In mice, vaccine stops urinary tract infections linked to catheters

To adhere to catheters and start urinary tract infections, bacteria extend microscopic fibers with sticky proteins at their ends. Scientists have developed a vaccine that blocks the EbpA protein, visible as a white bulge above, and stops infections in mice. Credit: John Heuser

The most common type of hospital-associated infection may be preventable with a vaccine, new research in mice suggests.

The experimental vaccine, developed by researchers at Washington University School of Medicine in St. Louis, prevented urinary tract infections associated with catheters, the tubes used in hospitals and other care facilities to drain urine from a patient's bladder.

Each day a catheter is present in the urethra and the bladder, the risk of urinary tract infection increases. Nearly every patient who has a catheter for more than 30 days acquires a urinary tract infection. The infections make urination painful and can damage the bladder. If untreated, bacteria can cross into the bloodstream and cause sepsis, a potentially life-threatening complication.

"Catheter-associated urinary tract infections are very common," said first author Ana Lidia Flores-Mireles, PhD, a postdoctoral research associate at the School of Medicine. "Antibiotic resistance is increasing rapidly in the bacteria that cause these infections, so developing new treatments is a priority."

The study is available online Sept. 17 in Science Translational Medicine.

Manufacturers typically coat catheters with antibiotics to reduce the risk of infection. But Flores-Mireles and her colleagues in the laboratory of Scott Hultgren, PhD, showed that inserting catheters into the bladder provokes an inflammatory response that results in the catheter being covered with fibrinogen, a blood-clotting protein.

Fibrinogen shields bacteria from the antibiotics and provides bacteria with a landing pad to adhere to and food to consume as they establish an infection, the research revealed.

"The bacteria use long, thin hairs known as pili to anchor themselves to the fibrinogen, and then they can start to form biofilms, which are slimy coatings on the surface of the catheter composed of many bacteria," said co-author Michael Caparon Jr., PhD, professor of molecular microbiology. "The biofilms protect the bacteria from antibiotics and immune cells, further prevent them from being washed from the body by the flow of urine, and make it possible for bacteria to seed the lining of the bladder with infections."

The urethra and bladder of a mouse are too small to insert a full catheter into, but the scientists showed that surgically implanting a small segment of catheter into the bladder via the urethra increased vulnerability to infection in a similar fashion.

Working with Enterococcus faecalis, a common cause of catheter-associated urinary tract infections, Flores-Mireles showed that a protein on the end of the pili, EbpA, binds to fibrinogen and makes it possible for the bacteria to begin forming biofilms.

When Flores-Mireles prevented the bacteria from making EbpA, they couldn't start infections.

"This protein is like the anchor of a boat," she said. "Without the anchor, the infection is at the mercy of the waves and gets washed away."

Next, the researchers injected the mice with a vaccine containing EbpA. The vaccine caused the animal's immune systems to produce antibodies that blocked EbpA and stopped the infectious process.

The scientists are testing to see if the vaccine helps mice clear established infections of E. faecalis. They also are working to develop a monoclonal antibody that blocks EbpA to prevent catheter-associated infections in the urinary tract and elsewhere in the body.

"We took a closer look at this protein and found that one-half of it is essential for binding to fibrinogen to induce infections," Flores-Mireles said. "The segment of genetic code that makes this part of the protein is also found in the genes of many other bacteria that cause urinary tract infections, so a vaccine, antibody or drug that blocks this part of the protein may help prevent other infections linked to catheters in the urinary tract and in other parts of the body."

Source: Washington University in St. Louis

Trial confirms Ebola vaccine candidate safe, equally immunogenic in Africa

"This is the first study to show comparable safety and immune response of an experimental Ebola vaccine in an African population," says lead author Dr Julie Ledgerwood. "This is particularly encouraging because those at greatest risk of Ebola live primarily in Africa, and diminished vaccine protection in African populations has been seen for other diseases." Credit: © nito / Fotolia

Two experimental DNA vaccines to prevent Ebola virus and the closely related Marburg virus are safe, and generated a similar immune response in healthy Ugandan adults as reported in healthy US adults earlier this year. The findings, from the first trial of filovirus vaccines in Africa, are published in The Lancet.

"This is the first study to show comparable safety and immune response of an experimental Ebola vaccine in an African population," says lead author Dr Julie Ledgerwood from the National Institutes of Allergy and Infectious Diseases (NIAID) at the National Institutes of Health, USA. "This is particularly encouraging because those at greatest risk of Ebola live primarily in Africa, and diminished vaccine protection in African populations has been seen for other diseases."

Scientists from the NIAID developed the DNA vaccines that code for Ebola virus proteins from the Zaire and Sudan strains and the Marburg virus protein. The vaccines contain the construction plans for the proteins on the outer surface of the virus. Immune responses against these proteins have shown to be highly protective in non-human primate models.

In this phase 1 trial, the Makerere University Walter Reed Program enrolled 108 healthy adults aged between 18 and 50 from Kampala, Uganda between November, 2009 and April, 2010. Each volunteer was randomly assigned to receive an intramuscular injection of either the Ebola vaccine (30 volunteers), Marburg vaccine (30), both vaccines (30), or placebo (18) at the start of the study, and again 4 weeks and 8 weeks later.

The vaccines given separately and together were safe and stimulated an immune response in the form of neutralising antibodies and T-cells against the virus proteins. Four weeks after the third injection, just over half of the volunteers (57%; 17 of 30) had an antibody response to the Ebola Zaire protein as did 14 of 30 participants who received both the Ebola and Marburg vaccines. 

However, the antibodies were not long-lasting and returned to undetectable levels within 11 months of vaccination.

Both DNA vaccines were well tolerated in Ugandan adults with similar numbers of local and systemic reactions reported in all groups. Only one serious adverse event (neutropenia; low white blood cell count) was reported in a Marburg vaccine only recipient, but was not thought to be vaccine related.

According to Dr Ledgerwood, "These findings have already formed the basis of a more potent vaccine, delivered using a harmless chimpanzee cold virus, which is undergoing trials in the USA, UK, Mali, and Uganda in response to the ongoing Ebola virus outbreak."

Writing in a linked Comment, Dr Saranya Sridhar from the Jenner Institute at the University of Oxford in the UK says, "[This] study deserves to be the focal point around which the broader question of vaccine development, particularly for Africa, must be addressed. With the uncharitable benefit of hindsight in view of the evolving 2014 Ebola outbreak, we must ask ourselves whether a filovirus vaccine should have been in more advanced clinical development. The international response to the present Ebola outbreak is an exemplar of the speed and purpose with which clinical vaccine development can progress and has set the benchmark against which future vaccine development must be judged. This study is the first step on the aspirational road towards the deployment of filovirus vaccines in Africa and must serve to shake the metaphorical cobwebs that can stall our advance towards this destination."

Source: The Lancet

Experimental Ebola vaccine appears safe, prompts immune response

A 39-year-old woman, the first participant enrolled in VRC 207, receives a dose of the investigational NIAID/GSK Ebola vaccine at the NIH Clinical Center in Bethesda, Md. on September 2. Credit: NIAID

An experimental vaccine to prevent Ebola virus disease was well-tolerated and produced immune system responses in all 20 healthy adults who received it in a Phase 1 clinical trial conducted by researchers from the National Institutes of Health. The candidate vaccine, which was co-developed by the NIH's National Institute of Allergy and Infectious Diseases (NIAID) and GlaxoSmithKline (GSK), was tested at the NIH Clinical Center in Bethesda, Maryland. The interim results are reported online in advance of print in the New England Journal of Medicine.

"The unprecedented scale of the current Ebola outbreak in West Africa has intensified efforts to develop safe and effective vaccines, which may play a role in bringing this epidemic to an end and undoubtedly will be critically important in preventing future large outbreaks," said NIAID Director Anthony S. Fauci, M.D. "Based on these positive results from the first human trial of this candidate vaccine, we are continuing our accelerated plan for larger trials to determine if the vaccine is efficacious in preventing Ebola infection."

The candidate NIAID/GSK Ebola vaccine was developed collaboratively by scientists at the NIAID Vaccine Research Center (VRC) and at Okairos, a biotechnology company acquired by GSK. It contains segments of Ebola virus genetic material from two virus species, Sudan and Zaire. The Ebola virus genetic material is delivered by a carrier virus (chimpanzee-derived adenovirus 3 or cAd 3) that causes a common cold in chimpanzees but causes no illness in humans. The candidate vaccine does not contain Ebola virus and cannot cause Ebola virus disease.

The trial enrolled volunteers between the ages of 18 and 50. Ten volunteers received an intramuscular injection of vaccine at a lower dose and 10 received the same vaccine at a higher dose. At two weeks and four weeks following vaccination, the researchers tested the volunteers' blood to determine if anti-Ebola antibodies were generated. All 20 volunteers developed such antibodies within four weeks of receiving the vaccine. Antibody levels were higher in those who received the higher dose vaccine.

The investigators also analyzed the research participants' blood to learn whether the vaccine prompted production of immune system cells called T cells. A recent study by VRC scientist Nancy J. Sullivan, Ph.D., and colleagues showed that non-human primates inoculated with the candidate NIAID/GSK vaccine developed both antibody and T-cell responses, and that these were sufficient to protect vaccinated animals from disease when they were later exposed to high levels of Ebola virus.

The experimental NIAID/GSK vaccine did induce a T-cell response in many of the volunteers, including production of CD8 T cells, which may be an important part of immune protection against Ebola viruses. Four weeks after vaccination, CD8 T cells were detected in two volunteers who had received the lower dose vaccine and in seven of those who had received the higher dose.

"We know from previous studies in non-human primates that CD8 T cells played a crucial role in protecting animals that had been vaccinated with this NIAID/GSK vaccine and then exposed to otherwise lethal amounts of Ebola virus," said Julie E. Ledgerwood, D.O., a VRC researcher and the trial's principal investigator. "The size and quality of the CD8 T cell response we saw in this trial are similar to that observed in non-human primates vaccinated with the candidate vaccine."

There were no serious adverse effects observed in any of the volunteers, although two people who received the higher dose vaccine did develop a briefly lasting fever within a day of vaccination.

Source: NIH/National Institute of Allergy and Infectious Diseases

Researchers ferret out a flu clue

Professor Michael Jennings, Deputy Director of the Institute for Glycomics. Credit: Image courtesy of Griffith University

Research that provides a new understanding as to why ferrets are similar to humans is set to have major implications for the development of novel drugs and treatment strategies.

Published in the journal Nature Communications, the research is a collaboration between Professor Michael Jennings and other researchers from the Institute for Glycomics, Griffith University and collaborators at the University of Queensland and the University of Adelaide.

The team has shown for the first time that ferrets share a mutation that was previously thought to be unique to humans, among the mammals. This helps to explain why the molecular characteristics of ferrets so uniquely mimic human susceptibility, severity and transmission of influenza A virus strains.

Professor Michael Jennings, Deputy Director of the Institute for Glycomics, says these findings open up a completely novel approach to tackling human diseases from influenza through to cancer.

"For over 80 years we've known that ferrets are uniquely susceptible to human influenza A virus, but the precise reason was unknown," Professor Jennings said.

"We have shown that ferrets have a mutation in a gene required to make a crucial sugar called sialic acid. Most animals can make two types of sialic acid. Ferrets, like humans can make only one. Different flu strains have preferences for the type of sialic acid they bind to cause infection. Because ferrets can only make the human form of this sugar, they are naturally "humanized" for the receptors recognised by human strains of the flu virus."

Source: Griffith University

Flu virus key machine: First complete view of structure revealed

The complete structure allows researchers to understand how the polymerase uses host cell RNA (red) to kick-start the production of viral messenger RNA. Credit: © EMBL/P.Riedinger

Scientists looking to understand -- and potentially thwart -- the influenza virus now have a much more encompassing view, thanks to the first complete structure of one of the flu virus' key machines. The structure, obtained by scientists at EMBL Grenoble, allows researchers to finally understand how the machine works as a whole, and could prove instrumental in designing new drugs to treat serious flu infections and combat flu pandemics.

If you planned to sabotage a factory, a recon trip through the premises would probably be much more useful than just peeping in at the windows. Scientists looking to understand -- and potentially thwart -- the influenza virus have now gone from a similar window-based view to the full factory tour, thanks to the first complete structure of one of the flu virus' key machines. The structure, obtained by scientists at the European Molecular Biology Laboratory (EMBL) in Grenoble, France, allows researchers to finally understand how the machine works as a whole. Published in two papers in Nature, the work could prove instrumental in designing new drugs to treat serious flu infections and combat flu pandemics.

The machine in question, the influenza virus polymerase, carries out two vital tasks for the virus. It makes copies of the virus' genetic material -- the viral RNA -- to package into new viruses that can infect other cells; and it reads out the instructions in that genetic material to make viral messenger RNA, which directs the infected cell to produce the proteins the virus needs. Scientists -- including Cusack and collaborators -- had been able to determine the structure of several parts of the polymerase in the past. But how those parts came together to function as a whole, and how viral RNA being fed in to the polymerase could be treated in two different ways remained a mystery.

"The flu polymerase was discovered 40 years ago, so there are hundreds of papers out there trying to fathom how it works. But only now that we have the complete structure can we really begin to understand it," says Stephen Cusack, head of EMBL Grenoble, who led the work.

Using X-ray crystallography, performed at the European Synchrotron Radiation Facility (ESRF) in Grenoble, Cusack and colleagues were able to determine the atomic structure of the whole polymerase from two strains of influenza: influenza B, one of the strains that cause seasonal flu in humans, but which evolves slowly and therefore isn't considered a pandemic threat; and the strain of influenza A -- the fast-evolving strain that affects humans, birds and other animals and can cause pandemics -- that infects bats.

"The high-intensity X-ray beamlines at the ESRF, equipped with state-of-the-art Dectris detectors, were crucial for getting high quality crystallographic data from the weakly diffracting and radiation sensitive crystals of the large polymerase complex," says Cusack. "We couldn't have got the data at such a good resolution without them."

The structures reveal how the polymerase specifically recognises and binds to the viral RNA, rather than just any available RNA, and how that binding activates the machine. They also show that the three component proteins that make up the polymerase are very intertwined, which explains why it has been very difficult to piece together how this machine works based on structures of individual parts.

Although the structures of both viruses' polymerases were very similar, the scientists found one key difference, which showed that one part of the machine can swivel around to a large degree. That ability to swivel explains exactly how the polymerase uses host cell RNA to kick-start the production of viral proteins. The swivelling component takes the necessary piece of host cell RNA and directs it into a slot leading to the machine's heart, where it triggers the production of viral messenger RNA.

Now that they know exactly where each atom fits in this key viral machine, researchers aiming to design drugs to stop influenza in its tracks have a much wider range of potential targets at their disposal -- like would-be saboteurs who gain access to the whole production plant instead of just sneaking looks through the windows. And because this is such a fundamental piece of the viral machinery, not only are the versions in the different influenza strains very similar to each other, but they also hold many similarities to their counterparts in related viruses such as lassa, hanta, rabies or ebola.

The EMBL scientists aim to explore the new insights this structure provides for drug design, as well as continuing to try to determine the structure of the human version of influenza A, because although the bat version is close enough that it already provides remarkable insights, ultimately fine-tuning drugs for treating people would benefit from/require knowledge of the version of the virus that infects humans. And, since this viral machine has to be flexible and change shape to carry out its different tasks, Cusack and colleagues also want to get further snapshots of the polymerase in different states.

"This doesn't mean we now have all the answers," says Cusack, "In fact, we have as many new questions as answers, but at least now we have a solid basis on which to probe further."

Source: European Molecular Biology Laboratory (EMBL)

Study may help slow the spread of flu

A false color image of an influenza virus particle, or “virion.” Credit: Centers for Disease Control/Cynthia Goldsmith

An important study conducted in part at the Department of Energy's SLAC National Accelerator Laboratory may lead to new, more effective vaccines and medicines by revealing detailed information about how a flu antibody binds to a wide variety of flu viruses.

The flu virus infects millions of people each year. While for most this results in an unproductive and uncomfortable week or two, the flu also contributes to many deaths in the average flu season. And while vaccines are effective in preventing the flu, they require almost yearly reformulation to keep up with the constantly changing virus.

At SSRL and APS, a team of researchers from The Scripps Research Institute, Fujita Health University and Osaka University studied both samples of flu virus components and an anti-flu antibody. The antibody, called F045-092, was already known to neutralize the flu by connecting to the region of the flu virus that binds to host cells, so it can no longer bind to its target and cause infection.

"There are patches of the virus that are more hypervariable than others," said Peter Lee, a postdoctoral research associate at The Scripps Research Institute and first author of the paper. "But the flu always binds to host cells within the same region, and so that binding site needs to be functionally conserved. That makes it a site of vulnerability."

The team used the X-ray beams at SLAC's Stanford Synchrotron Radiation Lightsource (SSRL) and Argonne National Laboratory's Advanced Photon Source (APS), both DOE Office of Science User Facilities, to view the structure of the antibody bound to one subtype of the flu virus called H3N2. They discovered that the antibody inserts a loop into the binding site of the virus, which would otherwise attach to a receptor in a host cell. Additional experimental data showed that F045-092 binds a wide variety of strains and subtypes, including all H3 avian and human viruses from 1963 to 2011 that were tested.

This understanding of the antibody's structural details and binding modes offers new insight for future structure-based drug discovery and novel avenues for designing future vaccines.
But the only way to achieve those goals is for many groups of scientists to work together, Lee said. "Our lab is very focused on the structure of the virus and antibodies, while there are lots of other labs focused on everything from small protein design to vaccine design," he said. "Hopefully we can use this structural information and join together as one big team to tackle the flu."

Source: SLAC National Accelerator Laboratory