Download Middle East Respiratory Syndrome-Corona Virus (MERS-CoV)

DOI : 10.9780/2321-3485/1322013/74
Reviews Of Progress
ISSN:-2321-3485
ORIGINAL ARTICLE
th
Vol - 3, Issue - 1, May 4 2015
Middle East Respiratory Syndrome-Corona
Virus (MERS-CoV): An Overview
Sagar Aryal1, Pratiksha Pokhrel Pahadi2, Bikash Rana
and Rana Kausar
1
Department of Microbiology, St. Xavier’s College, Kathmandu, Nepal
Department of Microbiology, St. Xavier’s College, Kathmandu, Nepal
3
Department of Microbiology, St. Xavier’s College, Kathmandu, Nepal
4
Department of Biochemistry, FUUAST, Karachi, Pakistan
2
Abstract:
Middle East Respiratory Syndrome-Corona Virus (MERS-CoV) is an emerging
virus which causes respiratory infections in animals and humans caused by novel corona
virus (CoV). There is presence of Spike Glycoprotein in the virus which gets attached
with DPP4 of the target cell during replication. Fever, cough, shortness of breath and
gastrointestinal symptoms including diarrhea are some of the major symptoms of this
infection having the incubation period of 5.2 days. The first case was seen on 2012 in
Saudi Arabia where the patient died of acute respiratory and renal failure. As of 14
October 2014, 896 laboratory-confirmed cases of MERS-CoV have been confirmed
including 357 deaths as of 14 October 2014. Transmission can occur from animal to
human or from human to human. rRT-PCR is the best diagnostic method for the detection
of virus. There is no current treatment or vaccination available for MERS-CoV.
KEYWORDS: MERS-CoV, Review, Infections, Outbreaks, Transmission
INTRODUCTION:
Middle East Respiratory Syndrome (MERS) is the viral respiratory illness which is caused by a
corona virus called Middle East Respiratory Syndrome-Corona Virus (MERS-CoV) 6. MERS-CoV was
used to be called Novel Corona Virus 1. It was termed by the Coronavirus Study Group of the International
Committee on Taxonomy of Viruses which was announced in the “Journal of Virology” on 15 May 2013 2.
MERS-CoV is a beta coronavirus belonging to the Coronaviridae, a group of large, enveloped single
stranded RNA viruses 1. Infection is primarily zoonotic in nature but can be spread from human to human
too 2. According to WHO, as of 11 June 2014, there were 699 laboratory confirmed cases of human
infections, including at least 209 deaths 11. According to European Centre for Disease Prevention and
Control, as of 14 October 2014, there were 896 laboratory-confirmed cases of MERS-CoV and including
357 deaths 16.
Structure of MERS-CoV
They measure about 100 to 160 nm. MERS-CoV contains spike glycoprotein (S) which targets the
cellular receptor, dipeptidly peptidase 4 (DPP4) which is also known as CD26 5. Glycoprotein also shows an
antigenic action. It has neucleocapsid phosphoprotein for RNA-binding and membrane glycoprotein for
triple membrane spanning 7. MERS-CoV consists of a core and a receptor binding sub domain in where it
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Middle East Respiratory Syndrome........
interacts with DPP4 3,6. Spike glycoprotein consists of a globular SI domain at the N-terminal region,
followed by membrane proximal S2 domain, an intracellular domain and also a receptor-binding domain
(RBD) 4.
MERS-CoV RBD consists of a core subdomain and receptor binding subdomain 3.
a. Core Subdomain
The core subdomain is a five stranded antiparallel beta (B) sheets (B1, B2, B3, B4 and B9) with
two short alpha helicase in the connecting loops. There is presence of 3 disulfide bonds in the core
subdomain to maintain the fold 3.
b. Receptor Binding Subdomain
It is a four stranded antiparallel B sheets (B5, B6, B7 and B8) located between B4 and B9 of the
core subdomain. B7 and B9 is connected via a long loop that crosses perpendicular to the B sheet. There is
presence of one disulfide bond which connects the long loop with strand B5 3. This supports for the
interaction with DPP4.
Figure 1: Structure of MERS-CoV [14]
Genome
MERS-CoV genomes are phylogenetically categorized into 2 clades, clade A and B clusters. Full
genome deep sequencing can be done on nucleic acid extracted straight from PCR-confirmed clinical
samples 8. Genome of MERS-CoV is an unsegmented single stranded positive sense RNA having the size
of 29.9 kb. Genomes are capped and polyadenylated at 3’ end. Sequence analysis of the MERS-CoV
genome identified the emerging virus as being a member of the lineage C of the Betacoronaviridae, with the
closest relatives known are bat coronaviruses. RNA are structurally polycistronic but functionally
monocistronic.
The genome of MERS-CoV encodes both structural proteins as well as non-structural proteins.
The 3’ one-third of the genome encodes fours structural proteins – Spike (S), Membrane (M), Envelope (E)
and Nucleocapsid (N) along with the sets of accessory proteins which are required for the formation of viral
particles 9.
The Polymerase gene, 5’ two third of the RNA genome encodes a large polyprotein (ORF 1a/1b) whose
expression is controlled by ribosomal frame shifting 9. It is proteolytically cleaved to generate 15 or 16 nonstructural proteins (nsp’s) which is required for the viral replication as well as the modulation of the
antiviral reaction.
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Figure 2: MERS-CoV genomes
Figure 3: A schematic of the complete genome of the first fully sequenced MERS-CoV variant in
lineage C of the betacoronaviruses, HCoV 2c EMC/2012.1 [14]
Replication
The entire cycle of virus replication occurs in the cytoplasm. It involves various steps as follows:
1.Attachment
The glycoprotein spike (S) present in the envelope of the virus attached to the DDP4 receptor on
the target cell. At the interface a total of 14 residues of the MERS-CoV contact with the 15 residues of the
DPP4 with the distance cut off of 3.6A 3.
2. Entry of the genome
The hydrophobic core formed between MERS-CoV RBD and DPP4 plays a critical role in
mediating the viral binding and entry into the target cell 38. Virus gets into the host cell by receptor mediated
endocytosis or by the fusion of viral envelope with the cell membrane.
3. Uncoating and Release
After the penetration of the virus particle, uncoating of the genome takes place and released into
the cytoplasm.
4. Protein and Genomic RNA Formation
MERS-CoV has a single positive stranded RNA genome, which can directly produce their
proteins and new genome on the cytoplasm. At first, the viral genomic RNA translocates to produce virusspecific RNA-dependent RNA polymerase. This recognizes and produces viral RNAs. RNA polymerase
synthesize the minus (-ve) strand using the positive strand as template. Then this –ve strand RNA serves as
template to transcribe full length genomic RNA and sub genomic mRNAs, which are translated into a single
polypeptide 15.
5. Nucleocapsid Complex Formation
The newly synthesized genomic RNA interacts in the cytoplasm with the Nucleocapsid (N)
protein to form the Helical Nucleocapsid. Membrane (M) protein is inserted into the ER and anchored in the
Golgi apparatus. Nucleocapsid binds to the M protein at the budding site into the ER lumen. Envelope (E)
and Membrane (M) proteins interacts to trigger the budding of virions, enclosing the Nucleocapsid.
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Middle East Respiratory Syndrome........
6. Assembly and Maturation
Spike Glycoprotein (S) associate with the M protein and are incorporated into the maturing virus
particles and virus get fully matured now.
7. Release
These viral progeny are finally transported by Golgi vesicles to the cell membrane and released by
exocytosis like fusion of vesicles with plasma membrane.
Figure 4: Replication of MERS-CoV [13]
Pathogenesis
Symptoms of MERS-CoV infections include fever, cough, shortness of breath and
gastrointestinal symptoms including diarrhoea 29. Severe illness can cause respiratory failure that requires
mechanical ventilation and support in an intensive-care unit. Some patients develop organ failure,
especially of the kidneys, or septic shock. Approximately 27% of patients with MERS have died. The virus
seems to cause more severe disease in people with weakened immune systems, older peoples, and those
with such chronic diseases like diabetes, cancer, and chronic lung disease 31. The average incubation period
was found to be 5.2 days 29.
In the cell line susceptibility study, MERS-CoV infected human cell lines, with lower respiratory,
kidney, intestinal, and liver cells and histiocytes 40.
MERS-CoV prevent the secretion of interferon (IFN)-α and IFN-ß and persuade the expression of
pro-inflammatory tumor necrosis factor (TNF)-a and Interleukin-6, and inducing the inflammation of
surrounding tissue. MERS-CoV infects and replicate in human monocyte–derived macrophages (MDM)
and the induction of cytokines in these cells contribute to pathogenesis. Furthermore, in MDM, MERSCoV proliferates the expression of MHC-class I molecule and co-stimulatory leading to an activation of
immune responses 36.
In the case of MERS-CoV, in vivo target cells contain type II alveolar cells and non-ciliated cells
epithelial cells where MERS-CoV is able to infect endothelial cells as well.
DPP 4 has many other roles besides being the receptor, in glucose homeostasis, T-cell activation,
neurotransmitter purpose, and modulation of cardiac signaling, but the enzymatic function of DPP4 is not
essential for viral entry 37,38.
When there is entry of MERS-CoV into the host cell, type II transmembrane protease TMPRSS2 activates
the spike (S) protein by transforming the mature S protein into two subunits (S1 and S2) and increasing the
fusogenicity with the receptor of host cell 32, 35. The existence of both the receptors for MERS-CoV and S
cleaving protease determine the potential animal reservoir and the sources of recurring transmission to
humans from animals. Since the MERS-CoV receptors in human, horse and camel are similar, screening
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should be done in both camels and horses.
The person infected with MERS-CoV doesn’t show only respiratory infections but also acute renal failure
33, 34
. Infection and replication of kidneys with MERS-CoV might therefore not only lead to acute renal
failure but also to shedding and transmission of MERS-CoV in urine which leads to new cases not only via
airborne transmission but also under favorable conditions via contaminated drinking water 34.
Epidemiology
As the name suggests, it is mainly distributed in Middle East countries which includes Iran,
Jordan, Kuwait, Oman, Qatar, Saudi Arabia, United Arab Emirates and Yemen. It is distributed in Africa
and Europe too. This includes Algeria, Egypt and Tunisia in Africa whereas France, Germany, Greece, Italy,
the Netherlands and the United Kingdom (UK) in Europe. In Asia, Malaysia and Philippines are affected
with MERS-CoV 11,16. It is seen in USA too. There were 896 laboratory-confirmed cases of MERS-CoV
reported to the public health authorities worldwide, which includes 357 deaths, as of 14 October 2014 16.
MERS-CoV not only affects the human but also infects animals like Camel where the viruses has been
isolated from Egypt 17, Saudi Arabia 18,19,22,24, Qatar 21,23, Jordan 25, Oman 26, UAE 27 and Africa 20. Dromedaries
Camel is thought to be the possible viral reservoirs 27.
Transmission
The virus is transmitted by respiratory route in close contact. Transmission to human has been
thought to be transmitted from Camels due to high similarities of MERS-CoV carried by human and camels
18
. Airborne nosocomial transmission can occur in the room shared by the patients in the hospitals. There is
still the confusion of transmission through body fluids or clinical samples, including stools and a cross
transmission with medical devices or hands. The previous studies did not tell about the source of virus, nor
did they reveal whether MERS-CoV can be transmitted from human to human or not 28. Later it was found
that the virus transmission can probably occur in dialysis units, medical wards and ICUs 29.
Outbreaks
MERS-CoV was first identified in cell culture taken from the patients who died of pneumonia in
Saudi Arabia in 2012. After that, 896 laboratory-confirmed cases of MERS-CoV have been reported,
including 357 deaths as of 14 October 2014 16. Very Large number of cases and death has been reported in
Saudi Arabia. Table 1 shows the total number of confirmed cases and deaths, by country of reporting, from
March 2012 to 13 October 2014 16. Figure 5 and table 1 show the distribution of confirmed cases of MERSCoV reported from March 2012 to 13 October 2014. Many of the cases have been seen in the Middle East
Countries like Saudi Arabia, United Arab Emirates, Qatar, Jordan, Oman, Kuwait, Egypt, Yemen, Lebanon
and Iran. The latest confirmed case of MERS-CoV infection was seen on a 43-year-old male from Doha,
Qatar on 20 October, 2014 30. There were no cases of MERS-CoV infection during Hajj in 2012 or 2013 50.
Figure 5: Distribution of confirmed cases of MERS-CoV reported March 2012–14 October 2014, by
reporting country (n=896)
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Table 1: Number of confirmed cases and deaths, by country of reporting, March 2012–13 October
2014 [16]
S.No
Country
Cases
Death
Date of most Recent
Cases
1
Saudi Arabia
762
324
13/10/2014
2
United Arab Emirates
73
9
11/06/2014
3
Jordan
18
5
23/05/2014
4
Qatar
8
4
12/10/2014
5
United Kingdom
4
3
06/02/2013
6
Iran
5
2
25/06/2014
7
Oman
2
2
20/12/2013
8
Tunisia
3
1
01/05/2013
9
Kuwait
3
1
07/11/2013
10
Germany
2
1
08/03/2013
11
France
2
1
08/05/2013
12
Algeria
2
1
24/05/2014
13
Yemen
1
1
17/03/2014
14
Greece
1
1
08/04/2014
15
Malaysia
1
1
08/04/2014
16
Netherlands
2
0
05/05/2014
17
USA
2
0
01/05/2014
18
Egypt
1
0
22/04/2014
19
Lebanon
1
0
22/04/2012
20
Austria
1
0
29/09/2014
21
Italy
1
0
31/05/2013
22
Philippines
1
0
11/04/2014
23
Total
896
357
Laboratory Diagnosis
rRT-PCR can be done from lower respiratory tract specimens such as sputum, endotracheal
aspirate, or Broncho-alveolar lavage fluid. Acute and convalescent sera should be taken for serologic
testing. If only a single sample is to be obtained, it should be collected at least 14 days after onset of
symptoms. A stool sample or rectal swab can also be collected. If a negative result is obtained from a patient
with high risk group and suspicion, additional specimens can be taken and examined 41.
Specimens should be transported to the laboratory soon after collection. When there is a delay of
more than 48 hours for respiratory tract specimens, specimens should be frozen, preferably at -80°C, and
transported on dry ice. Serum should be separated from whole blood and then stored and transported at 4°C
or frozen to -20°C. Temperature fluctuation should be avoided for the storage of respiratory and serum
specimens 42.
1. Cell Culture
MERS-CoV can be recovered from Vero and LLC-MK2 cells 1. MERS-CoV strain can also be
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replicated in fully separated human airway bronchial epithelium (HAE) cultures grown at the air-liquid
interface (ALI) 54.
2. Polymerase chain reaction and sequencing
In a patient with multiple myeloma and MERS-CoV infection high concentrations of MERS-CoV
needs to be detected from respiratory specimens by rRT-PCR 45. MERS-CoV was also be detected from
nasal secretions, stool, urine and stool, but in low concentrations 46.
There are three rRT-PCR assays for routine detection of MERS-CoV 41. Currently described tests
are an assay targeting a region upstream of the E protein gene (upE) and assays targeting the open reading
frame 1b (ORF 1b) and the open reading frame 1a (ORF 1a) 43,44. In some cases, sequencing should be
performed for confirmation. Hemi-nested sequencing amplicons targeting RdRp (present in all corona
viruses) and N gene (specific to MERS-CoV) fragments can be generated for confirmation via sequencing.
3. Serology
Different serology test have been developed for the detection of MERS-CoV antibodies,
including immunofluorescence assays and a protein microarray assay. CDC has developed a two-stage
approach, which uses an enzyme-linked immunosorbent assay (ELISA) for screening followed by an
indirect immunofluorescence test or microneutralization test for confirmation. Any positive test by a single
serologic assay should be confirmed by a neutralization assay. Sensitivity and specificity of antibody tests
for MERS-CoV has not been confirmed. According to the WHO, cases with a positive serologic test in the
absence of PCR testing or sequencing are considered probable cases if they meet the other elements
comprising the case definition of a probable case 41.
Treatment
There is no treatment or vaccination available for MERS-CoV till now 39. People with MERS can
have medical care to help relieve symptoms. In cell culture and animal experiments, combination therapy
with interferon (IFN)-alpha-2b and ribavirin seems to be promising by limiting the viral replication 47.
Other experimental therapies have been investigated which includes convalescent plasma, monoclonal
antibodies, and inhibition of the viral protease 48.
Vaccines
Currently, there is no vaccine to prevent MERS-CoV infection 39 but some discussion is going on
with the partners of CDC for developing the vaccines. One company has developed an experimental
candidate MERS-CoV vaccine which is based on the major surface spike protein by means of recombinant
nanoparticle technology 51. Other candidate vaccines that are being studied include a full-length infectious
cDNA clone of the MERS-CoV genome in a bacterial artificial chromosome 52 and a recombinant Modified
Vaccine Ankara (MVA) vaccine expressing full-length MERS-CoV spike protein 53.
Prevention and Controls
CDC routinely advises that people help protect themselves from respiratory illnesses by taking
everyday preventive actions 49.
v
Wash the hands always with soap and water for 20 seconds, and help young children do the same. If soap
and water are not available, we should use an alcohol-based hand sanitizer.
v
Always cover your nose and mouth with a tissue when you cough or sneeze and throw them in the proper
place.
v
Avoid touching the eyes, nose and mouth with unwashed and dirty hands.
v
Avoid personal contact, such as kissing, or sharing cups or eating utensils, with sick and infected people.
v
Clean and disinfect the frequently touched surfaces such as toys and doorknobs.
CONCLUSION
MERS-CoV is the emerging respiratory diseases that have mostly infection the people of Middle
East Countries. Saudi Arabia is the most affected country among all. Since this virus has not treatment and
vaccines, we should try to minimize the transmission rate. Travelers should know the general hygiene
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Middle East Respiratory Syndrome........
measures, including hand washing before and after touching animals, and avoiding contact with the sick
animals. Travelers should also avoid consumption of raw or undercooked animal products. Due to the high
risk for severe MERS infections, people with diabetes, kidney failure, or chronic lung disease and people
who have weakened immune systems should try to avoid contact with camels and horses.
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