Single genetic test can identify multiple pathogens and can be used by the UCSF clinical laboratory team to help physicians identify difficult to diagnose diseases
Continuing improvements in gene sequencing technologies and analytical software tools are enabling clinical laboratorians to diagnosis patients who have challenging symptoms. One such example is a new genomic test developed by researchers at University California, San Francisco (UCSF). The single test analyzes both RNA and DNA to detect almost any type of pathogen that may be the cause of specific illnesses.
The test uses a genomic sequencing technique known as metagenomics next-generation sequencing (mNGS). It works by sequencing genetic material found in blood, tissue, or body fluid samples and compares the sequenced data against a broad database of known pathogens to seek a match. Instead of looking for just one pathogen at a time, mNGS analyzes all of the nucleic acids, RNA, and DNA present in a sample simultaneously to detect nearly all pathogens, including viruses, bacteria, fungi, and parasites.
The mNGS test is not intended to replace existing clinical laboratory tests, but to help physicians diagnose an illness in cases where patients are experiencing severe symptoms, and where initial, commonplace tests are ineffective. In such cases, medical professionals require additional information to achieve a proper diagnosis.
“Our technology is deceptively simple,” said Charles Chiu, MD, PhD (above), professor of laboratory medicine and infectious diseases at UCSF and senior author of the studies in a news release. “By replacing multiple tests with a single test, we can take the lengthy guesswork out of diagnosing and treating infections.” The new technology may help physicians diagnose patients who have challenging symptoms and where current clinical laboratory testing is ineffective at identifying specific pathogens. (Photo copyright: University California San Francisco.)
Diagnostic Armamentarium for Physicians
According to an article published by the American Society for Microbiology (ASM) titled, “Metagenomic Next Generation Sequencing: How Does It Work and Is It Coming to Your Clinical Microbiology Lab?” mNGS is “running all nucleic acids in a sample, which may contain mixed populations of microorganisms, and assigning these to their reference genomes to understand which microbes are present and in what proportions. The ability to sequence and identify nucleic acids from multiple different taxa [plural for taxon] for metagenomic analysis makes this a powerful new platform that can simultaneously identify genetic material from entirely different kingdoms of organisms.”
The researchers developed the mNGS test years ago and it has produced promising results, including:
Diagnosing cases of encephalitis in transplant recipients to yellow fever in their organ donors.
Helping to identify the cause of a meningitis outbreak in Mexico among surgical patients.
Detecting a case of leptospirosis in a patient who was in a medically induced coma, which prompted doctors to prescribe penicillin and resulted in the full recovery of the patient.
Identifying the cause of neurological infections such as meningitis and encephalitis. The test successfully diagnosed 86% of neurological infections in more than 4,800 spinal fluid samples.
“Our mNGS test performs better than any other category of test for neurologic infections,” said Charles Chiu, MD, PhD, professor of laboratory medicine and infectious diseases at UCSF and senior author of the two studies, in a UCSF news release. “The results support its use as a critical part of the diagnostic armamentarium for physicians who are working up patients with infectious diseases.”
FDA Breakthrough Device Designation
The UCSF test has not yet been approved by the federal Food and Drug Administration (FDA), but it was granted a “breakthrough device” designation by the agency. This classification authorizes labs to use the test as a valid diagnosis method due to its potential ability to benefit patients.
Chiu told NBC News that the test costs about $3,000 per sample and fewer than 10 labs routinely use it due to several issues.
“Traditionally, it’s been used as a test of last resort, but that’s primarily because of issues involving, for instance, the cost of the test, the fact that it’s only available in specialized reference laboratories, and it also is quite laborious to run,” he said.
This type of lab testing is not feasible for most hospitals as it is costly and complicated, and because physicians may need assistance from clinical laboratory personnel who have the appropriate expertise to properly read test results.
“This just is not something that a clinical lab will be doing until somebody commercially puts it in a box with an easy button,” Susan Butler-Wu, PhD, associate professor of clinical pathology at the University of Southern California (USC), told NBC News. “It’s not a one-stop shop. It just can be helpful as an additional tool.”
Although the technology has some limitations, Chiu says the research performed by his team “raises the possibility that we perhaps should be considering running this test earlier” in symptomatic patients. He hopes the test will be used on a widespread basis in hospitals to diagnose various illnesses in the future.
“We need to get the cost down and we need to get the turnaround times down as well,” he told NBC.
Definitive Tool for Pathogen Detection
To increase access to the technology, Chiu and his colleagues founded Delve Bio, which is now the exclusive provider of the mNGS tool created at UCSF. In December, the company announced the commercial launch of Delve Detect, a metagenomic test for infectious diseases. According to its website, Delve Detect “offers genomic testing of cerebrospinal fluid (CSF) for more than 68,000 pathogens, with 48-hour turnaround time and metagenomics experts readily available to discuss results.”
“These findings support including mNGS as a core tool in the clinical workup for CNS [central nervous system] infections,” said Steve Miller, MD, PhD, UCSF volunteer clinical professor, laboratory medicine, and chief medical officer of Delve Bio in the UCSF news release. “mNGS offers the single most unbiased, complete and definitive tool for pathogen detection. Thanks to its ability to quickly diagnose an infection, mNGS helps guide management decisions and treatment for patients with meningitis and encephalitis, potentially reducing healthcare costs down the line.”
This mNGS test may prove to have the potential to greatly improve medical care for some infections and possibly expedite the detection of new viral threats. It is probable that clinical laboratories will soon be learning about and performing more tests of this nature in the future.
Screening and analysis of ocean samples also identified a possible missing link in how the RNA viruses evolved
An international team of scientists has used genetic screening and machine learning techniques to identify more than 5,500 previously unknown species of marine RNA viruses and is proposing five new phyla (biological groups) of viruses. The latter would double the number of RNA virus phyla to 10, one of which may be a missing link in the early evolution of the microbes.
Though the newly-discovered viruses are not currently associated with human disease—and therefore do not drive any current medical laboratory testing—for virologists and other microbiologists, “a fuller catalog of these organisms is now available to advance scientific understanding of how viruses evolve,” said Dark Daily Editor-in-Chief Robert Michel.
“While scientists have cataloged hundreds of thousands of DNA viruses in their natural ecosystems, RNA viruses have been relatively unstudied,” wrote four microbiologists from Ohio State University (OSU) who participated in the study in an article they penned for The Conversation.
The OSU study authors included:
Ahmed Zayed, PhD, research scientist, Dept. of Microbiology, OSU.
Matthew Sullivan, PhD, Professor of Microbiology and Director of the Center of Microbiome Science at OSU.
“RNA viruses are clearly important in our world, but we usually only study a tiny slice of them—the few hundred that harm humans, plants and animals,” explained Matthew Sullivan, PhD (above), Director, Center of Microbiome Science, in an OSU news story. Sullivan led the OSU research team. “We wanted to systematically study them on a very big scale and explore an environment no one had looked at deeply, and we got lucky because virtually every species was new, and many were really new,” he added. (Photo copyright: University of Ohio.)
RNA versus DNA Viruses
In contrast to the better-understood DNA virus, an RNA virus contains RNA instead of DNA as its genetic material, according to Samanthi Udayangani, PhD, in an article she penned for Difference Between. Examples of RNA viruses include:
One major difference, she explains, is that RNA viruses mutate at a higher rate than do DNA viruses.
The OSU scientists identified the new species by analyzing a database of RNA sequences from plankton collected during a series of ocean expeditions aboard a French schooner owned by the Tara Ocean Foundation.
“Plankton are any aquatic organisms that are too small to swim against the current,” the authors explained in The Conversation. “They’re a vital part of ocean food webs and are common hosts for RNA viruses.”
The team’s screening process focused on the RNA-dependent RNA polymerase (RdRp) gene, “which has evolved for billions of years in RNA viruses, and is absent from other viruses or cells,” according to the OSU news story.
“RdRp is supposed to be one of the most ancient genes—it existed before there was a need for DNA,” Zayed said.
The RdRp gene “codes for a particular protein that allows a virus to replicate its genetic material. It is the only protein that all RNA viruses share because it plays an essential role in how they propagate themselves. Each RNA virus, however, has small differences in the gene that codes for the protein that can help distinguish one type of virus from another,” the study authors explained.
The screening “ultimately identified over 44,000 genes that code for the virus protein,” they wrote.
Identifying Five New Phyla
The researchers then turned to machine learning to organize the sequences and identify their evolutionary connections based on similarities in the RdRp genes.
“The more similar two genes were, the more likely viruses with those genes were closely related,” they wrote.
The technique classified many of the sequences within the five previously known phyla of RNA viruses:
But the researchers also identified five new phyla—including two dubbed “Taraviricota” and “Arctiviricota”—that “were particularly abundant across vast oceanic regions,” they wrote. Taraviricota is named after the Tara expeditions and Arctiviricota gets its name from the Arctic Ocean.
They speculated that Taraviricota “might be the missing link in the evolution of RNA viruses that researchers have long sought, connecting two different known branches of RNA viruses that diverged in how they replicate.”
In addition to the five new phyla, the researchers are proposing at least 11 new classes of RNA viruses, according to the OSU story. The scientists plan to issue a formal proposal to the International Committee on Taxonomy of Viruses (ICTV), the body responsible for classification and naming of viruses.
Studying RNA Viruses Outside of Disease Environments
“As the COVID-19 pandemic has shown, RNA viruses can cause deadly diseases. But RNA viruses also play a vital role in ecosystems because they can infect a wide array of organisms, including microbes that influence environments and food webs at the chemical level,” wrote the four study authors in The Conversation. “Mapping out where in the world these RNA viruses live can help clarify how they affect the organisms driving many of the ecological processes that run our planet. Our study also provides improved tools that can help researchers catalog new viruses as genetic databases grow.”
This remarkable study, which was partially funded by the US National Science Foundation, will be most intriguing to virologists and microbiologists. However, clinical laboratories also should be interested in the fact that the catalog of known viruses has just expanded by 5,500 types of RNA viruses.
Genomic analysis of pipes and sewers leading from the National Institutes of Health Clinical Care Center in Bethesda, Md., reveals the presence of carbapenem-resistant organisms; raises concern about the presence of multi-drug-resistant bacteria previously undetected in hospital settings
If hospitals and medical laboratories are battlegrounds, then microbiologists and clinical laboratory professionals are frontline soldiers in the ongoing fight against hospital-acquired infections (HAIs) and antibiotic resistance. These warriors, armed with advanced testing and diagnostic skills, bring expertise to antimicrobial stewardship programs that help block the spread of infectious disease. In this war, however, microbiologists and medical laboratory scientists (AKA, medical technologists) also often discover and identify new and potential strains of antibiotic resistance.
Potential Source of Superbugs and Hospital-Acquired Infections
According to the mBio study, “Carbapenemase-producing organisms (CPOs) are a global concern because of the morbidity and mortality associated with these resistant Gram-negative bacteria. Horizontal plasmid transfer spreads the resistance mechanism to new bacteria, and understanding the plasmid ecology of the hospital environment can assist in the design of control strategies to prevent nosocomial infections.”
Karen Frank, MD, PhD (above), is Chief of the Microbiology Service Department at the National Institutes of Health and past-president of the Academy of Clinical Laboratory Physicians and Scientists. She suggests hospitals begin tracking the spread of the bacteria. “In the big picture, the concern is the spread of these resistant organisms worldwide, and some regions of the world are not tracking the spread of the hospital isolates.” (Photo copyright: National Institutes of Health.)
Frank’s team used Illumina’s MiSeq next-generation sequencer and single-molecule real-time (SMRT) sequencing paired with genome libraries, genomics viewers, and software to analyze the genomic DNA of more than 700 samples from the plumbing and sewers. They discovered a “potential environmental reservoir of mobile elements that may contribute to the spread of resistance genes, and increase the risk of antibiotic resistant ‘superbugs’ and difficult to treat hospital-acquired infections (HAIs).”
Genomic Sequencing Identifies Silent Threat Lurking in Sewers
Frank’s study was motivated by a 2011 outbreak of antibiotic-resistant Klebsiella pneumoniae bacteria that spread through the NIHCC via plumbing in ICU, ultimately resulting in the deaths of 11 patients. Although the hospital, like many others, had dedicated teams working to reduce environmental spread of infectious materials, overlooked sinks and pipes were eventually determined to be a disease vector.
In an NBC News report on Frank’s study, Amy Mathers, MD, Director of The Sink Lab at the University of Virginia, noted that sinks are often a locus of infection. In a study published in Applied and Environmental Microbiology, another journal of the ASM, Mathers noted that bacteria in drains form a difficult to clean biofilm that spreads to neighboring sinks through pipes. Mathers told NBC News that despite cleaning, “bacteria stayed adherent to the wall of the pipe” and even “splashed out” into the rooms with sink use.
During the 2011-2012 outbreak, David Henderson, MD, Deputy Director for Clinical Care at the NIHCC, told the LA Times of the increased need for surveillance, and predicted that clinical laboratory methods like genome sequencing “will become a critical tool for epidemiology in the future.”
Frank’s research fulfilled Henderson’s prediction and proved the importance of genomic sequencing and analysis in tracking new potential sources of infection. Frank’s team used the latest tools in genomic sequencing to identify and profile microbes found in locations ranging from internal plumbing and floor drains to sink traps and even external manhole covers outside the hospital proper. It is through that analysis that they identified the vast collection of CPOs thriving in hospital wastewater.
In an article, GenomeWeb quoted Frank’s study, noting that “Over two dozen carbapenemase gene-containing plasmids were identified in the samples considered” and CPOs turned up in nearly all 700 surveillance samples, including “all seven of the wastewater samples taken from the hospital’s intensive care unit pipes.” Although the hospital environment, including “high-touch surfaces,” remained free of similar CPOs, Frank’s team noted potential associations between patient and environmental isolates. GenomeWeb noted Frank’s findings that CPO levels were in “contrast to the low positivity rate in both the patient population and the patient-accessible environment” at NIHCC, but still held the potential for transmission to vulnerable patients.
Since carbapenems are a “last resort” antibiotic for bacteria resistant to other antibiotics, the NIHCC “reservoir” of CPOs is a frightening discovery for physicians, clinical laboratory professionals, and the patients they serve.
The high CPO environment in NIHCC wastewater has the capability to spread resistance to bacteria even without the formal introduction of antibiotics. In an interview with Healthcare Finance News, Frank indicated that lateral gene transfer via plasmids was not only possible, but likely.
“The bacteria fight with each other and plasmids can carry genes that help them survive. As part of a complex bacterial community, they can transfer the plasmids carrying resistance genes to each other,” she noted. “That lateral gene transfer means bacteria can gain resistance, even without exposure to the antibiotics.”
The discovery of this new potential “reservoir” of CPOs may mean new focused genomic work for microbiologists and clinical laboratories. The knowledge gained by the discovery of CPOs in hospital waste water and sinks offers a new target for study and research that, as Frank concludes, will “benefit healthcare facilities worldwide” and “broaden our understanding of antimicrobial resistance genes in multi-drug resistant (MDR) bacteria in the environment and hospital settings.”
Obesity may be one of several health conditions and diseases where the human microbiome can be harnessed for diagnostic and therapeutic uses
Microbiologists could soon be the front lines in the nation’s fight against obesity and possibly other chronic diseases. New research underway at Vanderbilt University could lead to a host of new clinical laboratory tests that use engineered microbes.
This research is revealing how the human microbiome can be the source of new biomarkers for diagnostic tests and therapeutic drugs. In fact, early research findings point to the possibility that pathologists and clinical laboratories may eventually use the human microbiome in their daily work.
Engineering Bacteria to Battle Obesity
The human microbiome has remained largely unstudied. One reason why this is true is that it has been difficult to recreate, in the laboratory, the optimal conditions to allow these microbes to grow and thrive just as they do in the human body. However, as researchers continue to make new discoveries about this community of micro-organisms, there is optimism that elements of the human microbiome can be used to develop novel medical laboratory tests. (more…)
Research breakthrough heralded as key insight that can lead to more accurate clinical laboratory tests and more effective antibiotics for treating E. Coli infections
Antibiotic-resistant strains of bacteria are one of healthcare’s biggest threats to patient safety and improved patient outcomes. Now advanced gene sequencing has given researchers a startling new understanding of how Escherichia coli (E. coli) has developed resistance to antibiotics.
This discovery may have a major impact on microbiology labs in hospitals, because they do so much of the medical laboratory testing to detect and identify infections. These new research findings also demonstrate to pathologists how quickly genome analysis can generate new knowledge about diseases and their causes. (more…)
