 Thank you very much. It's an honor to be here with you. I think this is going to be a fast-paced presentation on the Zika virus in the post-epidemic phase, an evolving narrative. Next slide, Ruth, please. Yeah, so this is quickly my background. I come from Brazil, trained in infectious diseases there and got involved with a lot of research in genomics. And now I do a PhD on using genomics for diagnosis of infectious diseases. This is something that I will talk about very, very quickly here. Next, please, Ruth. Yeah, so the objectives. I think it's going to be really an overview to the detriment of getting too much of the detail on the pathology itself. But I think it will be interesting to see how this particularly epidemic evolved. And I think in your mind, I would ask you, and I think this is going to be intuitive, and you'll be doing this anyway, to kind of establish parallels with all the situations of epidemics or pandemics that we've been living. Okay, so of course, there will be the parallel with COVID-19 pandemic, and you can also think about the outbreaks of monkeypox infections. So I think this will be cool so that we, yeah, overview and where we are now. So I would hopefully provide you with insights. And then, depending on your interest, you can delve into that later. So surveillance systems and notification strategies, epidemics, communication strategy, and pathogenomics as well. And then, again, as I said, the parallel between the different epidemics. Next, Ruth, please. So I will start with this interesting post from ProMed. I don't know if you're all familiar with ProMed. I highly recommend you look at that. It's the largest publicly available surveillance system that kind of does some global reporting of infectious diseases outbreak throughout the world. You just can subscribe and you receive a feed of emails. And so I remember seeing this post in May of 2015, when the Minister of Health reported cases of a rash illness in the Northeast region of Brazil. So you see, Brazil is divided into states, and then the Northeast region, there are many states there. And in that particular region, there was this rash disease. And after three months, the total number of notifications reached 7,000. So very, very fast spreading. So the majority of cases had a really, really benign cause. The most common signs and symptoms, there were an itchy rash and less frequently low-grade fever, headache, pain, and swelling joints or muscle pain. And the most affected age group was a young population between 20 and 40 years of age. So what is the differential here? We know that Brazil since 2009 faces a quasi-permanent epidemic state for dengue. So loads of cases, okay? So between, in that year, between January and September, we had over 1,500,000 cases of dengue notified. So what do you think with that description? Wow, is it dengue? Next, Ruth, please. Yeah, so, but fine, fair enough. Really difficult to test. The majority of the proportion of those cases, so you're not, you really don't have any theology at the time. But then a few months later, a few months later, there's an unusual increase in babies born with microcephaly in Penambuco. P.E. Penambuco is one of the states in the northeast region of Brazil. This was flagged to the State Secretary of Health by one doctor that is specializing in doing a prenatal ultrasound. So that's very interesting, right? He says, wow. I mean, this week, I saw two or three cases, which is absolutely unusual, okay? So the State Secretary of Health issued a clinical protocol, okay? So let's start recording this. Immediately, 90 cases of microcephaly cases were registered. When the annual mean is nine, okay? So what is the differential here? So I think it's important to say that microcephaly, there is a difficulty in diagnosing it. And it's not only in Brazil or in lower middle-income countries. No, this is worldwide. Because we don't, first of all, we don't have a baseline prevalence of central nervous system anomalies, okay, in children. Microcephaly is very complicated because there is no standardization on how to measure. It's not as straightforward to measure as you would think. And there are so many different levels of ascertainment that vary in Europe and the United States that most countries actually, microcephaly, it's not a mandatory notification, okay? So after this pandemic, of course, it became so clear that you need to standardize things. So there was a study conducted worldwide to be able to try and get to what is the normal head to conference, right? It's such a, such a primal question. But yeah, there you go. Next Ruth, please. So, yeah, so in November, so many months later, what are we doing about that from a, from a, you know, like communication point of view or how the country is managing this? Or because it's, it's, I remember feeling I was living here at the time already, but I remember feeling it's very terrifying, isn't it? You have an epidemic of, of, of children being born with this really, really severe problem. So, and we don't know what is causing it. Okay, so politicians, of course, come into play. And then I think invites you to kind of make the parallel with, with the COVID-19 pandemic, whatever you lived the first year of the pandemic or so. So politicians, can they solve diagnostic conundrums? Well, not really, right? So what do you need when the situation is, is, is, it seems that it's so, it's always scary for the general public. Of course, for, for the medical community, it is as well, you need to establish the right studies to understand what that is. So epidemiological studies such as case control, you get the people that are affected and match with controls and try to identify the risk factors that are most likely contributing to that. And then you follow up cohorts. But then usually, you know, like politicians, the burden of a certain thing, it's too heavy. And it was particularly heavy in Brazil at the time, economic crisis and other problems. So there was a selection of one piece of evidence to fit with the assumptions. Okay, because I need to tell the people what this is. So at the moment, there had been one case of a positive Zika test in a child with who had microcephaly. Okay, so, yeah, the minister of health jumps in next please Ruth. And what do they do? They do a declaration of a public health emergency of national concern. Okay, this was in line with international sanitary regulation set up by the WHO. But of course, there was a lot of people who were bewildered. How can you say that when, you know, there's, there's only one case in the with where you could, you could prove the kind of microbiological test. In retrospect, this was absolutely correct. But at the time, I remember thinking that was a bit of a very, I don't know, it wasn't, it wasn't right from the from the government to have done that. So when you set up this public health emergency of national concern, you make the mandatory notifications of case of microcephaly, you put up guidance and protocols, you direct more money towards that. And then there is a plan to to national plan to fight microcephaly. And then you you kind of link different agencies and different ministers to coordinate the response. Next Ruth, please. And then it's very interesting, right, to see the this ever evolving adjustments and the risk perception from the point from the part of researchers and and the and the public health agencies and the public and the language, right, that is used. So so that was after three months from the Brazilian national emergency declaration, then the WHO declares the public health emergency of international concern. Okay, and what what gives ground to this decision is they say that the language is the link while strongly suspected between the virus Zika virus and case of microcephaly has not yet been proven. Okay, so but I mean, you need you need to to again COVID-19 pandemic, one of the WHO officers said, you need to get there fast and hard because when a situation like this comes up, there is no empty spaces. All right. So somebody wouldn't would dominate the narrative and it could be you could be really fine tuned risk communication. I think there is a science behind it. You know, of course, there is and what do you do and how do you convey this thing that we don't know and knowledge is being built as we speak. Right. So so this is I think in Brazil in the case they got there fast and it was a good thing in again. So but the WHO recommended increased research. And then what does this public health emergency of international concern again, really tell the country tells the country you need to get your act together and you need to then put resources in place to deal with this. Okay. So it's kind of it's mostly to bring awareness and to force countries to of course, it can't it's not binding. So of course, countries will choose how to deal with it. But yeah, there is a strong recommendation that something should be done next week. And the evidence builds up. Okay. So most of the data in the beginning still deriving from studies of very weak design, as you would expect, it's it's it's normal. The consistency of data is not ideal and the consistency of measuring measurements of head to say the least, right? Of head. Conferences. And then you have this guilty until proven innocent versus okay, circumstantial evidence only. Okay. So what happens at this stage? Countries. So they had never been, I will talk about this, but no cases in the Americas ever of Zika virus infection, but there had been outbreaks before. So in these countries where there had been outbreaks, people start looking into the data they had. And so there is this first paper here. There was an outbreak in French Polynesia. Okay. So there was an outbreak of Zika virus in loads of cases, an increase in case of glimbary syndrome diagnosed. So let's go and understand if there were a relationship then with our records. And then at the same time, there is a cohort of pregnant women in Rio and the January in Brazil that it started being followed to see the outcomes of pregnancy. So this is this is really interesting. Next Ruth, please. So quick overview of microcephaly. Okay. So what can cause microcephaly infections, toxins, or it could be of genetic cause. Okay. So by May 2016, so a year after the first case was flagged up by the by the ultrasoundographer in Pernambuco who was scanning too many babies or fetuses with small heads. So a year later, nearly 7000 reported cases of microcephaly or the congenital abnormalities of the central nervous system or both had been reported in Brazil. Okay. But then you need to confirm because of course you define confirmed case suspected cases and confirmed cases. So 7000 suspected, but then you need to go and confirm with with a physical examination and with the blood test. So it takes time. The state of Pernambuco, that region in Brazil, the leader of notified cases. I think nearly 2000 cases have been reported since the first epidemiological bulletin. Okay. It should a year late earlier. Next Ruth, please. Yeah. And so Guillain-Barre syndrome, if you don't remember the autoimmune neurological illness that causes muscle weakness. So it's immune system damaging the peripheral nervous system. Okay. And then a few countries started reporting that. Okay. Cases of Guillain-Barre syndrome associated with Zika virus infection. Next Ruth, please. And then other at the same time countries started notifying all the severe presentations of CNS involvement and known CNS involvement and Zika infection. So with patients in whom we diagnosed Zika infection, either indirectly with serology or with the PCR to detect the RNA of the virus. So cases of meningoencephalitis, acute myelitis, and then lots of eye abnormalities and eye issues such as UVitis. But up to then thankfully a very small number of fatal cases reported. Next please. Yeah. So at that point a new entity is described, which is the congenital Zika syndrome. Okay. So these babies, in terms of neurological findings, lots of signs and symptoms. So hypotonia, hypereflexia, extreme irritability, tremor, seizures, brainstem dysfunction and dysphagia. So of course it's a spectrum of how much the central nervous system is affected and you can have really, really tragic manifestations. You also have loads of eye abnormalities. So macrothalmia, length subluxation, cataracts, intraocular cosifications, different abnormalities of the optic nerve. Yeah. So there were so many cases described that then this syndrome was defined. Next one please Ruth. So what types of evidence have we got so far? So we've got the epidemiological evidence, right? So you see countries reporting suspected cases that fit established definitions. So you see how many countries reporting over a year later, nearly 30 countries. Next please. Another type of, you know, more epidemiological evidence. So Salvador is a city in Bahia, another state of the Northeast region of Brazil that it was beautiful. Look at these epidemic epidemiological curves here. So the number of suspected cases of microcephaly peaked after a lag of 30 to 33 weeks from an acute rash illness peak at the time and then it corresponds to time of potential infection of pregnant mothers during the first trimester of pregnancy. Okay. So this is observational evidence, epidemiological evidence. Next Ruth please. What else? So you have this cohort and you have the preliminary reports of studies being conducted. So you have this follow-up of a cohort of the 88 pregnant women in real with rash illness and then more data of course emerging from other countries. Colombia also had an important outbreak there. So with this really large groups of pregnant women being followed up and then there was at the time here as well the preliminary report from a case control study in Brazil with needing more numbers of course, but already indicating that the association was very plausible. Next Ruth please. So epidemiological evidence. What about the clinical evidence gathering? So you start having case reports of fetal infection where you have the clinical manifestations and you can prove microbiologically that the Zika virus was there. Okay. So yes, the association gets increasingly stronger. So you have the Zika virus RNA detected in the amniotic fluid of two children, the microcephaly, and then you see serological evidence of past Zika infection. So you find antibodies identified in Hawaii in a baby with congenital microcephaly born to a mother who had been in Brazil during pregnancy. Next Ruth please. And then if you do this, if you follow up this pregnant women, and you can do beautiful studies where you observe, you have data in terms of blood tests to detect the presence of immunological markers or to diagnose infection itself. You can follow up, you can know exactly what was the clinical manifestation in the mother, and you can also serially follow up with ultrasounds how the fetus is developing. So this is very interesting. Next please. And then yeah, and now in terms of this microbiological evidence, what do you do? You start going to Muran models. So you inoculate the virus and you see what happens. And then you can get tissues, different tissues and specimens and see if cells get infected. So using a Polynesian strain of the virus, it was proven that you can infect placental cells. And then you can see that there was damage to the brain of the mouse, the fetal mouse brain. Next Ruth. Yeah, so I will go mindful of time or just I will not get into details. But then there is similar studies doing with strains found in Cambodia as well. So you can, you see, so now you have to prove that in the presence of Zika, there is, there is cells are infected, and then there will be the consequence of that in different tissues. And then you start establishing what is the mechanism whereby the the virus will cause the problem. So next please Ruth. Similar study with Brazilian strains. So you can see how infection really has an impact in the development of of fetuses. Next please. And then finally you found you find evidence in human tissue, right? So this was a study in Puerto Rico that you can see, yes indeed. In fact, placental cells of humans as it did, in fact, on the on the neuron models. So yeah, this is the levels of strength. And I think here it gets pretty clear that yeah, there is association which is most likely causation. Next Ruth. So yeah, so that kind of the biological aspects of it in a nutshell. What about the dynamics of transmission then, right? So where was Zika before this massive outbreak in the Americas? So the virus itself was first identified and described in 1948, okay, in the Zika forest in Uganda. The first illness was described in Uganda decades later. Serological studies had indicators and widespread human infection in some countries in Africa, in India, in Southeast Asia, in Philippines. But up to 2007, there had been only 13 naturally occurring cases reported, okay. Next Ruth please. So 2007 and why is that? Because in 2007 and the Yap Island in the Macronesian archipelago, that was when infection in humans outside Africa and Asia were detected. And this island had a population of 6700 people. The estimate is that almost the entire island got infected, okay. Of course, here is suspected cases. Not all of them will have microbiological confirmation, but this is extraordinary, right? It's beautiful to see, not beautiful at the time for the island, but from an epidemiological perspective. You see an island and with a naive immunological population, how the virus can wipe all the population, which is really interesting. Next Ruth please. So there was that outbreak in 2013-2014, there was an outbreak across all the Pacific Islands, okay. So at this year, there were over 30,000 suspected cases. Next please. Yeah, so you see it seems like the virus is slowly moving, making its way towards increased expansion until it meets South America initially. So what could have brought this virus in? Okay, so think about COVID-19. It's the interconnected world. It's most likely was brought from travelers. So let's think about particular events that could have happened in Brazil at the time that would have brought, would have introduced the virus there. So there was a World Cup in Brazil in 2014, but no economic Pacific countries were competing and therefore, you know, the numbers of people coming from these countries would be very, very low. However, on the same year, it was in August. So I think it was a month after the World Cup. There was this canoe, canoeing race competition, and it was a world, so vast world springed championship. And of course, all these countries where we talked about Zika being there, they're really strong in canoeing. It's kind of a common practice in sports. So yeah, so it looks like you could have been introduced on that at that time. Next Ruth, please. And then this is the beauty of genomics. Somehow you can go back to these places because people will have will have either samples stores or of course you keep your your specimens there that you can go back and test in the lab. And then when you get more cases, you try to compare them right with the genomically so you can do the whole genome sequencing and you can see how all these different strains will relate to one another. You would have a timeline and you'll be able to tell established mutation rates and molecular clock and you can, the timings are clear. Okay, so yeah, it showed that all these strains that had been the types that were more common in Africa and then the ones that from Brazil, they share a common ancestor with the strain that circulated in French Polynesian in November 2013. Okay, so that that is it was interesting to see. Next Ruth, please. So wide explosive spread. Okay, so okay, fine. It was introduced in Brazil, but why was it so successful in establishing the outbreak and why is that? Because it was an immunologically naive population, first of all, but Brazil has is infested basically everywhere. There are loads of very and highly competent the highly competent vector, which is that it is a cheap time. Okay, genomic studies show that there was no viral mutation that made the virus much more infectious. Okay, no, it was just that beautiful, perfect situation for the virus, competent vector everywhere in the country and in a naive population. Next Ruth. Yeah, so beautiful isn't it? So classic epidemiology. So you have the international concern, the public health emergency of that international concern that was declared. Not a lot of knowledge at the time, but public health responses could not wait. Okay, so it's one of I think, again, learned with COVID-19, you need to act fast. All right. And then with the growing body of evidence, epidemiological and microbiological and the models, the strong suspicion has become a strong scientific consensus, which was really interesting. Next slide, Ruth, please. And I think now I have to go really fast. So this is just the final report. The preliminary report, sorry, of the case control study that was being conducted in Rio, okay, following up the, you get the babies affected with Zika and then you get babies that weren't affected, but were born at the same time and had very similar characteristics so they weren't affected and then you can establish different risk factors and odds ratio. Next, please. This is a map from this year. What are the countries and territories with current or previous Zika virus infection? Okay, so we have 89 countries reporting transmission from 2015 onwards and it continues up to now. All right, so I think over a million people I thought have been infected and approximately 5,000 cases confirmed congenital syndrome case associated with Zika. Next, please. Not going to delve into that. It's just the specificity of Brazil in terms of other countries with large epidemics. Brazil was by far the one with the highest incidence of congenital Zika syndrome. Okay, next, please. Yeah, so why was that? You know, we didn't really get the numbers right. So is it because we had a much, much higher baseline of infections overall and then this proportion of microcephaly will be in keeping with what other countries saw or there are other cofactors, co-infection with dengue or chikungunya that we have a lot as well. Anything else? Next, please. There were a few hypotheses, but we don't know that yet. It's not yet clear why the cases in Brazil were so dramatic. Next one, please. So what do we need? We need more numbers. We need to continue monitoring. You need to continue to establish other risk factors that could have been accountable for that. Next, please. This is then beautifully two years later. The final report of the case control study was published. And then this one, it was really nice. So the association, the first line, Interpretation to Association between microcephaly and congenital Zika virus infection, was confirmed with kind of massive odds ratio. And in this study, they were able to see whether there could have been a relationship with vaccination during pregnancy, or with the use of a specific pesticide in the region there, but they didn't really find strong evidence that there couldn't be association with that. So this was kind of, yeah, it is. From an epidemiological point of view, it is the virus. Next, Ruth. So 2016, end of the global emergency, okay, the problems were still there, but at the time, so what's the difference from the beginning? Now we know, we know more, we know where it comes from when we know there is still lots of unknowns, but from a point of view of, you know, the logistics of WHO, yeah, we can stop that. Okay. And I think that's the right thing to do. And of course, it might make it difficult for situations to keep up, to countries to keep up the momentum in terms of infrastructure. But yeah, that's what happens. So in November 2016, the end of the pandemic, sorry, pandemic, no, the public health and emergency of international concern. And finally, very briefly, some next slide please, Ruth. Some pertinent perspectives that go a bit beyond, you know, so now let's think about all these affected children. Okay. So first of all, human rights. So what happens here? We have a new generation of children born with the serious birth defects and disabilities that will last a lifetime. Putting this strain on families that the Northeast of Brazil is very deprived. We need to cater for the needs, the needs, okay, it's an obligation of the state and proper support must be put in place. And of course, this has not happened. Next, Ruth, please. Another really, this is a really, really sad thing. So this is recent study and it showed, it looked in Brazil, it's a cohort study that included all suspected cases of congenital Zika syndrome born in Brazil over three years. Okay. And then what it showed, it confirmed that cases had an almost two times higher mortality when compared with unconfirmed cases at 36 months of age. Right. Next, Ruth, please. Which is very sad. So this is this Kaplan-Meier curve showing survival. Okay. So you see the confirmed cases, the orange bit, the suspected cases, the blue, you see it's kind of worse than the yellow, but because there's much more children in there and it's the children on this case that wasn't confirmed. However, very high proportion of children with microcephaly there. So you see what we are doing, right? I mean, it's very sad. Next, Ruth, please. And it's the penultimate light. Climate change as well. So you know, diseases take flight with climate change. Why? Because it's providing opportunities for insects, it's providing infectious to new populations. So we know that our edges, albopictus, it's, which is the green, you see the map in the United States. Look, it goes up everywhere, everywhere, you know, up to North or and look at Europe when you have the albopictus as well and it's coming North and North. So regions where you wouldn't really see this before, you will probably start having autoctonous transmission and established community transmission of such viruses. And finally, the health economics aspects, right? So of course, there's new cases. There was a pandemic and this case is, you know, support for the families and their children. It's kind of forgotten. And yeah, it shouldn't really happen. And finally, to my last last slide, I think you went back, Ruth. And next. Yeah. And so I mean, there's so many competing interests and so many competing epidemics. And we kind of, we need to, we need to keep this, this, this people at heart and at the target of policy, right? So particularly the mothers. And here I put this mother with this child. Because in Brazil, specifically, there was some very kind of sad, common occurrence, the partners of these women, they would just abandon them. The woman and their families, because there's a lot of, I think anything that happens during pregnancy, there is a lot of burden and guilt attributed over the mothers. So very difficult. And I think this is a nice image to finish this and to kind of highlight who is at the heart of everything we do. Yeah, thank you. I think it was a rush. But as I said, it was to provide some insight and try to give you a kind of a a nice overview of this, of the Zika epidemic.