Ebola Virus A to Z

Dr. Y. Ashok Babu

Contents of the topic:

• About the virus (taxonomy)• History of Ebola of Viral Disease• Geographical Distribution and Epidemiological preamble• Molecular Biology of the Virus • Host pathogen interaction (Entry, Replication and Host immunity evasion) • Patho-physiology and clinical manifestations• Diagnosis• Treatment and complications• Disease out brake dynamics• Management of Ebola outbreak at Health care facilities and handling of the virus•Threat perception for India • Conclusion

• Ebola Virus is a ss(-)RNA Virus

• Belongs to order Mononegi virales, Family filovoviridae

• Current species in circulation causing major out breaks is Zaire ebola virus

• The other virus species which cause Ebola virus disease include Bundibugyo ebolavirus (BDBV) Reston ebolavirus (RESTV) –(Live Stock Infection) Sudan ebolavirus (SUDV) Taï Forest ebolavirus (TAFV) – Asia (Live stock Infection)

Marburg Virus is another genus of Filoviridae causing similar hemaregic fever with comparative less fatality (30%)

History of Ebola Virus Disease

Initially it was suspected as a species of Marburg Virus with high virulence recent genetic analysis confirmed as new genus with 70% homology to Marburg virus

Since no treatment exists and symptoms are similar to marburg virus the management of the disease followed same procedure

The first case of the infection reported in Zaire (1977) now called DRC with case fatality of 88%.

The spread of the disease attributed a single incidence of handling carcasses dead baboons by a single human which triggered human to human in close contacts.

Date Country Species Reported cases

Reported deaths Fatality

1976 Zaire ZEBOV 318 280 88%1976 Sudan SEBOV 284 151 53%1979 Sudan SEBOV 34 22 65%1994 Gabon ZEBOV 52 31 60%1995 Zaire ZEBOV 315 250 79%

1996 Jan–Apr Gabon ZEBOV 37 21 57%

1996–1997Jul–Jan Gabon ZEBOV 60 45 75%

2000–2001 Uganda SEBOV 425 224 53%

2001–2002 Oct–Jul GabonCongo ZEBOV 122 96 79%

2002–2003 Dec–Apr Congo ZEBOV 143 128 90%

2003 Nov–Dec Congo ZEBOV 35 29 83%

2007 DR Congo ZEBOV 264 187 71%

2007–2008 Dec–Jan Uganda BDBV 149 37 25%

2008–2009 Dec–Feb DR Congo ZEBOV 32 14 45%

2012 Jun–Aug Uganda SEBOV 24 17 71%

2012 Jun–Nov DR Congo BDBV 77 36 47%

2013–2014Dec–present

Guinea, LiberiaSierra LeoneNigeria

ZEBOV 1848 1013 64%

Molecular Biology of the Virus

The virus is a filamentus negative sense SS RNA virus with genome size of 19kb

The genome of Ebola virus is continues and codes for 7 structural genes

NP: Nucleoprotein…………….Nuclear capsid protein VP35: Virion Protein 35……. Polymerase complex VP 40: Virion Protein 40…… Matrix Protein Gp/sGP: Glyco Protein……….Trans membrane/secretory protein VP30: Virion Protein 30……..Transcription regulatory protein VP 24: Virion 24…………………. Matrix associated ??? . L: RNA dependent RNA polymerase

Schematic diagram showing a typical Ebola Virion

Host pathogen Interaction

The primary targets of Ebola virus is cell of lymphoid tissues including Dendritic cells, Monocytes, and Epithelial cells.

However, after initiation of infection in the host the virus can infect all type cells expressing NPC C1 protein.

The virus enters into the cytoplasm of host cell through Macropinocytosis.

Entry of the virus occurs in two phases, at both the phases involving interaction of GP protein of the virus with host receptors.

Multiple targets for binding of GP protein on cell surfaced was found such as Intigrins, Lectins, TAM

Virus internalizes into Macro Pinosome

GP1,2 protein cleaved by cathepsin B and L at the glycosylation rich region GP1 exposing Receptor binding domine.

Receptor Binding site interact with NPC C1 on Macropinosome leading to release of viral content into the cytoplasm

Being –ssRNA the virus carries along with it the RdRP (L-protein) to initiate transcription.

The whole genome of the virus is under control of a single promoter and ORF site at -1 of NP protein.

The peculiar feature of -ssRNA viruses is they have start and stop regions at the beginning and end of each gene, leading to high expression of proteins near to promoter than the gene located away.

Entry of Ebola Virus into the cell

Over view of life cycle of Ebola virus

Host Immunity evasion

The Ebola infection is fast acting, and highly replicating coupled with disturbing host protein synthesis

The virus rapid multiplication outperforms the cellular protection mechanism leading initial failure of humeral immunity to contain infection

The virus releases a secretory form of GP out of the infected cell in abundance which due to epitope cross reactivity to full GP competes to bind with neutralizing anti bodies leading to failed immune response.

The same reason is the leading concern in developing vaccine against Ebola infection.

Symptoms of Ebola Virus

Initial Signs • Fever (at least 102°F)• Weakness & exhaustion • Pain

– Severe headache– Muscles & joints– Abdominal pain

• Sore throat• Nausea• Dizziness

Progressed Symptoms • Vomiting• Diarrhea• Extensive bleeding

– Red eyes• hemorrhage of sclerotic arterioles

– From mouth, nose, eyes, rectum & mucouse membranes

• Maculopapular rash – Spreads over the body (often hemorrhagic)

• Other secondary symptoms – Hypotension , Hypovolemia , Tachycardia – Organ damage – Internal and external bleeding

Diagnosis: Tool and Techniques

The clinical samples of Ebola virus infection pose serious Bio Hazard to laboratory workers at par with the care givers.

Use of protocols involving possible generation of Aerosol such as pippetting, Centrifugation, overtaxing completely banned outside BSL4 facility

Treatment and complications

There is no treatment for the infection of Ebola Virus.

Symptomatic supportive care can be given to the Individuals including re hydration, supplementing with coagulation factors.

The virus pose high risk of Nosocomial transmission and hence bio hazardous to health care workers and doctors.

Various studies on protection against EBOV infection in non human primates with monoclonal antibodies, and si RNA against NP and VP40 protein shown encouraging results.

There is no FDA approved therapy available for either Pre- or Post exposure to the virus

However, very recently an American doctor and his assistant who acquired the Infection was given the experimental drug containing monoclonal antibodies against three EBOV.

Both of them recovered from the infection

On 6th August in the meeting of WHO it has recommended that a controlled treatment with experimental drug may be permitted in humans with learned consent as humanitarian crisis over looming over ethics of experimental drug use.

However, the WHO recommendations limited to 2014 Ebola outbreak only

Management of Ebola outbreak at Health care facilities

Ebola vial infection is highly contagious with rapid health deterioration in the infected patients

The virus get secreted in abundance in all body fluids including tears, saliva, blood, vomit, urine.

Extreme personal protection is required while handling the infected patients, and contaminated material.

WHO, CDC recommended personal protection protocol after the recent outbreak include full impermeable suite (Single use), N95 particulate filter max, impermeable boots, and gloves, protective goggles.

Minimum handling of the infected patient is recommended and patient care by the relatives may be totally avoided except in case of children with full protection

Frequent hand hygiene with detergent or alcohol based solutions is paramount to protect against infection in health care workers

The virus can be easily get destroyed by 70% alcohol , 0.05% Sodium Hypochlorite solution, bleaching powder or solution containing 500ppm chlorine.

Contaminated materials need to be treated with above solution before disposal , if available incineration of contaminated materials such as clothes, cotton swabs soiled with body fluids.

Funeral of dead people with confirmed or suspected Ebola infection should be strictly adhered to personal protection protocol, Un-protected handling of dead body should be fully avoided

Threat perception for India Ebola is a zoonotic infection transmitted through a direct or indirect contact with the reservoir

Initial contact with natural reservoir fruit bat or in contact with infected monkeys, other non human primates is the actual trigger for initiation of spread of disease

Persistence of the disease in the wild depends on abundance of the carrier in the area, and its initial spread largely depends on human and wild animal contact

In Sub-Saharan region predominantly of worlds poorest countries, majority of the population depend on forest, and consumption of wild animals which make these population vulnerable to repeated epidemic of the infection.

This single point shows how this virus remained an exotic pathogen to India.

However, In the recent decades with increased economic interaction with African countries, and presence of Indian UN peace keeping forces made transmission of the disease to India become a possible threat

Heightened monitoring public transiting from these countries at entry points is the most important to prevent entry of the virus.

With proper health care management in the event of out break the disease may be contained. The extent of causality is inversely proportional to the availability quality health care facilities.

Since the infection pose a serious Nosocomial bio-hazard, proper equipping with PPE, training of health care workers and doctors should be initiated.

Mass awareness about the disease and spread of disease may be issued to general public, which is sufficient at the moment.

With the reports of 4 Indian doctors want to return to India in fear of Ebola infection where doctors health workers of WHO, MSF, Samaritan Purse combating to contain the disease spread, there will requirement of sensitization of Indian health care workers, and doctors for their cooperation as the disease is a serious threat to health care givers.

Since entry point for the pathogen is major cities with international air ports, Establishing quarantine facility is easier than other places as such cities have better health care infra structure.

Conclusion The ebola virus infection causes severe hemorrhagic disease with case fatality ranging from 30% to 90% among different species.

The species causing the current out break is Zaire Ebola virus with high case fatality rate of above 70%

Morphologically Ebola virus resembles with that of other filoviride virus Marburg virus with a difference of filament size

With small molecule inhibitors NPC 1 (Cholesterol carrier) indentified to be the required for entry of virus into cytoplasm

The viral protein GP shown to be cytotoxic to epithelial cells causing inhibition of expression of cell adhesion molecules leading to vesicular leakage.

Ebola virus evade host immunity by antibody subversion, by producing antigen decoys, which render neutralizing antibodies useless.

Early diagnosis of the infection involves Antigen Capture ELISA, PCR, RT-PCR, IgM detection and electron microscopy, In the later stage of disease the infection may be detected by IgG ELISA apart from above methods.

Treatment with monoclonal antibodies against the virus may be possible as researchers shown their positive effect in mouse and no human primates.

Aseptic management and handling of infected persons and contaminated materials is the key to containment of the disease.

Repeated outbreak of the Ebola virus is associated with regular human and wild animal contact which is prevalent in African countries. Poor Health care infrastructure fans the spread of disease from human to human.

Prevention of entry of the pathogen is paramount for India.

Thanks