Clinical diagnosis of ebola virus

Access to Document Link to publication in Scopus. Fingerprint Dive into the research topics of 'Diagnosis and management of Ebola samples in the laboratory'. Together they form a unique fingerprint. View full fingerprint. Expert Review of Anti-Infective Therapy , 14 6 , WHO guidelines currently state that breast milk from lactating EVD survivors may be tested by RT-PCR if desired by the patient, with retesting every 48 h for those who test positive until two consecutive negative results are obtained The WHO recommends against testing vaginal fluid ; recommendations regarding the testing of other body fluids have not been provided.

Validation of EVD diagnostic tests on a wider variety of specimen types should be a priority, and guidelines are needed to address specimen collection and biosafety issues for testing of alternative body fluid specimens. Viral sequencing tools hold the potential to benefit EVD diagnostic efforts on multiple fronts, including i de novo diagnosis in the setting of an emerging outbreak, ii identification of viral strains responsible for new transmission chains in an ongoing outbreak, iii estimation of viral mutation rate, and iv analysis of the impact of viral mutations on the performance of molecular diagnostic tests.

During the epidemic, next-generation sequencing NGS platforms enabled the characterization of large numbers of viral genomes in a relatively short time frame days to weeks , and advances in portable sequencing tools have made it possible to acquire sequencing data acutely in field facilities.

Initial sample-processing steps for current sequencing strategies parallel those required for standard RT-PCR testing i.

Effective field deployment of portable sequencing devices has been achieved and will likely play an important role in surveillance and outbreak response efforts moving forward; however, these efforts remain challenged by the procedural complexity and the relatively low sensitivity for viral detection of current sequencing methods.

As part of this initial effort, whole-genome sequences were characterized from three specimens. Several large studies using NGS techniques were conducted over the course of the epidemic, representing an international effort that ultimately provided a rich database of hundreds of EBOV whole-genome sequences.

The first of these studies, carried out in a U. A total of additional EBOV genomes were sequenced in the United States Illumina platform from patient specimens collected from June to January in Sierra Leone ; importantly, the study demonstrated that no significant mutations had accumulated in known primer sites for RT-PCR diagnostic tests.

The most recent large sequencing study, performed in Europe Illumina platform , provided EBOV genomes from patient specimens collected in Guinea from March to January The broad time range captured in this data set allowed the authors to illustrate the emergence of two distinct EBOV lineages, one of which was restricted to Guinea in the early months of the epidemic and a second lineage which spread from Guinea to Liberia and Sierra Leone.

This new genomic capacity has allowed for ongoing monitoring of EBOV sequences within Liberia and provided key evidence for a case of sexually transmitted EVD Near-real-time sequencing was provided in Sierra Leone by Cambridge University using an Ion Torrent sequencer in the closing stages of the outbreak, and this facilitated rapid contact tracing by identifying the probable location of exposure in patients who had traveled home when sick A.

Arias, submitted for publication. Nanopore sequencing is a relatively new technology that offers advantages compared to standard NGS methods, including longer sequencing reads and the ability to perform real-time sequence analysis concurrently with data acquisition. The MinION device Oxford Nanopore Technologies , a portable nanopore sequencer, has been evaluated as a diagnostic tool for outbreak responses.

The potential for deployment of the MinION device in a resource-limited setting was demonstrated in a field diagnostic laboratory in Liberia, where it was utilized to sequence EBOV genomes from eight clinical specimens These studies lay an important foundation for the further development of field-ready sequencing tools with the capacity for rapid pathogen identification and in-depth characterization.

Diagnosis by standard real-time RT-PCR in an outbreak setting requires field laboratories with substantial infrastructure, operation and maintenance of complex equipment, and expertise in molecular techniques. While such resources were eventually deployed in the epidemic, their integration into sustainable regional laboratory capacities for ongoing surveillance and response to future outbreaks will be a great challenge.

Furthermore, the requirement for collection and transport of venipuncture blood will continue to confer additional safety and logistical hurdles.

In order to face these challenges, it is imperative that international partners work together with national health ministries to strengthen laboratory capacity in regions where Ebola is endemic, including the development of practical improvements to pre- and postanalytic processes and the training of local laboratory technicians in molecular diagnostic techniques, biosafety practices, and quality control.

Novel diagnostic platforms, such as automated NAATs and rapid antigen detection tests, that can be used in decentralized health care settings with minimal laboratory infrastructure will likely play a major role moving forward.

More field data are needed to establish the appropriate use of these novel tests. Existing evidence suggests that RDTs, if thoughtfully integrated into testing algorithms, could have an immediate impact as point-of-care tests in high-risk populations. Given the persistent challenges in obtaining expeditious RT-PCR results, a point-of-care RDT with imperfect sensitivity and specificity especially if used in combination with RT-PCR testing still stands to confer substantial benefits to case management and infection control efforts and should improve the utilization of limited clinical and public health resources Accordingly, current WHO guidelines recommend initial testing with an RDT when RT-PCR testing is not immediately available and to assist in triage and case management when clinical and laboratory resources are overwhelmed Several innovative RDT platforms are in development 53 , , , , and have shown promising results in laboratory-based evaluations.

Automated NAATs, such as the GeneXpert and Filmarray systems, that provide rapid, sample-to-answer results with minimal operator dependence or potential for cross-contamination have great potential impacts; however, their implementation in decentralized health care settings will require careful consideration of biosafety and operational challenges e. As more clinical validation data become available and practical experience is compiled, local regulatory agencies, in collaboration with the WHO, will be responsible for developing updated EVD testing algorithms specific for different health care settings in both high-prevalence outbreak and low-prevalence surveillance scenarios.

In addition to assay selection for acute diagnostic testing, further guidance is needed to optimize biosafety practices e. As testing algorithms and guidelines evolve, it remains a critical responsibility of health care agencies to ensure means for safe and efficient specimen management, tracking, and reporting. We thank David Heymann Public Health England for his helpful comments during the preparation of the manuscript.

Jana Broadhurst , M. In this role, Dr. Broadhurst collaborated with the Sierra Leone Ministry of Health and Sanitation and WHO to improve practices pertaining to specimen collection and handling for EVD diagnosis at Partners In Health patient care centers and worked extensively alongside local health care workers within these facilities.

Broadhurst collaborated with international laboratory partners to facilitate mobile laboratory deployment within Sierra Leone and served as the field lead for two studies evaluating novel EVD diagnostic tests.

Her long-term research interests include diagnostics development for parasitic diseases and immune regulation during chronic infection with helminth parasites. RIPL is a WHO collaborating laboratory for High Consequence Pathogens and provides diagnostic and clinical advice for a wide range of unusual bacterial and viral pathogens. The IFS offers hour service for acutely ill travellers arriving in the United Kingdom from anywhere in the world.

Brooks' research interests range from environmental detection of microorganisms and clinical diagnostics, through aerobiology and decontamination, to disease pathogenesis and work for the European Space Agency. He led the PHE Ebola laboratories in Sierra Leone and is working with local officials to establish regional laboratory services.

Nira R. Pollock , M. Pollock has an active research program focused on the development and evaluation of diagnostic tests for infectious diseases and related applications. Pollock's diagnostics research has spanned a range of diseases, including Clostridium difficile infection, active and latent tuberculosis, influenza, Lyme disease, and Ebola virus disease EVD , and has involved many different technologies, ranging from simple paper-based lateral flow and microfluidic platforms to novel automated platforms for protein and nucleic acid detection.

During the EVD outbreak, Dr. National Center for Biotechnology Information , U. Journal List Clin Microbiol Rev v. Clin Microbiol Rev. Published online Jul Jana Broadhurst. Tim J. Author information Copyright and License information Disclaimer. Corresponding author. Address correspondence to M. Jana Broadhurst, ude.

Pollock, ude. Diagnosis of Ebola virus disease: past, present, and future. Clin Microbiol Rev — All Rights Reserved. This article has been cited by other articles in PMC. SUMMARY Laboratory diagnosis of Ebola virus disease plays a critical role in outbreak response efforts; however, establishing safe and expeditious testing strategies for this high-biosafety-level pathogen in resource-poor environments remains extremely challenging. Open in a separate window. FIG 1. FIG 2. Cell Culture The traditional gold standard method to confirm the presence of Ebola virus is viral isolation in cell culture, typically using Vero E6 African Green monkey kidney cells.

Antibody Detection Serologic assays for the detection of specific antiviral antibodies in patient serum have been used to demonstrate current or prior infection with Ebola virus since the first outbreak investigations of this virus in 6 , 7.

Protein Antigen Detection The detection of viral protein antigens circulating in blood provides a reliable method for diagnosing acute EVD in symptomatic patients, as viral proteins typically accumulate to detectable levels within a few days of disease onset.

Field Diagnostic Laboratory Efforts in Prior Outbreaks Despite the challenges, successful field diagnostic laboratory operations have been carried out in several past Ebola virus outbreaks Fig. Rapid antigen detection tests.

Viral inactivation in diagnostic specimens. Detection of Viral Persistence in Nonblood Body Fluids As discussed above, the demonstration of viral clearance from blood is recommended for the discharge of EVD survivors back into the community.

Viral persistence in seminal fluid. Viral persistence in other body fluids. Viral Sequencing in the Epidemic Viral sequencing tools hold the potential to benefit EVD diagnostic efforts on multiple fronts, including i de novo diagnosis in the setting of an emerging outbreak, ii identification of viral strains responsible for new transmission chains in an ongoing outbreak, iii estimation of viral mutation rate, and iv analysis of the impact of viral mutations on the performance of molecular diagnostic tests.

Filoviruses and arenaviruses , p — Rapid diagnosis of Ebola hemorrhagic fever by reverse transcription-PCR in an outbreak setting and assessment of patient viral load as a predictor of outcome. J Virol 78 — Basic clinical and laboratory features of filoviral hemorrhagic fever. J Infect Dis :S—S Clinical illness and outcomes in patients with Ebola in Sierra Leone. N Engl J Med — Filoviridae: Marburg and Ebola viruses , p — Isolation and partial characterisation of a new virus causing acute haemorrhagic fever in Zaire.

Lancet — Ebola hemorrhagic fever in Sudan, Bull World Health Organ 56 — Wulff H, Lange J. Indirect immunofluorescence for the diagnosis of Lassa fever infection. Bull World Health Organ 52 — Laboratory diagnosis of Ebola and Marburg hemorrhagic fever. Bull Soc Pathol Exot 98 — Ebola viral load at diagnosis associates with patient outcome and outbreak evolution. J Clin Invest — Laboratory diagnostic systems for Ebola and Marburg hemorrhagic fevers developed with recombinant proteins.

Clin Vaccine Immunol 13 — Alpha Diagnostic International. Alpha Diagnostics International, Inc. Clinical management of Ebola virus disease in the United States and Europe. Clinical, virologic, and immunologic follow-up of convalescent Ebola hemorrhagic fever patients and their household contacts, Kikwit, Democratic Republic of the Congo.

J Infect Dis :S28—S Defective humoral responses and extensive intravascular apoptosis are associated with fatal outcomes in Ebola virus-infected patients. Nat Med 5 — Human asymptomatic Ebola infection and strong inflammatory response. Immunoglobulin G in Ebola outbreak survivors, Gabon.

Emerg Infect Dis 15 — Enzyme immunosorbent assay for Ebola virus antigens in tissues of infected primates. J Clin Microbiol 30 — Detection of Ebola viral antigen by enzyme-linked immunosorbent assay using a novel monoclonal antibody to nucleoprotein. J Clin Microbiol 39 — Development, characterization and use of monoclonal VPantibodies for the detection of Ebola virus.

J Virol Methods — Med Microbiol Immunol — Identification of essential outstanding questions for an adequate European laboratory response to Ebolavirus Zaire West Africa J Clin Virol 72 — Detection and molecular characterization of Ebola viruses causing disease in human and nonhuman primates.

J Med Virol 60 — Persistence and genetic stability of Ebola virus during the outbreak in Kikwit, Democratic Republic of Congo, Detection of Ebola virus in oral fluid specimens during outbreaks of Ebola virus hemorrhagic fever in the Republic of Congo. Clin Infect Dis 42 — Newly discovered Ebola virus associated with hemorrhagic fever outbreak in Uganda. PLoS Pathog 4 :e J Clin Microbiol 40 — Prognostic indicators for Ebola patient survival.

Emerg Infect Dis 22 — The contribution of Ebola viral load at admission and other patient characteristics to mortality in a Medecins Sans Frontieres Ebola case management centre, Kailahun, Sierra Leone, June-October J Infect Dis — Blood kinetics of Ebola virus in survivors and nonsurvivors. Age and Ebola viral load correlate with mortality and survival time in Ebola virus disease patients.

J Infect Dis 42 — Relationship between Ebola virus real-time quantitative polymerase chain reaction-based threshold cycle value and virus isolation from human plasma. World Health Organization. Laboratory diagnosis of Ebola virus disease. Centers for Disease Control and Prevention. Guidance for collection, transport and submission of specimens for Ebola virus testing.

J Clin Virol 77 :9— First international quality assurance study on the rapid detection of viral agents of bioterrorism. J Clin Microbiol 42 — Kelly JD. Make diagnostic centres a priority for Ebola crisis. Nature World Health Organization, Geneva, Switzerland. Mobile diagnostics in outbreak response, not only for Ebola: a blueprint for a modular and robust field laboratory.

Outbreak of Ebola haemorrhagic fever, Uganda, August January Wkl Epidemiol Rec 76 — Flexibility of mobile laboratory unit in support of patient management during the Ebola-Zaire outbreak in the Democratic Republic of Congo.

Zoonoses Public Health 59 — Centers for Disease Control and Prevention. Accessed Jan. Samai M, et al. The Journal of Infectious Diseases. Marburg hemorrhagic fever Marburg HF.

Bray M, et al. Epidemiology and pathogenesis of Ebola virus disease. Treatment and prevention of Ebola virus disease. Clinical manifestation and diagnosis of Ebola virus disease. Bray M. Marburg virus. Viral hemorrhagic fever.