Review

COVID-19 salivary signature: diagnostic and

J Clin Pathol: first published as 10.1136/jclinpath-2020-206834 on 7 August 2020. Downloaded from http://jcp.bmj.com/ on October 7, 2021 by guest. Protected by copyright.
research opportunities
Dipak Sapkota,1 Tine Merete Søland,1,2 Hilde Kanli Galtung,1 Lars Peter Sand,1
Simone Giannecchini,3 Kelvin K W To,4,5,6 Maria Cassia Mendes-­Correa,7
Daniel Giglio,8 Bengt Hasséus,9,10 Paulo Henrique Braz-­Silva  ‍ ‍7,11

For numbered affiliations see ABSTRACT more widely used nasopharyngeal/oropharyngeal

end of article. The COVID-19 (caused by severe acute respiratory swabs for COVID-19 diagnosis and disease moni-
syndrome coronavirus 2 (SARS-­CoV-2)) epidemic started toring. In addition, we discuss unique opportunities
Correspondence to and possible challenges related to the saliva-­based
in Wuhan (Hubei Province, China) in mid-­December 2019
Professor Paulo Henrique
Braz-­Silva, Division of General and quickly spread across the world as a pandemic. As research activities on COVID-19.
Pathology, Department of a key to tracing the disease and to implement strategies
Stomatology, Universidade aimed at breaking the chain of disease transmission, SALIVA AS A POTENTIAL DIAGNOSTIC FLUID
de Sao Paulo Faculdade de extensive testing for SARS-­CoV-2 was suggested.
Odontologia, Sao Paulo, FOR SARS-COV-2
Brazil; ​pbraz@​usp.​br and Although nasopharyngeal/oropharyngeal swabs are the The emergence of the COVID-19 pandemic has
Professor Dipak Sapkota, most commonly used biological samples for SARS-­CoV-2 highlighted the need for multiple diagnostic strat-
Department of Oral Biology, diagnosis, they have a number of limitations related to egies to efficiently evaluate potential cases in order
Faculty of Dentistry, Oslo, sample collection and healthcare personnel safety. In this
Norway; ​dipak.​sapkota@​odont.​ to provide information on population exposure
uio.​no
context, saliva is emerging as a promising alternative and immunity. These tools currently include virus
to nasopharyngeal/oropharyngeal swabs for COVID-19 molecular testing and rapid host immune response
Received 3 June 2020 diagnosis and monitoring. Saliva collection, being a assays.
Revised 16 July 2020 non-­invasive approach with possibility for self-­collection,
Accepted 21 July 2020
Saliva is a biological fluid in which SARS-­CoV-2
circumvents to a great extent the limitations associated can be found and for this reason saliva has been
Published Online First
7 August 2020 with the use of nasopharyngeal/oropharyngeal swabs. taken into consideration in the diagnosis of
In addition, various salivary biomarkers including the COVID-19. The presence of SARS-­ CoV-2 in
salivary metabolomics offer a high promise to be useful saliva may be related to different sources such as
for better understanding of COVID-19 and possibly in the i) virus entry to the oral cavity from lower/upper
identification of patients with various degrees of severity, respiratory tract,1 5 ii) access to the mouth via oral
including asymptomatic carriers. This review summarises cavity-­specific crevicular fluid or iii) release of viral
the clinical and scientific basis for the potential use of particles in the oral cavity via salivary ducts from
saliva for COVID-19 diagnosis and disease monitoring. the infected salivary glands6 (figures 1 and 2). The
Additionally, we discuss saliva-­based biomarkers and latter observation may explain how COVID-19
their potential clinical and research applications related transmission can occur through asymptomatic cases
to COVID-19. with no obvious infection in the respiratory tracts.
The major salivary glands (parotid, submandib-
ular and sublingual glands) are the major contrib-
utors of saliva secretion (figure 2). Approximately
INTRODUCTION 600–1000 mL of saliva, containing molecules such
An epidemic of a new coronavirus with pneumonia-­ as growth factors, cytokines and secretory IgA, is
like symptoms started in Wuhan (Hubei Province, secreted each day from the human salivary glands.7
China) in December of 2019. The COVID-19, Of note, the unique salivary glands structure
identified to be caused by severe acute respira- with rich surrounding blood circulation has been
tory syndrome coronavirus 2 (SARS-­CoV-2) infec- suggested to facilitate the exchange of molecules in
tion,1 2 spread very quickly across the world and the blood into the salivary acini and subsequently
was declared a pandemic by WHO. As of 24 May in the saliva.8 Saliva has been studied thoroughly
2020, the COVID-19 infection has accounted for as a potential diagnostic tool and it is expected to
>5 000 000 cases with >3 00 000 deaths reported become a substitute for other biological fluids such
worldwide.3 The fast spread of this disease is related as serum or urine in disease diagnosis.8 9 Compared
to its highly infectious nature, and the disease is with other diagnostic fluids, saliva sampling has
suggested to be transmitted through saliva droplets the advantages and disadvantages as mentioned in
© Author(s) (or their
employer(s)) 2021. No and nasal discharge.4 In order to trace the disease table 1.
commercial re-­use. See rights and to implement strategies aimed at breaking the The diagnostic potential of saliva was estab-
and permissions. Published chain of disease transmission, WHO has recom- lished by studies that revealed that, like serum,
by BMJ. mended extensive testing for COVID-19. This is saliva contains hormones, antibodies, growth
To cite: Sapkota D, particularly important as approximately 80% of factors, enzymes, microbes and their products that
Søland TM, Galtung HK, the disease transmission has been reported to be can enter saliva through blood via passive diffu-
et al. J Clin Pathol related to asymptomatic cases.4 Here, we suggest sion, active transport or extracellular ultra filtra-
2021;74:344–349. that saliva-­based testing can be an alternative to the tion. Therefore, saliva can be a reliable fluid for
344   Sapkota D, et al. J Clin Pathol 2021;74:344–349. doi:10.1136/jclinpath-2020-206834
 Review
gland provides pure saliva, but the the collection process is time

J Clin Pathol: first published as 10.1136/jclinpath-2020-206834 on 7 August 2020. Downloaded from http://jcp.bmj.com/ on October 7, 2021 by guest. Protected by copyright.
consuming and requires special equipment. Several protocols
and approaches are available for DNA and RNA extraction and
antibody detection, providing good performances regardless of
sampling technique.11
The diagnostic topic of saliva (called ‘Salivaomics’) includes
the study of salivary proteins (proteomics), the study of sali-
vary RNAs (transcriptomics), the study of salivary metabolites
(metabolomics), the study of salivary microRNAs (microRNA)
and the study of salivary microbiota (microbiome).12
To date, saliva is used for the diagnosis of several diseases
including hereditary diseases, autoimmune diseases, malignan-
cies, infections, dental caries and periodontal disease.13 14 Addi-
tionally, saliva can be used for diagnosing oral diseases with
relevance for systemic diseases or for monitoring of levels of
hormones, drugs and bone turnover markers.12
Diagnosis of saliva-­based viral infections depends on the pres-
Figure 1  Schematic illustration demonstrating clinical implications ence of viral DNA, RNA, microRNA, antigens or host antibodies
and various sources of severe acute respiratory syndrome coronavirus 2 in saliva. In this context, some viruses have been detected in
(SARS-­CoV-2) in saliva. Inset: a suggested mechanism for SARS-­CoV-2 saliva up to 29 days after infection, indicating that a saliva-­based
entry into the salivary gland cells—the viral spike protein binds with non-­invasive diagnostic platform can be useful for early diag-
the ACE2 receptor on the surface of the cell, followed by its priming nosis and for monitoring the disease and treatment.9 15
with serine protease TMPRSS2 and subsequent entry into the cells. After
replication and packaging, multiple new virus particles are released
SALIVA TESTING FOR SARS-COV-2
from the cells in saliva.
SARS-CoV-2 detection using reverse-transcription PCR
SARS-­CoV-2 is an enveloped, positive single-­stranded RNA virus
monitoring the physiological function of the body.10 Although consisting of a core of RNA genome associated with nucleocapsid
the low concentration of some analytes in saliva compared with protein (N) and surrounded by a phospholipid membrane with
the blood previously proved challenging, the advent of highly three main viral structural proteins, spike surface glycoprotein
sensitive molecular methods and nanotechnology have to a large (S), small envelope protein (E) and matrix protein (M).16 Nucle-
extent circumvented this limitation. otide sequences within a number of SARS-­CoV-2 genes such as
Collection of saliva can be done in several ways, such as spitting E, RdRp, N1 and N2 and S can be used as detection targets
out, collection with the help of sponge-­like device and directly for RT-­PCR-­based test methods.17 On the other hand, detection
from the salivary gland duct.7 The spitting out technique is the of SARS-­CoV-2 antigens and/or immunoglobulins against them
cheapest one, and the saliva sample thus collected also includes form the basis for enzyme immunoassays.17
nasopharyngeal/oropharyngeal/airway secretions. Sponge-­ like Presently, RT-­PCR is the most commonly used diagnostic test
devices provide relatively more pure saliva, but this technique for the detection of SARS-­CoV-2 RNA in the biological samples.
requires special equipment which is not always widely avail- For large-­scale testing as in the case of SARS-­CoV-2, proper
able. Saliva collected directly from the ducts of major salivary selection of the type and the site of biological specimen collec-
tion is crucial for obtaining reliable test results.18 Biological
samples from the upper (such as nasopharyngeal swabs, oropha-
ryngeal swabs, throat swabs, nasal swabs) and lower (such as
tracheal aspirates and brochoalveolar lavage) respiratory tracts
can be used for the detection of SARS-­ CoV-2 with varying
degree of test sensitivity19–21 (figure 2). Tracheal aspirates and
bronchoalveolar lavage, although more reliable for SARS-­CoV-2
detection, are the less preferred specimens as compared with
the nasopharyngeal/oropharyngeal swabs due to technical
complexity in obtaining these samples.21 Currently, nasopharyn-
geal/oropharyngeal swabs where virus samples are collected by
respectively rubbing the nasopharyngeal wall and the posterior
pharynx/tonsillar areas with minitip swabs, are routinely used
for SARS-­CoV-2 detection17 (figure 2).
Despite the widespread use, the collection of nasopharyngeal/
oropharyngeal swabs has a number of limitations.22 23 The collec-
tion of these swabs is less acceptable to patients as compared
with non-­invasive methods like saliva collection, as it tends to
cause patient discomfort and even bleeding. Patient acceptance
is highly desirable for test methods where multiple testing is
Figure 2  Schematic illustration demonstrating major salivary glands needed for disease monitoring and follow-­up, as in the case of
(parotid, submandibular and sublingual) and their respective ducts, COVID-19. Furthermore, the risk for disease transmission to
oropharynx and nasopharynx, and approximate anatomic locations for the healthcare personnel when collecting these samples is high
collection of oropharyngeal and nasopharyngeal swabs. as it requires active involvement of the test taker. Additionally,
Sapkota D, et al. J Clin Pathol 2021;74:344–349. doi:10.1136/jclinpath-2020-206834 345
 Review

Table 1  Advantages and disadvantages of saliva sampling

J Clin Pathol: first published as 10.1136/jclinpath-2020-206834 on 7 August 2020. Downloaded from http://jcp.bmj.com/ on October 7, 2021 by guest. Protected by copyright.
Advantages Disadvantages
Non-­invasive approach for disease diagnosis and monitoring of general health. Not always reliable for measurement of certain markers.
Painless (no patient discomfort and anxiety for sampling). Contents of saliva can be influenced by the method of collection, degree of stimulation
of salivary flow, interindividual variation and oral hygiene status.
Easy collection and applicable in remote areas. Serum markers can reach whole saliva in an unpredictable way.
Relatively cheap technology. Medications may affect salivary gland function and consequently the quantity and
composition of saliva.
Cost-­effective applicability for screening large populations. Possibility for degradation of salivary proteins due to presence of proteolytic enzymes.
Suitable for children, anxious/disabled/elderly patients.
Possible multisampling.
Safer collection for health professionals than other biological samples such as
nasopharyngeal swabs and blood.
Cheap to store and ship.
Easy to handle.
No need for expensive equipment/instruments (swabs, suction tubes or special collection
devices) for collection. Only needs a sterile container.

collection of these samples demands the use of personal protec- SARS-­ CoV-2 for cases with moderate-­ to-­severe symptoms,25
24
tive and healthcare resources, both of which tend to be in short and for asymptomatic or mild cases. The latter is particularly
supply in a pandemic like COVID-19. important for the screening of the suspicious/asymptomatic cases
and for the surveillance of the healthcare workers. Self-­sampling
Saliva as a biological fluid for molecular detection of SARS- of saliva could also be an option in large-­scale population-­based
CoV-2 point-­prevalence studies.
Saliva is emerging as a promising alternative to nasopharyn- Being a non-­invasive specimen type, saliva is well-­suited for
geal/oropharyngeal swabs for COVID-19 diagnosis and moni- serial viral load monitoring. The SARS-­CoV-2 load in the saliva
toring.24 25 Indeed, the use of saliva as a biological specimen for is reported to be highest after the first week of symptom onset,
SARS-­CoV-2 testing to a great extent circumvents the above-­ followed by a gradual decline.25 28 29 This underlines that saliva
mentioned limitations associated with the use of nasopharyn- is a good candidate for SARS-­CoV-2 detection in earlier disease
geal/oropharyngeal swabs. With clear instructions, patients can phase. The temporal profile of SARS-­CoV-2 load in saliva has
self-­collect saliva samples. This is highly desirable in an outbreak been reported to be more consistent25 as compared with that
in order to minimise the burden on healthcare personnel, the use of nasopharyngeal swabs, suggesting its suitability for disease
of personal protective equipment and to allow serial sampling monitoring. Furthermore, saliva can be used for monitoring
required for disease monitoring. A recent study has reported that the response to antivirals in clinical trials.34 Nonetheless, saliva
self-­collection of saliva sample for SARS-­CoV-2 testing is feasible can also be a potential source of viral transmission, thereby
and can produce reliable test results.24 requiring standard protocols for its collection and subsequent
The potential use of saliva for SARS-­ CoV-2 detection is handling. In the light of these promising results, a saliva-­
scientifically well founded. Saliva is considered to be a good based SARS-­CoV-2 RNA detection assay has already obtained
reservoir for viruses that originate from oral shedding, and approval through the US Food and Drug Administration emer-
secretions from the lower respiratory tract, nasopharynx and gency use authorisation.
possibly infected salivary glands22 23 (figure 1). Indeed, Chen et
al were able to detect SARS-­CoV-2 RNA in three out of four
saliva samples directly collected from the salivary gland ducts,
thereby precluding contamination from respiratory secretions, Table 2  Main findings of recent studies on SARS-­CoV-2 detection in
of critically ill cases.26 Together with the demonstration of ACE2 saliva samples by using RT-­PCR.
expression,27 a main surface receptor type for SARS-­CoV-2,1 in Authors Main finding(s) related to salivary specimens
the salivary gland, the above findings substantiate the idea that
To et al28 91.7% of nasopharyngeal swab-­diagnosed cases.
salivary gland could be one of the sources for SARS-­CoV-2 in
Live virus was detected in saliva using viral culture.
saliva. In line with this observation, recent studies by To et al
To et al29 87% of nasopharyngeal swab-­diagnosed cases.
demonstrated the presence of live SARS-­CoV-2 in saliva.28 29
Salivary viral load was highest during the first week of symptom.
Furthermore, the possible diagnostic use of saliva for several
respiratory viruses including coronavirus has been supported by Azzi et al32 Detected in all nasopharyngeal swab-­diagnosed cases.
studies demonstrating a high sensitivity and specificity of saliva-­ Kojima et al24 Self-­collected saliva and nasal swab had similar sensitivity as
compared with the clinician-­collected nasopharyngeal swabs
based tests, with >90% concordance between saliva and naso-
pharyngeal swabs.30 Wyllie et al25 Saliva is more sensitive and consistent than nasopharyngeal
swabs
Current studies from different groups have shown promising
Williams et al31 84.6% of nasopharyngeal swab-­diagnosed cases.
results on the possible use of saliva for detection of SARS-­CoV-2
Viral load was higher in the nasopharyngeal swab.
RNA24 25 28 29 31–33 (table 2). The sensitivity of saliva-­ based
SARS-­CoV-2 RNA detection methods seem to be comparable Pasomub et al33 84.2% of nasopharyngeal swab-­diagnosed cases.
to24 32 or better than that of nasopharyngeal swabs.25 Addition- Saliva might be an alternative specimen for COVID-19 diagnosis.
ally, saliva seems to be a good candidate for the detection of SARS-­CoV-2, severe acute respiratory syndrome coronavirus 2.

346 Sapkota D, et al. J Clin Pathol 2021;74:344–349. doi:10.1136/jclinpath-2020-206834

 Review
Antibodies against SARS-CoV-2 dehydrogenase, malic acid, guanosine monophosphate and

J Clin Pathol: first published as 10.1136/jclinpath-2020-206834 on 7 August 2020. Downloaded from http://jcp.bmj.com/ on October 7, 2021 by guest. Protected by copyright.
Besides RT-­PCR-­based RNA detection of SARS-­CoV-2, prelim- proteins associated with macrophage, platelet degranulation
inary studies have reported promising results for the detection and complement system pathways are shown to be present in
of IgM and IgG against SARS-­CoV-2 in serum/plasma samples saliva. These findings support the possible use of saliva-­based
of patients with SARS-­CoV-2.35 Interestingly, the production of metabolic/protein/lipid biomarkers as a non-­invasive approach
SARS-­CoV-­specific secretory IgA in the saliva of mice intrana- for patient stratification in COVID-19 disease.
sally immunised with SARS-­CoV virus-­like particles has been Metabolomics is a strategy used in the study of small mole-
documented.36 Hence, it is reasonable to speculate that anti-­ cules from the metabolic profile of cells, tissues or fluids, which
SARS-­CoV-2 antibodies might also be present in human saliva. help in the characterisation of a phenotype. These molecules,
Viral antibodies have been detected in saliva and the immu- called biomarkers, are fundamental in clinical practice for deter-
nisation status of measles, rubella, mumps and hepatitis can mining the state of a disease.46 Thus, metabolomics has helped
be verified by analysing IgG, IgM and IgA in oral fluids.37 38 to identify biomarkers with diagnostic potential and description
Regarding SARS-­CoV-2, to date only a study protocol aimed to of metabolic pathways in the most diverse clinical situations,
analyse IgG, IgM and IgA in different biological fluids including including those involving viral and bacterial pathogens, and
self-­collected saliva for rapid SARS-­CoV-2 diagnosis has been more specifically viruses that cause respiratory diseases such as
published.39 However, there are so far no results describing the influenza47 48 and SARS.49 In a study by Wu et al,49 patients
presence of antibodies against SARS-­ CoV-2 in human saliva. recovered from severe acute respiratory syndrome caused
This clearly warrants future studies on the potential use of sali- by SARS-­CoV were recruited after 12 years of infection for
vary immunoglobulins for COVID-19 in diagnostics, disease metabolic evaluation of the consequences of the disease. The
progression and immunisation monitoring. comparison of patients’ serum with healthy individuals showed
differences in organic acids, amino acids, phospholipids, carni-
tine and inositol derivatives.49 These results exemplify the prac-
Saliva biomarkers—perspectives for COVID-19 diagnosis and tical application of metabolomics in the evaluation of long-­term
prognosis outcomes.
Salivary biomarkers and their role in point-­of-­care applications MicroRNAs, non-­ coding RNAs of 20-­ nucleotide to
have highlighted the development of the use of more advanced 22-­nucleotide length, silencing gene expression by a transcript-­
technologies such as micro/nanoelectro-­ mechanical systems, specific target-­mediate inhibitory activity, play a key role in several
paper-­based technology, RNA-­sequencing, liquid biopsy, fluo- cellular processes including cell development and differentiation,
rescent biosensors, photometric and electrochemical methods, immunity, cell metabolism, proliferation, apoptosis and cancer.50
induced release and measurement method.12 40
electric field-­ The relevance of monitoring microRNA is related to the fact that
Contemporary available point-­of-­care can be delivered in form a single microRNA can be implicated in several cellular regula-
of small and portable smartphones or ‘lab-­on-­chips’.40 tory pathways, which involve different molecules. To date, there
Coronaviruses, such as SARS-­ CoV and Middle East respi- are studies reporting a particular microRNA upregulation and
ratory syndrome (MERS)-­CoV), have developed strategies to downregulation of nuclear factor-κB pathway and IFN pathway
decrease or delay the production of interferon (IFN), triggering associated with several viruses including respiratory virus infec-
exuberant inflammatory responses leading to severe pulmo- tion.51 In this context, studies reporting coronavirus (including
nary conditions.41 42 The host’s unregulated immune response SARS-­CoV) regulation of cellular microRNA showed the over-
and the production of inflammatory cytokines, known as ‘cyto- expression of miR-574-­5p and miR214 and regulation of miR-9
kine storm’, are believed to correlate with disease severity and and miR-98 with effect on apoptosis, cancer and autoimmune
poor prognosis during SARS-­CoV and MERS-­CoV infection.41 functions. Of note, SARS-­CoV has been reported in the direct
Several pro-­ inflammatory cytokines and chemokines, such as viral nucleocapsid downregulation of miR-223 and miR-98
chemokine (C-­C motif) ligand (CCL)-2, CCL-3, regulated on expression, with effect in pro-­inflammatory cytokine produc-
activation, normal T cell expressed and secreted (RANTES), tion.52 Additionally, in this context, since microRNAs associ-
interleukin (IL)-2 and IL-8, were highly expressed during ated with extracellular vesicles are known to be protected from
MERS-­CoV infection.41 Recent studies have reported that severe enzymatic degradation, several studies have been focused on the
cases of COVID-19 exhibit increased plasma levels of IL-2, IL-6, investigation of the expression of microRNAs in extracellular
IL-7, IL-10, granulocyte colony stimulating factor (GSCF), vesicles obtained from saliva as potential biomarkers. Thus, the
INF-γ-inducible protein-10 (IP-10), macrophage chemotactic fact that microRNA present in biological fluid can reproduce the
protein 1, macrophage inflammatory protein-­1A and tumour molecular event within the cellular context, make them a poten-
necrosis factor-α compared with mild cases, indicating that the tial exhaustive marker to check the cell-­infection status; this is
inflammatory response mediated by cytokine release is critical especially important in a low replicative condition in which virus
in the progression of COVID-19.42 43 Markers of the inflam- cannot be present in biological fluid,15 and provides an oppor-
matory process, such as cytokines and chemokines, can be tunity to assess virus pathological effect-­associated diseases as in
measured in saliva. Such information has been suggested to be COVID-19.
useful for the diagnosis and prognosis of both oral cavity and
systemic diseases.13 Hence, it is possible to establish an inflam-
matory profile of COVID-19 by analysing inflammation-­related Saliva mark of SARS-CoV-2 cell-receptor features in COVID-19
biomarkers in saliva. Its well known that SARS-­CoV-2 infects host cells, including those
Unique proteomic, metabolic and/or lipid profiles in serum/ in the respiratory tract lining, mainly using ACE2 receptor.53 It
plasma have been suggested to be useful in stratification of fatal/ is reported that SARS-­CoV spike protein S has a high affinity for
severe COVID-19 cases from the mild and healthy ones44; and the ACE2 receptor and is activated by host type II transmem-
to predict progression of COVID-19 patients from a milder brane serine protease TMPRSS2 on primary target cells to fulfil
from to severe stages.45 Interestingly, some of the identified viral entry54 (figure 1). However, other host proteases such as
biomarkers in these studies such as C reactive protein, lactate furin on the tongue may be implicated in cleaving SARS-­CoV-2
Sapkota D, et al. J Clin Pathol 2021;74:344–349. doi:10.1136/jclinpath-2020-206834 347
 Review
viral envelope glycoproteins in its furin-­like cleavage site and scientifically well founded. However, further studies will be key

J Clin Pathol: first published as 10.1136/jclinpath-2020-206834 on 7 August 2020. Downloaded from http://jcp.bmj.com/ on October 7, 2021 by guest. Protected by copyright.
enhancing infection with host cells.55 to understanding the mutual relationship between COVID-19
The use of ACE2 receptor by other coronaviruses to infect sali- and saliva, leading to the adoption of less invasive diagnostic
vary gland epithelial cells has been reported in rhesus macaques.6 techniques and facilitating the application of molecular tests on
Additionally, in vitro analysis of RNA-­seq profiles from four a large scale, a central strategy for controlling the epidemic.
public and consensus datasets revealed the expression of ACE2 The search for salivary biomarkers associated with the devel-
receptor in human granular cells in salivary glands.26 These opment and progression of COVID-19 could allow a better
observations suggest that the salivary glands can be a reservoir distinction between asymptomatic, mild, moderate or advanced
for SARS-­CoV-2 and contribute to the presence of the transmis- disease. Knowledge of this kind might lead to the development
sible form of viral infectious particles in saliva.56 Additionally, it of point-­ of-­
care devices, which can be extremely useful for
is possible that the salivary glands can harbour latent COVID-19 understanding of the evolution of contagions and immunolog-
infection with possibility for subsequent reactivation. The latter ical responses in population studies.
suggestion clearly warrants further studies.
In addition to the salivary glands, ACE2 is abundantly expressed Author affiliations
1
in the oral epithelial cells with highest expression in the tongue Department of Oral Biology, Faculty of Dentistry, University of Oslo, Oslo, Norway
2
when compared with buccal and gingival tissues, T cells, B cells Department of Pathology, Rikshospitalet University Hospital, Oslo, Norway
3
Department of Experimental and Clinical Medicine, Universita degli Studi di Firenze,
and oral fibroblasts.57 These results raise a possibility that oral Firenze, Toscana, Italy
epithelial cells can function as a host for SARS-­CoV-2. Epithe- 4
State Key Laboratory for Emerging Infectious Diseases, Department of Microbiology,
lial cells in the oral mucosa are protected by a viscous mucous Li KaShing Faculty of Medicine of the University of Hong Kong, Hong Kong, China
5
layer containing large glycoprotein macromolecules—mucins— Department of Microbiology, Queen Mary Hospital, Hong Kong, China
6
Department of Clinical Microbiology and Infection Control, The University of Hong
produced in the salivary glands, and water. Virus particles must
Kong – Shenzhen Hospital, Shenzhen, China
penetrate the mucous layer to be able to infect the cells in the 7
Laboratory of Virology (LIM-52), Institute of Tropical Medicine of São Paulo, School
epithelial lining. In the respiratory tract, mucins in the mucous of Medicine, University of São Paulo, São Paulo, Brazil
8
layer have been shown to play a significant role in protecting Department of Clinical Oncology, The Sahlgrenska Academy, University of
airway epithelium to influenza and respiratory syncytial virus.58 Gothenburg, Gothenburg, Sweden
9
Department of Oral Medicine and Pathology, Institute of Odontology, The
Surprisingly, few studies have looked at the role of mucous layer Sahlgrenska Academy, University of Gothenburg, Gothenburg, Sweden
covering human epithelial linings during SARS-­ CoV-2 infec- 10
Clinic of Oral Medicine, Region Västra Götaland, Gothenburg, Sweden
tion. Case reports have described mucous plugging in the lungs 11
Department of Stomatology, School of Dentistry, University of São Paulo, São Paulo,
in postmortem examinations of patients that have succumbed Brazil
to COVID-19.59 Assuming that oral epithelial cells can be a
possible route of entry for the SARS-­CoV-2, studies are needed Handling editor  Tahir S Pillay.
to understand the possible ways the virus particles penetrate the Acknowledgements  The authors would like to thank Evan Michael Vallenari for
mucous layer and infect the underlying epithelial cells. language correction of the manuscript.
Of note, it has been reported by using airway epithelial cells in Contributors  DS, SG, BH and PHB-­S contributed to conception, design, data
vitro that ACE2 is a human IFN-­stimulated gene suggesting that interpretation and drafted the manuscript; TMS, HKG, LPS, KKWT, MCM-­C and DG
SARS-­CoV-2 could exploit IFN-­driven upregulation of ACE2 as contributed to design and data interpretation, critically revised the manuscript. All
a mechanism to enhance viral infection and play a role in the authors gave their final approval and agreed to be accountable for all aspects of the
work.
development of COVID-19 pathogenesis.60 All of these find-
ings point out the potential interest in the investigation of saliva Funding  The authors have not declared a specific grant for this research from any
funding agency in the public, commercial or not-­for-­profit sectors.
viral and host biomarkers as an opportunity to obtain a more
complete molecular view of clinical relevance in the COVID-19 Competing interests  None declared.
risk assessment as well as to develop new therapeutic antiviral Patient consent for publication  Not required.
treatments. Provenance and peer review  Not commissioned; externally peer reviewed.

ORCID iD
CONCLUSION AND FUTURE PERSPECTIVES Paulo Henrique Braz-­Silva http://​orcid.​org/​0000-​0002-​1842-​9521
Saliva-­based testing can be an alternative to the more widely
used nasopharyngeal/oropharyngeal swabs for COVID-19
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