Powerful Sequencing Tool Helps Identify Infectious Diseases in Mali

An advanced diagnostic tool used in an observational clinical study in Bamako, Mali, helped identify infectious viruses in hospital patients that normally would have required many traditional tests. Scientists, led by the National Institute of Allergy and Infectious Diseases (NIAID), designed the study to help physicians identify the causes of unexplained fever in patients and to bring awareness to new technology in a resource-limited region.

Because malaria is the most common fever-causing illness in rural sub-Saharan Africa, most medical workers in the region presume patients with a fever have malaria. But recent NIAID work has identified dengue, Zika and chikungunya viruses – like malaria, all spread by mosquitos – in some Malian residents.

The observational study of 108 patients, published recently in The American Journal of Tropical Medicine and Hygiene, added the advanced diagnostic test, known as VirCapSeq-VERT, to traditional testing methods to identify cases of measles, SARS-CoV-2, HIV, and other viral diseases in patients. Surprisingly, more than 40% of patients were found to have more than one infection.

VirCapSeq-VERT is the virome capture-sequencing platform for vertebrate viruses, a powerful DNA sequencing technique capable of finding all viruses known to infect humans and animals in specimens, such as plasma. VirCapSeq-VERT uses special probes that capture all virus DNA and RNA in a specimen, even if the researcher does not know which specific virus to look for. Scientists then sequence the captured DNA and RNA to identify viruses present to solve the mystery of which viral infection(s) a patient has.

In the study, the researchers recommend that combining VirCapSeq-VERT with traditional diagnostic tests could greatly assist physicians “in settings with large disease burdens or high rates of coinfections and may lead to better outcomes for patients.”

Scientists from NIAID’s Division of Clinical Research collaborated on the project from July 2020 to October 2022 with colleagues from the University of Sciences, Techniques, and Technologies of Bamako, Mali, and Columbia University.

Reference: A Koné, et al. Adding Virome Capture Metagenomic Sequencing to Conventional Laboratory Testing Increases Unknown Fever Etiology Determination in Bamako, Mali. The American Journal of Tropical Medicine and Hygiene DOI: https://doi.org/10.4269/ajtmh.24-0449 (2024).

This blog is the fifth in a series about the future of NIAID's HIV clinical research enterprise. For more information, please visit the HIV Clinical Research Enterprise page.

The outcomes of HIV clinical trials are often determined by precisely and accurately measuring how specific interventions work biologically in people. Whether tracking immune responses to a preventive vaccine candidate, monitoring changes to the amount of virus in the body, or screening for certain adverse events after administering a novel therapeutic, study teams routinely interact with clinical trial participants to safely obtain, store, transport, and analyze tissue and bodily fluid samples to answer important scientific questions about the impact of an HIV intervention in a laboratory. High quality, reliable laboratory infrastructure is critical to the accuracy and validity of clinical trial results. 

More than 150 NIAID-supported laboratories in 20 countries are addressing the diverse scientific programs of the four clinical trials networks in the Institute’s HIV clinical research enterprise. Since the start of HIV clinical research, laboratory capacities have grown in scope to support an increasing number of global clinical trials, emerging complexities in study protocol design and laboratory testing demands and evolving regulatory requirements for research and licensure.

NIAID is engaging research partners, community representatives, and other public health stakeholders in a multidisciplinary evaluation of its HIV clinical trials networks’ progress toward short- and long-term scientific goals. This process assesses knowledge gained since the networks were last awarded in 2020 to identify an essential path forward based on the latest laboratory and clinical evidence. Future NIAID HIV clinical research investments build on the conclusions of these discussions. 

In the next iteration of HIV clinical trials networks, laboratory functions will continue to evolve to align with scientific priorities and research approaches. Networks will support small early-phase trials, large registrational trials and implementation science research to examine preventive vaccine candidates and non-vaccine prevention interventions, antiviral treatments, HIV curative strategies, and therapies to improve the clinical outcomes of people affected by and living with HIV. Selected studies also will rely on high quality laboratory resources to examine interventions for tuberculosis, hepatitis, mpox and other infectious diseases. Clinical trial networks will need to employ a variety of laboratory types to achieve these objectives.  To increase flexibility and ensure the timeliness and the high quality standards the HIV field relies on for evidence that informs science, licensure and equitable practice, NIAID will have the ultimate authority for laboratory selection and approval.

Efficiency and Versatility 

Laboratory assays for HIV clinical trials continue to expand in quantity and complexity and require proportionate technical expertise and management. Future clinical research needs will include immunologic, microbiologic, and molecular testing, as well as standard chemistries and hematologic assays, with fluctuating volumes across a global collection of research sites. Balancing capacity, efficiency, scalability, and cost will require a mixed methods approach. These may include centralized laboratory testing where feasible and advantageous for protocol-specified tests; standardized processes for rapid assessment and approval of new network laboratories; and validated third-party outsourcing of routine assays to ensure timely turnaround when demands surge. 

Quality and Standardization

Ensuring consistent laboratory operations and high quality laboratory data will require continued compliance with the NIAID Division of AIDS Good Clinical Laboratory Practices and other applicable regulatory guidelines, ongoing external quality assurance monitoring, strong inventory management, importation and exportation expertise, and data and specimen management.

The research community plays an essential role in shaping NIAID’s scientific direction and research enterprise operations. We want to hear from you. Please share your questions and comments at NextNIAIDHIVNetworks@mail.nih.gov.

About NIAID’s HIV Clinical Trials Networks

The clinical trials networks are supported through grants from NIAID, with co-funding from and scientific partnerships with NIH’s National Institute of Mental Health, National Institute on Drug Abuse, National Institute on Aging, and other NIH institutes and centers. There are four networks—Advancing Clinical Therapeutics Globally for HIV/AIDS and Other Infections, the HIV Vaccine Trials Network, the HIV Prevention Trials Network, and the International Maternal Pediatric Adolescent AIDS Clinical Trials Network.

Findings From NIH-Funded Study Could Facilitate Early Treatment of Neonatal Sepsis

Scientists have identified a four-gene signature detectable in newborns’ blood at birth that predicts before symptom onset whether a baby will develop sepsis during the first week of life, according to a study co-funded by the National Institutes of Health’s National Institute of Allergy and Infectious Diseases (NIAID). Sepsis is a potentially life-threatening condition that arises when the body's response to infection injures its own tissues and organs. Using the newly discovered genetic signature to identify newborns who will develop sepsis could facilitate early treatment and obviate the need to give antibiotics to all newborns with suspected sepsis but lacking a definitive diagnosis. The findings were published today in the journal eBioMedicine. 

Two to 3% of newborns globally develop sepsis, and 17.6% of those babies die. The signs of sepsis in newborns—such as irritability, poor feeding and respiratory distress—are common to many illnesses. Consequently, clinicians sometimes misdiagnose newborn sepsis or suspect it too late, leading to death. If a clinician does suspect that a newborn has sepsis, they give the baby antibiotics pending confirmatory laboratory diagnosis of infection. The most common diagnostic technique takes several days, however, and is often inconclusive. As a result, clinicians often must decide between stopping antibiotics early and risking under-treatment, or giving antibiotics based only on a clinical diagnosis and risking serious side effects and development of antimicrobial resistance.

The NIAID-supported study aimed to find a way to accurately predict sepsis in newborns so it can be diagnosed and treated early while avoiding unnecessary antibiotic use. The researchers conducted their study in a subset of 720 initially healthy, full-term newborns who were enrolled in a larger clinical trial at two community health centers in The Gambia, West Africa. Blood was collected from all babies at birth.

Thirty-three infants were hospitalized within the first month of life for clinical signs suggestive of sepsis. Of those, 21 babies were diagnosed with sepsis, including 15 within the first week of life, which is considered early-onset sepsis. Twelve babies were diagnosed with non-septic localized infections. The researchers matched these 33 babies with 33 healthy controls and analyzed their blood to identify genes that were comparatively more active or less active at birth in each of the four groups. Using machine learning methods, the researchers detected four genes that were comparatively more active at birth only in those newborns who developed early-onset sepsis. The four-gene signature was 92.5% accurate at predicting at birth which of the 66 infants would develop early-onset sepsis. 

The researchers tested the predictive accuracy of this gene signature in a different group of 12 infants whose blood had been collected soon after birth. Half had developed early-onset sepsis, while the other half had remained healthy. The four-gene signature predicted sepsis with 83% accuracy in this group. Further research is needed to determine how well the gene signature predicts early-onset sepsis in much larger groups of newborns.

The study was led by Robert E. W. Hancock, Ph.D.; Tobias R. Kollmann, M.D., Ph.D.; Beate Kampmann, M.D., Ph.D.; and Amy H. Lee, Ph.D. NIAID co-funded the study through its Human Immunology Project Consortium (HIPC) and Immune Development in Early Life program. 

The larger study that enrolled the 720 newborns was called Systems Biology to Identify Biomarkers of Neonatal Vaccine Immunogenicity, sponsored by Boston Children's Hospital and funded by NIAID through the HIPC. More information is available in ClinicalTrials.gov at study identifier NCT03246230.

Reference: An et al. Predictive gene expression signature diagnoses neonatal sepsis before clinical presentation. eBioMedicine DOI: 10.1016/j.ebiom.2024.105411 (2024).

Recently, the UN General Assembly held a High-Level Meeting to discuss an urgent, global public health problem: Antimicrobial resistance (AMR), which occurs when bacteria, viruses, fungi and parasites evolve to evade antibiotics and other medicines that are meant to kill them. Currently, antimicrobial resistance results in approximately 1.3 million deaths worldwide, including 35,000 in the United States each year, according to the Centers for Disease Control and Prevention. Antibiotic resistance can make infections more difficult to treat—and, unfortunately, due in part to overuse and misuse of antibiotics, antimicrobial-resistant infections are becoming more common.

The National Institute of Allergy and Infectious Diseases (NIAID), part of the National Institutes of Health, has prioritized AMR research for many years. For instance, through the Antibacterial Resistance Leadership Group (ARLG), the Institute has supported more than 60 AMR clinical research projects, involving more than 20,000 participants at sites around the world. The results of this work, combined with 10 years of coordinated efforts from across the U.S. government, have led to significant improvements in how public health officials consider antibiotics and how physicians can tackle difficult infections. As a global, evolving problem, AMR continues to demand new research and innovations from the public health sector.

One ongoing clinical trial is examining a particularly thorny facet of treating antimicrobial resistant (AMR) infections: when dealing with antibiotic-resistant infections, healthcare providers may need to try several different treatments before they identify an effective one, and tests to verify whether treatments will work can take days to complete. This delay can slow the patient’s recovery or lead to death. For example, ineffective therapy due to treatment delay is associated with high mortality among patients with bloodstream infections caused by Gram-negative bacteria. 

A new clinical trial sponsored by NIAID is currently evaluating whether the use of a rapid test of antibiotic susceptibility for bacteria growing in blood cultures improves clinical outcomes for patients with sepsis in settings that have high rates of antibiotic-resistant bacterial infections. The trial is being conducted by ARLG and will ultimately enroll roughly 900 hospitalized participants at seven locations around the world. 

The Fast Antibiotic Susceptibility Testing for Gram-Negative Bacteremia Trial (FAST) will test whether use of the VITEK^® REVEAL™ AST System, a direct-from-positive-blood-culture fast phenotypic susceptibility test (manufactured by Specific Diagnostics, San Jose, CA, a wholly owned subsidiary of bioMérieux, Inc., Salt Lake City, UT), can speed up the identification of effective antibiotics to treat bloodstream infections and lead to better patient outcomes. The VITEK^® REVEAL™ system is already available for clinical use in the European Union (and recently received FDA 510(k) Clearance (K230675) on June 20, 2024, and can identify phenotypic susceptibility of 10 different bacteria to 23 different antibiotics. Unlike standard of care susceptibility testing, which typically takes several days to provide results, the VITEK^® REVEAL™ test results are available in an average of 5.5 hours.

A recent clinical trial supported by NIAID through the ARLG showed that using a similar rapid test for antimicrobial susceptibility helped healthcare providers identify and use effective antibiotics faster than standard-of-care testing. However, that trial was unable to show whether using a rapid test led to better patient outcomes—possibly because the trial was carried out in areas with relatively low rates of antimicrobial-resistant infections. The researchers hope that data from the FAST trial will show whether the test improves outcomes in regions where many patients have antimicrobial-resistant infections.

The FAST trial has enrolled hospitalized participants with Gram-negative bacteria identified in their blood. Such bacteria include Klebsiella species and Escherichia coli. They have been randomized to one of two groups: half the participants are having their blood cultures tested with VITEK^® REVEAL™ as well as standard bacterial culture and antibiotic susceptibility testing. The other participants are serving as a control group and their blood cultures will undergo standard bacterial culture and susceptibility testing. The participants’ progress is being monitored for 30 days, as clinical staff record how quickly participants recover and whether they experience any negative effects, such as worsening or relapsing while still in the hospital, requiring readmission to the hospital later for the same problem, acquiring a new infection while in the hospital, or death.

Improved testing alone will not eliminate the threat of AMR infections. For that, healthcare providers will need better therapeutics, better means of keeping the infections from spreading, and other tools. However, when patients present to the hospital with severe infections, the initial hours of waiting for test results can make all the difference. Confirming that rapid tests actually improve patient outcomes is an important step on the road to fighting back against ever-changing pathogens.

To read more about this trial, search ClinicalTrials.gov using the identifier NCT06174649.