Malian scientists at the University of Sciences, Techniques, and Technologies of Bamako, Mali, perform VirCapSeq-VERT testing on plasma specimens from patients with unknown fevers. Shown are Dr. Fousseyni Kane (left) and Dr. Dramane Diallo (right).
Credit:Credit: Amadou Kone (CDC)
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).
Magnified image of Aedes aegypti mosquito taking a blood meal. Captured by Dr. Laura Willen, an author of the study, at NIAID’s Laboratory of Malaria and Vector Research in Bethesda, Maryland.
Our research focuses on the complex interactions between the human immune system and insect-derived molecules, and how these interactions can influence the outcomes of vector-borne diseases such as dengue, Zika, Chikungunya, and leishmaniasis. When an insect bites, it injects hundreds of arthropod molecules into the host's skin, alerting our immune system to these foreign agents. If the insect is infected with a pathogen, the microorganism is delivered along with these insect-derived molecules. Our immune response to these molecules over time can either help or hinder pathogen establishment, ultimately affecting the disease outcome.
Our work is conducted at two primary locations: the Laboratory of Malaria and Vector Research (LMVR) in Rockville, which is equipped with cutting-edge technologies, and the NIAID International Center of Excellence in Research (ICER) in Cambodia, where we conduct field observations and studies.
At LMVR-Rockville, we use advanced technologies and methodologies to explore the molecular and immunological mechanisms underlying the human response to arthropod bites and the pathogens they transmit. In Cambodia, at the NIAID ICER, we engage in extensive fieldwork to gather critical data and observations directly from affected populations. By integrating field data with laboratory findings, we aim to develop robust hypotheses that can lead to effective strategies for disease mitigation and control.
Our multidisciplinary approach allows us to bridge the gap between laboratory research and field applications. By understanding how the human immune system responds to arthropod molecules, we can identify potential targets for vaccines, therapeutics, and diagnostic tools. Additionally, our research contributes to the development of innovative vector control strategies that can reduce the incidence of these debilitating diseases.
Through collaboration with local communities, healthcare providers, and international partners, we strive to translate our scientific discoveries into practical solutions that can improve public health outcomes. Our ultimate goal is to reduce the burden of vector-borne diseases and enhance the quality of life for people living in endemic regions.
Our research aims to improve dengue prevention and treatment strategies for U.S. travelers, personnel in endemic areas, and regions with reported dengue cases, such as Hawaii, Florida, Texas, Puerto Rico, the U.S. Virgin Islands, and Guam. Enhanced predictive, management, diagnostic, and preventive measures for dengue outbreaks are particularly crucial for these at-risk regions. The development and use of prophylactic therapeutics targeting specific immune responses to mosquito bites could reduce the transmission of arboviruses, including eastern equine encephalitis, Jamestown Canyon, La Crosse, Powassan, St. Louis encephalitis, and West Nile viruses. Improved diagnostic capabilities for vector-borne diseases and emerging infections will lead to better patient outcomes.
Characterization of human immune response to ticks, mosquito, and sand fly saliva in the context of medically significant vector-borne diseases (Lyme disease, Powassan, dengue, malaria, and leishmaniasis)
Clinical and field epidemiology of the impact of mosquito saliva immunity on the outcome of dengue, Zika, and other diseases carried by mosquitos
Strategies to block vector-borne diseases by targeting the arthropod vector and interruption transmission to the human host
NIH-Funded Clinical Trial Will Evaluate New Dengue Therapeutic
A Phase 2 clinical trial will test the safety and efficacy of an experimental treatment for dengue, a viral disease transmitted by mosquitoes.
An Evaluation of Repeated Oral Doses of JNJ-64281802 Against DENV-3 Challenge
This study is hypothesizing that the highest dose of the investigational study drug is superior to receiving a placebo with respect to its antiviral activity in healthy adult participants inoculated with Dengue Serotype 3.
NIH Awards Establish Pandemic Preparedness Research Network
The Research and Development of Vaccines and Monoclonal Antibodies for Pandemic Preparedness network—called ReVAMPP—will focus its research efforts on “prototype pathogens,” representative pathogens from virus families known to infect humans, and high-priority pathogens that have the potential to cause deadly diseases. The pandemic preparedness research network will conduct research on high-priority pathogens most likely to threaten human health with the goal of developing effective vaccines and monoclonal antibodies.
An Aedes mosquito, similar to those studied by Dr. Patricia Scaraffia.
Credit:NIAID
Mosquitoes are considered one of the most dangerous animals on earth because of their broad distribution and the many pathogens they transmit to humans. Some of the most important human diseases in tropical and temperate regions of the planet are caused by mosquito-borne pathogens. Malaria, dengue, and filariasis, among other mosquito-borne diseases, kill or sicken millions of people worldwide every year.
Mosquito-borne pathogens are transmitted to the vertebrate host, such as a human, when the mosquito bites the host in search of blood. The proteins found in blood are essential for female mosquitoes: without it, they lack the resources to create eggs. Greater knowledge of the biological processes involved in the mosquito life cycle could lead to new or improved strategies to control mosquito populations.
Dr. Patricia Scaraffia, Associate Professor at the Tulane University School of Public Health and Tropical Medicine, has dedicated her career to understanding the metabolism of the mosquito Aedes aegypti that carries the pathogens responsible for dengue, Zika, chikungunya, and yellow fever to humans.NIAID reached out to Dr. Scaraffia about her team’s research.
What got you interested in studying mosquito metabolism?
I have studied the metabolism of insects that are vectors of pathogens causing human diseases since I was a graduate student at the Universidad Nacional de Cordoba, in Argentina. My Ph.D. dissertation was focused on the energy metabolism in Triatomine insects, vectors of Trypanosoma cruzi, the etiological agent of Chagas´ disease. After my dissertation, I participated as a speaker in a two-week course for PhD students entitled Biochemistry and molecular biology of insects of importance for public health. During the course, Argentinian professors encouraged me to contact the late Dr. Michael A. Wells, a leader in insect metabolism, and apply for a postdoctoral training in his lab. Soon after, I joined Dr. Wells´s lab at the University of Arizona as a research associate and opened a new line of investigation in his lab. Since then, I have never stopped working on A. aegypti mosquito metabolism. I am passionate and curious about the tremendous complexity of mosquito metabolism. It is a fascinating puzzle to work on. It constantly challenges me and my research team to think outside the box when trying to decipher the unknowns related to mosquito metabolism.
Dr. Patricia Scaraffia's work focuses on the secrets of mosquito metabolism.
Credit:Dr. Patricia Scaraffia
What are the metabolic challenges faced by mosquitoes after feeding on blood?
Female mosquitoes are a very captivating biological system. It is during blood feeding that female mosquitoes can transmit dangerous, and sometimes lethal, pathogens to humans. Interestingly, the blood that the females take could be twice their body weight, which is impressive. Female mosquitoes have evolved efficient mechanisms to digest blood meals, eliminate excess water, absorb and transport nutrients, synthesize new molecules, metabolize excess nitrogen, remove nitrogen waste, and successfully lay eggs within 72 hours! Despite significant progress in understanding how females overcome these metabolic challenges, we have not yet fully elucidated the intricate metabolic pathways, networks, and signaling cascades, nor the molecular and biochemical bases underlying the multiple regulatory mechanisms that may exist in blood-fed female mosquitoes.
What are the greatest potential benefits of understanding mosquito metabolism?
Metabolism is a complicated process that involves the entire set of chemical transformations present in an organism. A metabolic challenge faced by mosquitoes is how to break down ammonia that results from digesting a blood meal and is toxic to the mosquito. With NIAID support, we found that in the absence of a functional metabolic cycle to detoxify ammonia, A. aegypti mosquitoes use specific metabolic pathways that were believed to be non-existent in insects. This discovery has opened a new field of study.
A better understanding of mosquito metabolism and its mechanisms of regulation in A. aegypti and other mosquito species could lead us to the discovery of common and novel metabolic targets and/or metabolic regulators. It would also provide a strong foundation for the development and implementation of more effective biological, chemical and/or genetic strategies to control mosquito populations around the world.
What are the biggest challenges to studying mosquito metabolism?
We have often observed that genetic silencing or knockdown—a technique to prevent or reduce gene expression—of one or more genes encoding specific proteins involved in mosquito nitrogen metabolism results in a variety of unpredictable phenotypes based on our knowledge of vertebrate nitrogen metabolism. Notably, female mosquitoes get control of the deficiency of certain key proteins by downregulating or upregulating one or multiple metabolic pathways simultaneously and at a very high speed. This highlights the tremendous adaptive capacity of blood-fed mosquitoes to avoid deleterious effects and survive.
We have been collaborating closely with scientists that work at the University of Texas MD Anderson Cancer Center Metabolomics Core Facility, and more recently, with bioanalytical chemists that work in the Microbiome Center’s Metabolomics and Proteomics Mass Spectrometry Laboratory in Texas Children’s Hospital in Houston. Our projects are not turn-key type of projects with quick turn-round times. We have to invest considerable time and effort to successfully develop and/or optimize methods before analyzing mosquito samples. Despite these challenges, our research work keeps motivating us to unlock the metabolic mysteries that female mosquitoes hold.
Your research has focused on Aedes aegypti, the main vector of dengue, Zika, etc.Why did you choose to study this mosquito species rather than others that are also important vectors of malaria and other diseases?
My research has focused on Aedes aegypti not only because it is a vector of pathogens that pose public health threats, but also because it is genetically one of the best-characterized insect species. The availability of the Aedes aegypti genome is a great resource for a wide range of investigations. In addition, Aedes aegypti is relatively simple to rear and maintain in the lab. In my lab, we are interested in expanding our metabolic studies to other mosquito species by working in collaboration with scientists with expertise in the biology of different vectors.
What important questions remain unanswered about mosquito metabolism?
Many important questions remain unanswered about mosquito metabolism. I’d like to highlight a few of them that may help us enhance our knowledge of the mosquito as a whole organism rather than as a linear sum of its parts. For example, what are the genetic and biochemical mechanisms that drive metabolic fluxes in mosquitoes in response to internal or external alterations? How do key proteins interact with each other, and how are they post-translationally regulated to maintain mosquito metabolism? How are the metabolic networks regulated in noninfected and pathogen-infected mosquitoes? What are the critical regulatory points within the mosquito metabolism and the vector-host-pathogen interface?
While basic science will continue to be crucial in answering these questions, to successfully fight against mosquitoes, we must work together as part of a multidisciplinary team of scientists to tightly coordinate our efforts and close the gap between basic and applied science.
An Aedes mosquito and colorized 3D renderings of dengue (green), Zika (yellow), and chikungunya (red) virus capsids. Renderings based on x-ray crystallography data.
Credit:NIAID
NIAID Raises Awareness to Malaria-like Diseases in W. Africa
Dengue, Zika, Chikungunya Viruses in Mali; Disease Likely Misdiagnosed
NIAID scientists and colleagues have identified dengue, Zika and chikungunya viruses in the West African country of Mali, where health care providers likely misdiagnose patients with illness from those viruses due to unavailable diagnostic tools. Because malaria is the most common fever-causing illness in rural sub-Saharan Africa, most medical workers presume patients with a fever have malaria. The primary cause of all four infectious diseases is a mosquito bite.
Records from the Malian Health Information System show that about one-third of all patient visits to health care facilities are related to malaria, with 2.37 million clinical cases.
A new study from NIAID’s Rocky Mountain Laboratories and the University of Sciences, Techniques and Technologies in Mali aims to help spread information to medical workers about the existence of the additional viral diseases. Ideally, circulating the information will help them obtain the necessary diagnostics.
The study, published in The American Journal of Tropical Medicine and Hygiene, involved 600 residents, 200 from each of the southern Malian communities of Soromba, Bamba and Banzana. The scientists detected antibodies to dengue virus in the blood of 77.2% of the residents tested; to Zika virus in 31.2%, and to chikungunya virus in 25.8%. They detected at least one of the three viruses in 84.9% of participants, meaning just 15.1% tested negative to any of the three viruses.
Evidence of the parasites that cause malaria was found in 44.5% of those tested. Unlike malaria, however, where most cases are found in children under age 14, residents over age 50 were most likely to have been exposed to dengue, Zika or chikungunya viruses.
“Despite the high exposure risk to dengue virus in southern Mali, dengue fever cases have rarely been reported,” the researchers write. “This is likely due to the lack of diagnostic testing and the biased clinical focus on malaria in the region. Awareness of dengue virus as a cause of febrile illness needs to be urgently established in medical communities as an important public health measure.”
The scientists are hoping data from a more in-depth clinical study that just ended will provide additional details about the prevalence of these viruses in Mali. They also are planning to examine patients who have undiagnosed fevers to establish infection rates.
NIAID scientists are investigating dengue, Zika and chikungunya viruses to try and develop preventive and therapeutic treatment options, none of which exist.
