The Role of Saliva in Examining for SARS-CoV-2
1Department of Oral Biology, Faculty of Dental Medicine, Universitas Airlangga, Surabaya, Indonesia.
2Department of Oral Pathology and Maxillofacial, Faculty of Dental Medicine,
Universitas Airlangga, Surabaya, Indonesia.
3Master Student of Dental Health Science, Faculty of Dental Medicine,
Universitas Airlangga, Surabaya, Indonesia.
4Student of Conservative Dentistry Specialist of Dental Medicine, Faculty of Dental Medicine,
Universitas Airlangga, Surabaya, Indonesia.
5Undergraduate Student, Faculty of Dental Medicine, Universitas Airlangga, Surabaya, Indonesia.
*Corresponding Author E-mail: arundinaira@gmail.com
ABSTRACT:
SARS-CoV-2 causes COVID-19 (Corona Virus Infection Disease-19), an infectious disease (Severe Acute Respiratory Syndrome Corona Virus-2)virus which was first confirmed on December 2, Wuhan, China, in 2019 and very quickly world-wide. World Health Organization (WHO) declared it on March 11th, 2020,fever, cough, and breathing difficulties are all common symptoms. The gold standard for diagnosing SARS-CoV-2 infection is the real-time reverse transcription polymerase chain reaction (rRT-PCR)by detecting viral RNA using nasopharyngeal swabs and other respiratory specimens. However, this process necessitates the use of specialist medical staff, centralized laboratory facilities, and a lengthy period of time for results to be obtained. Besides that, the possibility of virus transmission is not negligiblefor procedure's operator. Due to this reason, several investigations have shown that other bodily fluids, like as saliva, could be used to identify SARS-CoV-2. Saliva is used as a diagnostic sample has many advantages: it is easy to collect samples by the patient themselves with a fairly comfortable procedure, does not require health personnel with special expertise to manage it, and reduces the risk of transmitting the virus to the operator. Currently, there are several alternative tests for SARS-CoV-2 which have simpler and more efficient procedures, such as antigen based rapid diagnostic tests, antibody based rapid diagnostics, and RT LAMP tests. This examination is suitable for screening and mass examination, but cannot be used as a primary diagnostic tool because the sensitivity and specificity are not as high as rRT-PCR.
KEYWORDS: Good Health and Well-Being, Diagnostic test, Immunoassay, Saliva, SARS-CoV-2.
INTRODUCTION:
By the year's endof 2019, precisely in December, an outbreak of pneumonia appeared with characteristic symptoms of fever, dry cough, fatigue, and indigestion, the exact cause of them was unknown. The outbreak was first remarkedin the Chinese city of Wuhan.
Most of these pneumonia patients came from vendors at the Huanan market that sells live animals in Wuhan, China. Researchers discovered the culprit of this pneumonia on January 7, 2020, and it was the new coronavirus. The sickness has been dubbed COVID-19 (Corona Virus Disease 2019) by the World Health Organization, and SARS-CoV-2 is the virus's name(Severe Acute Respiratory Syndrome Coronavirus 2)1–3.
This virus is spread by way of droplets released resulting from coughing, sneezing, or speaking according to Wuhan literature, and has a 5.2-day incubation period. While the period of symptom onset to death for people infected with COVID-19 is around days 6 through 41, with a 14-day average4. At the beginning of the pandemic, there were several diagnostic tests recommended by WHO for the diagnosis of COVID-195. Viral RNA examination using rT-PCR through a nasopharyngeal swab is considered to be the "gold standard". However, there are some limitations in this examination, namely it takes up to 1 day to get results, making it difficult to use those that require fast results, such as the need for mass screening. At the height of the pandemic, laboratory sample processing for rT-PCR was overloaded, reducing the effectiveness of other diagnostic tests, such as examining patients with chronic diseases. Besides that, sampling using the nasopharyngeal swab method requires special expertise from medical personnel and there is a risk of transmitting the virus to the operator, and can cause irritation to the nasopharynx, discomfort, sneezing, and coughing5–9.
Based on these reasons, several studies have stated that there are other body fluids that can be alternative samples to detect SARS-CoV-2, including urine, feces, tears, and saliva. Among several body fluids, saliva is one of the components that become the main alternative for SARS-CoV-2 diagnostic examination after nasopharyngeal swab. The advantage of using a saliva sample is easy, without causing discomfort, does not require special skills of medical personnel, and minimizes the transmission of the virus to medical personnel. Several peer-reviewed journals, the media, and companies have reported on the latest breakthroughs in salivary testing to identify infection with SARS-CoV-2 in the recent few months6,10,11.
SARS-CoV-2:
Coronavirus is an enclosed using a single virus positive RNA strain. Corona viruses belong to the Orthocoronavirinae subfamily, with characteristic spikes such as "crowns"on the virus's surface2. Protein S, often known as spike, is a viral antigen protein that contains the primary structure for gene writing. The attachment and penetration of viruses into host cells are aided by protein S. (interaction of protein S with its receptors on host cells).
Pathogenesis:
Coronaviruses can only multiply via infecting their host cells. This virus is incapable of surviving without the presence of host cells. The initial step in the coronavirus cycle is the virus's attachment and entry into cell of the host, which has been mediated by the virus's surface protein S. Protein S is the major determinant in infecting the host species as well as the tropical determinant. The initial step in the coronavirus cycle is the virus's attachment and entry into the host cell, which is mediated by the protein S on the virus's surface. Protein S is the major determinant in infecting the host species as well as the tropical determinant. Besides that, protein S binds to receptors on host cells, namely the enzyme ACE-2 (Angiotensin-Converting Enzyme 2). Mucosa of the mouth and nose, nasopharynx, lungs, stomach, small and large intestines, skin, thymus, bone marrow, spleen, liver, kidney, brain, pulmonary alveolar epithelial cells, small intestinal enterocytes, arteriovenous endothelial cells, and smooth muscle cells are all known to contain ACE-21,3,12,13.
This virus will duplicate the genetic material and proteins required inside the cell before forming new versions on the cell surface. This virus, like SARS-CoV, releases its RNA genome into the cytoplasm after entering the cell, and the Golgi cells are then translated into two lipoproteins and structural proteins in order to replicate. The severity of this COVID-19 infection is determined by the viral component and the immunological response. The virus's cytotoxic effect and ability to overwhelm the immune response are factors in the severity of viral infection. Inadequate immune system in response to infection also determine the severity. On the other hand, excessive immune response also contributes to tissue damage. When the virus enters the cell, the viral antigen will be presented to the Antigen Presenting Cell (APC). Cell presentation to APC will respond to humoral and cellular immune systems mediated by T cells and B cells. IgM and IgG are formed from the humoral immune system. In SARS-CoV, IgM will disappear on day 12 and IgG will last longer. Viruses can escape from the immune system by inducing double membrane vesicles that do not have pattern recognition receptors (PRRs) and can replicate in these vesicles thus they cannot be recognized by immune cells5,14–16.
Figure 1: Illustration of the pathogenesis of SARS CoV 19.6
Transmission:
In COVID-19, it is not known with certainty the process of transmission from animals to humans, but phylogenetic data allows COVID-19 to also be a zoonosis. The development of further data shows that transmission between humans (human to human), which is in most cases, the virus enters the open mucosa through droplets and contact with the virus. SARS-CoV-2 is distributed mostly via respiratory droplets derived from infected persons' mucosal secretions, which are aerosolized while coughing, sneezing, or talking and can be transferred through the air and contaminated surfaces. The investigation revealed that one patient may infect up to three persons around him, but the probability of transmission during the incubation phase made the patient's contact duration with those around him longer, increasing the risk of multiple contacts contracting from one patient17,18.
Clinical Symptoms:
The intensity of symptoms encountered by COVID-19 patients varies greatly. Some people may be asymptomatic or have little symptoms, while others may develop a severe and life-threatening respiratory crisis. Sore throat, fever, cough, muscle aches, headache, and loss of smell (anosmia) and taste are also common symptoms (dysgeusia). Progressive pulmonary pathology can develop in severe cases, commencing with shortness of breath and advancing to pneumonia or acute respiratory distress syndrome (ARDS), which frequently necessitates intubation and mechanical lung ventilation. Progressive pulmonary pathology can develop in severe cases, commencing with shortness of breath and advancing to pneumonia or acute respiratory distress syndrome (ARDS), which frequently necessitates intubation and mechanical lung ventilation 19,20.
COVID-19 Check Flow:
Rapid collection of appropriate specimens from patients with strong suspicion of COVID-19 infection and accurate laboratory diagnosis of these patients are two priorities that underpin infection control and clinical treatment of patients are two topics that come up frequently. Ensure that health professionals collecting clinical specimens from suspected patients follow infection prevention and control (IPPC) recommendations and use adequate PPE (PPE). A person infected with a virus may test positive for viral nucleic acid or viral proteins without displaying symptoms (asymptomatic) or before displaying symptoms (presymptomatic), as well as during periods of disease (symptomatic)21,22.
Nucleic Acid Amplification Test (NAAT) is the standard's gold for confirming COVID-19 is a virus that has been identified. The viral genes that are generally sought are the N, E, S and RdRO genes. NAATs that can do viral RNA nucleic acid sequencing include real-time reverse-transcription polymerase chain reaction (rRT-PCR). The type of sample for NAAT examination can come from lower respiratory system, such as aspiration, sputum, and lavage; or upper respiratory tract, such as a nasopharyngeal, oropharyngeal swab, or nasopharyngeal wash with aspiration23,24.
A negative RT-PCR result cannot rule out infection with the COVID-19 virus. Several factors such as poor quality of specimens, timing of specimen collection that is too slow or too fast, improper storage or shipping of specimens, inappropriate sampling techniques, viral mutation, and polymerase chain reaction (PCR) inhibition can cause false negative results25.
Figure 2: Flow of Antigen Examination with NAAT11
Antigen Rapid Diagnostic Test (Ag-RDT) is used for the purpose of cost and time efficiency. Most of the Ag-RDT for COVID-19 use the sandwich immunodetection method with a compact lateral flow test format. The target of the analyte is the SARS-CoV-2 virus's nucleocapsid protein. Specimens used nasal or nasopharyngeal swabs. According to research, Ag-RDT has a sensitivity of 0-94% and a specificity of >97%. Patients with a lot of viruses (Ct values of 25 or >106 genomic viral copies/mL are considered high) who are in the early stages of their illness (1-3 days before onset) and approaching the symptomatic phase benefit from Ag-RDT (5-7 days at the first day of illness). Although Ag-RDT is not indicated for clinical patient care, it can aid in patient management, public health policy development, and surveillance26. Examination of viral sequencing tests aims to confirm the virus and monitor viral genome mutations.
There are commercial and non-commercial assays available that measure antibody binding (total immunoglobulin (Ig), IgG, IgM, and/or IgA in various combinations) with techniques such as LFI, ELISA, and CLIA. The gold standard for identifying the presence of functional antibodies is the assay. This test necessitates highly trained personnel and a BSL 3 culture facility, and it is not appropriate for regular diagnostic tests.Some of the supporting examinations to establish the diagnosis of COVID-19 are complete blood tests (hematologic, clinical chemistry, immunology/ infection), blood gas analysis, CT scan of the thorax, chest X-ray, ultrasound of the lungs25,26
Immune Components in Saliva:
The oral epithelium and lamina propria operate as a physical barrier, preventing microorganisms from entering the mouth from invading the underlying tissue and prevents some environmental dangers from penetrating. Macrophages, dendritic cells, naturally occurring killer cells, polymorph nuclear leukocytes, and soluble immunoinflammatory mediators such as cytokines, chemokines, antimicrobial peptides, and complement components system protect the mucosa. Oral mucosal immunity is enhanced by salivary secretory immunoglobulin A (sIgA), biologic mediators produced from oral keratinocytes, and components of the crevicular fluid of the gingiva27.
· Secretory Ig-A (SIgA):
Secretory immunoglobulin A (sIgA) antibodies in the oral mucosa help to maintain oral immunity by preventing germs from colonizing and invading the epithelium as a form of humoral defense. The switch from Ig to IgA isotype class can be T helper dependent (dependent) or without T helper (independent) and is dendritic cells and monocytes produce cytokines that aid in the process28.
Figure 3: Factors related to oral immunity14
Saliva contains the oral mucosa, which is protected from bacterial invasion by sIgA, mucins, and enzymes. Oral immunity is aided by leukocytes, IgG, IgM, IgA, and other substancesin the gingival crevicular fluid that reaches the oral cavity. After collecting foreign antigens, Langerhans cells and other myeloid dendritic cells travel to sites that induce immunological responses (Waldeyer ring, regional lymph nodes), where they become key effector cells of the immune system. These immune effector cells then move to the propria lamina (mucosal lymphoid foci), where they control the active immunological response or tolerance to the immunological system28.
· Gingival Fluid:
Oral immunity is aided by leukocytes, IgG, IgM, IgA, and other substancesin the gingival crevicular fluid that gets into the mouth. Gingival fluid has been considered an immunoglobulin. The IgG, IgA and IgM molecules contained in the gingival fluid may contain some specific antibodies against members of the microflora in the gingival crevice29
Saliva as a Test Sample:
Because SARS-CoV-2 mostly attacks the nasal passages, it's important to understand how it spreads, possibly through the eyes, with draining into the URT following, With the appearance of SIgA antibodies in URT secretion, this can be used as a basis for anticipating that the initial immune response at work is via the mucosal immune system, as well as saliva and tear fluid. It is also feasible to produce serum IgG and IgA antibodies at the same time30.
Human saliva was the first place where "natural" polyreactive SIgA was identified31. Mucosal immunity, especially secretory IgA, is thought to be important in the host's initial line of defense against respiratory infections, and findings suggest it is present in COVID-19. Individual and community immune responses, illness severity, clinical risk, and herd immunity may all be indicators of salivary IgA, which is accessible as a mucosal immunity measure19.
Besides that, as a comparison with samples taken from the Nosopharyngeal Swab (NPS), in one of the study showed that the sensitivity and specificity of this saliva sample was quite high, where in the study evaluated 1924 people who might be infected with COVID-19, both saliva samples (86% and 99.96%) and NPS samples (86% and 99.96%) were used (92% and 99.93 % respectively)32. This indicates that saliva samples are effective enough to be used as an alternative method for detecting COVID-19.
Some of the reasons for using saliva as a test sample to detect COVID-19 can also be seen from the technical advantages in taking in the field15, such as:
a. An strategy to illness diagnosis and general health monitoring that is non-invasive.
b. Effortless (no patient discomfort and anxiety for sampling).
c. It is simple to collect and can be used in remote regions.
d. Technology that is relatively inexpensive.
e. Application that can be used to screen huge groups of people at a low cost.
f. Children, anxious/disabled/elderly patients will benefit from this treatment.
g. Multisampling is a possibility.
h. The collection of biological samples such as nasopharyngeal swabs and blood samples is safer for medical professionals.
i. Nasopharyngeal swabs and blood are two biological samples that are more dangerous for medical practitioners to gather.
Sample Collection Method:
There were several ways of sampling in an effort to SARS-CoV-2 detection. Upper respiratory tract samples such as swabs from the nasopharynx, lower respiratory swabs, throat swabs, nasal swabs, and oropharyngeal swabs tract procedures such as tracheal aspiration and brochoalveolar lavage can be employed with variable degrees of sensitivity to SARS-CoV-2 detection. Bronchoalveolar lavage and tracheal aspiration, because of the technical difficulty of acquiring these samples, they are less preferable specimens than nasopharyngeal/ oropharyngeal swabs for SARS-CoV-2 detection33.
Currently, the nasopharyngeal/oropharyngeal swab is the most common approach for collecting viral samples SARS-CoV-2 has been detected., with viral specimens being gathered by scraping the posterior pharyngeal/ tonsillar region and the nasopharyngeal wall with a minitip swab34. Saliva has been offered as a solutionas an excellent COVID-19 diagnostic candidate, because it has specificity and sensitivity similar to swabs from the nasopharynx. The currently available methods for collecting saliva are draining, spitting, suction and swab methods34.
· Draining Method: Subjects were sitting calmly with their heads down and mouths open, allowing saliva to drip passively into a graded sterile tube from the lower lip.
· Spitting Method: The participant is asked to spit out any saliva that has accumulated on the floor of their mouth into reweighed or graduated test tubes.
· Suction Method: Saliva is allowed to accumulate on the floor of the mouth and aspirated continuously using a micropipette, syringe, saliva ejector or aspirator.
· Swabbing Method: This is conducted by inserting a synthetic gauze sponge, pre-weighed cotton swab or cotton swab into the mouth, in the orifice of the main salivary glands. Subjects were asked to chew so that the sponge was drenched in saliva.
Saliva collection can be performed in several ways above and collection with the help of tools such as sponges and straight from the ducts of the salivary glands35. The cheapest approach is to spit out, and the sample of saliva taken comprises nasopharyngeal, oropharyngeal, and airway secretions. Although this method of collecting saliva straight from the ducts of the primary salivary glands yields pure saliva, it is time consuming and requires specialized equipment15.
Figure 4: Schematic illustration showing the major salivary glands (parotid, submandibular and sublingual) and their respective ducts, oropharynx and nasopharynx, and approximate anatomic locations for oropharyngeal and nasopharyngeal swab collection6.
Saliva rRT-PCR test:
A real-time Reverse Transcription Polymerase Chain Reaction (rT-PCR) using samples of nasopharyngeal or oropharyngeal swabs, sputum, or bronchial lavage is currently the standard test for the identification of SARS-CoV-2. The use of reverse transcription polymerase chain reaction (rRT-PCR) requires a standard protocol, including Ribo Nucleic Acid (RNA) must be extracted and the presence of viral RNA confirmed by rRT-PCR. There are several target genes used to detect SARS-CoV-2, namely the E gene (Envelope), the N gene (nucleocapsid), the S gene (Spike), and the RdRp gene36.
A patient is said to have confirmed COVID-19 if detection by rRT-PCR found a unique sequence of viral RNA. A positive rRT-PCR result indicates that it is possible for someone to be infected with COVID-19, while a negative result cannot exclude someone from being infected with COVID-19, there is still the possibility of false negatives which can be caused by a low viral load, inaccurate sampling technique and timing, or possible mutation of the viral genome36.
Sampling using a nasopharyngeal swab requires special expertise from medical personnel and may pose a risk of aerosol exposure. Based on these reasons, several studies have stated that there are other body fluids that can be an alternative sample to detect SARS-CoV-2, one of them is saliva2,36. Based on the research of Hung, et al., 2020, compared with taking samples with nasopharyngeal and oropharyngeal swabs, SARS-CoV-2 can still be detected in saliva samples up to 20 days or more after being positive for SARS-CoV-2 infection. This shows that saliva is quite good as a specimen for screening patients infected with SARS-CoV-236.
Test RT-LAMP Saliva:
SARS-CoV-2 can be detected quickly and accurately using reverse transcription loop-mediated isothermal amplification (RT-LAMP). The RT-LAMP isothermal amplification approach is appealing from a technological standpoint since it allows for quick Reverse transcription of RNA and amplification of DNA without the need for a temperature cycle. The LAMP technology does not even necessitate the purchase of expensive equipment or the establishment of a laboratory infrastructure37.
The RT-LAMP sensitivity for SARS-CoV-2 in specimens from the upper and lower respiratory tracts, as well as saliva, has been shown to be on par with that of RT-PCR; it demonstrated a 95% match with RT-PCR 38. However, one study discovered that the RT-LAMP test was ineffective with saliva samples had a lower sensitivity than the RT-PCR test for detecting SARS-CoV-2 (70.9% RT-LAMP vs. 81.6% RT-PCR) 37. When compared to the RT-PCR test with nasopharyngeal swab samples, RT-LAMP showed a sensitivity of 72.7 percent and a specificity of 95.7 percent with saliva samples39. The benefit of employing RT-LAMP forCOVID-19 diagnosis is that the outcomes can be obtained in 30-60 minutes, even at the point of care (POC). More research is needed to validate the RT-LAMP test using a variety of respiratory and salivary specimens37–39.
Saliva Antigen Test:
The COVID-19 diagnostic test using the real-time polymerase chain reaction (RT-PCR) is a technique that to be the "gold standard" but this examination has drawbacks, namely the results take a long time, not all laboratories have sophisticated equipment (such as viral nucleic acid extraction tools) and not all laboratory medical personnel have the ability and expertise to do so. molecular skills to process samples. Antigen Based Rapid Diagnostic Test (Ag-RDT) technique uses nasal and nasopharyngeal swab methods to obtain oropharyngeal saliva samples to examine viral antigens (proteins on the surface of the virus). Nasal and nasopharyngeal swabs are considered to cause discomfort in patients when sampling is carried out, therefore further research is currently being carried out on the examination of SARS-CoV-2 with sampling methods through oral saliva, oral fluids, and other sampling systems to be able to expand usage options and to facilitate safe and efficient tests6,40.
Figure 5: Several kinds of saliva samples.4
Besides that, the SARS-CoV-2 examination using a saliva sample has several advantages, namely sampling can be done independently without the need for medical assistance, avoiding injury to the nasal and nasopharyngeal areas due to being exposed to swabs, and minimizing the transmission of the virus to medical personnel40.
Figure 6: Types of SARS-CoV-2 examination using saliva samples4
The difference between the Antigen Rapid Test and the Antibody Rapid Test is that the Antigen Rapid Test cannot see antibodies against SARS-CoV-2, both IgG and IgM, because the Antigen Rapid Test can only see the presence of the virus during the examination. In the Rapid Antigen Test using saliva samples, the results can be shown within 10 minutes (detecting Spike protein in saliva samples). Saliva is obtained from the patient, placed on a sample pad, then by means of capillary action, the saliva will be absorbed on the nitrocellulose membrane for about 5-10 minutes. If the capillary is fully operational, the results will appear, if there are 2 lines (control line and test line) then the results show positive for SARS-CoV-2. However, if only 1 line appears (control line only) then the results show negative SARS-CoV-2. This examination can be used for mass screening, although the sensitivity is not as good as with the RT-PCR examination, thus it cannot be used as an examination to establish a diagnosis6.
Figure 7: Interpretation of the Rapid Test Antigen results using saliva samples.4
SALIVA ANTIBODY TEST:
Antibody tests are not appropriate for detecting acute infection, but serological assays allow:
· Qualitative and quantitative research on the immunological SARS-CoV-2 reaction.
· Determining the accuracy of the rate of infection in an affected area as an essential variable in determining the fatality rate of infection.
· Identifying individuals with a strong antibody response to convalescent serum/plasma therapy donors.
· Assessing the success of vaccination policy implementation and provide information41.
Blood serum is the most common material used to detect antibodies, although saliva has been used as a substitute since it is easier to collect and can be utilized in a point-of-care (POC) scenario. Saliva can be used in antibody test kits as a sample using Lateral Flow Assay (LFA) technology on the terrain, or in a centralized laboratory using Enzyme Linked Immunosorbent Assay (ELISA)and/or Chemiluminescent Assay (CLIA) technology6,17,42.
In a study by Faustini et al that calculated the response of non-nucleocapsid anti-spike antibodies to IgG, IgA and IgM using the ELISA technique, among a sample of non-hospitalized patients who are both symptomatic and asymptomatic, it antibodies of all three types were discovered to be active could be detected in saliva specimens43.
In contrast, Pisanic et al.44 found that the specific IgG response to the SARS-CoV-2 antigen in saliva and serum samples, from 28 participants taken at the same time of visit, was significantly correlated. In saliva, the kinetic profile of IgG is similar to that found in serum. McMullan et al. reported the same result42 which adapted a commercial ELISA using saliva samples. It was found 84.2% sensitivity and 100% specificity were found in 149 saliva samples. Hettegger et al reported that in some cases, plasma and salivary IgI was nearly the same in the case of a significant number of antigens45.
A longitudinal investigation of COVID-19 patients searching for antibody duration in various collections found that IgG levels in blood and saliva were steady until the 105th day after symptom onset (PSO)46. IgA antibodies have been found inCOVID-19 patients' sera, and they seem to be detected before IgM orIgG, at least two days after symptom start. One of the advantages of saliva samples over blood serum is antibodies against IgA are present. Saliva is a good specimen to use for detecting IgA antibodies since its concentrations are higher in mucosal secretions than in blood47. Saliva has been effectively utilized to identify IgA in various cases of different viral diseases, SARS, MERS, HIV, RSV, and seasonal influenza are just a few examples. Varadhachary et al.17 devised an examination technique that had a 92 percent positive predictive value (PPV) and a 97 percent negative predictive value (NPV) for calculating salivary sIgA using the Brevitest IgA Salivary Mucosal Test [BRAVO].
Previous results suggest that saliva may be a viable option for antibody testing, especially for the detection of IgA. Blood samples (serum/plasma) are required for diagnostic purposes are still an option. Moreover, the results of antibody tests using saliva samples still need to be confirmed by antibody tests using blood samples. To identify the level, more research is required and antibodies of various kinds have different levels of persistence in saliva from time to time. Salivary antibody test validation is highly recommended.
CONCLUSION:
The use of saliva as a substitute sample has been a focus of research from the beginning of the COVID-19 epidemic. The gold standard diagnostic testing for SARS-CoV-2 is still a nasopharyngeal swab, saliva sample can help identify folks who are contagious in the community more quickly. Saliva is simple to collect, non-invasive, and both health experts and laypeople can simply alter the collection process. Hence, the required resources can be reduced (laboratory facilities, human resources, PPE and others). Independent sample collection can be conducted, thereby lowering the chance of being exposed to health workers.
Overall, research findings reveal that the detection sensitivity saliva contains less viral RNA than a nasopharyngeal or oropharyngeal swab taken at the same time as sample of saliva from the same patient, while other studies have demonstrated that saliva samples have slightly higher sensitivity. During times of high viral load, however, comparable sensitivity and adequate to reliably detect individual infections. The research suggests that saliva is beneficial can be utilized as an alternate RT-PCR sample material in symptomatic patients when a nasopharyngeal swab is unavailable, as well as for repeated monitoring of asymptomatic patients.Clinical confirmation of the sensitivity and utility of saliva as a sample source for rapid antigen detection is limited. A nasopharyngeal swab sample is recommended by the majority of fast antigen tests. RT-PCR has a higher sensitivity than rapid antigen assays. More research is needed to see if saliva can be used to identify SARS-CoV-2 RNA, antigens, and antibodies.
CONFLICT OF INTEREST:
The authors have no conflicts of interest regarding this investigation.
ACKNOWLEDGMENTS:
This work was supported by Airlangga Research Fund Universitas Airlangga Surabaya Indonesia in the Schema Penelitian Dasar Unggulan (PDU) Universitas Airlangga in 2023 with Grant Number: 77/UN3.1.2/PT/2023.
REFERENCES:
1. Levani Y, Prastya AD, Mawaddatunnadila S. Coronavirus Disease 2019 (COVID-19): Patogenesis, Manifestasi Klinis dan Pilihan Terapi. Jurnal Kedokteran dan Kesehatan. 2021; 17: 44–57. doi.org/10.24853/jkk.17.1.44-57
2. Wu Y-C, Chen C-S, Chan Y-J. The outbreak of COVID-19: An overview. Journal of the Chinese Medical Association 2020; 83: 217–220. doi.org/10.1097/JCMA.0000000000000270
3. Pinki, Rani D, Bajaj H, Singh R. SARS-COV-2 (COVID-19) and role of real time Reverse Transcription Polymerase Chain Reaction (RT-PCR) in its diagnosis. Research Journal of Pharmacy and Technology. 2021; 14: 3437–3440. doi.org/10.52711/0974-360X.2021.00598
4. Sucahya PK. Barriers to Covid-19 RT-PCR Testing in Indonesia: A Health Policy Perspective. Journal of Indonesian Health Policy and Administration. 2020; 5. doi.org/10.7454/ihpa.v5i2.3888
5. Salvamani S, Tan HZ, Thang WJ, et al. Understanding the dynamics of COVID-19; implications for therapeutic intervention, vaccine development and movement control. Br J Biomed Sci 2020; 77: 168–184.doi.org/10.1080/09674845.2020.1826136
6. Azzi L, Maurino V, Baj A, et al. Diagnostic Salivary Tests for SARS-CoV-2. J Dent Res. 2021; 100: 115–123. doi.org/10.1177/0022034520969670
7. Differential Analysis and Putative Roles of Genes, Cytokines and Apoptotic Proteins in Blood Samples of Patients with Respiratory Viral Infections: A Single Center Study. Journal of Pure and Applied Microbiology. 2021
8. Kumar V, Kancharla S, Jena MK. In silico screening of FDA approved drugs predicts the therapeutic potentials of Antibiotic drugs against the papain like protease of SARS-CoV-2. Research Journal of Pharmacy and Technology. 2021; 14: 4035–4039. doi.org/10.52711/0974-360X.2021.00699
9. Padmaharish V. Salivary Biomarkers of Oral Cancer – A Review. Research Journal of Pharmacy and Technology. 2016; 9: 1007–1012. doi.org/10.5958/0974-360X.2016.00190.6
10. Park H-R. Effect of Salivary Streptococci mutans and Lactobacilli levels after uptake of the Probiotic for Clinical Trial. Research Journal of Pharmacy and Technology. 2017; 10: 2984–2988. doi.org/10.5958/0974-360X.2017.00528.5
11. Harsha NS, Rivas-Santisteban J, Satish RT, Kumar GS. Analysis of the Evolutionary pattern of SARS-CoV-2 and its implications in the spread of the disease. Research Journal of Pharmacy and Technology. 2021; 14: 2229–2232. doi.org/10.52711/0974-360X.2021.00396
12. Ansori ANM, Kharisma VD, Fadholly A, Tacharina MR, Antonius Y, Parikesit AA. Severe Acute Respiratory Syndrome Coronavirus-2 Emergence and Its Treatment with Alternative Medicines: A Review. Research Journal of Pharmacy and Technology. 2021; 14: 5551–5557. doi.org/10.52711/0974-360X.2021.00967
13. Kandula UR, Veerabhadrappa KV, Goruntla N, et al. Knowledge, Concept on severe Acute Respiratory Syndrome Coronavirus-2(SARS-CoV-2). A Review of the Literature and Future perspective. Research Journal of Pharmacy and Technology. 2023; 16: 441–446. doi.org/10.52711/0974-360X.2023.00075
14. Yuliana Y. Corona virus diseases (Covid-19): Sebuah tinjauan literatur. Wellness And Healthy Magazine. 2020; 2: 187–192. doi.org/10.30604/well.95212020
15. Sapkota D, Søland TM, Galtung HK, et al. COVID-19 salivary signature: diagnostic and research opportunities. J Clin Pathol 2020:jclinpath-2020-206834.doi.org/10.1136/jclinpath-2020-206834
16. Anam K, Prabowo B, Kusuma MT, et al. Multi Epitopes Potential on Surface SARS-CoV-2 Protein as a Covid-19 Vaccine Candidate. Research Journal of Pharmacy and Technology. 2022; 15: 1437–1442. doi.org/10.52711/0974-360X.2022.00238
17. Handayani D, Hadi DR, Isbaniah F, Burhan E, Agustin H. Corona Virus Disease 2019. Jurnal Respirologi Indonesia. 2020; 40: 119–129. doi.org/10.36497/jri.v40i2.101
18. E N, Savitha G. A study of Salivary Lactate Dehydragenase (LDH) level in normal individuals and the Oral Cancer Patients. Research Journal of Pharmacy and Technology. 2015; 8:.932–934. doi.org/10.5958/0974-360X.2015.00155.9
19. Varadhachary A, Chatterjee D, Garza J, et al. Salivary anti-SARS-CoV-2 IgA as an accessible biomarker of mucosal immunity against COVID-19. medRxiv 2020: 2020.08.07.20170258.doi.org/10.1101/2020.08.07.20170258
20. Agarwal R, T L. Salivary Enzymes as Biomarkers for Periodontitis – An Update. Research Journal of Pharmacy and Technology. 2014; 7: 98–100.
21. Burhan E, Susanto AD, Nasution SA, et al. Perhimpunan Dokter Paru Indonesia (PDPI) Perhimpunan Dokter Spesialis Kardiovaskular Indonesia (PERKI) Perhimpunan Dokter Spesialis Penyakit Dalam Indonesia (PAPDI) Perhimpunan Dokter Anestesiologi dan Terapi Intensif Indonesia (PERDATIN) Ikatan Dokter Anak Indonesia (IDAI). 149.
22. Jain MR, Gheena .S, P G. Comparison of Aerobic Bacteria in Saliva Samples of Chronic and Aggressive Periodontitis Patients- An in Vitro Study. Research Journal of Pharmacy and Technology 2016; 9: 1183–1186.doi.org/10.5958/0974-360X.2016.00226.2
23. World Health Organization (WHO). Diagnostic Testing for SARS Cov-2: Interim Guidance. 2020
24. Brijesh K, Gadhvi V, Gupta A, Roopchandani K, Patel N. A Review: Production of Monoclonal Antibody. Research Journal of Pharmacy and Technology. 2013; 6: 701–705.
25. PDS P. Panduan Tatalaksana Pemeriksaan Antigen. 2020: 1–101.
26. World Health Organization (WHO). Antigen-detection in the diagnosis of SARS-CoV-2 infection. 2021
27. Fábián TK, Hermann P, Beck A, Fejérdy P, Fábián G. Salivary Defense Proteins: Their Network and Role in Innate and Acquired Oral Immunity. Int J Mol Sci. 2012; 13: 4295–4320. doi.org/10.3390/ijms13044295
28. Mestecky J, Russell M, Elson C. Intestinal IgA: Novel views on its function in the defence of the largest mucosal surface. Gut 1999; 44: 2–5. doi.org/10.1136/gut.44.1.2
29. Wilton M. A comparative study of circulating and cell-mediated immunity induced by the gingival and systemic administration of oral bacteria. J dent Res Suppl. 1969; 48
30. Quiding-Järbrink M, Nordström I, Granström G, et al. Differential expression of tissue-specific adhesion molecules on human circulating antibody-forming cells after systemic, enteric, and nasal immunizations. A molecular basis for the compartmentalization of effector B cell responses. J Clin Invest 1997; 99: 1281–1286.doi.org/10.1172/JCI119286
31. Quan CP, Berneman A, Pires R, Avrameas S, Bouvet JP. Natural polyreactive secretory immunoglobulin A autoantibodies as a possible barrier to infection in humans. Infect Immun. 1997; 65: 3997–4004.
32. Yokota I, Shane PY, Okada K, et al. Mass Screening of Asymptomatic Persons for Severe Acute Respiratory Syndrome Coronavirus 2 Using Saliva. Clin Infect Dis. 2021; 73: e559–e565. doi.org/10.1093/cid/ciaa1388
33. Liu R, Han H, Liu F, et al. Positive rate of RT-PCR detection of SARS-CoV-2 infection in 4880 cases from one hospital in Wuhan, China, from Jan to Feb 2020. Clin Chim Acta. 2020; 505: 172–175. doi.org/10.1016/j.cca.2020.03.009
34. Pang J, Wang MX, Ang IYH, et al. Potential Rapid Diagnostics, Vaccine and Therapeutics for 2019 Novel Coronavirus (2019-nCoV): A Systematic Review. Journal of Clinical Medicine. 2020; 9: 623. doi.org/10.3390/jcm9030623
35. Malamud D, Rodriguez-Chavez IR. Saliva as a Diagnostic Fluid. Dent Clin North Am. 2011; 55: 159–178. doi.org/10.1016/j.cden.2010.08.004
36. Hung K-F, Sun Y-C, Chen B-H, et al. New COVID-19 saliva-based test: How good is it compared with the current nasopharyngeal or throat swab test? J Chin Med Assoc. 2020; 83: 891–894. doi.org/10.1097/JCMA.0000000000000396
37. Nagura-Ikeda M, Imai K, Tabata S, et al. Clinical Evaluation of Self-Collected Saliva by Quantitative Reverse Transcription-PCR (RT-qPCR), Direct RT-qPCR, Reverse Transcription-Loop-Mediated Isothermal Amplification, and a Rapid Antigen Test To Diagnose COVID-19. J Clin Microbiol. 2020; 58: e01438-20. doi.org/10.1128/JCM.01438-20
38. Bikos DA, Hwang C, Brileya KA, et al. SLAMP: A Rapid Fluorometric RT-LAMP Assay for Sensitive and Specific Detection of SARS-CoV-2 from Human Saliva. 2021. doi.org/10.1101/2021.03.31.21254634
39. L’Helgouach N, Champigneux P, Schneider FS, et al. EasyCOV : LAMP Based Rapid Detection of SARS-CoV-2 in Saliva. 2020: 2020.05.30.20117291.doi.org/10.1101/2020.05.30.20117291
40. Basso D, Aita A, Padoan A, et al. Salivary SARS-CoV-2 antigen rapid detection: A prospective cohort study. Clin Chim Acta. 2021; 517: 54–59. doi.org/10.1016/j.cca.2021.02.014
41. Kapoor P, Chowdhry A, Kharbanda OP, Bablani Popli D, Gautam K, Saini V. Exploring salivary diagnostics in COVID-19: a scoping review and research suggestions. BDJ Open. 2021; 7:1–10. doi.org/10.1038/s41405-021-00064-7
42. MacMullan MA, Ibrayeva A, Trettner K, et al. ELISA detection of SARS-CoV-2 antibodies in saliva. Sci Rep. 2020; 10: 20818. doi.org/10.1038/s41598-020-77555-4
43. Faustini SE, Jossi SE, Perez-Toledo M, et al. Detection of antibodies to the SARS-CoV-2 spike glycoprotein in both serum and saliva enhances detection of infection. medRxiv. 2020:2020.06.16.20133025.doi.org/10.1101/2020.06.16.20133025
44. Pisanic N, Randad PR, Kruczynski K, et al. COVID-19 Serology at Population Scale: SARS-CoV-2-Specific Antibody Responses in Saliva. J Clin Microbiol. 2020; 59: e02204-20. doi.org/10.1128/JCM.02204-20
45. Hettegger P, Huber J, Paßecker K, et al. High similarity of IgG antibody profiles in blood and saliva opens opportunities for saliva based serology. PLoS One. 2019; 14: e0218456. doi.org/10.1371/journal.pone.0218456
46. Isho B, Abe KT, Zuo M, et al. Persistence of serum and saliva antibody responses to SARS-CoV-2 spike antigens in COVID-19 patients. Sci Immunol. 2020; 5: eabe5511. doi.org/10.1126/sciimmunol.abe5511
47. Yu H, Sun B, Fang Z, et al. Distinct features of SARS-CoV-2-specific IgA response in COVID-19 patients. Eur Respir J. 2020; 56: 2001526. doi.org/10.1183/13993003.01526-2020
Received on 10.02.2023 Modified on 16.06.2023
Accepted on 07.09.2023 © RJPT All right reserved
Research J. Pharm. and Tech 2024; 17(1):443-451.
DOI: 10.52711/0974-360X.2024.00070