Detection Of Viral Nucleic Acids Research Articles

Better design leads to better results - Importance of virological outcome design in clinical trials for antiviral treatment of coronavirus disease 2019.

Better design leads to better results - Importance of virological outcome design in clinical trials for antiviral treatment of coronavirus disease 2019.

Fast and direct identification of SARS‐CoV‐2 variants via 2D InSe field‐effect transistors

AbstractAs the COVID‐19 pandemic evolves and new variants emerge, the development of more efficient identification approaches of variants is urgent to prevent continuous outbreaks of SARS‐CoV‐2. Field‐effect transistors (FETs) with two‐dimensional (2D) materials are viable platforms for the detection of virus nucleic acids (NAs) but cannot yet provide accurate information on NA variations. Herein, 2D Indium selenide (InSe) FETs were used to identify SARS‐CoV‐2 variants. The device's mobility and stability were ensured by atomic layer deposition (ALD) of Al2O3. The resulting FETs exhibited sub‐fM detection limits ranging from 10–14 M to 10–8 M. The recognition of single‐nucleotide variations was achieved within 15 min to enable the fast and direct identification of two core mutations (L452R, R203M) in Delta genomes (p < 0.01). Such capability originated from the trap states in oxidized InSe (InSe1−xOx) after ALD, resulting in traps‐involved carrier transport responsive to the negative charges of NAs. In sum, the proposed approach might highly provide epidemiological information for timely surveillance of the COVID pandemic.image

A label-free electrochemical DNA biosensor used a printed circuit board gold electrode (PCBGE) to detect SARS-CoV-2 without amplification.

The emergence of coronavirus disease 2019 (COVID-19) motivates continuous efforts to develop robust and accurate diagnostic tests to detect severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). Detection of viral nucleic acids provides the highest sensitivity and selectivity for diagnosing early and asymptomatic infection because the human immune system may not be active at this stage. Therefore, this work aims to develop a label-free electrochemical DNA biosensor for SARS-CoV-2 detection using a printed circuit board-based gold substrate (PCBGE). The developed sensor used the nucleocapsid phosphoprotein (N) gene as a biomarker. The DNA sensor-based PCBGE was fabricated by self-assembling a thiolated single-stranded DNA (ssDNA) probe onto an Au surface, which performed as the working electrode (WE). The Au surface was then treated with 6-mercapto-1-hexanol (MCH) before detecting the target N gene to produce a well-oriented arrangement of the immobilized ssDNA chains. The successful fabrication of the biosensor was characterized using cyclic voltammetry (CV), electrochemical impedance spectroscopy (EIS), and atomic force microscopy (AFM). The DNA biosensor performances were evaluated using a synthetic SARS-CoV-2 genome and 20 clinical RNA samples from healthy and infected individuals through EIS. The developed DNA biosensor can detect as low as 1 copy per μL of the N gene within 5 minutes with a LOD of 0.50 μM. Interestingly, the proposed DNA sensor could distinguish the expression of SARS-CoV-2 RNA in a patient diagnosed with COVID-19 without any amplification technique. We believe that the proposed DNA sensor platform is a promising point-of-care (POC) device for COVID-19 viral infection since it offers a rapid detection time with a simple design and workflow detection system, as well as an affordable diagnostic assay.

Effectiveness of Oral Fluid in Pathogenic Surveillance of Acute Respiratory Infection

Objective: Oral fluid (OF) is a new safe, non-invasive, convenient, and efficient biological sample that can be used for virus nucleic acid and antibody detection. Because few studies have performed surveillance of multiple respiratory pathogens, this study sought to explore the application value of OF in this field. Methods: OF and throat swabs were collected from December 2020 to December 2021 in patients with acute respiratory tract infections in Beijing. Multiplex real-time PCR was performed, and the detection performance of two samples was compared. Results: A total of 769 OF and throat swab samples were collected. The detection rates of respiratory pathogens in throat swabs and OF were 29.26% (225/769) and 20.81% (160/769), respectively. The sensitivity and specificity of the OF assay, compared with the throat swab assay, were 71.11% (160/225) and 100% (544/544), respectively. The two assays had excellent agreement (kappa = 0.78). The detection consistency varied among pathogens. For OF samples, the most common pathogen was the influenza B virus, and the highest detection rate was in the ≤5-year-old group. The highest positivity rate was observed in December 2021. Conclusion: OF samples have excellent potential for the epidemiological surveillance of respiratory pathogens, and may have application prospects in preventing and controlling infectious diseases.

The Babylon River's common carp (Cyprinus carpio) gills were used for the histopathological examination and PCR detection of Koi herpesvirus disease (KHVD)

The first outbreak of koi herpesvirus in Iraq occurred in Babylon Province in the floating cages in the Euphrates River from the Musayyib thermal electric power station to the Al-Hindiya Dam. Fish were suffering from gills rot that did not respond to treatment and lesions and ulcers on the fish's body. The fatalities reached 80%. was water temperature 24°C, pH 6.85, Salinity 760 ppt. So the present study aims to highlight the pathological changes in gills after the Koi herpes virus infection in Babylon Province. The study started after the detection of (KHV) by polymerase chain reaction ( PCR) from the suspected samples (30 ) for the detection of virus nucleic acid. Also, study the water characterization during the periods of outbreaks. The gills specimen was collected from the positive cases for gross and histopathological examination. The result showed that ten samples were positive for (KHV) infection. The gross and histopathological examination results showed severe congestion with necrotic foci and sloughing of secondary lamella with increased mucus secretion. In addition, necrosis in the primary lamellae, Edam and inflammatory cells infiltration with hyperplasia of the secondary lamellae with the presence of intranuclear inclusion body in all exanimated slides. Keywords: Koi Herpes; Cyprinus Carpio; Euphrates River; KHV; secondary lamellae; floating cages

95. Prolonged respiratory viral infection associated with presence of coinfections in an urban birth cohort

Abstract Background Prolonged infection by respiratory viruses has been reported, especially in hospitalized or immunocompromised children. However, little is known of factors contributing to prolonged respiratory viral infection, particularly in asymptomatic and less severe infections. We examined characteristics associated with prolonged viral infection in a community-based birth cohort. Methods The PREVAIL cohort is a CDC-sponsored two-year birth cohort in Cincinnati, Ohio conducted during 4/2017 to 8/2020. Mid-turbinate nasal swabs were collected weekly from children and tested using the Luminex Respiratory Pathogen Panel. The primary outcome was prolonged viral infection, which was defined as a viral nucleic acid detection lasting 4 or more weeks. Proportions of prolonged viral infections were compared using Fisher’s exact test with Holms corrections. Adjusted odds ratios (aOR) and 95% confidence intervals were calculated using a mixed effects logistic regression model while controlling for within-subject clustering, viral species, child age, child sex, symptom status, and coinfection. This analysis was limited to subjects who provided at least 70% of weekly samples. Results Among 101 children, providing 7871 child-weeks of follow-up, we identified 780 viral infections. The median duration of infection across all respiratory viruses was 1 week, except for bocavirus and coronavirus NL63, each with 2 weeks; 40% of bocavirus and >10% of adenovirus, coronavirus NL63, RSV A, human metapneumovirus, and parainfluenza 1 infections were associated with prolonged infection (>4 weeks). No prolonged infections were detected for influenza A or B, coronavirus 229E or HKU1, or parainfluenza 2 or 4 infections. Viral coinfection (aOR=3.1, 95% CI 1.9, 5.0) and female sex (aOR 1.8, 95%CI 1.1, 2.9) were significantly associated with prolonged infection, while symptom status and child age were not. Conclusion In the PREVAIL cohort, detection of respiratory viruses lasting 4 weeks or longer was common for certain respiratory pathogens and was especially prolonged for bocavirus. Biological factors such as the presence of additional viral infections or child sex may affect the likelihood of prolonged infection. Disclosures All Authors: No reported disclosures.

A viral metagenomic protocol for nanopore sequencing of group A rotavirus

AimDevelopment of an unbiased methodology using Oxford Nanopore Technology (ONT) sequencing to obtain whole-genome sequences (WGS) of Rotavirus A (RVA) from clinical samples. Methods157 RVA qRT-PCR positive faecal samples were enriched by virus-like particle (VLP) purification and host nuclease digestion to enhance the detection of viral nucleic acids and cDNA generated as per the NetoVIR protocol. ONT sequencing was then performed using the ONT Native Barcoding kit (SQK-LSK-109) on the GridION platform. Data was basecalled, demultiplexed and assembled into near complete RVA genomes. The accuracy and quality of the obtained sequences was assessed by comparing to Sanger sequencing and RVA reference genomes. ResultsThe developed protocol generated 146 near-complete RVA WGS out of the 157 RVA-positive clinical samples. The quality of the assembled genomes was assessed by comparison against publicly-available sequences with results showing 98.76 % ± 0.03 % similarity and > 90 % genome coverage. A concordance assessment was performed comparing the identity of partial RVA VP7 and VP4 segments obtained by Sanger sequencing (n = 51) against corresponding nanopore sequences which demonstrated an overall identity of 100.0 % ± 0.02 %. ConclusionsThe nanopore protocol generated both high quality and accurate RVA WGS extracted from faecal samples. This protocol can be extended to other viral agents in other sample types.

Influencing factors of nucleic acid negative conversion in patients with mild and common COVID-19 induced by the Omicron variant of SARS-COV-2.

To observe the clinical characteristics of mild and common COVID-19 patients infected with the Omicron variant, and to analyze related factors affecting the time to negative conversion of viral nucleic acid detection. Clinical data of 1781 patients with coronavirus disease 2019 (COVID-19) admitted to a cabin hospital in Shanghai from April 12 to May 26, 2022, were retrospectively analyzed, including age, gender, height, weight, clinical symptoms, comorbid diseases, COVID-19 vaccination, treatment, and nucleic acid negative conversion time. Univariate and multivariate logistic regression analyses were used to analyze the influencing factors of nucleic acid negative conversion time. Among the 1781 patients, 995 were male and 786 were female, with a median age of 39 (30, 52) years. There were 727 patients (40.8%) with overweight and obesity [body mass index (BMI) > 24 kg/cm 2) and 413 patients (23.2%) had comorbid diseases. 205 cases (11.5%) were not vaccinated while 1576 cases were vaccinated. There were 1233 cases (69.2%) with one or more symptoms. The main clinical symptoms were cough (60.3%), expectoration (50.4%) and fever (36.9%). 1444 cases (81.0%) were treated with Chinese medicine, 78 cases (4.4%) were treated with western medicine, 14 cases (0.8%) were treated with integrated Chinese and western medicine, and 245 cases (13.8%) did not receive any medical treatment. All patients improved and were discharged. The median nucleic acid negative conversion time was 10.3 (7.4, 12.4) d. Univariate and multivariate analysis showed that, age ≥ 60 years ( OR＝1.537, 95% CI: 1.116 - 2.115, P<0.01), BMI > 24 kg/cm 2 ( OR＝1.344, 95% CI: 1.106 - 1.634, P<0.01 ) and hypertension ( OR＝1.518, 95% CI: 1.094 - 2.106, P<0.05) were independent risk factors for prolonged nucleic acid negative conversion. COVID-19 vaccination ( OR=0.548, 95% CI: 0.398 - 0.755, P<0.01) was a protective factor, that is, vaccination shortened the time for the nucleic acid test to become negative. The symptoms of the Omicron variant infection were relatively mild and occult. Age ≥ 60 years old, comorbid hypertension, no vaccination and BMI > 24 kg/cm 2 are independent influencing factors for prolonged nucleic acid negative conversion.

Visual and label-free ASFV and PCV2 detection by CRISPR-Cas12a combined with G-quadruplex.

African swine fever (ASF) and postweaning multisystemic wasting syndrome (PMWS) are acute infectious diseases caused by the African swine fever virus (ASFV) and porcine circovirus type 2 (PCV2). At present, there are no effective vaccines for the prevention of ASFV. PMWS, which is harmful to the domestic and even the world pig industry, is difficult to cure and has a high mortality. So, developing simple, inexpensive, and accurate analytical methods to detect and effectively diagnose ASFV and PCV2 can be conducive to avoid ASFV and PCV2 infection. CRISPR has become a potentially rapid diagnostic tool due to recent discoveries of the trans-cleavage properties of CRISPR type V effectors. Herein, we report the visual detection based on CRISPR-Cas12a (cpf1), which is more convenient than fluorescence detection. Through in vitro cleavage target DNA activation, Cas12a can trans-cleavage ssDNA G-quadruplex. TMB/H2O2 and Hemin cannot be catalyzed by cleavaged G-DNA to produce green color products. This protocol is useful for the detection of ASFV and PCV2 with high sensitivity. This method can enable the development of visual and label-free ASFV and PCV2 detection and can be carried out in the field without relying on instruments or power. This method can complete nucleic acid detection at 37 °C without using other instruments or energy. Our research has expanded the application of Cas12a and laid the foundation for the field's rapid detection of viral nucleic acid in future.

Prevalence and pathology of equine parvovirus-hepatitis in racehorses from New York racetracks

BackgroundTheiler’s disease, a.k.a. equine serum hepatitis, is a devastating, highly fatal disease of horses. Equine parvovirus-hepatitis (EqPV-H) has been identified as the likely cause of this disease. While the incidence of Theiler’s disease is low, the prevalence of EqPV-H DNA in horses is high, with up to 37% in some regions, suggesting that subclinical or persistent infection is common.MethodsTo determine the prevalence and pathogenicity of EqPV-H infection at New York racetracks, DNA was extracted from archived formalin-fixed, paraffin-embedded liver tissues from racehorses submitted for necropsy to the Animal Health Diagnostic Center as part of the New York State Gaming Commission-Cornell University postmortem examination program. A total of 191 liver samples from horses between 2 and 13 years old were evaluated. Extracted DNA was tested for EqPV-H using PCR and gel electrophoresis. PCR-positive samples were further assessed for tissue morphology using histology and detection of viral nucleic acid using in situ hybridization.ResultsForty-two samples were PCR positive (22%). Of those, 31 samples had positive viral nucleic acid hybridization in hepatocytes with 11 samples showing positive hybridization in necrotic hepatocytes associated with inflammatory cells, indicating active hepatitis. Both individual hepatocyte necrosis and hepatitis were positively associated with EqPV-H detection (p < 0.0001 and p = 0.0005, respectively).ConclusionThese findings indicate that presence of EqPV-H in the liver and parvoviral-associated hepatitis are prevalent in racehorses from New York racetracks, thus warranting additional studies examining potential associations between EqPV-H infection and racehorse performance.

(Digital Presentation) Biomimic Nanotemplating Assay Based on Molecularly Imprinted Polymers for the Impedimetric Detection of Sars-Cov-2 and Influenza A Spike Proteins in Untreated Saliva

With the continuously fluctuating incidence of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), critical gaps in the field of rapid diagnostic testing have been exposed, particularly in the diagnosis of viral respiratory infections. Current gold standard methods rely on real time quantitative polymerase chain reaction (RT-qPCR) for the detection of viral nucleic acids, but these tests are challenged with long turnaround times, costly centralized laboratory equipment and the need for trained personnel to execute the protocols. With the growing number of emerging variants that can evade both immune responses and inoculation, addressing current testing challenges is critical to manage the spread of viral infections. Here, we propose the design of a novel assay based on our previously studied gold nano/micro islands (NMIs) as a core for the fabrication of an ultrathin molecularly imprinted polymer (MIP) for the impedimetric detection of SARS-CoV-2 and Influenza A spike proteins (SPs) in untreated saliva within 10 minutes. The proposed electrofabrication protocol is rapidly adaptable to a diverse repertoire of protein biomarkers; in this work, we demonstrate the impedimetric detection of the SARS-CoV-2 original strain, Alpha B.1.1.7, Delta B.1.617.2 and Omicron B.1.1.529 variant SP, as well as the Influenza A SP within physiologically relevant ranges and at a low limit of detection to enable the diagnosis of acute infections. Validation was performed at two unique test sites with 51 SARS-CoV-2 patient samples to demonstrate an overall 100% sensitivity and 100% specificity of the NMIs/MIPs assay. Robust quantification of the electrochemical assay was confirmed against RT-qPCR, which effectively enabled statistically significant (p < .0005) viral load quantification on a rapid, miniaturized, and ultrasensitive platform. This novel technology presents the development of a quantitative and versatile electrochemical assay with the potential for the rapid detection of current and future viral respiratory infections, which can guide future electrochemical clinical and commercial point-of-care testing platforms.

A Remotely Controlled Oropharyngeal Swab Sampling Robot and Its Application in COVID-19 Prevention

Oropharyngeal swab sampling is the major viral nucleic acid detection method to diagnose COVID-19. Medical staff exposes themselves to the respiratory secretions of patients, which makes them vulnerable to infection. To protect medical staff, we summarize the clinical requirements for robot into five considerations (standardization, ergonomics, safety, isolation, and task allocation) and developed a remotely operated oropharyngeal swab sampling robot. With robot assistance, spatial isolation between medical staff and the patients can be achieved. We designed a hybrid force/position control scheme for the sampling robot to achieve intuitive operation and stable contact force. The experiment results on phantom tissue show that the sampling robot can achieve intuitive operation and stable contact on the soft posterior pharyngeal. Clinical trials for 20 volunteers and 2 patients diagnosed with COVID-19 are carried out. The results of the clinical trial indicated that the sampling robot can collect samples stably and effectively, and the contact force is gentler and more uniform. For two patients diagnosed with COVID-19, the robot sampling results are consistent with manual sampling.

Detection of SARS-CoV-2 Specific Antibodies in Saliva Samples.

Molecular assays on nasopharyngeal swabs act as a confirmatory test in coronavirus disease (COVID-19) diagnosis. However, the technical requirements of nasopharyngeal sampling and molecular assays limit the testing capabilities. Recent studies suggest the use of saliva for the COVID-19 diagnostic test. In this study, 44 patients diagnosed with COVID-19 in The Third People’s Hospital of Shenzhen were enrolled. Saliva and serum specimens were obtained at different time points and the immunoglobulins against SARS-CoV-2 were measured. The results showed that saliva IgA presented a higher COI value than IgG and IgM. In matched saliva and serum samples, all saliva samples presented lower IgG levels than serum samples, and only one saliva sample presented a higher IgM level. The conversion rates of saliva IgA and the detection of viral nucleic acids were analyzed in the first and second weeks after hospitalization. The positive rates increased when combining saliva IgA and viral nucleic acid detection. In conclusion, our results provide evidence that saliva IgA could serve as a useful index for the early diagnosis of COVID-19.

Real-Time Detection of LAMP Products of African Swine Fever Virus Using Fluorescence and Surface Plasmon Resonance Method

African swine fever (ASF) is a swine disease with a very high fatality rate caused by a complex double-stranded DNA virus. The fluorescence PCR detection method is widely used for virus nucleic acid detection. Surface plasmon resonance (SPR) is a label-free and real-time detection method, unlike the fluorescence PCR detection method. In this research, we detected the loop-mediated isothermal amplification (LAMP) products of the African swine fever virus by using the SPR and fluorescence methods separately and simultaneously. By comparing the positive and negative control results, we found that the SPR response unit is completely different before and after the LAMP process. In addition, the fluorescence results on a chip showed that with an increase in the concentration of the sample, the cycle threshold (CT) value decreased, which is consistent with commercial instruments. Both the decline rate of the SPR response unit and the CT value of the fluorescence realized were used to distinguish the positive control from the negative control and water, which indicates that the SPR method can be combined with fluorescence to detect LAMP products. This research provides a label-free and simple method for detecting LAMP products.

A Novel Virus Detection Strategy Enabled by TR512-Peptide-Based Bioorthogonal Capture and Enrichment of Preamplified Nucleic Acid.

High-cost viral nucleic acid detection devices (e.g., qPCR system) are limited resources for developing counties and rural areas, leading to underdiagnosis or even pandemics of viral infectious diseases. Herein, a novel virus detection strategy is reported. Such detection method is enabled by TR512-peptide-based biorthogonal capture and enrichment of commercially available Texas red fluorophore labeled nucleic acid on the functionalized paper. The GST-TR512 fusion protein electrostatically immobilized on the paper is constructed to retain the binding affinity of TR512-peptide toward Texas red fluorophore labeled nucleic acid released in the preamplification process, then the enrichment of analytes enhances fluorescence signal for rapid detection as volume of sample filters through the paper. The method is generally applicable to different nucleic acid preamplification strategies (PCR, RAA, CRISPR) and different virus types (Hepatitis B virus (HBV), African swine fever virus (ASFV), human papillomavirus (HPV), and severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2 or 2019 nCoV)). Finally, a full-set virus detection device is developed in house to detect the presence of Hepatitis B virus (HBV) viral gene in patients' blood samples. Taken together, we first apply TR512-peptide in the signal enrichment and the novel detection strategy may offer an inexpensive, rapid, and portable solution for areas with limited access to a standard diagnosis laboratory.

Chikungunya virus: An emerging public health challenge for Pakistan

Chikungunya virus: An emerging public health challenge for Pakistan

Applying Machine Learning with Localized Surface Plasmon Resonance Sensors to Detect SARS-CoV-2 Particles.

The sudden outbreak of COVID-19 rapidly developed into a global pandemic, which caused tens of millions of infections and millions of deaths. Although SARS-CoV-2 is known to cause COVID-19, effective approaches to detect SARS-CoV-2 using a convenient, rapid, accurate, and low-cost method are lacking. To date, most of the diagnostic methods for patients with early infections are limited to the detection of viral nucleic acids via polymerase chain reaction (PCR), or antigens, using an enzyme-linked immunosorbent assay or a chemiluminescence immunoassay. This study developed a novel method that uses localized surface plasmon resonance (LSPR) sensors, optical imaging, and artificial intelligence methods to directly detect the SARS-CoV-2 virus particles without any sample preparation. The virus concentration can be qualitatively and quantitatively detected in the range of 125.28 to 106 vp/mL through a few steps within 12 min with a limit of detection (LOD) of 100 vp/mL. The accuracy of the SARS-CoV-2 positive or negative assessment was found to be greater than 97%, and this was demonstrated by establishing a regression machine learning model for the virus concentration prediction (R2 > 0.95).

Epidemiology of Echovirus 30 Infections Detected in a University Hospital in Catalonia, Spain, in 1995-2020.

There is a growing interest in echovirus 30 (E30), an enterovirus responsible for neurological disease and hospitalization. There are multiple studies of outbreaks, but few that study the epidemiology over long periods of time. Our study aims to describe the clinical, epidemiological and microbiological characteristics of a series of E30 infections detected over 26 years. Data were retrospectively collected from a database of all enterovirus infections identified in our laboratory. They were detected by viral isolation or nucleic acid detection in patients presenting with respiratory or neurological infections, rash, sepsis-like syndrome, or gastroenteritis. Enterovirus genotyping was performed by amplification of the VP1 gene using RT-nested PCR, followed by sequencing and BLAST analysis. Of the 2402 enterovirus infections detected, 1619 were linked to at least one genotype and 173 were caused by E30. Clinical information was available for 158 (91.3%) patients. E30 was associated with neurological infection in 107 (67.8%) cases and it was detected almost every year. Phylogenetic analysis was performed with 67 sequences. We observed that E30 strains circulating in Catalonia from 1996 to 2016 belong to two lineages (E and F), although the majority cluster was in F. In 2018, lineage I emerged as the dominant lineage.

A systematic review and meta-analysis of the accuracy of SARS-COV-2 IGM and IGG tests in individuals with COVID-19.

Introduction: Active SARS-CoV-2 infection is confirmed mainly through the detection of viral nucleic acid via the reverse transcriptase polymerase chain reaction (RT-PCR) technique. Methods to assess humoral responses contribute to the monitoring of the disease and confirmation of exposure to the virus.Objective: To evaluate the accuracy of tests for IgM and IgG antibodies for SARS-CoV-2 infection confirmed by RT-PCR and utility as complementary data for immunosurveillance.Methods: Literature research was performed by searching the terms “COVID-19”, “COVID-19 diagnostic testing” and “test” in the databases MEDLINE, EMBASE, Cochrane Library, Web of Science and Cumulative Index to Nursing and Allied Health Literature to search for potentially eligible observational studies without language restrictions published up to September 2020.Results: The pooled sensitivity and specificity, regardless of collection moment, was 80.0% (CI 95% 72.0–86.0) and 97.0% (CI 95% 94.0–98.0) for “IgM and/or IgG”, respectively. Serology considering immunoglobulins M and G together had a high accuracy performance on “fifteenth day and after”: sensitivity and specificity was 91.0% (CI 95% 85.0–94.0) and 98.0% (CI 95% 95.0–99.0) respectively, DOR 461 and AUC 0.98.Conclusion: This study shows that serology is a group of tests with high accuracy, mainly following the second week after infection.

Food for thought: Eating before saliva collection and interference with SARS-CoV-2 detection.

Saliva is a promising specimen for the detection of viruses that cause upper respiratory infections including severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) due to its cost‐effectiveness and noninvasive collection. However, together with intrinsic enzymes and oral microbiota, children's unique dietary habits may introduce substances that interfere with diagnostic testing. To determine whether children's dietary choices impact SARS‐CoV‐2 molecular detection in saliva, we performed a diagnostic study that simulates testing of real‐life specimens provided from healthy children (n = 5) who self‐collected saliva at home before and at 0, 20, and 60 min after eating 20 foods they selected. Each of 72 specimens was split into two volumes and spiked with SARS‐CoV‐2‐negative or SARS‐CoV‐2‐positive clinical standards before side‐by‐side testing by reverse‐transcription polymerase chain reaction matrix‐assisted laser desorption ionization time‐of‐flight (RT‐PCR/MALDI‐TOF) assay. Detection of internal extraction control and SARS‐CoV‐2 nucleic acids was reduced in replicates of saliva collected at 0 min after eating 11 of 20 foods. Interference resolved at 20 and 60 min after eating all foods except hot dogs in one participant. This represented a significant improvement in the detection of nucleic acids compared to saliva collected at 0 min after eating (p = 0.0005). We demonstrate successful detection of viral nucleic acids in saliva self‐collected by children before and after eating a variety of foods. Fasting is not required before saliva collection for SARS‐CoV‐2 testing by RT‐PCR/MALDI‐TOF, but waiting for 20 min after eating is sufficient for accurate testing. These findings should be considered for SARS‐CoV‐2 testing and broader viral diagnostics in saliva specimens.