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Nanotechnology against malaria parasites

December 13, 2014 Leave a comment

After maturation, malaria parasites (yellow) are leaving an infected red blood cell and are efficiently blocked by nanomimics (blue). (Fig: Modified with permission from ACS)

Malaria parasites invade human red blood cells, they then disrupt them and infect others. Researchers at the University of Basel and the Swiss Tropical and Public Health Institute have now developed so-called nanomimics of host cell membranes that trick the parasites. This could lead to novel treatment and vaccination strategies in the fight against malaria and other infectious diseases. Their research results have been published in the scientific journal ACS Nano.

For many infectious diseases no vaccine currently exists. In addition, resistance against currently used drugs is spreading rapidly. To fight these diseases, innovative strategies using new mechanisms of action are needed. The  Plasmodium falciparum that is transmitted by the Anopheles mosquito is such an example. Malaria is still responsible for more than 600,000 deaths annually, especially affecting children in Africa (WHO, 2012).

Artificial bubbles with receptors

Malaria parasites normally invade human red  in which they hide and reproduce. They then make the host cell burst and infect new cells. Using nanomimics, this cycle can now be effectively disrupted: The egressing parasites now bind to the nanomimics instead of the red blood cells.

Researchers of groups led by Prof. Wolfgang Meier, Prof. Cornelia Palivan (both at the University of Basel) and Prof. Hans-Peter Beck (Swiss TPH) have successfully designed and tested host cell nanomimics. For this, they developed a simple procedure to produce polymer vesicles – small artificial bubbles – with host cell receptors on the surface. The preparation of such polymer vesicles with water-soluble host receptors was done by using a mixture of two different block copolymers. In aqueous solution, the nanomimics spontaneously form by self-assembly.

Blocking parasites efficiently

Usually, the malaria parasites destroy their host cells after 48 hours and then infect new . At this stage, they have to bind specific host cell receptors. Nanomimics are now able to bind the egressing parasites, thus blocking the invasion of new cells. The parasites are no longer able to invade host cells, however, they are fully accessible to the immune system.

The researchers examined the interaction of nanomimics with malaria parasites in detail by using fluorescence and electron microscopy. A large number of nanomimics were able to bind to the parasites and the reduction of infection through the nanomimics was 100-fold higher when compared to a soluble form of the host cell receptors. In other words: In order to block all , a 100 times higher concentration of soluble host  is needed, than when the receptors are presented on the surface of nanomimics.

“Our results could lead to new alternative treatment and vaccines strategies in the future”, says Adrian Najer first-author of the study. Since many other pathogens use the same  receptor for invasion, the nanomimics might also be used against other . The research project was funded by the Swiss National Science Foundation and the NCCR “Molecular Systems Engineering”.

More information: Adrian Najer, Dalin Wu, Andrej Bieri, Françoise Brand, Cornelia G. Palivan, Hans-Peter Beck, and Wolfgang Meier. “Nanomimics of Host Cell Membranes Block Invasion and Expose Invasive Malaria Parasites.” ACS Nano, Publication Date (Web): November 29, 2014 | DOI: 10.1021/nn5054206

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Common Parasite May Trigger Suicide Attempts

September 5, 2012 Leave a comment

A parasite thought to be harmless and found in many people may actually be causing subtle changes in the brain, leading to suicide attempts.

New research appearing in the August issue of The Journal of Clinical Psychiatry adds to the growing work linking an infection caused by the Toxoplasma gondii parasite to suicide attempts. Michigan State University’s Lena Brundin was one of the lead researchers on the team.

About 10-20 percent of people in the United States have Toxoplasma gondii, or T. gondii, in their bodies, but in most it was thought to lie dormant, said Brundin, an associate professor of experimental psychiatry in MSU’s College of Human Medicine. In fact, it appears the parasite can cause inflammation over time, which produces harmful metabolites that can damage brain cells.

“Previous research has found signs of inflammation in the brains of suicide victims and people battling depression, and there also are previous reports linking Toxoplasma gondii to suicide attempts,” she said. “In our study we found that if you are positive for the parasite, you are seven times more likely to attempt suicide.”

The work by Brundin and colleagues is the first to measure scores on a suicide assessment scale from people infected with the parasite, some of whom had attempted suicide.

The Toxoplasma gondii parasite has been linked to inflammation in the brain, damaging cells. Image adapted from MSU press release image. (up)

The results found those infected with T. gondii scored significantly higher on the scale, indicative of a more severe disease and greater risk for future suicide attempts. However, Brundin stresses the majority of those infected with the parasite will not attempt suicide: “Some individuals may for some reason be more susceptible to develop symptoms,” she said.

“Suicide is major health problem,” said Brundin, noting the 36,909 deaths in 2009 in America, or one every 14 minutes. “It is estimated 90 percent of people who attempt suicide have a diagnosed psychiatric disorder. If we could identify those people infected with this parasite, it could help us predict who is at a higher risk.”

T. gondii is a parasite found in cells that reproduces in its primary host, any member of the cat family. It is transmitted to humans primarily through ingesting water and food contaminated with the eggs of the parasite, or, since the parasite can be present in other mammals as well, through consuming undercooked raw meat or food.

Brundin has been looking at the link between depression and inflammation in the brain for a decade, beginning with work she did on Parkinson’s disease. Typically, a class of antidepressants called selective serotonin re-uptake inhibitors, or SSRIs, have been the preferred treatment for depression. SSRIs are believed to increase the level of a neurotransmitter called serotonin but are effective in only about half of depressed patients.

Brundin’s research indicates a reduction in the brain’s serotonin might be a symptom rather than the root cause of depression. Inflammation, possibly from an infection or a parasite, likely causes changes in the brain’s chemistry, leading to depression and, in some cases, thoughts of suicide, she said.

“I think it’s very positive that we are finding biological changes in suicidal patients,” she said. “It means we can develop new treatments to prevent suicides, and patients can feel hope that maybe we can help them.

“It’s a great opportunity to develop new treatments tailored at specific biological mechanisms.”

References:

Source: Michigan State University press release
Image Source: T. gondii image adapted from Michigan State University press release image
Original Research: Abstract and full paper from MSU (PDF file) for “Toxoplasma gondii Immunoglobulin G Antibodies and Nonfatal Suicidal Self-Directed Violence” by Yuanfen Zhang, MD, PhD; Lil Träskman-Bendz, MD, PhD; Shorena Janelidze, PhD; Patricia Langenberg, PhD; Ahmed Saleh, PhD; Niel Constantine, PhD; Olaoluwa Okusaga, MD; Cecilie Bay-Richter, PhD; Lena Brundin, MD, PhD; and Teodor T. Postolache, MD in Journal of Clinical Psychiatry online July 2012 73(8):1069–1076 doi: 10.4088/JCP.11m07532

Drug Found for Parasite That Is Major Cause of Death Worldwide


Research by a collaborative group of scientists from UC San Diego School of Medicine, UC San Francisco and Wake Forest School of Medicine has led to identification of an existing drug that is effective against Entamoeba histolytica. This parasite causes amebic dysentery and liver abscesses and results in the death of more than 70,000 people worldwide each year.

(up) Entamoeba histolytica cyst. (Credit: UC San Diego School of Medicine)

Using a high-throughput screen for drugs developed by the research team, they discovered that auranofin — a drug approved by the US Food and Drug Administration 25 years ago for rheumatoid arthritis — is very effective in targeting an enzyme that protects amebae from oxygen attack (thus enhancing sensitivity of the amebae to reactive oxygen-mediated killing).

The results of the work, led by Sharon L. Reed, MD, professor in the UCSD Departments of Pathology and Medicine and James McKerrow, MD, PhD, professor of Pathology in the UCSF Sandler Center for Drug Discovery, will be published in the May 20, 2012 issue of Nature Medicine.

Entamoeba histolytica is a protozoan intestinal parasite that causes human amebiasis, the world’s fourth leading cause of death from protozoan parasites. It is listed by the National Institutes of Health as a category B priority biodefense pathogen. Current treatment relies on metronidazole, which has adverse effects, and potential resistance to the drug is an increasing concern.

“Because auranofin has already been approved by the FDA for use in humans, we can save years of expensive development,” said Reed. “In our studies in animal models, auranofin was ten times more potent against this parasite than metronidazole.”

In a mouse model of amebic colitis and a hamster model of amebic liver abscess, the drug markedly decreased the number of parasites, damage from inflammation, and size of liver abscesses.

“This new use of an old drug represents a promising therapy for a major health threat, and highlights how research funded by the National Institutes of Health can benefit people around the world,” said Reed. The drug has been granted “orphan-drug” status (which identifies a significant, newly developed or recognized treatment for a disease which affects fewer than 200,000 persons in the United States) and UC San Diego hopes to conduct clinical trials in the near future.

This work was supported by the Sandler Foundation and US National Institute of Allergy and Infectious Diseases grant 5U01AI077822-02, with additional support from R01 GM050389.

Story Source:

The above story is reprinted from materials provided byUniversity of California, San Diego Health Sciences, via Newswise.

Note: Materials may be edited for content and length. For further information, please contact the source cited above.

Journal Reference:

Anjan Debnath, Derek Parsonage, Rosa M Andrade, Chen He, Eduardo R Cobo, Ken Hirata, Steven Chen, Guillermina García-Rivera, Esther Orozco, Máximo B Martínez, Shamila S Gunatilleke, Amy M Barrios, Michelle R Arkin, Leslie B Poole, James H McKerrow, Sharon L Reed. A high-throughput drug screen for Entamoeba histolytica identifies a new lead and targetNature Medicine, 2012; DOI: 10.1038/nm.2758

Notre Dame researchers report fundamental malaria discovery

February 11, 2012 1 comment

A team of researchers led by Kasturi Haldar and Souvik Bhattacharjee of the University of Notre Dame’s Center for Rare and Neglected Diseases has made a fundamental discovery in understanding how malaria parasites cause deadly disease.

The researchers show how parasites target proteins to the surface of the red blood cell that enables sticking to and blocking blood vessels. Strategies that prevent this host-targeting process will block disease.

The research findings appear in the Jan. 20 edition of the journal Cell, the leading journal in the life sciences. The study was supported by the National Institutes of Health.

Malaria is a blood disease that kills nearly 1-3 million people each year. It is caused by a parasite that infects red cells in the blood. Once inside the cell, the parasite exports proteins beyond its own plasma membrane border into the blood cell. These proteins function as adhesins that help the infected red blood cells stick to the walls of blood vessels in the brain and cause cerebral malaria, a deadly form of the disease that kills over half a million children each year.

In all cells, proteins are made in a specialized cell compartment called the endoplasmic reticulum (ER) from where they are delivered to other parts of the cell. Haldar and Bhattacharjee and collaborators Robert Stahelin at the Indiana University School of Medicine – South Bend (who also is an adjunct faculty member in Notre Dame’s Department of Chemistry and Biochemistry), and David and Kaye Speicher at the University of Pennsylvania’s Wistar Institute discovered that for host-targeted malaria proteins the very first step is binding to the lipid phosphatidylinositol 3-phosphate, PI(3)P, in the ER.

This was surprising for two reasons. Previous studies suggested an enzyme called Plasmepsin V that released the proteins into the ER was also the export mechanism. However, Haldar, Bhattacharjee and colleagues discovered that binding to PI(3)P lipid which occurs first is the gate keeper to control export and that export can occur without Plasmepsin V action. Further, in higher eukaryotic cells (such as in humans), the lipid PI(3)P is not usually found within the ER membrane but rather is exposed to the cellular cytoplasm.

Haldar and Bhattacharjee are experts in malaria parasite biology and pathogenesis. Stahelin is an expert in PI(3)P lipid biology, and David and Kaye Speicher are experts in proteomics and a method called mass spectrometry.

Story Source:

The above story is reprinted (with editorial adaptations by MedicalXpress staff) from materials provided by University of Notre Dame (news : web)

Scientists Characterize Protein Essential to Survival of Malaria Parasite

January 11, 2012 Leave a comment

A biology lab at Washington University has just cracked the structure and function of a protein that plays a key role in the life of a parasite that killed 655,000 people in 2010.

A cartoon based on the electron density map makes it easier to see the protein’s structure and figure out how it works. The enzyme’s job is to add a methyl group — three times — to a starting molecule as part of a process for making cell membranes. In this cartoon, the phosphate is a stand-in for the starting molecule and the green molecule is the one that donates a methyl group. Both are positioned in the active site of the enzyme, the pocket where the chemistry takes place. (Credit: Jez)

The protein is an enzyme that Plasmodium falciparum, the protozoan that causes the most lethal form of malaria, uses to make cell membrane.

The protozoan cannot survive without this enzyme, but even though the enzyme has many lookalikes in other organisms, people do not make it. Together these characteristics make the enzyme an ideal target for new antimalarial drugs.

The research was published in the January 6 issue of the Journal of Biological Chemistry (JBC) as “Paper of the Week” for that issue.

The work also will be featured in ASBMB Today (the newsletter of the American Society for Biological Molecular Biology, which publishes JBC).

Sweating the cold room

The protein’s structure might have remained an enigma, had it not been the “unreasonable optimism” of Joseph Jez, PhD, associate professor of biology in Arts & Sciences, which carried his team through a six-year-long obstacle course of failures and setbacks.

“What my lab does is crystallize proteins so that we can see what they look like in three dimensions,” Jez says. “The idea is that if we know a protein’s structure, it will be easier to design chemicals that would target the protein’s active site and shut it down,” Jez says.

The latest discovery is the culmination of a project that began years before when Jez was working at the Danforth Plant Science Center in St. Louis and collaborating with scientists at the local biotech startup Divergence. “At the time, C. elegans had just been sequenced and the Divergence scientists were looking at using it as an easy model to work out the biochemistry of parasitic nematodes,” Jez says.

C. elegans is a free-living nematode, or microscopic roundworm, but many nematodes are parasitic and cause disease in plants, livestock and people.

During this project, Lavanya Palavalli, a summer intern working with Jez, crystallized the C. elegans version of the enzyme. The job of the enzyme, phosphoethanolamine methyltransferase, thankfully abbreviated to PMT, is to add methyl groups to a starting molecule, phosophoethanolamine.

“When Soon Goo Lee later took up the project,” says Jez, “the plan was to try to grow better crystals of the C. elegans protein, ones good enough to get readable X-ray diffraction patterns.

Two years later, the crystals were looking better but still not good enough.

So Jez suggested that Lee go after homologous (look-alike) proteins in other organisms. “Even though the proteins are homologous, each has a different amino acid sequence and so will behave differently in the crystallizations,” Jez says. “Lee went from working with two C. elegans proteins to three plant proteins, two other nematode proteins and then the Plasmodium protein,” Jez says.

“He took all six of those PMT versions into the crystallization trials to maximize his odds,” Jez says.

“To crystallize a protein,” Jez says, “we put a solution of a salt or something else that might work as a desiccant in the bottom of a small well. And then we put a drop of our liquid protein on a microscope cover slip and flip it over the top of the well, so the drop of protein is hanging upside down in the well.”

“What we’re trying to do is to slowly withdraw water from the protein. It’s exactly like making rock candy, only in that case, the string hanging into the jar of sugar solution helps to withdraw water,” he says.

The difference is that sugar wants to form crystals and proteins are reluctant to do so.

“There are 24 wells to a tray, and we usually screen 500 wells per protein at first,” Jez says. “Lee had eight proteins and so his first pass was to screen 4,000 conditions. And then he had to try different combinations of ligands to the proteins and crystallize those. This is why it took a few years to finally get where he needed to go.”

Road trip!

 The scientists need crystals — preferably nice, big ones — to stick in the path of an X-ray beam at Argonne National Laboratory in Chicago. (If the crystal is a good one, and all the atoms are lined up in a repeating array, the scattered X-rays will produce a clear pattern of spots.)

Embedded in that pattern is the mathematical information needed to back-calculate to the position of the atoms in the protein, a process a bit like throwing a handful of pebbles in a lake and then calculating where they landed by the pattern of waves arriving at the shoreline.

Lee got the PMT from Haemonchus contortus to crystallize first, but there were technical issues with the diffraction pattern that would have made solving it technically and computationally very demanding.

“When the Plasmodium enzyme finally crystallized, Soon got four crystals kind of stacked on top of each other and each of them was paper thin,” Jez says.

“I never thought it would work, but we took them to Argonne anyway and he actually did surgery under the microscope and cracked off a little tiny piece of it.”

To everyone’s surprise, he got a clean diffraction pattern from the crystal. “Because the Plasmodium enzyme was the smallest one and the easiest to work on, we pushed that one first,” Jez says.

The moment of truth

“Once we had a Plasmodium crystal that was diffracting really well, we could try back-calculating to see whether we could extract the atom positions from the data,” Jez says.

After the computer finished its calculations, Lee clicked a mouse button to see the results, which would reveal whether his years of work finally would pay off.

When Lee clicked the mouse, he got an electron density map in exceptionally sharp focus.

“When you see a map like that, it’s like suddenly the wind has kicked up and you’re sailing free,” Jez says, “because there’s this moment, like, before you click that button, no one has ever seen how this protein is put together in three dimensions. You’re the first person to ever see it.

“The irony of it is we got such good quality diffraction pattern and electron density maps off such an ugly crystal,” he says.

Lock and load

“Once you have the electron density map, the task is to build a structure that matches the amino acid sequence of the protein,” Jez says.

“The first thing you do is put in the amino acid backbones and connect them together to form a chain. It’s like having a long thread, each inch of which is an amino acid, and your job is to take that thread and move it in three dimensions through that electron density map.”

The next step is to add the side chains that make one amino acid different from another, Jez says. “The amino acid sequence is known,” he says. “Your goal is to match the way you string together the amino acids in the electron density map to that sequence.”

“Once you have the overall structure, you can start to figure out how the enzyme works. The PMT enzyme is trying to join two molecules,” Jez says. “To do that, it has to lock them in place so that the chemistry can happen, and then it has to let go of them.

“We think the protein has a lid that opens and closes,” he says. “The active site stays open until the substrates enter, and then the lid clamps down, and when it clamps down it actually puts the substrates together.”

Calling Bill and Melinda Gates

Not only do infections by Plasmodium falciparum cause the most severe form of malaria, about 40 percent of the human population lives in areas where the parasite is endemic. Moreover, drugs that used to be effective against malaria are beginning to fail, in part because widespread drug counterfeiting has led to resistance.

New anti-malarial drugs are desperately needed, and the PMT protein is an ideal target. If PMT is disabled, the protozoan can’t make cell membranes and it dies. Moreover, a drug that would kill Plasmodium might have minimal side effects on patients.

Although the process of identifying compounds that would target PMT is in the early stages, a handful of anti-parasitical compounds used to treat diseases are known to block PMT as well.

As for Lee, he has had a hard go of it, but now things are breaking his way. Plasmodium PMT is giving up its secrets, and the plant and nematode PMTs are coming along as well.

When he clicked the mouse button and a clean electron density map came up, he says, it was like seeing “the light at the end of a five-year-long tunnel.”

Story Source:

The above story is reprinted (with editorial adaptations by ScienceDaily staff) from materials provided by Washington University in St. Louis.

Journal Reference:

S. G. Lee, Y. Kim, T. D. Alpert, A. Nagata, J. M. Jez.Structure and Reaction Mechanism of Phosphoethanolamine Methyltransferase from the Malaria Parasite Plasmodium falciparum: AN ANTIPARASITIC DRUG TARGETJournal of Biological Chemistry, 2011; 287 (2): 1426 DOI:10.1074/jbc.M111.315267

‘Virus-eater’ discovered in Antarctic lake


First of the parasitic parasites to be discovered in a natural environment points to hidden diversity.

Viruses from Organic lake, including the virophage (bottom left) and its prey (top). From reference 1 (left)

 

A genomic survey of the microbial life in an Antarctic lake has revealed a new virophage — a virus that attacks viruses. The discovery suggests that these life forms are more common, and have a larger role in the environment, than was once thought.

An Australian research team found the virophage while surveying the extremely salty Organic Lake in eastern Antarctica. While sequencing the collective genome of microbes living in the surface waters, they discovered the virus, which they dubbed the Organic Lake Virophage (OLV).

The OLV genome was identified nestling within the sequences of phycodnaviruses — a group of giant viruses that attack algae. Evidence of gene exchange, and possible co-evolution, between the two suggests that OLV preys on the phycodnavirus. Although OLV is the dominant virophage in the lake, the work suggests others might be present.

By killing phycodnaviruses, the OLV might allow algae to thrive. Ricardo Cavicchioli, a microbiologist at the University of New South Wales in Sydney, Australia, and his colleagues found that mathematical models of the Organic Lake system that took account of the virophage’s toll on its host showed lower algal mortality and more blooms during the lake’s two ice-free summer months.

“Our work reveals not only an amazing diversity in microbial life in this lake, but also how little we understand about the complexity of the biological functions at work,” says Cavicchioli. The findings are published in the Proceedings of the National Academies of Science1.

Giant killer

Another virophage described this month has similar ecological effects. The marine Mavirus attacks the giant Cafeteria roenbergensis virus, which preys on Cafeteria roenbergensis, one of the world’s most widespread species of zooplankton2.

“The Mavirus is able to rescue the infected zooplankton — which, in a way, confers immunity from infection,” says Curtis Suttle, a marine microbiologist at the University of British Columbia in Vancouver, Canada, and leader of the team that discovered the Mavirus.

“We unknowingly had Mavirus in culture with our Cafeteria system since the early 1990s,” says Suttle. But the virophage was not identified until the Cafeteria genome was sequenced.

The Mavirus genome is similar to DNA sequences called eukaryotic transposons, which insert themselves within the genomes of multicellular organisms such as plants and animals. These ‘jumping genes’ may be descended from a virophage, says Suttle. “One can imagine evolutionary pressure for hosts to somehow cultivate virophages to protect themselves from infection by giant viruses,” he says.

French Sputnik

The first virophage, dubbed Sputnik, was discovered in a water-cooling tower in Paris in 20083.

“We have been waiting for others to find virophages, to confirm our discovery wasn’t an artefact,” says Christelle Desnues, a microbiologist at the National Centre of Scientific Research in Marseilles, France, and a member of the team that described Sputnik. She now anticipates “an exponential discovery of virophages”.

The hosts of all three known virophages belong to a group of giant viruses known as nucleocytoplasmic large DNA viruses (NCLDV). “NCLDV viruses have large and complex genomes that allow them to incorporate the smaller virophages, something smaller viruses may not be able to do,” says Desnues.

The OLV was discovered when Cavicchioli’s graduate student, Sheree Yau, noticed that some of the sequences from microbes in Organic Lake were similar to those encoding Sputnik’s protein shell. Mavirus has similar sequences, so the trend might help to identify other virophages.

OLV, or virophages like it, may be widespread. The gene for its protein shell matches sequences already found in a host of other aquatic environments, including nearby Ace Lake in Antarctica, a saline lagoon in the Galapagos, an oceanic upwelling zone near the Galapagos, an estuary in New Jersey, and a freshwater lake in Panama.

The high number of matches reflects the fact that the OLV is the first virophage to be found in its natural environment, says Federico Lauro, also a molecular biologist at the University of New South Wales and a co-author of the paper.

Organic Lake, formed 6,000 years ago when sea levels were higher, is a natural laboratory, says Lauro. “These marine-derived lakes are great labs to work in because they are isolated, yet dynamic systems.”

 

References

Yau, S. et al. Proc. Natl Acad. Sci. USA advance online publication doi: 10.1073/pnas.1018221108 (2011).

Fischer, M. G. et al. Science advance online publication doi: 10.1126/science.1199412 (2011).

La Scola, B. et al. Nature 455, 100-104 (2008). | ArticleISIChemPort |