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Abstract. – COVID-19 pandemic can cause ir-
reparable damage to the involved society. This
study aimed to provide a summary of the up-to-dat-
ed clinical display, diagnostics, molecular and ge-
netic implications for COVID-19 infected patients.
In this review, 73 research articles published
before 25 March 2020 were analyzed to better
understand the clinical characteristics of pa-
tients and to introduce the available serologi-
cal, hematology and molecular diagnostic meth-
ods. Apart from articles extracted from PubMed
and Google Scholar, WHO (https://www.who.
int/ ), NHC (National Health Commission of the
People’s Republic of China (http://www.nhc.gov.
cn/ ), NICE (National Institute for Health and Clin-
ical Excellence, https://www.nice.org.uk/ ), CDC
(Centers for Disease Control and Prevention,
https://www.cdc.gov/), and National Administra-
tion of Traditional Chinese Medicine (http://www.
satcm.gov.cn/ ) were also accessed to search for
eligible studies. Papers published between Jan-
uary 1, 2020, and 25 March 2020 were searched
in English and the terms “2019-nCoV, Covid-19,
Clinical Characteristics OR manifestation, meth-
od of detection, COVID-19 Genome and molecu-
lar test” were used.
As the pandemic continues to evolve, there
have been reports about the possibility of as-
ymptomatic transmission of this newly emerged
pneumonia virus. We highlighted the role of HLA
haplotype in virus infection as HLA typing will
provide susceptibility information for personal-
ized prevention, diagnosis, and treatment in fu-
ture studies. All the data in this article will as-
sist researchers and clinicians to develop their
clinical views regarding infected patients and
to emphasize the origin of SARS-CoV-2 for di-
agnostics.
Key Words:
Diagnosis, Genetic, Epidemiology, Coronavirus,
SARS-CoV-2, COVID-19, Detection, Clinical display,
Novel coronavirus.
Introduction
Respiratory tract viral infection caused
by viruses is deemed one of the most
common diseases in human in the world.
2019-nCoV, divergent from SARS-CoV, belongs
to the coronavirus family and emerged in Decem-
ber 2019 caused pneumonia outbreak in Wuhan,
China1. The outbreak of this novel coronavirus
disease (COVID-19) quickly spread all over China
and to more than 184 other countries and territo-
ries worldwide2. This virus can cause the disease
named coronavirus disease 2019 (COVID-19) re-
sulting in multi-organ failures and has posed major
threats to global public health3-6. At rst, this virus
named 2019 novel coronavirus (2019-nCoV); how-
ever, the International Committee of Taxonomy
of Viruses (ICTV) attributed the name of Severe
acute respiratory syndrome coronavirus 2 (SARS-
CoV-2) to this newly developed virus7. Based on
varied clinical presentations in patients infected by
European Review for Medical and Pharmacological Sciences 2020; 24: 4607-4615
M. FOROUZESH1, A. RAHIMI2, R. VALIZADEH3,4,
N. DADASHZADEH1, A. MIRZAZADEH5,6
1Specialist in Forensic Medicine, Assistant Professor of Forensic Medicine, Legal Medicine Research
Center, Iranian Legal Medicine Organization, Tehran, Iran
2Department of Genetics and Molecular Biology, Faculty of Medicine, Isfahan University of Medical
Sciences, Isfahan, Iran
3Student Research Committee, Department of Epidemiology, School of Public Health, Iran
University of Medical Sciences, Tehran, Iran
4Nickan Research Institute, Isfahan, Iran
5Department of Medical Genetics, Shahid Sadoughi University of Medical Sciences, Yazd, Iran
6Joint Bioinformatics Graduate Program, University of Arkansas Little Rock and University of
Arkansas for Medical Sciences, Little Rock, AR, USA
Corresponding Author: Azin Mirzazadeh, MD; e-mail: amirzazadeh@ualr.edu,
azin.mirzazadeh68@gmail.com
Clinical display, diagnostics and genetic
implication of Novel Coronavirus (COVID-19)
Epidemic
Art. 1.9378-PM 18925
M. Forouzesh, A. Rahimi, R. Valizadeh, N. Dadashzadeh, A. Mirzazadeh
4608
COVID-19, the differentiation process of causative
agents without presenting an accurate and ef-
cient detection method is leading to false patient
management and unnecessary use of antibiotics.
Recommendation of the best diagnosis protocol is
almost controversial because of overlapped clinical
displays of involved patients.
Specimen Collection of Patients
In March 2020, the USA Centers for Disease
Control and Prevention (CDC) published interim
guidelines regarding the collection, handling, and
testing of clinical specimens for the diagnosis
of coronavirus disease 2019 (COVID-19). Ac-
cordingly, laboratories which test for COVID-19
virus should be strictly aligning with appropriate
biosafety practices. Rapid collection and testing
of appropriate specimens from suspect cases for
COVID-19 is a priority for patient management,
as well as outbreak control, and should be carried
out by a laboratory expert8.
According to the World Health Organization,
recommended respiratory material should be se-
lected from upper respiratory specimens such as
Oropharyngeal (OP) and Nasopharyngeal (NP)
swabs, Nasopharyngeal wash/aspirate and for
lower respiratory specimens including Sputum,
tracheal aspirate, bronchoalveolar lavage (BAL)
uid in patients with the severe condition and
pleural uid (Table I). Additional clinical spec-
imens of COVID-19, as for other coronaviruses
responsible for SARS and MERS can also be
detected in blood and stool9.
Autopsy of Deceased Cases by
Forensic Medicine
Performing an autopsy on a patient with con-
rmed COVID-19 infection is sophisticated from
both spiritually and physically aspects; however,
it is fundamental for giving a forensic certicate
to specify a mortality cause. Forensic medicine
is legally responsible for performing the autopsy.
The question of whether the autopsy of a dead
body with an infectious disease should be per-
formed is the issue of controversy10. The World
Health Organization (WHO) developed Interim
guidance on 24 March 2020 for the safe manage-
ment of a dead body in patients with COVID-19
disease11.
The presence of a manager who is aware of
staff interacting with the dead bodies is high-
ly needed. The specialists of forensic medicine
should try the best to transfer the dead body to
the mortuary area. It should be noted that there
is no need to disinfect it; however, facilitating the
process of burial setting is recommended. Addi-
tionally, there is no need to supply specic tools
to transfer the dead body10 .
It is important to be vigilant of the dead body
with an infectious disease at the time of expi-
ration since the virus can be alive in the lung
and other parts. Having mask and protection
equipment is strongly recommended. Further-
more, air conditioners should be equipped to at
least 160L/s/patient airow or having negative
pressure with at least 12 air changes per hour10.
All parts of the autopsy room must be disinfected
because the virus can remain active for 9 days12.
Clinical and Physical Display of
COVID-19 Patients
One of the key immediate clinical features
used in detection is u antigens. Based upon re-
cent ndings, routinely detected u antigens are
A, B, and H7N-subtypes. One factor for early
rapid screening of u is sampling of throat swabs;
however, it has relatively high false-negative rate7.
In addition to the fever which is still the typ-
ical symptom of 2019-nCoV infection1,3,13 , the
2019-nCoV infected cases have symptoms like
fatigue, dry cough, dyspnea, etc., with or with-
out nasal congestion, runny nose or other upper
respiratory symptoms that also conrmed by
the analysis of 262 infected cases to determine
the clinical and epidemiological characteristics
of COVID-19 in Beijing. They were categorized
into severe and common group respectively, and
the most common symptoms of illness onset re-
ported were fever (82.1%), cough (45.8%), fatigue
(26.3%), headache (6.5%), and dyspnea (6.9%,
and severe cases with dyspnea 32.6%)14. In an-
other retrospective study in Wuhan Jinyintan
Hospital from Jan 1 to Jan 20, 202015, patients
*Patients in severe condition.
Table I. Types of specimen collection.
Upper respiratory specimens
• Nasopharyngeal swab (NP)
• Oropharyngeal swab (OP)
• Nasopharyngeal wash/aspirate
Lower respiratory specimens
• Bronchoalveolar lavage (BAL) and Pleural uid*
• Sputum
• Endotracheal aspirate
Stools, whole blood, urine and infectious material
from autopsy in deceased cases
Clinical display, diagnostics and genetic implication of Novel Coronavirus (COVID-19) Epidemic
4609
had clinical manifestations of fever (83%), cough
(82%), shortness of breath (31%), confusion (9%),
headache (8%) sore throat (5%), rhinorrhea (4%),
chest pain (2%), diarrhea (2%), nausea and vom-
iting (1%) and other minor symptoms including
wheeze in lungs, weakened breath sounds, dull-
ness in percussion, and increased or decreased
tactile speech tremor and muscle ache (Figure
1). This study also showed that some patients de-
veloped acute respiratory distress syndrome and,
among them, (11%) of patients exacerbated in a
short period of time and died of multiple organ
failure. Additionally, the 2019-nCoV infection is
more likely to affect older males with additional
disorders. Given all data extracted from Chinese
hospitals16, generalizing these clinical and physi-
cal symptoms statistics is somewhat controversial
due to the varied nature of the genetic pool and
therefore, different immune responses in each
individual population. Table II illustrates the clin-
ical display of COVID-19.
Clinical Display of Cardiovascular
Disease (CVD) Combined with COVID-19
According to a study on 112 patients in the
Western district of Union Hospital in Wuhan on
their blood samples17, COVID-19 patients com-
bined with CVD were reported to have a higher
risk of mortality and patients posed lower lym-
phocyte counts and a higher pulmonary CT,
C-reactive protein CRP and procalcitonin (PCT)
in blood samples18. This survey also indicates that
contributing factors in the death of this group of
patients include fulminant inammation, lactic
acid accumulation, and thrombotic events. He
et al19 have reported that severe or critically
ill COVID-19 patients accompanied by myo-
cardial injury are associated with a higher risk
of in-hospital mortality. Additionally, Li et al20
have announced that COVID-19 patients with
previous cardiovascular metabolic diseases are at
increased risk for developing a severe illness with
COV I D -19.
Clinical Display of Liver and Kidney
Injuries Combined with COVID-19
Several studies have demonstrated that 2-11%
of COVID-19 patients had liver comorbidities,
and also abnormal levels of alanine aminotrans-
ferase and aspartate aminotransferase (AST)
were seen in 14-53% cases reported during dis-
ease progression. Further, liver injury was more
widespread in severe cases compared to mild
cases of COVID-1921. In addition to liver injuries,
Cheng et al22 have indicated that mortality rates
of COVID-19 patients with acute kidney injury
(AKI) were higher than other patients. Diao et
al23 suggested that the human kidney could be
considered as a particular target for SARS-CoV-2
infection leading to AKI and viral dissemination
Figur e 1. The most common symptoms of COVID-19 (11).
M. Forouzesh, A. Rahimi, R. Valizadeh, N. Dadashzadeh, A. Mirzazadeh
4610
in the body23. Likewise, another study showed
that about 40% of patients admitted to the hospi-
tal had proteinuria and hematuria24.
Clinical Display of Lungs Combined
With COVID-19
The various studies have demonstrated that
the lungs of most patients with novel coronavirus
were involved bilaterally and pathological fea-
tures comprising the alveolar exudative inam-
mation and interstitial inammation, alveolar
epithelium proliferation and hyaline membrane
formation have been obser ved25,26.
Clinical Display of Diabetes Mellitus
Patients With Coronavirus Infection
Results of different studies about the role of
diabetes or raised blood glucose as a risk factor
leading to death in COVID-19 patients are contro-
versial. For instance, a report of 72,314 cases of
COVID-19 published by the Chinese Centre for
Disease Control and Prevention indicated that the
case-fatality rate was raised among COVID-19
patients with diabetes (7.3%)27. Although, in an-
other study, diabetes has not been identied as a
risk factor for disease severity and progression in
140 patients with COVID-19 in Wuhan, China28.
Clinical Display of Pregnant Women
With New Coronavirus Infection
To investigate the clinical characteristics and
placental pathology of 2019-nCoV infection in preg-
nancy, as well as the assessment of intrauterine
vertical transmission potential of this virus, Chen
et al16 performed a study on three pregnant cases.
The clinical manifestations of pregnant women with
2019-nCoV infection in the last trimester pregnan-
cy were similar to those of non-pregnant patients,
and no severe detrimental pregnancy outcome was
found in the 3 case studies. Furthermore, no mor-
phological changes related to infection in the three
placentas were observed. Accordingly, there was no
evidence for intrauterine vertical transmission of
2019-nCoV in the three observed infected women
in their late pregnancy29.
COVID-19 Diagnostics
Methods of Detection
Imaging Examination (CT Imaging)
For both initial evaluation and follow-up, due
to the primary involvement of the respiratory sys-
tem, chest CT is strongly recommended in sus-
Table II. Clinical display of various conditions combined with COVID-19.
Condition Comment
Cardiovascular disease (CVD) • Higher risk of mortality
• Lower lymphocyte counts
• Higher pulmonary CT, C-reactive protein CRP and procalcitonin (PCT) in blood
samples
• Increased risk for developing a severe illness with COVID-19 in patients with
previous cardiovascular metabolic diseases
Liver injury • Abnor mal levels of alanine aminotransferase and aspartate aminotransferase (AST)
• Common in severe cases
Kidney injury • Higher risk of mortality
• Proteinuria and hemat uria
• SARS-CoV-2 infection leading to AKI
Lung • Bilateral lung involvement
• Alveolar exudative inammation
• Interstitial inammation
• Alveolar epithelium proliferation
• Hyaline membrane formation
Diabetes • The uncer tain effect on mortality
Neurologic • Hysgeusia
• Hyposmia
• Anosmia
Pregnant women • Similarity of clinical presentation in pregnant women with SARS-CoV-2 infection
in the last t rimester pregnancy to non-pregnant patients
• No morphological changes in placenta
Clinical display, diagnostics and genetic implication of Novel Coronavirus (COVID-19) Epidemic
4611
pected COVID-19 cases. In the intermediate to
advanced stages of the disease, chest radiographs
may show a progression of features of acute respi-
ratory distress syndrome (ARDS). In contrast, in
early stages, chest radiographs characterized by a
low diagnostic value, while CT ndings may be
present even before symptom onset. According to
several studies on myriads of cases, CT ndings
have proven to have a high diagnostic value in
a number of cases with an initial false-negative
reverse transcription-polymerase chain reaction
(RTPCR) screening test30.
Hematology and Serology Tests
Data extracted from the SARS epidemic prove
that serological responses, including viral-specif-
ic IgM and IgG, can allow for serologic diagno-
sis31, 32. It was shown that patients with 2019-nCov
pneumonia also had similar acute serological re-
sponses and enzyme-linked immunoassay (ELI-
SA) for specic IgM and IgG antibodies as a con-
ventional serological assay provide a high-yield
alternative, and it can be utilized as a uniform test
for all suspected patients and can facilitate more
complete identication of infected cases. In spite
of the fact that the nucleocapsid protein can serve
as a sensitive antigen, other 2019-nCoV-specic
antigens or epitopes should be explored for use
in the serology assay, thus, the use of the whole
N protein as the antigen for the serological assay
can lead to potentially specicity and sensitivity
issues33-3 7.
Autopsy material including lung tissue is highly
recommended in case of deceased patients, while
in surviving patients, paired serum, both acute
and convalescent, can be useful to retrospectively
dene cases. Most recently, it was proved that in
the early stage of the disease, the total number of
lymphocyte count is highly decreased, with de-
creased or increased or normal monocytes. When
the absolute value of lymphocyte is less than 0.8
× 109/L, or the numbers of CD4 and CD8 T cells
are signicantly decreased, high attention is re-
quired generally emphasizing a recheck for the
blood changes after 3 days. To highlight the role
of cytokine, Chen et al36 carried out a study on
29 patients with 2019 novel coronavirus and they
showed an increased level of expression in IL-10,
IL-2R, and IL-6 in serum. These results hallmark
the important role of interleukins as a marker for
prediction of the severity of the 2019-nCoV pneu-
monia and the prognosis of patients38.
There are other serological indicators for
the probability of 2019-nCoV infection, includ-
ing myoglobin, erythrocyte sedimentation rate
(ESR), C-reactive protein (CRP), Procalcitonin
(PCT), liver and kidney function, myocardial
enzyme, lactate, D-dimer, coagulation image,
inammatory factors (TNF - α, interleukin (IL-
6, IL-10), complement, anti-acid staining, urine
routine test, and blood gas analysis. Blood gas
analysis is helpful to determine the oxygen-
ation of moderately to severe infected patients.
Combining this analysis with the observation
of increased lactic acid provides the screen of
patients with high-risk of oxygenation disorder.
Some infected patients have increased D-dimer
with microthrombotic formation and frequent
clotting disorders in blood vessels, increased
liver enzymes, Erythrocyte sedimentation rate
ESR, muscle enzyme and myoglobin. To detect
whether there was bacterial infection in the
lung, the detection of CRP and PCT is of promi-
nent value. Detection of other inammatory fac-
tors may help to a fast evaluation of the immune
status of patients39,40 . Table III demonstrates a
summary of provisional available testing for the
management of patients with conrmed corona-
virus (COVID-19) disease.
Genetics of SARS-CoV-2
Notable Features of the
SARS-CoV-2 Genome
The SARS-CoV-2 genome contains two ank-
ing untranslated regions and a single long open
reading frame encoding a polyprotein includ-
ing replicase complex (orf1ab), S gene, E gene,
M gene, and N gene. According to a recent-
ly published study in Nature on the origin of
SA R S - C oV- 239, there are two prominent genome
characteristics attributed to SARS-CoV-2. This
virus holds a structure that makes this virus
optimum for binding to the human receptor An-
giotensin-converting enzyme, ACE2. Further-
more, polybasic cleavage site and the three adja-
cent predicted O-linked glycans of SARS-CoV-2
structure were not previously seen in lineage B
betacoronavirus family. However, in the newest
computational study performed41, it was shown
that the receptor binding domain RBD sequence
available in this new version of virus does not
bind ideally to ACE2 and this high-afnity bind-
ing of SARS-CoV-2 spike protein to human ACE2
is most likely the result of natural selection which
rules out the role of human manipulation for cre-
ation of this virus.
M. Forouzesh, A. Rahimi, R. Valizadeh, N. Dadashzadeh, A. Mirzazadeh
4612
The Role of the nsp2 and nsp3 in
COVID-19 Pathogenesis
In a homology modeling study performed by
SwissModel and HHPred servers, the transmem-
brane helical segments in coronavirus ORF1ab
nonstructural protein 2 (nsp2) and nsp3 were
tested and the results showed that both stabilizing
and destabilizing mutation of the endosome-as-
sociated-protein-like domain of the nsp2 protein
and nsp3 can explain for the high ability of con-
tagious in COVID-201942.
HLA Haplotypes and
SARS-CoV-2 Infection
One important factor in genetic suscepti-
bility to infectious diseases is the major-his-
tocompatibility-complex antigen loci (HLA).
During co-evaluation with pathogens, selec-
tive HLA-loci variability results from selec-
tive pressure. Immunologists have found that
different HLA haplotypes are associated with
distinct disease susceptibilities. It is mostly due
to T-cell antigen receptors with their ability to
recognize the conformational structure of the
antigen-binding-grove with the associated an-
tigen peptides43. Accordingly, it seems advan-
tageous to have HLA molecules with increased
binding specicities to the SARS-CoV-2 virus
peptides on the cell surface of antigen-present-
ing cells and identication of dominant alleles
will be conducive for the development of detec-
tion kits. Therefore, it is recommended to study
HLA haplotypes to see if specic HLA loci
either class I or II induce protective immunity
in the population.
Nucleic Acid Detection of SARS-CoV-2
There are updated diagnostic methods includ-
ing rapid diagnostic tests (RDT), NAATs, multi-
plex respiratory virus infection assays, CRISPR,
and metagenomics NGS, and the time required
for RT-PCR, CRISPR and metagenomics NGS
diagnostic tests is about 3, 2 and 24 h, respec-
tively44.
*ICU patients had higher plasma levels of IL2, IL7, IL10, GSCF, IP10, MCP1, MIP1A, and TNF-α.
Table III. Techniques for diagnosis of COVID-19 patients.
CT Scan
Hematology and serology tests • Complete blood count (CBC)
• Viral specic IgM and IgG antibodies
• Whole N protein
• Level of expression in IL-2R, IL-6 IL-7, IL-10, GCSF, IP10, MCP1, MIP1A and
TNF-α*
• Prothrombin time
• LDH
• Amylase and D-dimer
• A LT
• C-reactive protein (CRP)
• Procalcitonin (PCT)
• Muscle enzyme and myoglobin
• Myocardial enzyme
• Lactate
• Complement
• Creatine kinase
• ESR
Autopsy material
Molecular Test • Rapid diagnostic tests (RDT)
• Mesa’s test (updated 25 March)
• Coronavir us test: UK to make 15-minute at-home kits available ‘within days50.
• Nucleic acid amplication tests & point-of-care tests (POCT)
• RT-PCR and rRT-PCR
• Loop-mediated isother mal amplication (LAMP)
• Clustered regularly interspaced shor t palindromic repeats (CRISPR)
• Multiplex respiratory virus infection assays
• Shotgun metagenomics sequencing, NGS
Clinical display, diagnostics and genetic implication of Novel Coronavirus (COVID-19) Epidemic
4613
Various culture-independent nucleic acid am-
plication tests (NAATs), as well as point-of-care
tests (POCT), have contributed to the diagno-
sis of unexplained pneumonia. These techniques
are including polymerase chain reaction (PCR),
loop-mediated isothermal amplication (LAMP),
and clustered regularly interspaced short pal-
indromic repeats (CRISPR), etc. Laboratories
participating in the evaluation regularly use the
TaqMan Fast Virus 1-Step Master Mix (Thermo
Fisher Scientic, Waltham, MA, USA), as well
as QIAGEN One-Step RT-PCR Kit with the
specic cycling conditions and concentrations.
All SARS-CoV-2 commercially available or in
development tests for the diagnosis of COVID-19
is listed on this website (https://www.nddx.org/
covid-19/pipeline/).
The current nucleic acid amplication test
methods mainly targeted the open reading
frames of the replicase complex (orf1ab), S,
E, M and N genes. For primer designing two
sequence regions (ORF1b and N) that are high-
ly conserved among sarbecovirus, are often
selected for primer and probe designs Chu et
al37 showed that N gene assay is about 10 times
more sensitive than the ORF-1b gene assay in
detecting positive clinical specimens. Although
the virus (SARS-Cov-2) nucleic acid RT-PCR
test has become the standard method for di-
agnosis of this infection, these test kits have
many limitations. Apart from contamination
and technical problem, the false-negative rates
have been reported due to high genetic diver-
sity of the SARS-CoV-2 genome, it is therefore
difcult to deduce a meaningful evaluation of
what proportion of symptomatic cases are in-
fected and Shotgun metagenomics sequencing
is suggested for conrmation in some negative
NAATs cases45,46.
Shotgun metagenomics sequencing (mNGS)
including short-read and long-read sequencing
could obtain genomic data from both known
and novel pathogens. The rst full genomic
sequence of this coronavirus was released on
January 10, 2020. Many public access databas-
es for deposition of genetic sequence data are
available, including GISAID. The next-genera-
tion sequencing (NGS), and electron microscope
technology play a role in the early diagnosis, and
it can tell about the possible mutation; however,
because of the scarcity of information regarding
this new virus, the combination of RT-PCR,
CRISPR, and mNGS can assure clinical diagno-
sis for COVID-1945- 49.
Conclusions
Currently, there is a big challenge regarding re-
porting asymptomatically infected cases or very
mild cases of infection who are not tested for
viral RNA and they compose a large group of
patients. Therefore, the true rate of infection in
the population still remains unknown.
Conflict of Interest
The Authors declare that they have no conict of interests.
Acknowledgments
The authors thank the European Review for Medical and
Pharmacological Sciences for allowing the quick publica-
tion of this ar ticle without a fee.
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