Advanced technologies have been used to solve a long-standing mystery about why some people develop serious illness when they are infected with the malaria parasite, while others carry the infection asymptomatically.

An international team used mass cytometry – an in-depth way of characterising individual cells – and machine learning to discover ‘immune signatures’ associated with symptomatic or asymptomatic infections in people infected with the Plasmodium vivax parasite. This uncovered an unexpected role for immune T cells in protection against malaria, a finding that could help to improve the development of much-needed malaria vaccines.

The research, which was published in the journal JCI Insight was led by WEHI’s Dr Lisa Ioannidis and Associate Professor Diana Hansen, in collaboration with Professor Ric Price from the Menzies School of Health Research, Darwin, and Dr Rintis Noviyanti from the Eijkman Institute for Molecular Biology, Indonesia.

At a glance

• Advanced technologies have revealed ‘immune signatures’ that differentiate immune responses that drive symptomatic or asymptomatic Plasmodium vivax malaria infections.
• The international collaboration revealed a previously unrecognised role for immune CD4 T cells in preventing serious disease and controlling asymptomatic infection of low parasite burden.
• The findings could guide to the development of better vaccines against malaria, a disease that kills hundreds of thousands of people around the world each year.

Variable immune responses

Malaria is a parasitic disease impacting hundreds of millions of people each year. After infection, people develop immunity to the Plasmodium parasite that causes malaria – but this immunity only reduces the disease severity rather than preventing infection altogether. Despite the immense global impact of malaria, there are not yet vaccines in clinical use to prevent this disease.

The immune response to malaria is a ‘double-edged sword’, Associate Professor Hansen said. “While an immune response to the parasite can prevent severe disease, in some people it is an excessive immune response – driving severe inflammation – that exacerbates malaria, causing the most severe, and potentially fatal, symptoms,” she said.

“Our research has investigated the longstanding question of how immune responses differ between people with symptomatic and asymptomatic malaria infections. We focussed on the Plasmodium vivax form of malaria, which is most common in the Asia-Pacific and Latin America. This species is a particular challenge to control as infected people can carry it for many months in the liver without symptoms.”

Using the University of Melbourne’s mass cytometry facility, the research team were able to undertake in-depth, multi-dimensional assessments of the immune cells in blood samples provided by people living in a vivax malaria-endemic region of Indonesia. Dr Ioannidis said the team compared many aspects of immunity in samples from people who were uninfected, asymptomatically infected, or symptomatically infected with P. vivax.

“In collaboration with a WEHI bioinformatics team led by Professor Gordon Smyth, we used machine learning to develop an ‘immune signature’ that distinguised between these three categories of samples. These signatures could be applied to new blood samples from people infected with malaria, to accurately predict the severity of their infection,” Dr Ioannidis said.

Enhancing malaria control

Dr Ioannidis said the immune signatures revealed the key components of the immune response that drive immunity to malaria. “Antibodies produced by B cells were one important component, especially in people with high parasite loads and symptomatic disease, but we also discovered that certain types of CD4 T cells were critical to keep infections in check, preventing symptoms,” she said.

“This is the first time CD4 T cells have been shown to be important for controlling asymptomatic P. vivax infections.”

Associate Professor Hansen said the discovery could lead to better approaches to controlling – or even eliminating – malaria. “Malaria vaccine development has focussed almost entirely on measuring antibody responses as a marker of vaccine success. Our research has revealed the important role of CD4 T cells in controlling malaria infections – and we think these cells need much more consideration when designing malaria vaccines. Because vivax malaria can persist in asymptomatic people, it is critical that vaccines activate CD4 T cells to control these low-grade infections,” Associate Professor Hansen said.

The research was supported by the Australian National Health and Medical Research Council, the Australian Academy of Science, the Wellcome Trust, the Indonesian Ministry of Research and Technology and the Victorian Government.

Featured image: Dr Lisa Ioannidis (left) and Associate Professor Diana
Hansen (right) have led a study into why some people
develop serious illness as a result of malaria infections © WEHI

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Provided by WEHI

Finding Published in Nature Offer Potential for Use by Travelers and Prevention of Malaria in African Populations in Near Future

Researchers from Sanaria® Inc. and the National Institutes of Health (NIH) are making progress in the development of highly protective malaria vaccines.

In an article published today in Nature, Sanaria’s PfSPZ-CVac (CQ) vaccine is reported as being safe and protecting 100% of six subjects against a variant malaria parasite three months after their last dose in the company’s Phase 1 safety and efficacy trial. This is the first time complete protection against a variant malaria parasite has ever been achieved that long after vaccine administration.

The variant parasite used in the trial is a Brazilian malaria parasite genetically more variant from the African parasites in the vaccine than 700 malaria parasites from Africa. Protection was achieved at a dose that is 20% of the company’s first-generation malaria vaccine dosage.

“These results represent extremely important progress, unanticipated by most malaria experts,” said Professor Martin Grobusch, Head of the Center of Tropical Medicine and Travel Medicine, Amsterdam University Medical Centers. “Until recently, malaria vaccine developers sought to achieve high-level protection against non-variant malaria parasites, often only two to three weeks after vaccination, with immunity waning thereafter. The finding of 100% protection against variant parasites that are so divergent from the vaccine parasites at three months is unprecedented. This vaccine approach should be advanced now as a potential tool to protect travelers to Africa and further developed for the prevention of malaria in African populations.”

The Nature paper also includes results of a second study using PfSPZ-CVac (PYR), which combines Sanaria’s PfSPZ with pyrimethine (PYR), a drug used for seasonal malaria prevention in African preschoolers. This vaccine was well tolerated and protected 82% of the 17 subjects to whom it was administered from the Brazilian variant parasites or the African vaccine parasites three months after their last dose.

“We are encouraged by the significant findings reported in this seminal paper, which justify our investment in Sanaria and its systematic, scientifically sound approach to developing the highly protective, cost effective vaccines required to eliminate malaria, a scourge of humanity, particularly for the most underserved on our planet,” said Holm Keller, Co-Managing Director, the EU Malaria Fund.

“Sanaria’s vaccine development program is designed to produce safe, cost-effective vaccines that provide high-level protection against malaria parasites that cause more than 400,000 deaths annually, primarily in Africa,” said Stephen L. Hoffman, Sanaria’s CEO. “With this goal in mind, Sanaria and our partners in the International PfSPZ Consortium have pursued a step-by-step approach to maintaining safety, increasing efficacy toward 100% against variant parasites, increasing the durability of efficacy, and decreasing the required vaccine dosage. This study reports huge progress in all four areas.”

Sanaria® PfSPZ-CVac is a chemo-attenuated, live whole parasite vaccine in which an anti-malarial drug is co-administered with parasite cells (PfSPZ) to kill them before a clinical infection develops. In the trial reported in Nature, the anti-malarial was either chloroquine (CQ) or PYR and efficacy was measured by controlled human malaria infection (CHMI). In addition to exposure in natural settings in Africa, the company has relied on CHMI of vaccinated and unvaccinated adults to assess vaccine efficacy. This is a rigorous test of malaria vaccines that can be conducted with small numbers of trial participants since 100% of unvaccinated subjects develop malaria.

The clinical trial was sponsored by Sanaria Inc. and conducted by the LMIV, NIAID and NIH. The full paper in Nature can be accessed at nature.com here.

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Reference: Mwakingwe-Omari, A., Healy, S.A., Lane, J. et al. Two chemoattenuated PfSPZ malaria vaccines induce sterile hepatic immunity. Nature (2021). https://doi.org/10.1038/s41586-021-03684-z

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Provided by Sanaria

Scientists at the University of Nottingham have made a major breakthrough in discovering how the malaria parasite is able to multiply rapidly in the mosquito gut, and how targeting it at this stage may stop the transmission of the disease.

The research, funded by the Medical Research Council and led by Rita Tewari, Professor of Parasite Cell Biology in the School of Life Sciences at the University, could pave the way in helping to eradicate the disease.

The study, published in Communications Biology, was a collaborative effort by scientists from the Universities of Leicester, Warwick, Oxford and the Francis Crick Institute in the UK, the King Abdullah University of Science and Technology (KAUST), Saudi Arabia, University of Geneva, Switzerland, University of Leuven, Belgium, Hokkaido University, Japan.

Malaria is still one of the biggest killer infections worldwide, with nearly half a million people dying of the disease last year – particularly in sub-Saharan Africa. The disease is caused by a single-celled parasite called Plasmodium, which is transmitted between people by the female Anopheles mosquito when they bite to take blood.

In this new study, researchers wanted to uncover a role for a protein that is well known in humans and other species to co-ordinate the activity of other proteins called Protein Phosphatase 1, also known as PP1.

PP1 regulates how cells multiply through processes known as mitosis and meiosis. However, little is known about the role of PP1 in the malaria parasite, so the team looked at where PP1 is located in the cell and how blocking its activity affects Plasmodium growth.

Professor Tewari said: “We have uncovered how PP1 regulates the unusual multiplication process the male sex cell undergoes within the mosquito gut. We found that this key molecule is present at specific points during cell multiplication and only on certain parts of the chromosome called the kinetochore, which is crucial for chromosome separation.

“We wanted to understand how and when does the parasite use this protein for its rapid phase of cell multiplication in the male sex cells that gives rise to parasite male sperm cell? How does PP1 orchestrate parasite development during this stage? It is fascinating how a single cell can carry out DNA multiplication so rapidly, and we need to understand how it does this. This work commenced with Dr David Guttery when he was working at University of Nottingham and is presently working at University of Leicester.“

By analysing the parasite at specific stages present in the mosquito gut, the team of researchers found a new role for PP1 during Plasmodium mitosis in the male sex cells. They also found a surprising potential role in how the parasite changes from a round cell (zygote) into a banana-shaped ookinete, which is primed for moving through the mosquito gut ready for the next stage of development for transmission.

Dr Mohammed Zeeshan, from the School of Life Sciences at the University of Nottingham and lead scientist of the study said: “By artificially removing PP1 from the parasite by genetic manipulation, we discovered that this dramatically affects development of the male sex cells and the ookinetes looked very abnormal. When we looked at the location of PP1 during ookinete development, we found there is a very intense localisation at the exact point where the round zygote cell develops a protrusion from its body when it starts to change its shape and become like a banana. This is fascinating, but we need to look into this more to understand how we can use this knowledge to follow drug discovery.”

A full copy of the study can be found here.

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Provided by University of Nottingham

State-of-the-art video microscopy has enabled WEHI researchers to see the molecular details of how malaria parasites invade red blood cells – a key step in the disease.

The researchers used a custom-built lattice light sheet microscope – the first in Australia – to capture high-resolution videos of individual parasites invading red blood cells, and visualise the molecular and cellular changes that occur throughout this process. The research has provided critical new information about malaria parasite biology that may have applications for the development of much-needed new antimalarial medicines.

The research, which was published today in Nature Communications, was led by Ms Cindy Evelyn, Dr Niall Geoghegan, Dr Lachlan Whitehead, Professor Alan Cowman and Dr Kelly Rogers.

At a glance

• An advanced microscopy platform, called lattice light sheet microscopy, has been used to obtain detailed, real-time videos of the malaria parasite invading red blood cells.
• The research has revealed key steps in the parasite invasion process, which is a critical point of the malaria life-cycle and underpins many symptoms of malaria.
• The team’s discoveries could advance the development of much-needed new antimalarial medicines.

Focusing on a deadly parasite

Malaria is a mosquito-borne disease that kills around 400,000 people globally each year. Many of the serious symptoms of malaria occur because of the invasion and growth of the Plasmodium parasite in an infected person’s red blood cells, said Dr Rogers, who is the head of WEHI’s Centre for Dynamic Imaging.

“Understanding in better detail exactly how the parasite invades red blood cells may reveal new ways to stop this stage of the parasite life cycle, potentially leading to much-needed new therapies,” she said.

“We used microscopy – specifically a state-of-the-art approach, lattice light sheet microscopy (LLSM) – to follow the intricate cellular and molecular changes that occur when the malaria parasite invades red blood cells. We captured the first ever high-resolution, real time and dynamic views of the parasite in action.”

Ms Evelyn, who began the research as an Honours student, said the research revealed many previously unrecognised aspects of parasite invasion.

“The videos we recorded showed the ‘push and pull’ interactions as the parasite landed on the red blood cell, and then entered the cell in an enclosed chamber – called a vacuole – where it grew and multiplied.There has long been contention in the field about whether the vacuole is derived from the parasite or the host cell. Our research resolved this question, revealing it was created from the red blood cell’s membrane,” she said.

Most antimalarial therapies and vaccines target the initial binding of malaria to red blood cells.

“By visualising these processes in more detail, our research may contribute in several ways to the development of new antimalarial therapies. For example, now that we know that the parasite vacuole relies on components of the red blood cell membrane, it might be possible to target these components with medicines to disrupt the parasite life cycle. This host-directed approach could be one way to bypass the malaria parasite’s propensity to rapidly develop drug resistance,” Dr Rogers said.

“LLSM may also have applications for observing the specific steps of parasite invasion that are blocked by potential new drugs – an area we are now very interested in pursuing.” 

New views of cells

LLSM is an advanced imaging technology that enables researchers to visualise cells and organs in unprecedented detail and in real time. Dr Geoghegan said LLSM had changed how cells could be studied.

“In the past, the choice of microscope for an experiment had to be a compromise between capturing a lower resolution video, which revealed dynamic processes like shape changes or movement, and capturing much higher-definition still images, which provided much more detail about how the cells and molecules are functioning,” he said.

“LLSM allows us to obtain high-resolution videos of cells, which has been a game-changer for many fields of biological research. We custom built a LLSM at WEHI – the first version of this technology in Australia. This groundbreaking microscopy has enabled us to progress multiple areas of research, including this malaria study. To achieve this, we brought together a multidisciplinary team with expertise in physics, engineering and biology – and the results of this current study have vindicated our approach.”
 
The research was supported by the Australian National Health and Medical Research Council, an EMBO Long Term Fellowship, a Sir Henry Wellcome Fellowship and the Victorian Government.

WEHI Authors: Dr Niall Geoghegan, Ms Cindy Evelyn, Dr Lachlan Whitehead, Dr Michal Pasternak, Ms Phoebe McDonald, Mr Tony Triglia, Dr Danushka Marapana, Ms Jennifer Thompson, Dr Michael Mlodzianoski, Dr Julie Healer, Professor Alan Cowman, Dr Kelly Rogers

Featured image: Imaging researchers (from left) Ms Cindy Evelyn, 
Dr Niall Geoghegan, Dr Kelly Rogers and
Dr Lachlan Whitehead © WEHI

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Reference: Geoghegan, N.D., Evelyn, C., Whitehead, L.W. et al. 4D analysis of malaria parasite invasion offers insights into erythrocyte membrane remodeling and parasitophorous vacuole formation. Nat Commun 12, 3620 (2021). https://doi.org/10.1038/s41467-021-23626-7

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Provided by WEHI

Researchers have mapped in fine detail the genetic changes malaria parasites go through as they prepare to infect people.

The atlas maps the malaria parasite Plasmodium falciparum in unprecedented cellular detail as it develops inside a mosquito and prepares to infect humans through a bite. This detailed investigation could lead to new ways to block key stages in the parasite’s development and prevent transmission through future drugs or vaccines.

“Our new data have clear implications for malaria control, both in terms of drugs that kill the parasite as it moves between stages and protective vaccines.”

— Dr Eliana Real

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Mosquitoes are increasingly resistant to pesticides, and the parasite that causes malaria is also becoming increasingly resistant to antimalarial drugs. This has created an urgent need for new ways to fight malaria, which in 2019 caused an estimated 229 million cases and 409,000 deaths, most of which were young children in sub-Saharan Africa.

To reinvigorate efforts in drug or vaccine discovery, a team from the lab of Professor Jake Baum at Imperial College London and the lab of Dr Mara Lawniczak from the Wellcome Sanger Institute have examined the human malaria parasite P. falciparum in unprecedented detail. Their results are published today in Nature Communications.

Mapping genes

P. falciparum develops in the midgut of a mosquito before travelling to the mosquito’s saliva glands, ready to infect a human when the bug bites. During these phases, the parasite goes through many stages important for its development and ability to transmit, including changing into different forms.

The team tracked how these stages were controlled by analysing the activity of genes throughout the process. They isolated the different forms of the parasite and produced 1467 ‘transcriptomes’ – maps of which genes in single cells are turned on or off during the different stages.

When genes are turned on, they instruct the cell to make different proteins and drive developmental changes, such as causing the parasite to exit the midgut and colonise the salivary gland of the mosquito, or to travel through human cells to reach the liver, where the parasite prepares to invade more human cells.

Knowing how these processes work in detail at the cellular level reveals to researchers new targets that could be blocked to stop development, preventing transmission of the parasite.

New strategies

Dr Eliana Real, from the Department of Life Sciences at Imperial, said: “Being directly based on the human-infective parasite, our new data have clear implications for malaria control, which has an increasing focus on transmission blocking strategies both in terms of drugs that kill the parasite as it moves between stages and protective vaccines.

“Understanding how parasites behave transcriptionally within the mosquito vector provides a foundation from which new strategies will surely arise.”

https://giphy.com/embed/Xhc5sljZa9axeGKxzM

via GIPHY

^{3D plot of gene activity in human malaria parasites at different stages of the life cycle}

As well as surveying the whole transmission cycle of the parasite, the team focused on what is called the sporozoite stage: the form released into the human skin during a mosquito bite.

They sorted parasites from within the mosquito during their development, and isolated sporozoites after an infectious bite as they interact with human skin cells. In doing so, they were able to find specific patterns of gene expression that define each of the critical stages in these processes.

Dr Virginia Howick, previously from the Wellcome Sanger Institute and now based at the University of Glasgow, said: “This fine granularity enables us to trace sporozoite developmental processes and to propose new mechanistic targets essential for each step and future vaccine targets for blocking malaria infection.”

Invaluable resource

The team were also able to compare their data with a similar set from the related parasite Plasmodium berghei, a rodent malaria parasite that is often used as a model for studying malaria disease in the lab. This showed which genes are common between species, and which are specific to the human version of the parasite.

Dr Farah Dahalan, from the Department of Life Sciences at Imperial, said: “This level of gene surveillance at the individual parasite level throughout its life cycle will provide an invaluable resource for researchers to discover previously unexplored elements of Plasmodium cell biology, comparative Plasmodium species biology and the development of control methods that target particular pathways or lay the foundations for improving vaccines.”

The researchers have made all their data available on an interactive website, where the transcriptional profile of any gene across any stage of the Plasmodium life cycle can be easily and freely viewed.

The research was funded by Wellcome, the Bill & Melinda Gates Foundation, and the Royal Society.

‘A single-cell atlas of Plasmodium falciparum transmission through the mosquito,’ by Eliana Real et al. is published in Nature Communications.

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Provided by Imperial College London

Researchers at the Francis Crick Institute and the Latvian Institute of Organic Synthesis have designed a drug-like compound which effectively blocks a critical step in the malaria parasite life cycle and are working to develop this compound into a potential first of its kind malaria treatment. 

While drugs and mosquito control have reduced levels of malaria over recent decades, the parasite still kills over 400,000 people every year, infecting many more. Worryingly, it has now developed resistance to many existing antimalarial drugs, meaning new treatments that work in different ways are urgently needed.

“If we can effectively trap malaria in the cell by blocking the parasite’s exit route, we could stop the disease in its tracks and halt its devastating cycle of invading cells.”

— Mike Blackman

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In their research, published in PNAS, the scientists developed a set of compounds designed to stop the parasite being able to burst out of red blood cells, a process vital to its replication and life cycle. They found one compound in particular was highly effective in human cell tests. 

“Malaria parasites invade red blood cells where they replicate many times, before bursting out into the bloodstream to repeat the process. It’s this cycle and build-up of infected red blood cells which causes the symptoms and sometimes fatal effects of the disease,” says Mike Blackman, lead author and group leader of the Malaria Biochemistry Laboratory at the Crick.  

“If we can effectively trap malaria in the cell by blocking the parasite’s exit route, we could stop the disease in its tracks and halt its devastating cycle of invading cells.”

The compound works by blocking an enzyme called SUB1, which is critical for malaria to burst out of red blood cells. Existing antimalarials work by killing the parasite within the cell, so the researchers hope this alternative drug action will overcome the resistance the parasite has acquired. 

Importantly the compound is also able to pass through the membranes of the red blood cell and of the compartment within the cell where the parasites reside. 

The team is continuing to optimise the compound, making it smaller and more potent. If successful, it will need to be tested in further experiments and in animal and human trials to show it is safe and effective, before being made available to people. 

Chrislaine Withers-Martinez, author and researcher in the Malaria Biochemistry Laboratory, says: “Many existing antimalarial drugs are plant derived and while they’re incredibly effective, we don’t know the precise mechanisms behind how they work. Our decades of research have helped us identify and understand pathways crucial to the malaria life cycle allowing us to rationally design new drug compounds based on the structure and mechanism of critical enzymes like SUB1. 

“This approach, which has already been highly successful at finding new treatments for diseases including HIV and Hepatitis C, could be key to sustained and effective malaria control for many years to come.” 

Featured image: The optimised boronic acid inhibitor shown covalently bound into the active site of SUB1. Hydrogen bonds between the inhibitor and the enzyme are shown as dotted lines.- Chrislaine Withers-Martinez

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Provided by Francis Crick Institute