Introduction:
The extensive outbreak of Ebola recently in the West African regions has put the people in a place where an accurate and rapid diagnosis has become necessary. Initially, during the outbreak in 2014 to 2015, diagnosis completely relied on collecting blood samples from patients who were showing symptoms that had difficulties of their own, like working In a biocontainment laboratory setting (an isolated temporary laboratory setting) along with the difficulties in sample collection, storage, and data management with prolonged time spent on waiting for results. And hence there was a need for the development of newer methods of diagnosing, which gave faster and most accurate results. But as newer diagnostic technologies are emerging, it is important for clinicians to understand the analytics, strength, and limitations present in the methods. Overall an optimal method of diagnosis depends on various factors that include the nature of the population, the prevalence of the disease, health care infrastructure, biosafety, and infection control methods, local or regional laboratory capacity, training facilities, cost, and regulatory factors.
What Is an Ebola Virus and What Are Its Types?
The Ebola virus is an RNA (ribonucleic acid) virus that contains a single-stranded RNA genome that encodes several proteins like nucleoproteins, glycoproteins, and polymerase enzymes. There are five different types of Ebola strains detected so far, and they are:
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Zaire Ebola virus (EBOV).
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Sudan Ebola virus (SUDV).
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Tai forest Ebola virus (TAFV).
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Reston (RESTV).
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Bundibugyo (BDBV).
What Is the Concept Behind Ebola Diagnosis?
Various methods have been used for the past twenty-five years in the detection and clinical study of the Ebola virus. The overall outline of the tests can be divided into three categories:
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Serological tests to detect antibodies formed in patients to fight against the virus.
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Antigen tests to detect the presence of viral proteins.
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Molecular tests to detect a sequence of the viral RNA.
But however, the diagnostic tools needed to be more competent in the case of serological assays. At the same time, antigen detection and molecular tests have proven to be very effective for diagnosing acute infections, as the level of viruses tends to shoot up in the blood soon after the onset of symptoms. The incubation period of an Ebola infection typically ranges from three to thirteen days and may even last up to twenty-one days. So the diagnostic tests developed for Ebola are of two types:
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Used to detect the Ebola virus
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Used to differentiate the types of Ebola viruses.
What Are the Methods Used for Diagnosing Ebola Virus?
These are five diagnostic tests done for the detection and confirmation of Ebola, and they are:
1) Cell Culture Method: This is the most traditional and gold-standard test for assessing Ebola. There are two serum-based standard methods used to confirm the presence of the Ebola virus by isolating and culturing it in the kidney cells of Vero E6 African green monkeys. The isolated and inoculated viruses of one to five days can be directly viewed under an electron microscope or indirectly using immunofluorescence microscopy. Although this is a definitive method for detecting Ebola, these tests need a biosafety level 4 (BSL-4) containment which is generally restricted only to research and public health laboratories.
2) Antibody Detection: Serological assays for detecting specific antibodies in the patient's blood have been one of the most important methods of investigation since the first Ebola outbreak in 1976. There are two serum-based tests done in this method. They are:
- Indirect Fluorescent Antibody Detection Test (IFAT): The indirect fluorescent antibody detection test was the first test used in 1977 to differentiate the newly found Ebola virus from the Marburg virus. Based on the theory of antigen specificity, while the serum is in a convalescent phase after the patient has recovered from the infection. This method is done by cell culture with infecting virus. It is irradiated and fixed on the slide and incubated with the sera of the possibly infected patients and is seen under the immunofluorescence microscope. But although IFAT played an important role in the early ages of the Ebola outbreak, it is still considered a suboptimal test for specifying the virus and proving its sensitivity.
- Enzyme-Linked Immunosorbent Assay (ELISA): The enzyme-linked immunosorbent assay is a test used to detect antibodies IgM and IgG that is specifically seen with this virus, and this method offers a much more quickly and more accurate output when testing the blood serum. This test was first developed in the US Army Medical Research Institute of Infectious Diseases in the year 1990.
3) Protein Antigen Detection: This method aims to detect protein antigens of the virus circulating in the blood, which gives a trustworthy source for diagnosing the Ebola virus disease. This test was first performed in the year 1995 using eight mouse antibodies. In this method, the viral antigen is detected at the earliest of the first day of the symptoms and shows a very prominent result in all the patients with three days of illness. The antigen levels rapidly increase from the first seven to ten days of the infection and markedly decline from the sixteenth day. This antigen detection test has become one of the most standard diagnostic testing used in recent outbreaks.
4) Conventional RT-PCR: The conventional RT-PCR (reverse transcription-polymerase chain reaction) test was first done in the year 1996 by the Center International de Researchers Medicales de Franceville (CIRMF). The RT-PCR test for diagnosing the Ebola virus was developed by CDC where the first test was done using the serum of the patient with Ebola collected during the 1995 Ebola outbreak. The conventional RT-PCR was more sensitive to the detection of antigens and antibodies than ELISA and is recommended to be used along with ELISA for diagnosing acute infections of the Ebola virus. This is done by collecting infected fluid samples from saliva or semen.
5) Real-Time RT - PCR: The real-time RT-PCR tests are done using color-changing or fluorogenic probes, which were pre-designed to detect the Ebola virus. This procedure was performed in field laboratories during the year 2014 to 2015 epidemic outbreak. The test detects the sample should be done within 72 hours of infection, and patients who show a false positive result are asked to retest after the next 72 hours. Compared to conventional RT-PCR, real-time RT-PCR tests help in specific detection with the help of the probe with rapid results within two to three hours.
Conclusion:
In conclusion, these are the tools used for detecting Ebola virus disease during an outbreak, and they detect not only the presence of the virus but also the exact type, which helps and know the severity of the type and provides a proper treatment plan.
