Experts Mull H7N9 Bird Market Findings, Assess Rapid Tests

Chinese officials are wrapping up the bird sampling part of the investigation into the novel H7N9 avian flu virus, as international experts weigh the effect of live-bird market closures assess diagnostic tests, and explore candidate vaccine viruses.

In the meanwhile, no human cases have been reported since May 8, keeping the total number of cases at 131. No new fatalities have been reported since May 15, holding that number to 36.

China's agriculture ministry today said it has "basically" finished collecting and testing 899,758 samples from across the country, Xinhua, China's state news agency, reported today. Of that number 53 were positive for the H7N9 virus, all but two of them from 18 different live-poultry markets in the city of Shanghai and eight provinces: Anhui, Zhejiang, Jiangsu, Henan, Shandong, Guangdong, Jiangxi, and Fujian.

The remaining positive samples were both from pigeons. One was a wild pigeon from Nanjing, the capital of Jiangsu province, and the other was a domestic one from another city in the same province.

None of the positive samples were from farms or pigs.

The agriculture ministry statement said poultry market exposure poses the most danger to humans and that farms should boost efforts to guard against the virus, according to the Xinhua report.

Bird market findings

Meanwhile, a team from China and the World Health Organization (WHO) Western Pacific Regional Office (WPRO) surveyed the impact of live-poultry market closures in the wake of the H7N9 outbreak and concluded that human cases declined sharply after officials closed markets. Their findings appeared in the latest issue of the Western Pacific Surveillance and Response Journal.

The researchers focused their view on the market closures in Shanghai, plus Zhejiang and Jiangsu provinces. For example, Shanghai officials launched several measures on Apr 6, which also included culling birds in wholesale markets and disinfecting poultry environments. Researchers noted that the illness onset from the last infected patient from the city was Apr 13.

They found a similar pattern when they looked at market closures in three cities in Zhejiang province and four in Jiangsu province. The last case reported from the cities that had market closures was Apr 17, according to the report.

The drop-off in human cases supports the hypothesis that live-bird markets were the main source of the virus in the areas, but the group said surveillance data in the coming days will show if the control measure was helpful in other provinces.

Researchers also noted that more studies are needed to gauge if certain behaviors in poultry market workers or visitors put them at greater risk of H7N9 infection.

Health officials will have to weigh the benefits and drawbacks of extending or making permanent the live-poultry market closures, with decisions tailored to local situations and needs, the team concluded.

Rapid tests come up short for H7N9

In other developments, researchers from Australia and the Netherlands tested six widely available rapid flu tests to see how useful they would be for detecting H7N9 infections in clinical settings and found that they probably won't detect most cases, especially late in the illness course.

The team reported its findings yesterday in Eurosurveillance.

Currently real-time polymerase chain reaction (RT-PCR) is the gold standard for detecting the new virus, but the assays for H7N9 might not be available in settings where infections occur. They also require a fairly high level of laboratory expertise.

The researchers, therefore, were eager to see if the rapid test would be a useful adjunct for identifying the infections, especially in limited-resource areas.

The group tested the ability of the tests to detect the H7N9 virus and compared the performance at detecting several other flu viruses, including three other low-pathogenic avian H7 strains, two human seasonal H3N2 viruses, and two 2009 H1N1 viruses.

Testing found that the sensitivity of four of the six tests was lower for H7N9 than for the seasonal flu viruses and the other H7 avian flu strains. For all viruses tested, the Sofia point-of-care test, which uses an automated reader, had the highest sensitivity. The Clearview and SD Bioline tests had the poorest sensitivity.

They concluded that RT-PCR is still the best method for testing suspected H7N9 patients.

Candidate vaccine viruses

On the preparedness front, the WHO said yesterday that two more candidate H7N9 vaccine viruses are now available, raising the total to four. Candidate vaccine viruses are routinely developed for flu viruses that have pandemic potential.

Though no decisions have been made to produce a vaccine against the new virus, scientists can use the candidate viruses to start the first steps in making a vaccine, which can speed the process in case one is needed.

Source: Center for Infectious Disease Research & Policy, University of Minnesota. 

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Read About the Latest Advances in Rapid Methods in our Current Newsletter

In the current issue of our rapid methods newsletter, we discuss how PCR can help reduce the time to detection for methicillin-resistant Staphylococcus aureus, and review a dissertation abstract from Auburn University on the use of phage-based magnetoelastic (ME) biosensors.

We have also posted many rapid method news articles and press releases since our last newsletter, as well as interesting technological developments from our blog. The newsletter contains links to every news and blog article published over the past month.

Our Newsletter is FREE and is provided via email. Please sign up for our Newsletter at the following web address: http://rapidmicromethods.com/newsletter/.

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New Biosensor Catches 'Superbug' MRSA In Record Time

Scientists have developed a new test for antibiotic-resistant bacteria that could reduce the time for diagnosis from hours to minutes. This discovery will be invaluable to clinicians trying to treat patients and disinfect hospital facilities.

Antibiotic resistance is rapidly spreading in the world of microbes, which poses serious dangers to public health. Following a major outbreak this spring of the superbug Carbapenem-resistant Enterobacteriaceae (CRE), the director of the U.S. Centers for Disease Control (CDC) labelled emerging antibiotic resistance a "nightmare."

"Our strongest antibiotics don't work and patients are left with potentially untreatable infections," said CDC director Tom Frieden.

Until new treatments are developed, surveillance is the best method for stemming the spread of superbugs. However, current diagnostics for antibiotic resistance can take hours - sometimes days - to complete, increasing the likelihood that another person will catch the disease.

"Our [new] method can determine bacterial antibiotic resistance in 10-12 minutes, while other methods take hours," said senior author Dr. Vodyanoy, a physiologist at Auburn University. The project was a joint collaboration between Auburn and the Keesler Air Force Base with funding from the U.S. Air Force.

To create this method, the researchers relied on a natural enemy of these germs: bacteriophages.

A bacteriophage is a simple virus that can target and kill bacteria. It's name literally means "to eat bacteria."

Another common superbug - methicillin-resistant Staphylococcus aureus (MRSA) - was used to validate their system.

Tiny gold strips, the size and shape of sticks of gum, were placed in petri dishes and coated with modified bacteriophages. These tiny plates can ensnare staph bacteria from a biological sample - spit or blood.

Special markers for MRSA were then added to the dish to detect the presence of antibiotic-resistant strains.

While only samples of MRSA were used in this study, "it can also be applied to other antibiotic resistant bacteria," said co-author Iryna Sorokulova, Ph.D., professor of microbiology at Auburn University.

The study was published in the video journal JOVE, so that other scientists can learn these techniques.

"We felt it would be very beneficial and educational to publish [in JoVE]," said Dr. Vodyanoy. "We are interested in someone else reproducing our results; this technology can be used on a larger scale and for antibiotic resistance other than Staphylococcus."

Source: Medical Daily 

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Student's Fast Start in Research Contributes to Life-Saving Rapid Test

As if the story of breakthrough research were not enough, there is the story of the researcher. The accomplishment, in this case, is a rapid test for detecting devastating bacterial infections. The researcher is undergraduate pre-med student Kristin Wiggins, finishing her junior year at Northern Arizona University while working as an intern at Translational Genomics Research Institute North in Flagstaff.

At the heart of the story is someone who wants to make a difference in the world by saving people's lives—a motivation rooted in instinct and a family tradition of public service and health care. 

"I have always been fascinated with biology and I've had a lot of exposure to people in the medical and science professions," said Wiggins, whose great-uncle was a pharmacogenomics researcher and father was a firefighter. "All of my experiences with them have affirmed my passion to help others with science and medicine." 

From the age of 16, Wiggins has applied her passion by getting into the lab and going to work. As a junior in high school, she received a Helios Internship that placed her in the Cancer and Cell Biology Division of TGen in Phoenix. 

"I worked in the Brain Tumor Unit on Medulloblastoma and I absolutely loved it," Wiggins said. The summer internship stoked her passion for research. After her freshman year at NAU, she contacted TGen North and was invited to interview, which resulted in an internship in the Center for Public Health and Clinical Pathogens. 

"Pathogen research is extremely different from cancer research," Wiggins said. "I was really excited to be offered the opportunity to participate in the revolutionary pathogen research that goes on every day at TGen North." 

Wiggins explained that her research group handles six different dangerous agents. "We work with pathogens that are locally relevant but have a global impact," she said. 

One of those pathogens, Klebsiella pneumonia, is emerging worldwide as a public health crisis, Wiggins said. The pathogen's resistance to antibiotics fuels its danger as a secondary infection in hospital environments. The ability to identify it quickly—in a process known as a rapid KPC test—can save lives. Still experimental, the rapid test can accomplish in 90 minutes what now takes physicians two–three days.

In the lab, Wiggins' role was to grow Klebsiella isolates and extract their DNA for validation purposes. The task involved extreme repetition in the pursuit of virtual perfection. "It is an amazing experience seeing the research process from beginning to end, and all the hard work in between," Wiggins said. "I am thrilled that as an undergraduate intern, I can play a role in every single one of these steps." 

It's a role that has gotten noticed. According to David Engelthaler, director of programs and operations at TGen North, "Kristin is the exact model of what we hope to see in our interns—someone who has immediate dedication to making herself a better and smarter researcher, so she can achieve her long term vision of making a difference in the world." 

Wiggins said the research group hopes to begin publishing papers on its findings soon. "I think that after some more validation and research, we can start translating this into the clinic so that doctors can start using it to diagnose their patients," she said. "Physicians could start using our assay to shorten detection time, reduce the risk of false positives and enable improved treatment." 

While continuing the internship and the research project, Wiggins plans to finish her biomedical science major, with minors in chemistry, philosophy and mathematics. Then she intends to become a full-time research associate at TGen North while pursuing a master's degree and possibly even a Ph.D. before turning to a medical career. 

"One of the things that has surprised me the most is how much of an impact we have," Wiggins said. "We do not do research for the sake of research. Our mission is to advance public health and clinical medicine, and we strive for this every single day."

Source: Northern Arizona University

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U of Alberta Researchers Seek Ways to Rapidly Diagnose Diseases in Developing World

A pair of University of Alberta researchers have each received a $100,000 grant to develop new tests that can be used easily and cheaply to detect diseases in developing nations.

The grants come from the federally funded Grand Challenges Canada, which provides money to develop what it calls “bold new global health ideas.”

The two U of A recipients, John Davis and Darryl Adamko, are from different fields, yet their projects revolve around the same idea of using metabolites — small molecules associated with a biological process in the body — as a method of detection for diseases.

“There’s a lot of literature and more coming out every day that says various metabolites are associated with various diseases, sometimes in blood, sometimes in urine and sometimes in breath,” said Davis, a physicist.

Davis’s project is to create a breath test that would be delivered through a hand-held device similar to a breathalyzer used by police. Patients would blow into the box, which would have the ability to sense a number of different metabolites indicating the presence of diabetes, various cancers, tuberculosis, cholera and other illnesses.

“We have been working on these tiny little vibrating nanomechanical devices that are really sensitive to their external environment and their intrinsic properties,” he said.

Davis uses the analogy of guitar string to describe how the devices work. A guitar string’s sound can be changed several ways, such as by increasing its thickness or tension, or by doing more extreme things like plunging it under water. In the same way, the nanomechanical devices can be altered to become sensitive to various types of molecules, he said.

“If a cop pulls you over and gives you a breathalyzer, he is testing for one thing, ethanol, but what we would like to have is array of devices that are functionalized differently so each string is sensitive to a different thing,” Davis said.

“One of things we still have to do is make these things chemically functional so they respond to particular molecules, because right now they just kind of respond to everything. And we’re trying to make it all miniaturized to be hand held. Right now it’s big, laboratory scale of apparatus.”

Once the development is further along, Davis’s team plans to connect with a researcher in Bangladesh to test the device.

He said for diagnostic technology to work in the developing world, it needs to be portable, rapid and rugged. Many tests currently are done by taking swabs and then growing cultures, which is slow. This is problematic as it often requires patients to travel long distances to a clinic and endure lengthy waits to get results when they need to be at home or work.

A hand-held device could provide results right away, and could even be taken into remote villages rather than having patients come to the clinic.

“Just like the breath analysis a cop does, he knows right away, he doesn’t take a sample, culture it for a few days and then tell you if you can leave the crime scene,” Davis said. “If we can develop this rapid test, even if it were not 100-per-cent accurate, it would tell someone right away whether they needed to do a secondary test or if people were free to go home.”

As for Adamko, a children’s lung disease expert, his project involves using a simple urine test to look for respiratory diseases such as asthma, bronchitis, tuberculosis and pneumonia.

There are currently few effective measures of airway diseases that can be done in a family doctor’s office, so the idea is to create something that can be done easily and non-invasively, he said. That’s where metabolite science is handy, because each type of breathing disorder involves a different kind of that can be seen the fluids patients excrete.

“As a pediatrician, when I heard they were doing metabolic analysis in adults using sputum, I said that isn’t going to work for kids because kids aren’t going to spit, but what about urine tests?”

The idea is that children visiting a doctor’s office would provide a urine sample, which would then be sent to a lab. The lab would then send back a metabolic profile of the patient using a roadmap Adamko is developing. His information would tell the doctor that the presence of a particular molecule or set of molecules indicates a particular disease.

“It’s the pattern of metabolites,” he said. “Based on a urine test, you should be able to get an idea of whether they are sick or not sick, what type of breathing disease they have and also whether it’s getting better or worse.”

Adamko, who recently moved back to the University of Saskatchewan, started a company called Respirlyte and maintains a lab at the U of A to further develop the concept. Eventually the plan is to test the project in India.

“I would like to do a clinical trial with a lot of people and going to a place like India would be awesome,” he said.

Davis and Adamko were among 102 researchers selected to receive a Grand Challenges grant out of 436 applications.


Source: Edmonton Journal 

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Portable Nanodevice Provides Rapid, Accurate Diagnosis of TB and Other Pathogens

A handheld diagnostic device that Massachusetts General Hospital (MGH) investigators first developed to diagnose cancer has been adapted to rapidly diagnose tuberculosis (TB) and other important infectious bacteria. Two papers appearing in the journals Nature Communications and Nature Nanotechnology describe portable devices that combine microfluidic technology with nuclear magnetic resonance (NMR) to not only diagnose these important infections but also determine the presence of antibiotic-resistant bacterial strains.

"Rapidly identifying the pathogen responsible for an infection and testing for the presence of resistance are critical not only for diagnosis but also for deciding which antibiotics to give a patient," says Ralph Weissleder, MD, PhD, director of the MGH Center for Systems Biology (CSB) and co-senior author of both papers. "These described methods allow us to do this in two to three hours, a vast improvement over standard culturing practice, which can take as much as two weeks to provide a diagnosis."

Investigators at the MGH CSB previously developed portable devices capable of detecting cancer biomarkers in the blood or in very small tissue samples. Target cells or molecules are first labeled with magnetic nanoparticles, and the sample is then passed through a micro NMR system capable of detecting and quantifying levels of the target. But initial efforts to adapt the system to bacterial diagnosis had trouble finding antibodies – the detection method used in the earlier studies – that would accurately detect the specific bacteria. Instead the team switched to targeting specific nucleic acid sequences.

The system described in the Nature Communications paper, published on April 23, detects DNA from the tuberculosis bacteria in small sputum samples. After DNA is extracted from the sample, any of the target sequence that is present is amplified using a standard procedure, then captured by polymer beads containing complementary nucleic acid sequences and labeled with magnetic nanoparticles with sequences that bind to other portions of the target DNA. The miniature NMR coil incorporated into the device – which is about the size of a standard laboratory slide – detects any TB bacterial DNA present in the sample.

On the 2.5- by 7.5-cm cartridge (see above picture), DNA extracted from sputum samples is amplified in the chambers on the left. TB-specific sequences are magnetically labeled in the microfluidic mixing channels in the center and detected by passage through the micro-NMR coil on the right. Credit: Center for Systems Biology, Massachusetts General Hospital

Tests of the device on samples from patients known to have TB and from healthy controls identified all positive samples with no false positives in less than three hours. Existing diagnostic procedures can take weeks to provide results and can miss up to 40 percent of infected patients. Results were even stronger for patients infected with both TB and HIV – probably because infection with both pathogens leads to high levels of the TB bacteria – and specialized nucleic acid probes developed by the research team were able to distinguish treatment-resistant bacterial strains.

The Nature Nanotechnology paper, being issued online today, describes a similar system using ribosomal RNA (rRNA) – already in use as a bacterial biomarker – as a target for nanoparticle labeling. The investigators developed both a universal nucleic acid probe that detects an rRNA region common to many bacterial species and a set of probes that target sequences specific to 13 clinically important pathogens, including Streptococcus pneumoniae, Escherichia coli and methicillin-resistant Staphylococcus aureus (MRSA).

The device was sensitive enough to detect as few as one or two bacteria in a 10 ml blood sample and to accurately estimate bacterial load. Testing the system on blood samples from patients with known infections accurately identified the particular bacterial species in less than two hours and also detected two species that had not been identified with standard culture techniques.

While both systems require further development to incorporate all steps into sealed, stand-alone devices, reducing the risk of contamination, Weissleder notes that the small size and ease of use of these devices make them ideal for use in developing countries. "The magnetic interactions that pathogen detection is based on are very reliable, regardless of the quality of the sample, meaning that extensive purification – which would be difficult in resource-limited setting – is not necessary. The ability to diagnose TB in a matter of hours could allow testing and treatment decisions within the same clinic visit, which can be crucial to controlling the spread of TB in developing countries."

Hakho Lee, PhD, of the MGH Center for Systems Biology. co-senior author of both papers, notes that the system will alsohave important applications in developed countries. "The capacity of the system not only to identify bacterial species but also to differentiate factors such as antibiotic resistance will help clinicians treat patients with the 'right' drugs from the start, which also helps reduce the emergence of treatment-resistant strains. The fact that this device requires only a tiny drop of the sample to be tested will be helpful in instances when specimens can be hard to obtain, such as treating children or seniors."

Source: http://phys.org

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