Nanopore Detection of Single Flu Viruses to Control Outbreaks

Influenza is a highly contagious respiratory disease of global importance, which causes millions of infections annually with the ever-present risk of a serious outbreak. Passive vaccination is the only method available for partial control of the virus. Rapid diagnosis of influenza has been explored to prevent outbreaks by enabling medication at very early stages of infection; however, diagnostic sensitivity has not been high enough, until now.

In a new study published in Scientific Reports, a team of researchers led by Osaka University explored the usefulness of combining a single-particle nanopore sensor with artificial intelligence technology, and found that this approach created a new virus typing method that can be used to identify single influenza virions.

Genetic methods can identify many virus species, but require time-intensive processes and specialized staff. Therefore, these methods are unsuitable for point-of-care screening. In a novel approach, the researchers designed a sensor that could assess distinct nanoscale properties of influenza virions within physiological samples.

“We used machine-learning analysis of the electrical signatures of the virions,” says corresponding author Makusu Tsutsui. “Using this artificial intelligence approach to signal analysis, our method can recognize a slight current waveform difference, which cannot be discerned by human eyes. This enables high-precision identification of viruses.”

In testing this sensor, the research team found that electroosmotic flow (liquid motion induced by an electric current across the nanopore) through the pore channel could block the passage of non-virus particles. This ensured that the only particles evaluated by the sensor were virus particles, regardless of the complexity of the sample that contained those viruses.

“Our testing revealed that this new sensor may be suitable for use in a viral test kit that is both quick and simple,” says lead author Akihide Arima, “Importantly, use of this sensor does not require specialized human expertise, so it can readily be applied as a point-of-care screening approach by a wide variety of healthcare personnel.”

In addition to enabling early detection of influenza, this nanosensor method could be modified to enable early detection of other viral particles. This would enable rapid prevention and tracking for a variety of local epidemics and potential pandemics.

The article, "Selective detections of single-viruses using solid-state nanopores," was published in Scientific Reports at DOI: https://www.nature.com/articles/s41598-018-34665-4

Source: Osaka University 

Abstract:

Rapid diagnosis of flu before symptom onsets can revolutionize our health through diminishing a risk for serious complication as well as preventing infectious disease outbreak. Sensor sensitivity and selectivity are key to accomplish this goal as the number of virus is quite small at the early stage of infection. Here we report on label-free electrical diagnostics of influenza based on nanopore analytics that distinguishes individual virions by their distinct physical features. We accomplish selective resistive-pulse sensing of single flu virus having negative surface charges in a physiological media by exploiting electroosmotic flow to filter contaminants at the Si3N4 pore orifice. We demonstrate identifications of allotypes with 68% accuracy at the single-virus level via pattern classifications of the ionic current signatures. We also show that this discriminability becomes >95% under a binomial distribution theorem by ensembling the pulse data of >20 virions. This simple mechanism is versatile for point-of-care tests of a wide range of flu types.

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New Chlamydia Test Delivers Results in About 30 Minutes

NIBIB-funded researchers recently validated a rapid STD test that delivered accurate results in about 30 minutes for chlamydia, allowing patients to receive treatment immediately, thereby stemming the further spread of disease. Other analyses showed most women preferred the easy self-collection method the test offers.

Point-of-care (POC) testing eliminates the need for follow-up appointments since patients receive treatment at the time of diagnosis. Chlamydia has been identified by the World Health Organization as a sexually transmitted disease (STD) without an available POC test. "POC tests for STDs are making significant progress towards being highly sensitive, specific and easy to read within a short wait time." said Tiffani Bailey Lash, Ph.D., director of the National Institute of Biomedical Imaging and Bioengineering (NIBIB) programs in Point-of-Care Technologies and Connected Health (mHealth and Telehealth).

To help fill this healthcare void, a team at the Johns Hopkins University (JHU) Center for the Development of POC Tests for STDs, led by director Charlotte Gaydos, Dr. P.H., tested the binx health POC system at two clinics, in Maryland and Ohio. The study determined this POC test to accurately give a positive result 93 percent and a negative result 99 percent of the time.

With funding from NIBIB, the JHU Center selected binx health (formerly Atlas Genetics) as one of its first subgrantees to support the development and translation of this promising technology. Dr. Gaydos explained, "It takes time and money for POC technologies to get to the market. The goal of POC technology is to deliver quick results on a mobile platform, giving patients more options. A patient should be able to choose if he/she comes into a clinic, goes to a pharmacy, or takes a test at home for STD diagnosis; the bottom line is to encourage people to get tested."

It is common for STDs to present without visible symptoms, so patients think they are healthy while, in fact, they are spreading STDs. Currently, patients who are tested for STDs typically receive their results anywhere from 2-14 days later, which contributes to the onward transmission and rapid spread of STDs. Studies have shown that even patients who have been tested have a low probability of returning to receive results, counseling, and proper treatment. Many patients don't come back to the clinic for a follow-up appointment due to work or other obligations.

Another concern of doctors is the high number of patients who do not seek treatment for STDs due to privacy issues. In the future, POC tests may allow patients to collect a sample at home and send it in for testing. Dr. Gaydos said she was encouraged to observe that the majority (86%) of women in this study found self-collection in the clinic to be easy, and 70 percent preferred that method.

In this study, the samples were self-collected on a swab in the clinic and then placed into a liquid by a lab technician. The swabs were transported to a clinical lab and loaded into a disposable, hand-held cartridge. The lab technician inserted the cartridge into an instrument which has a user-friendly interface for processing and analysis. Each cartridge contains all the components necessary to complete the screening and deliver a positive negative, or indeterminate result in about 30 minutes.

In addition to the need for accurate and rapid test results, it is also important to consider a patient's attitude toward a POC testing. In the study, researchers received completed questionnaires from 273 women who self-collected their sample for the POC test. If treatment was available before leaving the clinic, only 25 percent of women were willing to wait up to 40 minutes for their test result, but 61 percent were willing to wait 20 minutes or less. Most women (81%) were willing to pay $20 or less for a POC test. Almost all women (96%) would tell their partner the result of their test. "It was promising to see how well-received the test was among patients. I think the world has been waiting for a POC STD test and I am eager to be a part of continuing to develop new POC technology," said Lash.

There is an urgent need to address the record high numbers of STDs reported by the Centers for Disease Control. In the U.S. alone there was a 10% increase in STDs between 2016 and 2017. Comprehensive studies like this one show the importance of understanding a patient's attitude toward POC tests and identifying barriers. The results from this study show the need for a reasonably priced test and results delivered in about half an hour or less so patients get treatment before leaving the clinic. JHU and binx health have recently launched a clinical trial and anticipate the data obtained in the trial will help them achieve FDA-clearance for a Chlamydia and Gonorrhea POC test in the U.S. Gaydos envisions a mobile/digital health approach for patients who do not wish to attend a clinic or doctors' office and are happy to self-collect a sample at home.

More information: Lea E. Widdice et al. Performance of the Atlas Genetics Rapid Test for Chlamydia trachomatis and Womenʼs Attitudes Toward Point-Of-Care Testing, Sexually Transmitted Diseases (2018). DOI: 10.1097/OLQ.0000000000000865 

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Researchers Develop Tool for Speedy Diagnosis of Bacterial Infections

Using a small and inexpensive biosensor, researchers at UBC Okanagan, in collaboration with the University of Calgary, have built a diagnostic tool that provides health care practitioners almost instant diagnosis of a bacterial infection.

The tool is able to provide accurate and reliable results in real-time rather than the two-to-five days required for existing processes that test infections and antibiotic susceptibility.

“Advances in lab-on-a-chip microfluidic technology are allowing us to build smaller and more intricate devices that, in the medical research space, can provide more information for health care practitioners while requiring less invasive sampling from patients,” explains Mohammad Zarifi, an assistant professor at UBC Okanagan.

According to health care statistics from 2017, every hour of delay in antibiotic treatment increases mortality rates by nearly eight per cent due to infection complications in the bloodstream.

Zarifi, and his research group in the School of Engineering’s Microelectronics and Advanced Sensors Laboratory, tested their device by tracking the amount of bacteria present in a variety of samples under various scenarios. The scenarios resembled those encountered in clinical microbiological laboratories.

By sending a microwave signal through the sample, the device quickly and accurately analyzes and then generates a profile of existing bacteria.

The diagnostic tool not only provides a rapid, label-free and contactless diagnostic tool for clinical analysis but it also goes further, says Zarifi.

“The device is able to rapidly detect bacteria and in addition, it screens the interaction of that bacteria with antibiotics,” he adds. “The combined results give health care practitioners more information than they currently have available, helping them move forward to determine accurate treatments.”

This biosensor, explains Zarifi is a significant step forward in improving the complex antibiotic susceptibility testing workflow and provides a rapid and automated detection of bacteria as well as screening the bacteria proliferation in response to antibiotics.

The research was published in the journal Nature Scientific Reports with financial support from CMC Microsystems and the Natural Sciences and Engineering Council of Canada.

Abstract

Sensitive, Real-time and Non-Intrusive Detection of Concentration and Growth of Pathogenic Bacteria using Microfluidic-Microwave Ring Resonator Biosensor

Infection diagnosis and antibiotic susceptibility testing (AST) are time-consuming and often laborious clinical practices. This paper presents a microwave-microfluidic biosensor for rapid, contactless and non-invasive device for testing the concentration and growth of Escherichia Coli (E. Coli) in medium solutions of different pH to increase the efficacy of clinical microbiology practices. The thin layer interface between the microfluidic channel and the microwave resonator significantly enhanced the detection sensitivity. The microfluidic chip, fabricated using standard soft lithography, was injected with bacterial samples and incorporated with a microwave microstrip ring resonator sensor with an operation frequency of 2.5 GHz and initial quality factor of 83 for detecting the concentration and growth of bacteria. The resonator had a coupling gap area on of 1.5 × 1.5 mm2 as of its sensitive region. The presence of different concentrations of bacteria in different pH solutions were detected via screening the changes in resonant amplitude and frequency responses of the microwave system. The sensor device demonstrated near immediate response to changes in the concentration of bacteria and maximum sensitivity of 3.4 MHz compared to a logarithm value of bacteria concentration. The minimum prepared optical transparency of bacteria was tested at an OD600 value of 0.003. The sensor’s resonant frequency and amplitude parameters were utilized to monitor bacteria growth during a 500-minute time frame, which demonstrated a stable response with respect to detecting the bacterial proliferation. A highly linear response was demonstrated for detecting bacteria concentration at various pH values. The growth of bacteria analyzed over the resonator showed an exponential growth curve with respect to time and concurred with the lag-log-stationary-death model of cell growth. This biosensor is one step forward to automate the complex AST workflow of clinical microbiology laboratories for rapid and automated detection of bacteria as well as screening the bacteria proliferation in response to antibiotics.

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Pregnancy Style Test for E. coli and Romaine Lettuce

With seven deaths, 31 suffering kidney failure, 159 hospitalised and at least 339 Americans and Canadians sickened since 2017 after eating romaine lettuce contaminated with E. coli, news of a faster E. coli test is welcome.

Western University, Ontario, has developed a kit that detects a protein unique to the pathogenic E. coli O157 bacteria and, using flow through technology, is able show results in hours rather than days.

Food samples to be tested are incubated for a few hours then a sample is placed on a pad. After 15 minutes, the pad displays one red line to show it worked properly — and a second if the sample contains E. coli O157; much like how pregnancy tests show their results.

The developers claim their kit also makes the process of testing for E. coli O157 cheaper than existing technology, which could make smaller-scale producers more amenable to testing their products.

Current food testing methods for E. coli typically rely on culture and by the time results are available some days later fresh produce has been shipped to retailers, sold and eaten. With results available in hours, rather than days or weeks, this new test will enable food processors to ensure food is uncontaminated before it is shipped.

Use of a cheaper, rapid test will mean processors can test more frequently.

The romaine lettuce crisis

A common source of E. coli illness is raw fruits and vegetables that have come in contact with faeces from infected animals.

Leafy greens, such as lettuce, can become contaminated in the field by soil, water, animals or improperly composted manure. Lettuce can also be infected by bacteria during and after harvest from handling, storing and transporting the produce.

Contamination in lettuce is also possible at the retail environment, in the refrigerator or from counters and cutting boards through cross-contamination with bacteria from raw meat, poultry or seafood.

Since 1 April 2017, the Canadian Food Inspection Agency has tested more than 2000 samples of imported fresh vegetables and salads, including romaine lettuce and pre-packaged salads containing romaine lettuce, as part of its regular microbiological surveillance program.

None of these samples have proven positive for E. coli O157. This means that contamination levels are low and most romaine lettuces are uncontaminated, but the real need is to identify those lettuces that are contaminated. More tests, with low cost but high accuracy, will help public health officials and produce marketers weed out the affected produce. Western’s new test will be invaluable in this circumstance.

In his blog Publisher’s Platform: How does this “Romaine” acceptable?, Bill Marler asks: “What will growers, processors, shippers, grocery stores, restaurants, consumers, regulators, and politicians do? Good readers – ideas?”. Possibly the best response was from Sue: “This should fix it: Move the growers’ focus from their wallets (greed) to their amygdala (fear.) Host a weekly luncheon for all Western Growers executives - attendance mandatory. Menu: Giant Caesar salad made with romaine lettuce from each grower. Eat it or close down.

The rapid test was developed at Western University’s Schulich School of Medicine & Dentistry and Robarts Research Institute with support from Mitacs, a federal non-profit that encourages academic and industrial collaboration with the food industry.

Source: https://www.foodprocessing.com.au

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Mesa Biotech Receives FDA 510(k) Clearance and CLIA Waiver for Its Accula™ RSV Molecular Point of Care Test

Mesa Biotech Inc., is a privately held, molecular diagnostic company that has developed an affordable and easy to operate PCR (polymerase chain reaction) testing platform designed specifically for point-of-care (POC) infectious disease diagnosis. Today, Mesa Biotech announced that it received 510(k) clearance and Clinical Laboratory Improvements Amendments (CLIA) waiver from the US Food and Drug Administration (FDA) for its Accula™ RSV test. The respiratory syncytial virus or RSV test cassette is cleared for diagnosing both children and adult populations.

The Accula RSV test is the second molecular POC diagnostic on the Accula platform to receive FDA clearance and CLIA waiver, following the company's Flu A/Flu B test, cleared earlier this year. The Center for Disease Control (CDC) acknowledges that the specificity and sensitivity of rapid influenza diagnostic tests (RIDT) are lower than for viral culture and RT-PCR and vary by test. The Accula molecular test system provides the superior sensitivity, specificity and information content of laboratory-based PCR testing, while offering the simplicity, convenience and procedural familiarity of traditional POC rapid immunoassays, with its palm-sized, reusable dock and disposable test cassettes. Furthermore, the Accula RSV test is indicated for use with nasal swab collection, which is less invasive than nasopharyngeal swabs and provides a more comfortable specimen collection experience for the patient, especially children.

"The addition of Accula RSV test to our Flu A/Flu B test significantly enhances our upper respiratory rapid, molecular diagnostic offerings," said Dr. Hong Cai, co-founder and CEO of Mesa Biotech. "It is important to have a distinct in-office diagnosis for RSV and flu, especially in young children and the elderly to provide timely, accurate information to clinicians to ensure appropriate treatment and improved patient management."

According to the CDC, young children and older adults are more likely to get serious complications if they get sick with RSV. Each year in the US, an estimated 57,000 children younger than five years old are hospitalized due to RSV infection. Additionally, it is estimated that more than 177,000 older adults are hospitalized and 14,000 of them die in the US due to RSV infection. Adults at the highest risk for severe RSV infection include those 65 years and older, adults with chronic heart or lung disease and/or adults with weakened immune systems.

For rapid distribution of both tests, Mesa Biotech entered into a strategic alliance with Sekisui Diagnostics for exclusive product distribution rights in the U.S. and Canada. The Accula RSV test, along with the company's Flu A/Flu B test, is marketed by Sekisui Diagnostics under the Silaris™ brand.

About Respiratory Syncytial Virus

Respiratory syncytial virus (RSV) is an RNA virus attributable to outbreaks of respiratory tract infections. RSV infections can occur throughout the year, but typically peak during the winter months. RSV most commonly causes a cold-like illness but can also cause lower respiratory infections like bronchiolitis and pneumonia, which often become severe in infants, young children and the elderly with underlying diseases. Diagnosis of RSV is difficult because the initial symptoms can be similar to those caused by other infectious agents. Considering that the RSV virus is highly contagious, accurate diagnosis and prompt treatment of patients can have a positive effect on public health.

About Mesa Biotech Inc.

Mesa Biotech designs, develops, manufactures and commercializes next generation molecular diagnostic tests, bringing the superior diagnostic performance of nucleic acid PCR amplification to the point-of-care (POC). Mesa Biotech's Accula™ System consists of a portable, palm-sized dock and disposable, assay-specific test cassettes. This patented system enables healthcare professionals to access actionable, laboratory-quality results at the POC with greater sensitivity and specificity than current infectious disease rapid immunoassay tests. The Accula Flu A/Flu B and the Accula RSV tests have obtained CE Mark in the EU and 510(k) clearance and Clinical Laboratory Improvements Amendments (CLIA) waiver from the US Food and Drug Administration (FDA). Both products are distributed in the US and Canada by Sekisui Diagnostics under the Silaris™ brand. Mesa Biotech has secured a number of strategic agreements for distribution in Europe and Asia.

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Indian Scientists Develop Highly Sensitive and Rapid Tests for Detection of Tuberculosis

Tuberculosis (TB) continues to be one of the leading causes of death worldwide. According to the World Health Organisation (WHO), some 10 million people across the world fell ill with TB in 2017, and 1.6 million died from the disease. India, which saw an estimated 28 lakh cases occurred and 4.8 lakh people died due to the disease in 2015, accounts for one-fourth of the global TB burden. Offering hope of finding tuberculosis early, scientists at the Translational Health Science and Technology Institute (THSTI) and All India Institute of Medical Sciences, New Delhi, have jointly developed highly sensitive and rapid tests for detection of TB infection in lungs and surrounding membranes, says a report in India Science Wire. Early detection and treatment are crucial to prevent the spread, outbreaks, and development of resistance.

Tuberculosis is an infectious disease caused by Mycobacterium tuberculosis. It commonly affects the lungs, but can also affect other parts of the body. It spreads from person to person through inhalation of infected air - when people who are infected with TB cough, sneeze they propel the germs into the air. According to the report, when the bacteria attack the lungs, the disease manifests itself as pulmonary form or pulmonary TB - the most common form of TB. But in 2016,  about 15% of new patients were found infected with extrapulmonary TB where organs other than the lungs may be affected.

Till date, the detection of all forms of TB is mostly based on sputum smear microscopy and culture tests. Although smear microscopy is inexpensive and simple, it has low sensitivity. A culture test is highly sensitive but takes 2 to 8 weeks to get results. For detection of bacterial proteins in sputum samples, conventional diagnostic tests use antibodies. And such tests suffer from limitations including batch-to-batch variability, limited shelf-life, and cost said the report.

However, to help address the shortcomings in the present testing protocol, the researchers developed two DNA aptamer-based tests - Aptamer Linked Immobilized Sorbent Assay (ALISA) and Electrochemical Sensor (ECS) for detecting a bacterial protein in the sputum. Aptamers are DNA, RNA or peptide molecules that bind to a specific target molecule. They can be used for both basic research and clinical purposes as macromolecular drugs. Aptamers bind the right target (which defines sensitivity) and at the same time rule out any non-specific binding to other targets.

In the present work, the researchers compared the performance of the newly developed tests with antibody-based tests in 314 sputum samples. The found that ALISA produced 92% sensitivity compared to the antibody-based method that showed 68% sensitive. The team used ALISA to detect a bacterial protein, HspX. But it took five hours for the researchers to yield results as the method requires sputum immobilisation which is a time-consuming step. So the team developed a simplified ECS test. In this technique, the team immobilized the aptamer with an electrode, and upon binding to HspX in the sputum sample, a drop in the electrical signal was recorded.

According to the researchers, the ECS test can be used for screening of samples in the field. It takes as less as 30 minutes to deliver results and is highly sensitive. The ECS test could also detect HspX protein in 91% of the samples tested in the study. In addition, there is no need for sputum sample preparation which is a complex and time-consuming process, said the report.

“We aim to develop aptamers for detection of multiple bacterial proteins simultaneously which is expected to lead a more robust test,” said Tarun Sharma, a member of the research team.

“The aptamer-based screening tests for pulmonary TB, pleural TB, and TB meningitis hold immense promise for a country like India, where the disease burden in high and primary health care is only a dream for many. The ECS platform could be used in a mobile screening van at the point-of-care,” added Jaya Tyagi, who led the research team AIIMS. “We hope that these tests are adopted by the TB programs in the country,” she added.

In the study, the team used the aptamer-based test also for detection of pleural TB, the second most prevalent form of extrapulmonary TB. The researchers note the early diagnosis of pleural TB is limited by the availability of a sensitive and rapid test. The performance of existing DNA-based tests varies widely due to low bacterial load in the pleural fluid sample.

“There is no test for making a confirmed diagnosis of pleural TB. Even WHO-endorsed Gene Xpert has a poor sensitivity of 22%. On the contrary, our aptamer-based test for pleural TB showed 93% sensitivity and is cost-effective,” explained Sagarika Haldar, a member of the research team, was quoted as saying by India Science Wire.

The results were published in the journals ACS Infectious Diseases and Analytical Biochemistry.

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LuminUltra Concludes Acquisition Of Aqua-Tools’ Rapid Microbial & Ballast Water Monitoring Solutions

LuminUltra, the Canada-headquartered specialist in microbiological monitoring tools, has completed its acquisition of the Rapid Microbial Solutions (RMS) division of France-based aqua-tools.

The acquisition, announced in October, includes the award-winning B-QUA ballast water test kit and aqua-tools’ European customer base for its 2nd Generation ATP® (Adenosine Triphosphate) measurement products.

Pat Whalen, President and CEO of LuminUltra, said: “The acquisition of aqua-tools’ B-QUA line of ballast water monitoring solutions along with its European customer base is a key development in our strategy to enter the international maritime sector. Our goal is to expand our global footprint through a wider network of partners across the European continent and we look forward to bringing our maritime customers closer to the LuminUltra family.”

aqua-tools employees dedicated to the Rapid Microbial Solutions division have joined the LuminUltra team while remaining in Paris, France. aqua-tools continues to operate as a separate business, focusing solely on its Infection Control Solutions.

Marc Raymond, Managing Director aqua-tools, said: “We are delighted the acquisition has concluded with the successful migration of our rapid microbial solutions and staff to LuminUltra.

“For LuminUltra, the technology transfer is something of a homecoming since the rapid ballast water test kit was initially developed as a collaborative project between aqua-tools, SGS Group (Switzerland) and LuminUltra. The 2nd generation ATP technique used in the B-QUA test kit is proven to be the most reliable and effective indicative monitoring solution on the market.”

The addition of B-QUA compliments LuminUltra’s portfolio of 2nd Generation ATP microbial testing solutions, all of which have significant relevance for the commercial and offshore marine segments.
The company’s Quench-Gone Organic Modified (QGO–M™) test kit, for instance, has been designed specifically to monitor the bacteria content of fuel systems to curtail biofouling and microbial influenced corrosion of pipework. Its water test kit (QGA™) can be used to monitor the microbial quality of drinking water onboard. Both of these analyses can be performed with the same equipment as the one used for ballast water.

“Effective microbiological control is critical for maintaining efficient processes in a number of maritime and offshore applications,” said Whalen.

“LuminUltra’s 2nd Generation ATP testing provides results within minutes to give operators a clear indication of the total microbial activity and health of their water, wastewater and fuel systems. Microbiology is rarely considered a problem until it becomes a problem.”

The acquisition of aqua-tools’ RMS division follows the September purchase of US-based InstantLabs, a developer of DNA-based measurement technologies.

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Two Pore Guys Receives Grant to Develop Sensitive, Low-Cost Molecular Diagnostic Tests for Tuberculosis

Two Pore Guys, Inc. (2PG) announced that the company has received a $2.8 million grant from the Bill & Melinda Gates Foundation to develop sensitive, low-cost molecular diagnostic tests for tuberculosis (TB) using its solid-state nanopore technology. The 18-month grant will fund proof-of-concept work to determine if 2PG technology could provide an easy-to-use, sensitive and low-cost point-of-care device that would be compatible with automated, wireless reporting for resource-poor settings.

TB is one of the top 10 causes of death worldwide, and the leading cause from a single infectious agent. Millions of people continue to fall sick with TB each year. WHO estimates 10 million people developed TB last year worldwide.

The WHO 2018 Global TB Report, released in September, calls for an unprecedented mobilization of national and international commitments to meet the global target of ending TB by 2030. Underdiagnosis of TB is a major challenge: only about 6.4 million of the 10 million people who contracted TB in 2017 were officially recorded by national reporting systems, leaving 3.6 million people undiagnosed or detected but not reported. Late detection of Mycobacterium tuberculosis (MTB), the infectious organism that causes TB, increases the risk of transmission, delays treatment, and burdens the healthcare system. Lack of affordable, sensitive and rapid detection of MTB is a major hurdle in low- and middle-income countries. Implementing a low-cost, sensitive and portable molecular diagnostic test is expected to save thousands of dollars per patient.

“Our point-of-care technology platform offers tremendous value to address medical testing needs in resource-limited areas,” said William Dunbar, Co-founder and interim CEO of Two Pore Guys. “We are grateful to the Bill & Melinda Gates Foundation for this support and for their enthusiasm about the promise of our technology. We hope to make a difference in the lives of millions of people around the world who need better access to affordable TB care.”

About the 2PG Technology

Unlike biological nanopores used in some DNA sequencing technologies, 2PG’s devices employ silicon nanopore chips, which are robust sensors that examine a wide range of individual analytes with high sensitivity. Additionally, 2PG workflows can support multiplexed and multi-modal assays, i.e., to test for DNA/RNA and analyte/protein from a single sample source. 2PG’s patented methods also permit genotyping, e.g., for detecting drug resistance.

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FluChip-8G Tested in US and China for Rapid Characterization of Influenza Viruses

Scientists at Duke Kunshan University and Duke University are collaborating with InDevR Inc. to evaluate the FluChip-8G Insight system for the rapid identification of the influenza virus subtype from human and animal samples.

The FluChip-8G test promises to be a faster and easier way to determine the subtype of seasonal and non-seasonal influenza viruses. It has the potential to reduce influenza A and B characterization times from two weeks to eight hours.

"Knowing that a patient is infected with a dangerous influenza virus could lead to early antiviral treatment, isolation of the patient, and early identification and mitigation of the virus source, which could save lives," said Professor Gregory Gray, the Principal Investigator for Duke Kunshan and Duke University.

Study teams at Duke Kunshan and Duke are employing the FluChip-8G test alongside standardized WHO/CDC rRT-PCR tests that are currently used for Gray's research in zoonotic influenza virus epidemiology.

The study will use specimens collected by the Duke Kunshan team and collaborators in Vietnam, China, Malaysia and the U.S.

In the United States, specimens will come from Duke University Hospital's Department of Clinical Virology, Wake Forest University's Department of Clinical Virology, and the North Carolina Veterinary Diagnostic Laboratory System, as well as a swine veterinarian from the U.S. Midwest.

"We are excited to participate in the multi-month field validation of this technology," said Haiyan Gao, vice chancellor for academic affairs at Duke Kunshan and Henry Newson Professor of Physics at Duke University.

"This collaboration shows the research strength of Duke Kunshan is widely recognized. It is also a typical example of how joint research projects between China and the United States can make people's lives better," Gao said.

Influenza -- commonly known as the flu -- is a respiratory infection caused by a virus. Influenza A and B viruses are the two types that most commonly infect humans. Influenza A viruses spread between animals and humans and sometimes cause epidemics.

Based in Boulder, Colorado USA InDevR is a leader in progressive new diagnostic and analytical technologies that enable accelerated development and manufacturing of vaccines and other bio-therapeutics.

Advanced development of this new molecular test, which was designed to improve pandemic preparedness, was funded by the Biomedical Advanced Research and Development Authority (HHSO100201400010C, HHSO100201400024C), which is part of the U.S. Department of Health and Human Services.

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Mesa Biotech Obtains CE Mark for its Accula RSV Molecular Point of Care Test

Mesa Biotech Inc., a privately-held, molecular diagnostic company that has developed an affordable and easy to operate PCR (polymerase chain reaction) testing platform designed specifically for point-of-care (POC), today announced it has obtained CE Mark in the European Union (EU) for its Accula™ RSV Test. The company's RSV test will be marketed by select distributors in the EU at once. Accula RSV Test is pending FDA clearance in the US.

"We are excited to introduce the second test in our PCR-based infectious disease diagnostic portfolio in the EU," said Hong Cai, Co-founder and Chief Executive Officer, Mesa Biotech, Inc. "The Accula RSV brings reference laboratory performance to the point of care, enabling clinicians to quickly and accurately diagnosis and treat their patients."

Respiratory syncytial virus (RSV) is an RNA virus attributable to outbreaks of respiratory tract infections. RSV infections can occur throughout the year, but typically peak during the winter months. Studies indicate that nearly 100 percent of children will suffer from RSV infections by age two. RSV viruses not only cause upper respiratory tract infections but also bronchiolitis of the lower respiratory tract, which often becomes severe in infants and toddlers with underlying diseases. Diagnosis of RSV is difficult because the initial symptoms can be similar to those caused by other infectious agents. Considering that the RSV virus is highly contagious, accurate diagnosis and prompt treatment of patients can have a positive effect on public health.

According to the World Health Organization (WHO), the global burden of RSV-associated acute lower respiratory infection is estimated at 33 million annually, resulting in more than 3 million hospitalizations and almost 60,000 in-hospital deaths in children under five years of age.

The Center for Disease Control (CDC) acknowledges that the specificity and sensitivity of rapid influenza diagnostic tests (RIDT) are lower than for viral culture and RT-PCR and vary by test.  The Accula System, a palm-sized, reusable dock with disposable test cassettes, offers the simplicity, convenience and procedural familiarity of traditional POC rapid immunoassays, while providing the superior sensitivity, specificity and information content of laboratory-based PCR testing. The Accula RSV test is indicated for use with nasal swab collection, which is less invasive than nasopharyngeal swabs and provides a more comfortable specimen collection experience for the patient, especially children.

About Mesa Biotech Inc.

Mesa Biotech designs, develops, manufactures and commercializes next generation molecular diagnostic tests, bringing the superior diagnostic performance of nucleic acid PCR amplification to the point-of-care (POC). Mesa Biotech's Accula™ System consists of a portable, palm-sized dock and disposable, assay-specific test cassettes. This patented system enables healthcare professionals to access actionable, laboratory-quality results at the POC with greater sensitivity and specificity than current infectious disease rapid immunoassay tests. The Accula Flu A/Flu B Test, Mesa Biotech's first product to market, has obtained CE Mark in the EU and 510(k) clearance and Clinical Laboratory Improvements Amendments (CLIA) waiver from the U.S. Food and Drug Administration (FDA).

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Bio-Rad Wins USDA Contract for Its Pathogen Detection Testing Products

Bio-Rad Laboratories, Inc., a global leader of life science research and clinical diagnostic products, today announced that the company has been awarded a contract for iQ-Check real-time PCR pathogen detection test kits and the iQ-Check Prep Automation System from the United States Department of Agriculture’s Food Safety and Inspection Service (USDA FSIS).

The contract includes real-time PCR-based tests for pathogens that include Salmonella spp., Listeria monocytogenes, Campylobacter, Escherichia coli O157:H7, and Shiga toxin producing E. coli (STEC) that may be found in raw meat and poultry, ready to eat meat and poultry, processed egg products, and other food products and environmental samples. Bio-Rad’s iQ-Check real-time PCR test kits were selected based on their proven real-time PCR technology that uses highly specific patented DNA probes to detect pathogenic bacteria in a sample. USDA FSIS will also incorporate Bio-Rad’s iQ-Check Prep automation system for high throughput sample processing and elevated traceability.

“We are pleased to partner with the USDA to help protect the food supply,” said Annette Tumolo, Bio-Rad President, Life Science Group. “This is an exciting development for Bio-Rad, reflecting our strong performance in the area of rapid food pathogen testing solutions.”

The iQ-Check kits for pathogen detection are routinely used in food safety programs worldwide, and are recognized by several renowned international validation organizations.

iQ-Check and Bio-Rad are trademarks of Bio-Rad Laboratories, Inc. in certain jurisdictions.

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FDA Authorizes Emergency Use of First Ebola Fingerstick Test with Portable Reader

The U.S. Food and Drug Administration announced that an emergency use authorization (EUA) has been issued for a rapid, single-use test for the detection of Ebola virus (Zaire ebolavirus). This is the second Ebola rapid antigen fingerstick test available under EUA, but the first that uses a portable battery-operated reader, which can help provide clear diagnostic results outside of laboratories and in areas where patients are likely to be treated.
The test, called the DPP Ebola Antigen System, is used with blood specimens, including capillary “fingerstick” whole blood, from individuals with signs and symptoms of Ebola virus disease (EVD) in addition to other risk factors, such as living in an area with large numbers of EVD cases and/or having contact with other individuals exhibiting signs and symptoms of EVD.

“The scourge of Ebola tragically demonstrates that we’re a global community when it comes to public health protection. Infectious disease doesn’t recognize nation states. Bacteria and viruses don’t respect territorial boundaries. It takes a sustained, robust and globally coordinated effort to protect our nation and the global community from various infectious disease threats. We’re all in this together. To that end, our FDA team of experts in drugs, vaccines and diagnostics continue to collaborate with our Federal, international and industry partners to employ our collective expertise, experiences from previous incidents, and resources to assist in the global response to the Ebola outbreak in the Democratic Republic of Congo,” said FDA Commissioner Scott Gottlieb, M.D. “This EUA is part of the agency’s ongoing efforts to help mitigate potential, future threats by making medical products that have the potential to prevent, diagnosis or treat available as quickly as possible. We’re committed to helping the people of the DRC effectively confront and end the current Ebola outbreak. By authorizing the first fingerstick test with a portable reader, we hope to better arm health care providers in the field to more quickly detect the virus in patients and improve patient outcomes.”

The FDA’s EUA authority allows the agency to authorize the use of an unapproved medical product, or the unapproved use of an approved medical product when, among other circumstances, there are no adequate, approved and available alternatives. When circumstances exist justifying authorization, the EUA becomes an important mechanism that allows broader access to medical products that have not been FDA cleared or approved and are instead only authorized for use for the duration of an emergency declaration. The FDA’s criteria for issuing an EUA for a diagnostic test includes making an assessment that it is reasonable to believe, based on the totality of evidence available to the agency, that the test may be effective and the known and potential benefits of using the test outweigh its known and potential risks.

In 2014, during the Ebola outbreak in West Africa, an emergency was declared by the Secretary of Health and Human Services. While that outbreak has ended, ongoing, smaller Ebola outbreaks have continued, and the emergency declaration is still in place. Recent outbreaks in remote areas with limited resources can benefit from rapid diagnostic tools, and the issuance of an EUA for the DPP Ebola Antigen System is an important step in addressing these outbreaks.

The DPP Ebola Antigen System provides rapid diagnostic results with tests that can be performed in locations where a healthcare provider does not have access to authorized Ebola virus nucleic acid tests (PCR testing), which are highly sensitive but can only be performed in certain laboratory settings that are adequately equipped. The DPP Ebola Antigen System has been authorized for use with capillary “fingerstick” whole blood, ethylenediaminetetraacetic acid (EDTA, an anticoagulant added to whole blood to prevent coagulation) venous whole blood and EDTA plasma. The DPP Ebola Antigen System should only be run in facilities, including treatment centers and public health clinics where patients are likely to be treated, and laboratories that are adequately equipped, trained and capable of such testing.

While today’s action will increase access to diagnostic tools for healthcare providers who may not have otherwise been equipped to perform tests, it is important to note that a negative result from the DPP Ebola Antigen System, especially in patients with signs and symptoms of EVD, should not be used as the sole basis for patient management decisions. The diagnosis of EVD must be made based on multiple factors such as, history, signs, symptoms, exposure likelihood and other laboratory evidence in addition to the detection of Ebola virus.

The FDA remains committed to using its authorities and resources to advance the development of countermeasures to address emerging threats and recently outlined its efforts to help address Ebola virus outbreaks. The FDA will continue to work with its federal partners and potential commercial product manufacturers in the most expedited manner to increase the availability of authorized diagnostic tests for Ebola virus disease for emergency use during this and any future outbreak.

With the issuance of the EUA for the DPP Ebola Antigen System to Chembio Diagnostic Systems Inc., the FDA has now issued EUAs for nine nucleic acid tests and two rapid diagnostic tests for Ebola virus detection in human specimens.

The FDA, an agency within the U.S. Department of Health and Human Services, protects the public health by assuring the safety, effectiveness, and security of human and veterinary drugs, vaccines and other biological products for human use, and medical devices. The agency also is responsible for the safety and security of our nation’s food supply, cosmetics, dietary supplements, products that give off electronic radiation, and for regulating tobacco products.

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Cellphone Technology Developed to Detect HIV

The management of human immunodeficiency virus 1 (HIV), an autoimmune disorder that cripples the immune system by attacking healthy cells, remains a major global health challenge in developing countries that lack infrastructure and trained medical professionals. Investigators from Brigham and Women’s Hospital have designed a portable and affordable mobile diagnostic tool, utilizing a cellphone and nanotechnology, with the ability to detect HIV viruses and monitor its management in resource-limited regions. The novel platform is described in a paper published recently in Nature Communications.

“Early detection of HIV is critical to prevent disease progression and transmission, and it requires long-term monitoring, which can be a burden for families that have to travel to reach a clinic or hospital,” said senior author Hadi ­­Shafiee, PhD, a principal investigator in the Division of Engineering in Medicine and Renal Division of Medicine at the Brigham. “This rapid and low-cost cellphone system represents a new method for detecting acute infection, which would reduce the risk of virus transmission and could also be used to detect early treatment failure.”

Traditional virus monitoring methods for HIV are expensive, requiring the use of polymerase chain reaction (PCR). Shafiee and his colleagues sought to design an affordable, simple tool that makes HIV testing and monitoring possible for individuals in developing countries with less access to medical care.

Utilizing nanotechnology, a microchip, a cellphone and a 3D-printed phone attachment, the researchers created a platform that can detect the RNA nucleic acids of the virus from a single drop of blood. The device detects the amplified HIV nucleic acids through on-phone monitoring of the motion of DNA-engineered beads without using bulky or expensive equipment. The detection precision was evaluated for specificity and sensitivity.

Researchers found that the platform allowed the detection of HIV with 99.1 percent specificity and 94.6 percent sensitivity at a clinically relevant threshold value of 1,000 virus particles/ml, with results within one hour. Notably, the total material cost of the microchip, phone attachment and reagents was less than $5 per test.

“Health workers in developing countries could easily use these devices when they travel to perform HIV testing and monitoring. Because the test is so quick, critical decisions about the next medical step could be made right there,” said Shafiee. “This would eliminate the burden of trips to the medical clinic and provide individuals with a more efficient means for managing their HIV.”

“We could use this same technology as a rapid and low-cost diagnostic tool for other viruses and bacteria as well,” said lead author Mohamed Shehata Draz, ­­PhD, an instructor in the Division of Engineering in Medicine and Renal Division of Medicine at the Brigham.. “This platform could help a lot of people worldwide.”

Funding for this work was provided by the National Institute of Health under award numbers R01AI118502, R21HD092828, and P30ES000002; Harvard T.H. Chan School of Public Health, Harvard Center for Environmental Health through Harvard NIEHS Grant; and American Board of Obstetrics and Gynecology, American College of Obstetricians and Gynecologists, American Society for Reproductive Medicine, Society for Reproductive Endocrinology and Infertility through ASRMAward, and Harvard University Center for AIDS Research (CFAR) under award number 5P30AI060354–14.

Draz, MS et al. “DNA engineered micromotors powered by metal nanoparticles for motion based cellphone diagnostics” Nature Communications DOI: 10.1038/s41467-018-06727-8

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FDA Approves First Drug Using the Recombinant Factor C Assay for Endotoxin Testing

Lonza announced today that the recombinant Factor C (rFC) Assay has been used for endotoxin testing of Eli Lilly’s Emgality™ (galcanezumab), the first drug approved by the U.S. Food and Drug Administration (FDA) to have been released using this method instead of traditional Limulus Amebocyte Lysate (LAL) - based  methods. Emgality™ is a monoclonal antibody drug treatment for the prevention of migraine in adults.

In 2003 Lonza launched the PyroGene™ recombinant Factor C Assay, the first endotoxin testing method to utilize a recombinant form of Factor C, the first component in the horseshoe-crab clotting cascade activated by the presence of endotoxins. As an animal-free method, the PyroGene™ Assay offers a more sustainable alternative to LAL-based tests that use the lysate obtained from horseshoe crab blood as their main ingredient.

“The recent FDA approval of Emgality™ marks a significant breakthrough in establishing the recombinant Factor C Assay as the non-animal method of choice for the identification of contaminating endotoxins in drug products,” said Lakiya Wimbish, Product Manager for Lonza’s PyroGene™ Recombinant Factor C Assay. “This development serves as an official confirmation that the recombinant Factor C method can be as accurate, sensitive and specific as LAL-based methods, while helping to secure the supply of natural resources.”

Allen Burgenson, Lonza’s Global Subject Matter Expert for Endotoxin Testing Solutions, said, “Lonza is the commercial innovator in bringing recombinant technology for endotoxin testing to market. This milestone accomplishment in advancing rFC technology will help to ensure the sustainability of the world’s horseshoe crab species. The three species of horseshoe crab found in Asia (Carcinoscorpius rotundicauda, Tachypleus tridentatus and Tachypleus gigas) are in serious decline due to overfishing, habitat loss and their use as a regional food. As these species become depleted due to a lack of regulatory oversight, countries such as China that use them to prepare Tachypleus Amebocyte Lysate (TAL) will look to the North American species, Limulus polyphemus, to fulfill their ever increasing needs in order to release non-pyrogenic pharmaceutical products to the market. Use of recombinant products, such as recombinant Factor C for endotoxin detection will greatly reduce the pressure on these species.”

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Detecting E. coli Strains Using Molecular Electronics

Finding a fast and inexpensive way to detect specific strains of bacteria and viruses is critical to food safety, water quality, environmental protection and human health. However, current methods for detecting illness-causing strains of bacteria such as E. coli require either time-intensive biological cell cultures or DNA amplification approaches that rely on expensive laboratory equipment.

Now, Josh Hihath, an associate professor of electrical and computer engineering at the University of California, Davis, and colleagues at the University of Washington and TOBB University of Economics and Technology in Ankara, Turkey have adapted a molecular electronic device called a single-molecule break junction to detect RNA from strains of E. coli known for causing illness. The findings were published online in the journal Nature Nanotechnology.

“The reliable, efficient and inexpensive detection and identification of specific strains of microorganisms such as E. coli is a grand challenge in biology and the health sciences,” said Hihath. “Our technique could pave the way for rapid, straightforward detection of pathogens, antimicrobial resistant bacterial strains and biomarkers for cancer.”

Hihath and his team focused on E. coli since it is a common pathogen that could easily be found in the food supply, but might not cause illness in a benign form. The worst strain of E. coli, called E. coli O157:H7, produces a toxic substance called Shiga toxin that causes bloody diarrhea, kidney failure and even death.

Single-molecule break junction devices consist of two metal electrodes with atomically sharp interfaces that are brought into contact in a liquid solution of interest, such as a solution containing RNA sequences from E.coli. As the electrodes are brought into contact and pulled apart, an electrical bias is applied and the current is measured. This process is repeated hundreds or thousands of times to determine the conductance of a single molecule.

“One of the questions we asked is how small of a change in the sequence is needed to cause a meaningful change in the electrical conductance?” said Hihath. “The smallest thing we can change is a single-base, so we decided to see if a single-base change can be measured.”

By testing short sequences of RNA bound to DNA with chemical linkers, the team examined an E. coli sequence that would produce Shiga toxin. Their findings showed that changes in the electrical resistance of RNA due to a single-base change could be measured, which would allow them to see not only if a sequence was E.coli, but the specific strain of E.coli that produces Shiga toxin.

“A system that could selectively identify short sequences of DNA or RNA opens up new avenues for developing an electronic sensor platform for a wide range of applications,” he adds. “Eventually, we want to get to the point where we can extract RNA samples from real organisms and measure their conductance on a sensing platform.”

This work was supported in part by the National Science Foundation.

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UCLA Engineers Invent Small, Inexpensive RNA Detector to Quickly Test for Infections

Today, most medical tests to detect bacteria or viruses in body fluids take hours or days to process. Now, researchers at the UCLA Samueli School of Engineering have invented a new technology to detect genetic material—such as that in microorganisms that cause disease—in a way that’s faster, cheaper, and does not require sophisticated equipment. A team led by Harold Monbouquette, professor of chemical and biomolecular engineering, and Jacob Schmidt, professor of bioengineering, patented their device, published the first description of the technology, and established a start-up company to begin developing prototypes of a commercial version of the detector.

“We’re working on developing a device that’s useful right at the point of care,” said Monbouquette, who is also the school’s associate dean for research and physical resources. “That means it can be easily run in a doctor’s office, clinic, or emergency room without samples being sent away to a lab.”
Nearly a decade ago, Monbouquette and Schmidt crossed paths—at a UCLA poster session—and discovered that they were both working on approaches to simplify the detection of nucleic acids, the DNA or RNA that defines cells’ and organisms’ identities. Rather than compete with each other, they teamed up and combined their ideas.

The new device relies on a 1-centimeter square glass slide that has an electric current flowing through it and is contacted by tiny plastic beads, each around 800 nanometers wide— less than one one-thousandth of a millimeter. Every plastic bead has molecules called peptide nucleic acid (PNA) coating its surface.

The PNAs are designed to bind specifically to whatever nucleic acid the test is detecting. If a doctor is interested in whether a patient has chlamydia, for instance, the PNA will only bind to a stretch of RNA that’s unique to chlamydia. If there’s even a tiny amount of that exact bit of RNA present, it will bind to the corresponding PNA and immediately interrupt the flow of the electric current, telling the doctor that the disease is present.

“It’s like a yes-or-no pregnancy test,” explained Monbouquette. “And that’s adequate for a lot of pathogens — you just want to know whether a patient is infected or not.”

In a recent paper, published in the journal Lab on a Chip in July, Monbouquette and Schmidt described the detection method’s high sensitivity— able to detect the equivalent of one bacterium in half a of a wine glass. Even when a mixture of bacteria was tested, the chip could detect the presence of Escherichia coli when there were 1 million-times more RNA molecules from another bacterium, Pseudomonas putida. Moreover, the technique had no false positive results during the initial experiments.

The researchers think the technology—once it’s adapted to a form that can be mass-produced—will give doctors the ability to run faster labs to diagnose a variety of infections.

“There are a lot of diseases where the doctor takes a sample, sends it off to the lab, and doesn’t get results for days,” said Schmidt. “We can do it in 15 minutes or less, which means you can sit in the waiting room, wait for an answer, and potentially start getting treated before you ever leave the doctor’s office.”

He envisions a small toaster-sized device in which different cartridges— each containing the glass slides for a specific pathogen— can be inserted depending on what test needs to be run. It’s hard to predict the cost since a manufacturable prototype hasn’t been developed yet, but Schmidt said each test will be “easily under $50.”

With that kind of application and price point in mind, Monbouquette and Schmidt have founded Electronucleics, a company that will oversee further development of the nucleic acid detection technology. Their first goal is to streamline the processing needed for samples to be run on the machine—in the lab, they’ve been using filters, pipets, test tubes and a handful of reagents to process samples before injecting them into the detector. Next, they will work with an outside firm to develop a planned prototype featuring push-button simplicity.

They plan to first optimize the technology for use in detecting gonorrhea and chlamydia infections—there’s an unmet need for rapid diagnosis of these sexually transmitted diseases, the researchers said. But they envision other applications in the future, including the diagnosis of respiratory illnesses and flu, or the detection of pathogens in food and water.

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