin commemorating one or two or partner a pair of AIDS patients so last time it was displayed in full because it got too big maybe it would fit on the Heathrow Airport there's been a lot more people involved in this work than myself Mitch Bush and David will were pioneers in helping develop the wildlife Laurie marker has told the story of cheetahs and worked with us for over 30 years melody roll key was the chief veterinarian the Serengeti study and also she was part of the cheetah study in the beginning for the SARS work I'd like to thank this all-girl cast of Ali Wilkerson Emma Jennifer Gila Gill melody Laurie and Meredith and for the FIV study it was led by Jennifer Troyer and melody rocky the plague story had a number of collaborators with ccr5 including Bob Fisher and clay Stevens and David Reich this is the team that did the HIV restriction discoveries this is the laboratory of viral of genomic diversity which I led at NIH until two year years ago and the informatics that we're doing now is led at the job Jansky center by myself and Pavel peasant or this is our group and last of all I'd like to thank you all for your attention [Applause] [Applause] [Music] [Applause] so Steven thank you I mean you've demonstrated so clearly them the major advances in understanding that genomic data make possible and the opportunities and tools and the challenges that follow from that understanding I mean all I can say is thank you really very very much for a truly wonderful lecture [Applause] so next week the Darwin college lectures continue with Professor Steven Emmett from Microsoft Research who will talk about the human plague in other words all of us so thank you we'll see you again then

Channel: Darwin College Lecture Series

Views: 4,508

Rating: 4.9595962 out of 5

Keywords: Plagues, Darwin College, Darwin College Lectures, University of Cambridge, genetic epidemiology, HIV /AIDS, retro-virology, bioinformatics biodiversity, Stephen J. O'Brien, human genome project, scourges, genomic archaeology

Id: QtkojdUysyQ

Length: 83min 19sec (4999 seconds)

Published: Sun Feb 23 2020