Dogs May Be More Sensitive at Detecting COVID-19 More Rapidly and Accurately than Current Tests

Our fur babies may provide a cheaper, faster and more effective way to detect COVID-19, and could be a key tool in future pandemics, a new review of recent research suggests. The review, published in De Gruyter’s Journal of Osteopathic Medicine, found that scent dogs are as effective, or even more effective, than conventional COVID-19 tests such as RT-PCR.

Dogs possess up to 300 million olfactory cells, compared to just 5 or 6 million in humans, and use one-third of their brains to process scent information, compared with just 5% for humans. Dogs trained to recognize specific volatile organic compounds created in the body during disease have successfully identified patients with certain cancers, Parkinson’s and diabetes.

Prof. Tommy Dickey of the University of California, Santa Barbara and Heather Junqueira of BioScent Detection Dogs reviewed 29 studies where dogs were used to detect COVID-19. The studies were performed using over 31,000 samples by over 400 scientists from more than 30 countries using 19 different dog breeds. In some studies, the scent dogs sniffed people directly, sometimes in public places as a health screening. In others, the dogs sniffed patient samples such as sweat, saliva or urine samples.

In the majority of studies, the scent dogs demonstrated similar or better sensitivity and specificity than the current gold-standard RT-PCR tests or antigen tests. In one study, four of the dogs could detect the equivalent of less than 2.6 x 10−12 copies of viral RNA per milliliter. This is equivalent to detecting one drop of any odorous substance dissolved in ten and a half Olympic-sized swimming pools and is three orders of magnitude better than modern scientific instruments. 

The dogs could detect COVID-19 in symptomatic, pre-symptomatic and asymptomatic patients, along with new COVID variants and even long COVID. A major benefit of using the dogs was their speed – they could provide a result in seconds to minutes, and did not require expensive lab equipment or create mountains of plastic waste, unlike conventional diagnostic approaches.

“Although many people have heard about the exceptional abilities of dogs to help humans, their value to the medical field has been considered fascinating, but not ready for real-world medical use,” said Prof. Dickey. “Having conducted this review, we believe that scent dogs deserve their place as a serious diagnostic methodology that could be particularly useful during pandemics, potentially as part of rapid health screenings in public spaces. We are confident that scent dogs will be useful in detecting a wide variety of diseases in the future."

Prof. Dickey and Heather Junqueira added that they feel that the impressive international COVID scent dog research described in their paper, perhaps for the first time, demonstrates that medical scent dogs are ready for mainstream medical applications.

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Blood culture vs. Metagenomic Next-Generation Sequencing for the Detection of Pathogenic Microbes in Patients with Bloodstream Infections

A recent study published in Scientific Reports compared the detection of pathogenic microbes between blood culture and metagenomic next-generation sequencing (mNGS) in patients suspected of bloodstream infections (BSIs).

Background

BSI can manifest as fungemia, viremia, and bacteremia, increasing hospitalization duration and costs. BSI incidence has increased over the past years. As such, research focus on the early identification of pathogens has been increasing. mNGS offers several advantages over a conventional blood culture assay, such as high speed and a wide range of pathogen detection. Nevertheless, mNGS is not commonly used for BSIs due to its high costs.

About the study

In the present study, researchers compared the pathogen detection consistency of blood culture assay and mNGS. They retrospectively evaluated patients with suspected BSIs admitted to the emergency department of a Chinese hospital between January 2020 and June 2022. Eligible patients aged 16 or older had chills and body temperature above 38.5 °C, with antibiotic usage longer than three days. mNGS was performed on the day of sampling.

Blood samples were collected from two anatomical sites and cultured for up to seven days using standard microbiological procedures. For mNGS, DNA was isolated, and DNA libraries were prepared. Quality control-passed libraries were sequenced. Short, adapter, low-complexity, and low-quality reads were removed. The remaining reads were aligned to microbial genome databases.

Unique reads had > 90% identity and > 80% alignment, with the ratio of sub-optimal to optimal alignment score lower than 0.8. Patients’ medical records were reviewed. The team obtained data on demographics, comorbidities, laboratory tests, mechanical ventilation, central venous intubation, sequential organ failure assessment (SOFA), and in-hospital death. Logistic regression was performed to identify risk factors for a positive blood culture or mNGS.

Findings

The study included 99 patients suspected of BSIs. They were predominantly males and aged 63 on average. Inflammatory indicators, such as C-reactive protein (CRP), white blood cells (WBCs), and procalcitonin (PCT), were elevated in patients. Mechanical ventilation and central venous intubation were required for 36.3% and 56.5% of patients, respectively.

The median SOFA score was six. In-hospital death occurred in 37 patients. There was a statistically significant difference in the number of patients positive on a blood culture assay and mNGS. Sixty-five patients were positive on mNGS compared to 12 on blood culture assays. mNGS detected a virus in 22 patients and fungi or bacteria in the remaining patients.

By contrast, blood cultures only detected fungi or bacteria. The most common pathogens detected through blood cultures were Staphylococcus haemolyticus, Klebsiella pneumoniae, and Enterococcus faecalis. Escherichia coli, K pneumoniae, and Salmonella enterica were the most commonly identified pathogens in mNGS.

The detection rate was significantly higher with mNGS than with blood culture; the concordance in detecting fungi and bacteria was 12% between mNGS and blood culture. Logistic regression identified lower WBC count, body mass index (BMI), and elevated CRP as risk factors for pathogen detection in mNGS.

Increased age and CRP, rheumatic diseases, and alcohol abuse were the risk factors for detecting fungi or bacteria in mNGS. Current smoking status and gender were identified as the risk factors for positive blood cultures. In-hospital mortality rates were 38.4% and 35.2% in mNGS-positive and -negative cases and 38.4% and 37.2% in blood culture-positive and -negative cases, respectively.

Conclusions

Early diagnosis of BSIs is crucial; blood cultures are currently the gold standard for detection, producing results within three to five days. Besides, other techniques based on multiplex real-time polymerase chain reaction (PCR) and metagenomics have been increasing. Of these, mNGS offers rapid results and could help improve patient management.

The rate of positive blood cultures was 13.1%, consistent with previous reports. While mNGS and blood cultures identified fungi and bacteria, only mNGS detected viruses. The positive rate of mNGS was 3.31-fold higher than that of blood cultures. The mortality rate was 38.4%, higher than in prior studies.

There were no significant differences in mortality rates between culture- or mNGS-positive and negative patients. The study revealed higher positivity rates with mNGS than with conventional blood cultures, and their combined use could maximize the detection rate of bloodstream pathogens.

Reference

Zhou, Y. et al. (2023) "Comparison of pathogen detection consistency between metagenomic next-generation sequencing and blood culture in patients with suspected bloodstream infection", Scientific Reports, 13(1). doi: 10.1038/s41598-023-36681-5 https://www.nature.com/articles/s41598-023-36681-5

Source: News-Medical.Net

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Tiny Nanopores Contribute to Faster Identification of Diseases

In a collaboration with Groningen University, Professor Jørgen Kjems and his research group at Aarhus University have achieved a remarkable breakthrough in developing tiny nano-sized pores that can contribute to better possibilities for, among other things, detecting diseases at an earlier stage.

Their work, recently published in the journal ACS Nano, shows a new innovative method for finding specific proteins in complex biological fluids, such as blood, without having to label the proteins chemically. The research is an important milestone in nanopore technology, and could revolutionize medical diagnostics.

Nanopores are tiny channels formed in materials, that can be used as sensors. The researchers, led by Jørgen Kjems and Giovanni Maglia (Groningen Univ.), have taken this a step further by developing a special type of nanopore called ClyA with scanner molecules, called nanobodies, attached to it.

These nanobodies, derived from antibodies, are capable of recognizing different proteins with astonishing accuracy. In this study, the researchers attached nanobodies to ClyA, using a DNA adapter. By using a series of nanobodies, they were able to create many different nanopore sensors, which could detect a variety of proteins of different sizes.

The research team created nanopores with specialized nanobodies attached, which have the ability to detect the Spike protein of SARS-CoV-2 (the virus that causes COVID-19) and a protein marker for breast cancer called urokinase-type plasminogen activator (uPA), respectively.

By measuring changes in electrical currents that are caused by the presence of these proteins, the researchers can find and identify individual proteins and even determine their concentrations. What makes this breakthrough even more remarkable is that the nanopores remained highly accurate and sensitive even when tested with complex samples like blood.

Although the nanopores are invisible to the naked eye, the signficance of this research is palpable. The existing technologies already allowed the integration of nanopores into a portable device that can utilize the nanopores' ability to scan liquids for specific molecules. Therefore, we can envision a future where patients can rapidly and accurately detect diseases like cancer or infectious diseases with a simple blood test. This could lead to earlier interventions, improved treatment outcomes, and overall improved healthcare.

Although further studies and validation are needed before this technology becomes widely available, the collaboration between these two universities brings us one step closer to this reality. The breakthrough exemplifies the power of scientific collaboration and innovation in transforming healthcare.

Reference

Xialin Zhang et al, Specific Detection of Proteins by a Nanobody-Functionalized Nanopore Sensor, ACS Nano (2023). DOI: 10.1021/acsnano.2c12733

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New Sensor Chip Advances Rapid, Cost-Effective Disease Diagnostics

Texas A&M AgriLife Research scientists and collaborators at Iowa State University have developed a sensor chip that can detect many disease pathogens with 10 times the sensitivity of currently available methods.The chip also eliminates the need for chemical dye reagents typically used in the diagnostic process. The new technology shows promise for rapid, low-cost point-of-care diagnostic capabilities in plants, foods, animals and humans, including detecting foodborne pathogens, bird flu and COVID-19.

An abstract diagram showing the basic configuration of the LAMP reaction chamber and nanopore film sensor containing immobilized LAMP products.

Results from the new sensor are available in about 30 minutes.

In their research, published in ASC Sensors, scientists used the new sensor to detect Phytophthora infestans. The pathogen causes globally devastating late blight disease — a particular threat to potato and tomato crops.

The research was co-led by Jinping Zhao, Ph.D., AgriLife Research postdoctoral research scientist in Dallas, and Subin Mao, a Ph.D. candidate in electrical and computer engineering at Iowa State University. Serving as corresponding authors were collaborators Junqi Song, Ph.D., associate professor and plant immunity research lead with AgriLife Research in Dallas, and Long Que, Ph.D., professor of electrical engineering at Iowa State University. Seed grants from each university funded the research.

“This research advances technologies that have emerged as some of our greatest opportunities for improving agriculture, food safety and human health,” Song said. “Our publication represents a step toward realizing these powerful tools against diseases.”

Building on existing technologies

The new sensor improves upon a technique known as loop-mediated isothermal amplification, or LAMP, which is widely used to detect pathogens by amplifying their DNA.

Detection of LAMP products amplified from templates, such as pathogen DNA, often requires that the products be “labeled” by using fluorescence dyes — a costly process with low sensitivity. The new sensor diagnoses pathogens without such reagents and at high sensitivity. It also eliminates a lengthy DNA purification process that creates challenges for point-of-care use.

The new chip consists of a nanopore thin-film sensor inside a special reaction chamber. Primers are uniquely designed to be immobilized on the nanofilm, causing amplified LAMP products to become bound to the sensor, which produces signals that can be directly and easily measured with a portable spectrometer.

What’s next

The LAMP chip offers a new portable platform to detect pathogens using label-free sensors with ultrasensitivity. The research team will now work to further enhance sensitivity to a subattomolar or even lower level.

The team aims to offset current challenges to detecting and distinguishing pathogen species and strains with high-sequence similarities. They will also work to improve the specificity of detections and establish quantitative detection by integrating artificial intelligence and CRISPR gene-editing technologies.

Their goal is to achieve a viable product for broad adoption in plant, animal and human health point-of-care applications.

Source: Texas A&M University AgriLife TODAY

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Lab-on-a-chip Device Enables Fast, Cheap Tuberculosis

Researchers have used laboratory-on-a-chip technology to create a low cost, sensitive tuberculosis (TB) test that could improve disease detection in high-endemic, under-resourced areas.

Writing in The Journal of Molecular Diagnostics, collaborators at St George’s University of London and QuantuMDx describe the limitations of existing TB tests. Smear microscopy, in which sputum samples are smeared onto microscope slides for examination, is cheap and requires minimal facilities but suffers from poor sensitivity, and the quality of the results can vary between sites and operators.

Growing the bacteria in sputum samples improves sensitivity; however, TB grows slowly, delaying diagnosis, and biosafety level 3 facilities are required to perform the work. Those factors are limitations in parts of the world where resources are restricted.

The St George’s and QuantuMDx collaborators identified dielectrophoresis as a way to improve testing for TB. Dielectrophoresis enables the selective attraction or repulsion of specific particles or cells based on their properties. Using the technique, the researchers isolated the bacteria that cause TB from the rest of the content of sputum samples. 

“This chip-based technology exploits the physiological property of the TB bacteria to be specifically collected onto the device so that small numbers can be visualized on the chip electrodes and act as a visual readout to replace the lab-based sputum smear/microscopic methods, which typically have low detection rates and require training laboratory staff, at molecular-like sensitivities and at a fraction of the price,” QuantuMDx CEO Jonathan O’Halloran, PhD, said in a statement (see below).

Purification of the bacterial population serves two purposes. First, visual analysis of the purified sample may enable the diagnosis of TB infection, serving as a substitute for smear microscopy. Second, the sample is then ready for further testing such as quantitative PCR and genotypic drug-susceptibility analysis that can confirm TB infection and show whether the pathogen is resistant to certain treatments. 

The researchers used a panel of 50 characterized sputum samples to optimize the prototype, and then compared its performance to culture diagnosis. A blinded screening of 100 characterized sputum samples found the prototype reported the same result as culture diagnosis for all of the smear-negative samples and for 87% of the smear-positive samples. Limiting the analysis to samples with high bacterial burdens increased the smear-positive concordance to 100%.

Work is underway to improve the prototype. Increasing sample throughput could improve sensitivity and accelerate testing, and refinements to biological processing and device design are expected to further enhance performance. 

Statement from Elsevier Press Release:

Lead investigator Philip D. Butcher, PhD, St George’s, University of London, explained, “The global burden of TB is not improving. Although it is infectious, it is highly treatable. However, affordable diagnostics suitable for implementation at the point of care are needed to reach the ‘missing millions.’ Our TB research group at St George’s has a longstanding appreciation of the global imperative for improved diagnostics for TB, and we realized that novel technologies may provide an answer. We saw an opportunity by collaborating on a new chip-based technology using dielectrophoresis to selectively isolate Mtb bacilli from sputum samples.”

The investigators describe a prototype microfluidic lab-on-a-chip system called CAPTURE-XT® from QuantuMDx that can process solubilized sputum from suspected TB patients, capture Mtb bacilli for visual analysis (as a substitute for smear microscopy), and provide a purified sample for molecular confirmation by quantitative PCR (qPCR) and ultimately for genotypic drug-susceptibility analysis. CAPTURE-XT® technology relies on the principle of dielectrophoresis – a little-utilized technique that can be tuned to selectively attract or repel specific particles or cells based upon their dielectric properties. In this case, it is the Mtb bacteria that cause TB that are specifically captured and concentrated, while the other sputum contents are washed away.

After optimization using a panel of 50 characterized sputum samples, the performance of the prototype was assessed by a blinded screening of 100 characterized and bio-banked sputum samples provided by the Foundation for Innovative New Diagnostics (FIND).

Concordance with culture diagnosis was 100% for smear negative samples and 87% for smear positive samples. Of the smear positive samples, the high burden sample concordance was 100%. These results demonstrate the potential of the technology to provide a powerful sample preparation tool that could function as a front-end platform for enhanced molecular detection. This versatile tool could equally be applied as a visual detection diagnostic, potentially associated with bacterial identification for low-cost screening.

Jonathan O’Halloran, PhD, founder and Chief Executive Officer of QuantuMDx Group Ltd said, “The CAPTURE-XT® technology is truly revolutionary and will have an impact in many different diseases from sepsis to oncology (circulating tumor cells). This application in Mtb is truly exceptional as its ultra-low cost and ultra-high sensitivity will profoundly improve equitable access to quality diagnoses for hundreds of millions of people. This chip-based technology exploits the physiological property of the TB bacteria to be specifically collected onto the device so that small numbers can be visualized on the chip electrodes and act as a visual readout to replace the lab-based sputum smear/microscopic methods, which typically have low detection rates and require training laboratory staff, at molecular-like sensitivities and at a fraction of the price. Moreover, when used as a front-end to downstream cellular, protein, and molecular devices, the possibilities are almost limitless.”

Professor Butcher concluded, “Collaboration between University-based academic researchers and biotechnology industry scientists presents a way forward to develop new approaches for some of the world’s greatest healthcare challenges, such as TB. This new chip-based technology could bring diagnostics to the patients that need it and also, by more accessible case-finding, prevent the further spread of this disease.”

Co-author Heather Murton, PhD, LEX Diagnostics, Melbourn, UK, and formerly of QuantuMDx Group Ltd, said, “Tuberculosis is one of the oldest challenges faced in human healthcare. This technology has the potential to meet the expectations for a mobile TB diagnostic, and it is exciting to see a seemingly abstract physics principle successfully applied to a neglected disease area.”

TB is the 13th leading cause of death worldwide, and until COVID-19 was the leading cause of death from a single infectious disease – more than malaria and HIV. Globally it infects 10 million individuals and kills 1.4 million individuals every year, of whom 230,000 are children.

Diagnostics with increased sensitivity and expanded drug susceptibility testing are also needed to address drug resistance and diagnose low-bacterial burden cases.

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Air Monitor can Detect COVID-19 Virus Variants in 5 Minutes

Now that the emergency phase of the COVID-19 pandemic has ended, scientists are looking at ways to surveil indoor environments in real time for viruses. By combining recent advances in aerosol sampling technology and an ultrasensitive biosensing technique, researchers at Washington University in St. Louis have created a real-time monitor that can detect any of the SARS-CoV-2 virus variants in a room in about 5 minutes.

The inexpensive, proof-of-concept device could be used in hospitals and health care facilities, schools and public places to help detect CoV-2 and potentially monitor for other respiratory virus aerosols, such as influenza and respiratory syncytial virus (RSV). Results of their work on the monitor, which they say is the most sensitive detector available, are published July 10 in Nature Communications.

The interdisciplinary team of researchers from the McKelvey School of Engineering and the School of Medicine consists of Rajan Chakrabarty, the Harold D. Jolley Career Development Associate Professor of energy, environmental & chemical engineering in McKelvey Engineering; Joseph Puthussery, a postdoctoral research associate in Chakrabarty’s lab; John Cirrito, a professor of neurology at the School of Medicine; and Carla Yuede, an associate professor of psychiatry at the School of Medicine.

“There is nothing at the moment that tells us how safe a room is,” Cirrito said. “If you are in a room with 100 people, you don’t want to find out five days later whether you could be sick or not. The idea with this device is that you can know essentially in real time, or every 5 minutes, if there is a live virus in the air.”

Cirrito and Yuede had previously developed a micro-immunoelectrode (MIE) biosensor that detects amyloid beta as a biomarker for Alzheimer’s disease and wondered if it could be converted into a detector for SARS-CoV-2. They reached out to Chakrabarty, who assembled a team that included Puthussery, who had expertise in building real-time instruments to measure the toxicity of air.

To convert the biosensor from detecting amyloid beta to coronavirus, the researchers exchanged the antibody that recognizes amyloid beta for a nanobody from llamas that recognizes the spike protein from the SARS-CoV-2 virus. David Brody, MD, PhD, a former faculty member in the Department of Neurology at the School of Medicine and an author on the paper, developed the nanobody in his lab at the National Institutes of Health (NIH). The nanobody is small, easy to reproduce and modify and inexpensive to make, the researchers said.

“The nanobody-based electrochemical approach is faster at detecting the virus because it doesn’t need a reagent or a lot of processing steps,” Yuede said. “SARS-CoV-2 binds to the nanobodies on the surface, and we can induce oxidation of tyrosines on the surface of the virus using a technique called square wave voltammetry to get a measurement of the amount of virus in the sample.”

Chakrabarty and Puthussery integrated the biosensor into an air sampler that operates based on the wet cyclone technology. Air enters the sampler at very high velocities and gets mixed centrifugally with the fluid that lines the walls of the sampler to create a surface vortex, thereby trapping the virus aerosols. The wet cyclone sampler has an automated pump that collects the fluid and sends it to the biosensor for seamless detection of the virus using electrochemistry.

“The challenge with airborne aerosol detectors is that the level of virus in the indoor air is so diluted that it even pushes toward the limit of detection of polymerase chain reaction (PCR) and is like finding a needle in a haystack,” Chakrabarty said. “The high virus recovery by the wet cyclone can be attributed to its extremely high flow rate, which allows it to sample a larger volume of air over a 5-minute sample collection compared with commercially available samplers.”

Most commercial bioaerosol samplers operate at relatively low flow rates, Puthussery said, while the team’s monitor has a flow rate of about 1,000 liters per minute, making it one of the highest flow-rate devices available. It is also compact at about 1 foot wide and 10 inches tall and lights up when a virus is detected, alerting administrators to increase airflow or circulation in the room.

The team tested the monitor in the apartments of two COVID-positive patients. The real-time PCR results of air samples from the bedrooms were compared with air samples collected from a virus-free control room. The devices detected RNA of the virus in the air samples from the bedrooms but did not detect any in the control air samples.

In laboratory experiments that aerosolized SARS-CoV-2 into a room-sized chamber, the wet cyclone and biosensor were able to detect varying levels of airborne virus concentrations after only a few minutes of sampling.

“We are starting with SARS-CoV-2, but there are plans to also measure influenza, RSV, rhinovirus and other top pathogens that routinely infect people,” Cirrito said. “In a hospital setting, the monitor could be used to measure for staph or strep, which cause all kinds of complications for patients. This could really have a major impact on people’s health.”

The team is working to commercialize the air quality monitor.

Reference

Puthussery J V, Ghumra DP, McBrearty KR, Doherty BM, Sumlin BJ, Sarabandi A, Mandal AG, Shetty NJ, Gardiner WD, Magrecki JP, Brody DL, Esparza TJ, Bricker TL, Boon ACM, Yuede CM, Cirrito JR, Chakrabarty RK. Real-time environmental surveillance of SARS-CoV-2 aerosols. Nature Communications, July 10, 2023. DOI: 10.1038/s41467-023-39419-z

Funding

This research was funded by the National Institutes of Health (NIH) RADx-Rad program (U01 AA029331, U01 AA029331-S1); NIH National Institute of Neurological Disorders and Stroke Intramural Research Program, SARS-CoV-2 Assessment of Viral Evolution (SAVE) Program; and WashU-IITB Joint Master’s Program.

Source: The Source, Washington University in St. Louis

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