Identifying and Understanding the Problem

When one pulls up a map of the global spread of dengue in 2020 like the one released by the CDC, it is easy to see that South Asia, along with other tropical countries, is one of the major regions to be affected by dengue. It is the most widely spread mosquito-borne disease, with an estimated 100 to 400 million cases and tens of thousands of deaths occurring every year, and the numbers are only expected to increase with climate change.

According to the World Health Organization, dengue fever is one of the top ten global health threats – it's also the most rapidly spreading. There has been a 30-fold increase in global incidence over the past 50 years.

It has been estimated that there are roughly 390 million dengue infections per year, of which 96 million have clinical or subclinical manifestations. Which means that only roughly a quarter of the total infections are symptomatic and are reported. This means that 75% of infections remain unreported or undiagnosed.

Dengue wreaks havoc in our country, India, every year. In one of our first team bonding sessions, we found out that almost half of our team has had dengue at least once, and all of us have family members who have been affected by dengue - the issue is personal to us. We had also been reading multiple reports on Dengue as the hidden epidemic during the COVID-19 pandemic, especially in Pune.

When we started looking into dengue, we found out that the disease has no cure - it is only treated symptomatically. Most times, the disease is asymptomatic, which could inadvertently lead to a worse second infection.

We sought the help of our PI Dr Srinivas Hotha, Dr Siddhesh Kamat, Dr Nishad Matange, Dr Vineeta Bal, Dr Gayathri Pananghat, who are all acclaimed experts in various fields currently working at IISER, Pune, during this early phase of ideation through multiple meetings trying to pinpoint the problem and coming up with a viable solution.
Our conversations led to the conclusion that the problem to be tackled here is the severe Dengue Hemorrhagic fever (DHF) that usually results from a second infection.
DHF could result in low platelet count, internal bleeding, shock, and even death. The worst part is that DHF does not even have a specific treatment aimed at decreasing the viral load. We were determined to come up with a solution for this.
We identified the main cause behind the heightened secondary infection to be non-neutralizing antibodies due to a heterologous primary infection or antibodies present in sub-neutralizing concentrations acting as trojan horses with the heterologous secondary infection (infection with a different serotype) leading to an increased viral load. Keeping with the Trojan horse analogy, the gates of Troy here would be the Fc gamma receptors present on cell surfaces. This phenomenon is known as Antibody-Dependent Enhancement.

In order to understand the problem and the gaps in science we needed to fill, we needed to understand the ground reality first. Early on in our cycle, we spoke to Dr Madhu Bashini M, a General Physician and Diabetologist with experience in treating Dengue Fever and Viral Fever at Chettinad Hospital in Chennai, to understand the problem better from the perspective of an experienced healthcare worker. She told us most Dengue cases go undiagnosed, which could lead to a worse secondary infection disguised as a primary infection later. She told us that the current treatments in place, aimed toward treating the symptoms and not the virus, are sufficient for a primary infection and so our solution had to be equally effective for a secondary infection as well as a primary infection.

Further along in our cycle, we spoke to Dr Prema Raghunathan, who is a professor of Pediatrics at Rajrajeshwari Medical College, Bangalore and Dr Harsha Patel, who is a postgraduate student at the Bowring Hospital, Bangalore. We presented our project to them and sought inputs. Dr Raghunathan told us that from her experience, most Indians tend not to visit doctors until the last minute. Dengue almost always goes undiagnosed, and confirmed that during the first wave of the pandemic, several cases of dengue were misdiagnosed as COVID-19. Additionally, she cautioned us against using antibody therapy for a progressed Dengue infection.

Dr Patel informed us of the stark reality of Dengue patients and cases in India. He said that during an outbreak one can expect upto 80-90 cases daily and that they, and most tertiary areas, face a shortage of beds and man-power. He also taught us about the 3 main phases of Dengue and advised on which phase to administer our therapeutic. He elaborated upon why the problem of Dengue is particularly unique and dangerous in India and said that it is very rarely true that clinicians can identify the different phases of dengue, and this is crucial since the type of fluid treatment depends on the phase of the disease.

In June, we spoke to Dr Shobha Rao, who is a Director of Research & Training at Society for Initiatives in Nutrition & Development, and Dr Smita Jog, who is a President at Rotary Club, Shivaji Nagar, Pune during the blood donation camp that we co-conducted. They pointed out a crucial point that we had missed about Dengue being passed on from mothers to unborn fetuses in pregnant individuals and that such cases are rarely documented because of lack of awareness and diagnostic centres.

To further get a detailed breakdown of cases in different Indian cities to understand the scope of the problem better, we spoke with local municipal bodies - Pune Municipal Corporation (PMC) and the Bruhut Bengaluru Mahanagara Palike (BBMP). At the PMC, we met Shri. Sanjiv Vavare from the Health Department. He spoke of the general trend seen in Pune over the years and said that due to the intermittent rains that cause an increase in stagnant water-filled years which in turn see a rise in the incidences of Dengue during the months of July-October. He was of the opinion that during COVID-19, the number of cases went down since people weren’t in closed spaces as much and added that it could also be due to Aedes primarily being a day-biter. He also made it a point to mention that the prevention of the spread of Dengue is entirely dependent on public participation and that their main control measures were holding rallies, society meetings for vector control, demonstrations and distributing pamphlets. At the BBMP, we met Dr Madhusudan from the Health Department. He gave us extensive data on the ward-wise cases of dengue in the last few years and commented on the decrease in cases seen over the last couple of years and agreed that it might be due to the pandemic. He asked us to reach out to Hospitals to gain more insight into case-statistics. We were unable to do so on our own, and sought the help of a team we have been working very closely with.
Our friends from the iGEM MIT_MAHE team reached out to doctors, on our behalf, from the Infectious Disease Department in a government medical college located in Kerala, India and spoke with Dr. Veeresh Kumar, MBBS MD, Arun Clinic, Guntakal, Anantapur, located in Andhra Pradesh, India managed to obtain data regarding the daily cases, hospital measures taken to treat patients, and their opinion on the government initiatives taken. The detailed information can be found

here

Fig: Yearly report of the confirmed cases of Dengue in Pune city.

Fig: Percentage yearly increase in the confirmed cases of Dengue in Pune city.

Fig: Yearly report of the confirmed cases of Dengue in Bangalore city.

Fig: Percentage yearly increase in the confirmed cases of Dengue in Bangalore city.

Here is the detailed data of the age-wise and sex-wise cases, the total number of confirmed cases in Pune, and the total number of cases in Bangalore we were given by PMC and BBMP.

The ones most affected by our solution is the general public. They are the ones who are at risk of contracting Dengue and the ones whom our therapeutic would be administered to. It is important to understand the extent of the problem from their perspective and to fit our project in accordance with their needs.
We visited a ‘vasti’ near our campus. A vasti is a small, slum-like settlement with very poor living conditions. This particular vasti, at the time of visit, was water-logged, had multiple swamps and extremely unhygienic toilets and contaminated water supply. We were appalled by their circumstances, and even more so when found out that it gets much worse as the monsoon progresses. Their surroundings were filled with mosquitoes and other insects. Our conversations with them led us to try and reach out to our institute to come up with a solution. We also realised that the primary consumer of our therapeutic would be people from rural and slum areas, and therefore it was crucial for us to minimise the cost of production and make it accessible and affordable to all.

We also spoke to several workers at our institute. One among them was Santosh Kadam, who is 34 years old and has been educated only till the 8th grade but has picked up the skills of operating the autoclave and protocols of decontamination and discard of waste biological materials. He raised several important points during our discussion. He talked at length about the low levels of medical care in the village, and the dependence on homemade remedies.
When we asked him about how a therapeutic for Dengue fever could be received in his community, he brought up the point of awareness. He drew parallels with the low COVID vaccination rates in his village, and erroneous beliefs like how COVID would vanish because of the high heat in the summer. We realised that such low levels of awareness need to be tackled from the ground-up. As a starting point, we came up with an informative survey designed to dispense tips on dealing with Dengue, vector control measures, knowledge on the disease pathology etc.

Further along our cycle, we spoke to local construction workers, Traffic police and local vendors. One major point that stood out in all these meetings was the lack of awareness among the local people and mistrust of the Indian medical system. They told us that most Indians, especially in rural areas, tend to never visit health clinics (if there are clinics in their villages) and instead prefer self-treatments with indigenous medicines. We hope that our efforts to spread awareness would go a long way in removing erroneous beliefs.

Determined to ensure that our stakeholders’ needs are met, we proceeded with coming up with a solution and evaluating it^{[1][2][3][4][5][6][7]}.

Coming up with a Solution

What makes a good therapeutic? - is a question that we asked ourselves and our stakeholders over the course of our project. After talking to our stakeholders, we identified the hallmarks of a good therapeutic.

An ideal therapeutic for Dengue should:

Incorporating these properties into our solution would enable it to be a viable option for the treatment of Dengue.
Over the course of our project, we’ve done our best to meet the above requirements and have constantly edited our project to fit into our vision of an ideal therapeutic.
However, we have been constrained by the nature of our solution in certain areas like having a low cost of production. Using a bacterial system for the production of our antibodies is probably more expensive than the traditional method of immunizing animals and extracting antibodies from them. However, we prioritised using a safe, ethical and modular method over having a low cost of production. Even so, our current method of using a bacterial chassis is relatively cheaper and easier than using eukaryotic cells, as outlined in detail below.
Additionally, we have also prioritised having a therapeutic that causes minimal immunogenic response over a minimally invasive mode of drug delivery as outlined in detail below.

When we first sat down to think of a solution, an obvious direction for us was to find a molecule that would render the virus inactive but not bind with the Fc receptors. After a literature study, we hit upon the avian class of antibodies - IgY, which checked all of our requirements. Being a non-mammalian antibody, it is incompatible with the mammalian Fc receptor and they could be engineered to bind and neutralize dengue virus or any virus with multiple virulent serotypes, for that matter, as long as we had the sequence.

We faced our first roadblock with acquiring the sequence and realized that the 3D structure of IgY had not yet been publicly resolved. With the help of Dr David Bradley and Juho Choi, IgY researchers, who sent us the sequence of the constant region, we put together the construct of an IgY-IgG hybrid with the constant region of IgY and the variable region of IgG and even resolved a 3D image of it. As a placeholder, our hybrid antibody was targeting the Maltose Binding Protein. We were extremely lucky to have had in-depth discussions with Dr Alexander I Taylor, who had worked extensively with IgYs before and our secondary PI Dr Mehmet Berkmen, a bacterial geneticist from NEB, regarding the properties of IgY. Dr Taylor advised against using an IV mode of drug delivery and warned us about the possible immunogenic response that could result from exposing our body to avian antibodies. We also had multiple discussions with Dr Satyajit Rath and Dr Vineeta Bal, who are both immunologists at our institution, regarding the possible immune response of our body against IgY and what we could do to circumvent it. While the prospect of deimmunisation by removing the T-cell epitopes on our construct was considered, the risk of inadvertently destabilizing the structure and thereby compromising the efficacy of our antibody was too much. Dr Rath also told us that our IgY-IgG construct was simply a very high-tech solution to a very low-tech industry. We also learnt from Dr Supriya Kashikar, the founder of Genext Genomics, who told us that the half-life of IgYs in the body would be low since it cannot bind to the FcRn receptor in our body and therefore cannot be recycled. Further, Dr Gavin Screaton, an Immunologist at the University of Oxford, advised us not to continue with IgY and instead opt for an alternative.

We wanted our drug to offer a solution to ADE, to effectively neutralise the disease, for the virus-antibody complex to be cycled out of our body with a minimal immune reaction from our body, and for the dosage levels to be non-toxic. Since our solution did not check all of these boxes, we decided to shift away from it. This came as a setback for us, but we were determined to have a realistic, simple, and effective solution for ADE and a good drug. Following advice given by Dr Screaton and Dr Rath, we looked into the functionality of Fc mutated IgGs and aglycosylated IgGs - it had all the advantages of using IgY with the added benefit of not causing an immunogenic response. However, circling back to Dr Kashikar’s warning and our subsequent literature review^[8], Fc mutated antibodies would still have meant that its half-life in our body would be low and the cost of use high.

In a bid to improve affordability and to simplify our solution further, we landed on single chain fragment variable (scFv).

ScFvs are small and have no constant regions (and thereby no effector functions). Their size makes them cost effective and efficient. However their half-life in the human body is quite low. To solve this problem, we decided to insert a peptide extension to the scFv that would increase its half-life by binding to the FcRn receptor. We decided to call this structure NeoFvs. We were very fortunate in successfully reaching out to Dr Shannon Sirk and Mr Vince Kelly, who have both worked extensively with scFvs before, to gain their inputs on working with scFvs with the FcRn binding peptide for dengue. Mr Kelly informed us that they were unable to generate all the variants with the binding peptide in different regions of the scFv due to time constraints and resources and asked us to look into it. We tried to surpass this by using HADDOCK simulations. He even suggested a possible future implementation to engineer bacteria and release them into the gut such that they produce these NeoFvs when needed. He commended us for taking forward their initial project of using these scFvs for breast cancer into viruses, calling it a true test of modularity.

Producing the Solution

A chassis is the platform that acts as a framework and support for biological parts and components.

When we looked into the existing production methods for IgY, we found that the popular method was to extract it from purified egg yolk by infecting chickens with the virus. We felt that this method was unethical. Moreover, it had issues with batch standardisation and sustained production. It also generated polyclonal antibodies, which increase chances of cross-reactivity, leading to false positives and non-specific interactions with the antigen, which could be non-neutralising.

We began looking for an alternative which ticked off our expectations for a good chassis and locked onto SHuffle. The SHuffle B strain derived from the E. coli B strain was a fit for most of our requirements. We reached out to Dr Mehmet Berkmen, who works at New England Biolabs (NEB) and was instrumental in the creation and development of the SHuffle strain of bacteria and was the first to produce full-length IgG antibodies with it. What started out as a zoom meeting ended up being a lengthy and fruitful relationship with him agreeing to be our secondary PI. He was delighted at the prospect of mentoring us and was incredibly helpful in ironing out the myriad questions we had regarding the culturing of SHuffle, production conditions and optimization.

Dr Raghavan Varadarajan, a biophysicist from IISc, Bangalore, urged us to look into alternative production methods like standardized CHO cell lines and yeast systems when presented with our project. Following his advice, we did a thorough literature search and found that while CHO cells produce accurate complex proteins with high yields, they are expensive to culture and are slow growing (both points independently reiterated by Dr Mugdha Gadgil, a CHO cell expert at NCL, Pune, they can also have unwanted post-translational modifications like glycosylation.
Furthermore, Dr Vinod Jyothikumar, a consultant for operational risk management at DSS+, during a question and answer session after a webinar, pointed out that CHO cell lines do not retain stable protein expression over long-term culture and have a high cost of production. Additionally, Dr Mayurika Lahiri, from our own institute, told us that most animal house facilities don’t keep chickens since they are only used for producing IgY. This traditional method of using chickens is removed from research and therefore our method of using a modular bacterial system could help promote further research into antibodies and antibody therapeutics.

A similar search regarding yeast systems and a conversation with Dr Umesh Shaligram, director of R&D at the Serum Institute, India (intoduced to us by the GATES Foundation, India), showed us that while it is widely used in recombinant protein production, they can mannosylate the antibody, which could make it immunogenic. Bacteria lack the machinery for N-linked glycosylation, and cannot do the above. Dr Ratnesh Jain, involved in recombinant protein production from ICT, also advised against using eukaryotic cell lines since Dengue is primarily a tropical disease and requires the therapeutic to be affordable to all.

Selecting our Epitope

Dengue virus belongs to the Flaviviridae family of viruses. It has 4 serotypes that can infect humans. India and Brazil have shown incidences of all four serotypes. Choosing an appropriate epitope that is equally conserved across all four serotypes, essential to the virulence of the virus and easily accessible was crucial to overcome ADE and neutralize the virus. After discussions with Dr Nishad Matange, a bacterial geneticist at IISER Pune, we looked into NS1 as a possible candidate based on his suggestions. NS1 protein is the only non-structural protein to be released extracellularly, which makes it an easy target for antibodies. Additionally, it is hypothesized to be crucial for the replication of the virus inside the host cells.

We then considered targeting the Fusion Loop epitope, which is essential for the virus to establish contact with the host cell and enter it. It is also highly conserved across serotypes. However, the FLE is partially hidden below EDI and ED-II and is exposed for a very short time, leading ot potential issues with accessibility. Meetings with Dr Varadarajan Sundaramurthy and Dr Shashank Tripathi, both host-pathogen interaction experts from NCBS, Bangalore and IISc, Bangalore respectively, yielded the same conclusion. They advised us to look into alternative epitopes.

Our conclusion was reached in a meeting with Dr Vidya Mangala Prasad, a Dengue structural expert from IISc, Bangalore. We presented the candidates we had in mind - NS1 protein, Fusion Loop epitope, EDI and ED-II to her and sought her inputs. She suggested that we target the E dimer Epitope, an epitope that is rarely targeted by the natural antibodies produced by our body, lessening competition with the therapeutic. It is also highly conserved across all four serotypes. Additionally, as EDE is present largely during the pre-mature stage, it will prevent binding and entry of the virus into the cell. She told us that the NS1 antibodies are only produced in greater concentrations after the viral load crosses a certain threshold and asked us to look into the C-10 class of antibodies, which had been isolated from human dengue patients. Following her suggestions, we did a thorough review of the literature on the E dimer Epitope and the stages of the viral cycle it existed in^[9][10]. We did a sequence comparison that showed that EDE was conserved across serotypes. We finally derived our scFv sequence from the C-10 antibody.
We looked into acquiring the E dimer epitope and considered obtaining the Envelope protein and dimerising it in the solution. However, Dr Milind Gore, a highly-acclaimed immunologist and the former director of the National Institute of Virology (NIV), made us aware of the fact that the Envelope protein only dimerises and folds accurately in the presence of the pRM protein. This is the reason why vaccines that use just the E protein tend to fail to elicit neutralizing antibodies against EDE.
Due to safety concerns, we could not use the entire virus and instead chose to work with a Dengue VLP whose cassette had been deposited by Dr Stephen Harrisson, Harvard, in Addgene. We then had several discussions with Dr Rahul Roy who is a Dengue Virologist at IISc, Bangalore, who was strongly supportive of going with the EDE and advised us against using a cocktail of antibodies, since that could lessen chances of neutralization. He also offered to let us borrow his Dengue VLP construct and offered his help and lab space to work on neutralization assays for our proof of concept. Debayani Chakraborty, from his lab, very graciously gave us her time and helped us perform our assay^[9][11].

Designing our Experiments

Once we settled on our project after integrating and evaluating our project based on all the advice we received, we proceeded with designing our model and testing it out with our experiments. Our Dry Lab and Wet Lab components are truly entwined throughout our cycle, and we received constant support, advice and guidance from several professors and PhD mentors at IISER who were always ready to troubleshoot and offer alternative solutions. We also spoke to several experts from across the world to ensure that our project design was viable.

Dry Lab

Modeling was an important part of our project. Our Dry Lab journey began with trying to find sequences for a full-length IgY. We managed to put together the constant region of IgY with the variable region of IgG, with the help extended by Dr David Bradley, and Juho Choi was kind enough to give us the sequence of the constant region.
During this process, we realized that a 3D structure of IgY had never been publicly resolved and so sought to determine the 3D structure for our IgY-IgG construct. Dr M.S. Madhusudhan, an Associate Professor at IISER, Pune, working in the field of Bioinformatics and Structure, was a great help to us. He guided us through the usage of AlphaFold for structure prediction and was key in helping us interpret the results. With his help, we managed to obtain a predicted structure for IgY-IgG.

The majority of our dry lab falls under two domains:
MOLECULAR MODELING: This is an umbrella domain for -

1. AlphaFold structure prediction
2. Molecular Docking
3. Molecular Dynamics

Dr Madhusudhan helped us extensively in getting started with AlphaFold and in interpreting the results. He helped us with an overview of molecular docking and suggested us to set up differential protonation of histidine residues in molecular dynamics simulations in testing out the functionality of our NeoFvs in different pH conditions.

Dr Arnab Mukherjee, who is a professor working in the fields of Chemistry, Data Science, Theoretical and computational chemistry and biophysics at IISER, Pune, helped us with the different techniques in the molecular dynamics in our screening workflow.

Statistical Design of Experiments (DOE):

We had several meetings with the inventor of SHuffle, Dr Mehmet (Memo) Berkmen, who was incredibly patient and gracious enough to go over the nitty-gritties of working with SHuffle and helped us design our plasmid construct for producing IgY-IgG and NeoFvs.
SHuffle bacterial strain was novel to us and largely to iGEM as well, so wanted to optimize the working conditions of SHuffle to produce complex proteins. Memo suggested that we try statistical design of experiments (DOE) to find the optimal working conditions for SHuffle. He put us in touch with Dr Thomas Howard, an international expert in DOE and synthetic biology. Dr Howard was very helpful in getting us started with the technique, introducing us to the philosophy of DOE and the various softwares used. He also advised us to have a BSA calibrated SDS-PAGE as a quantitative estimate of yield of protein as a tool to measure the success of DOE.
Dr Mugdha Gadgil, an expert in CHO cells and DOE, currently working at the National Chemical Laboratory (NCL), Pune, India, met with us multiple times and helped us in adapting our project and factors to the DOE workflow. Rushik Bhatti, a PhD student at IISER, Pune, helped us finalise our protocols, list of experiments and factors for DOE. He advised us against using RPM as a factor and told us to normalize the OD measurements for all our cultures.

For further details on our DryLab front, do check out our Modeling Page!

Wet Lab

Our wet lab work began in early April. Most of us were largely inexperienced with molecular biology, and our PhD mentors played an irreplaceable role, guiding us and showing us how to perform each experiment. They were constantly present and ready to help us troubleshoot. Dr Mehmet Berkmen was instrumental in getting us started and helped us chart out our early experiments. He helped us design our plasmid constructs and even went on to send us both strains of SHuffle E.coli - SHuffle B and SHuffle K-12, plasmids for cytoplasmic protein chaperones, proteins and antibodies for ELISA and the cyclonal plasmid from his own work.

Dr Rahul Roy, as previously mentioned, had offered us VLPs for a VLP fusion assay and we needed properly folded, purified proteins to complete the assay. To test the proper functionality of SHuffle with regard to protein folding, we approached several experts.

Dr Sunish Radhakrishnan, who is an Associate Professor at IISER, Pune, cautioned us against trusting an ELISA for proper protein folding since an ELISA only requires the binding sequence in the correct conformation. He suggested that we perform a maleimide-PEG assay in conjunction with ELISA to determine if the correct disulphide bonds have been formed. We were also looking for an assay to measure dissolved oxygen in our culture without using a bioreactor and he suggested that we go for the Clark electrode method.

Dr Siddhesh Kamat, an Associate Professor at IISER, Pune, suggested that we perform a Thermal shift assay with SYPRO Orange to test for proper protein folding. He suggested that we perform a Thermal shift assay with SYPRO Orange to test for proper protein folding.

After folding, we needed to determine the overall strength of binding of our antibody or its avidity and the subsequent neutralization capacity. Dr Gayathri Pananghat, Associate Professor at IISER Pune, offered to help us out by making mutations in our antibody that would increase its binding affinity. Dr Varadarajan Sundaramurthy, a host-pathogen interaction expert from NCBS, Bangalore, suggested that we try phagocytosis as a method for determining the neutralization extent of our antibodies. Since one of our antibodies had a peptide linker to increase its half-life, we were recommended an FcRn binding assay by Dr Sundaramurthy to check if our antibody could be recycled.
Dr Sanjeev Galande, a renowned epigeneticist at SNU, Delhi, helped us out with our wet lab organization and reagents.

For further details about our wetlab plans and list of experiments we conducted, check out our Experiments Page!

Devising the Drug Delivery System

Our literature review^[12] suggested three common modes of transport for antibody treatments - subcutaneous, intramuscular and intravenous. While subcutaneous, followed by intramuscular, would be best for ease of administration and compliance, the primary concerns in making this choice are that of safety and efficacy, which must be judged in preclinical and clinical trials. During our search for a viable mode of drug delivery we received inputs from several experts - Dr Supriya Kashikar told us about IgY being made into an oral prophylactic treatment for Covid-19 and advised us to do something similar, as it can easily be absorbed into the digestive tract.

Dr Harsha Patel instead advised us to go for an IV mode of treatment since it would be easier to administer in a government hospital set-up and would have greater accountability since the patient would have to come to the hospital to get treated and wouldn’t be responsible for their own treatment. This is especially important in a country like India where the majority of those infected by Dengue would be from poor economic backgrounds, and likely to be mistrustful and ignorant of medicines.
Dr Vandana Patravale, a drug-delivery expert from ICT, Mumbai and Prof Nishikant Subhedar, a pharmacology expert from IISER, Pune, on the other hand, advised us to go for an encapsulated mode of delivery and asked us to look into sustained release to prolong drug titre in circulation.
We intend to look into all the possible modes of drug delivery to come up with the best possible product.

Spreading Awareness

Our conversations with the people around us who made up a major component of our stakeholders, showed that the majority of the people were unaware about Dengue or worse had superstitions and flawed perceptions about the disease, doctors and medicine in general. In a bid to tackle this problem, we decided to come up with an informative survey that would serve the dual purpose of understanding the distribution of awareness about Dengue and dispense information regarding the disease, its pathology, prevention methods and current treatment. In order to make our survey more effective, we took the help of Dr Pooja Sancheti, an Assistant Professor in the field of Humanities and Social Sciences at IISER, Pune. She asked us to put ourselves in the shoes of our survey takers and analyse the kind of questions they would or would not prefer to answer. She suggested that we should ask if our survey taker was from a tropical or non-tropical country since that might affect their level of knowledge, which we incorporated into our survey.

Here is the link to our

Survey Questions

Fig: Pie chart showing the percentage of survey takers residing in tropical and non-tropical countries.

Fig: The distribution of responses to some of our questions.

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