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1.
PLoS One ; 16(4): e0249069, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-33848293

RESUMEN

BACKGROUND: The novel coronavirus disease (COVID-19) pandemic, caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), continues to remain a global challenge. There is emerging evidence of SARS-CoV-2 virus found in the blood of patients from China and some developed countries. However, there is inadequate data reported in Ghana and other parts of Africa, where blood transfusion service heavily relies on voluntary and replacement blood donors. This study aimed to investigate whether plasma of infected individuals could pose significant transfusion transmitted risk of COVID-19 in Ghanaian populations. METHODS: This cross-sectional retrospective study was conducted at the Kumasi Centre for Collaborative Research into Tropical Medicine (KCCR), KNUST, Ghana. Study subjects comprised contacts of COVID-19 individuals, those with classical symptoms of COVID-19 and individuals who had recovered based on the new Ghana discharge criteria. Whole blood, sputum or deep coughed saliva samples were collected and transported to KCCR for SARS-CoV-2 testing. Viral nucleic acid was extracted from sputum/nasopharyngeal samples using Da An Gene column based kit and from plasma using LBP nucleic acid extraction kit. Real-Time PCR was performed specifically targeting the ORF1ab and Nucleocapsid (N) genomic regions of the virus. RESULTS: A total of 97 individuals were recruited into the study, with more than half being males (58; 59.7%). The mean age of all subjects was 33 years (SD = 7.7) with minimum being 22 years and maximum 56 years. Majority (76; 78.4%) of all the subjects were asymptomatic, and among the few symptomatic subjects, cough (10; 10.3%) was the most predominant symptom. Of the 97 sputum samples tested, 79 (81.4%) were positive for SARS-CoV-2. We identified SARS-CoV-2 viral RNA in the plasma of 1 (1.03%) subject who had clinically recovered. CONCLUSION: This study reports the identification of SARS-CoV-2 viral RNA in a convalescent individual in Ghana. Due to the low prevalence observed and the marginal cycling thresholds associated, the risk of transfusion transmission of SARS-CoV-2 is negligible. Well-powered studies and advanced diagnostics to determine infectious viremia is recommended to further evaluate the potential risk of hematogenous transmission among recovered patients.


Asunto(s)
Transfusión Sanguínea , /patología , Adulto , /virología , Estudios Transversales , Femenino , Humanos , Masculino , Persona de Mediana Edad , ARN Viral/sangre , Reacción en Cadena en Tiempo Real de la Polimerasa , Estudios Retrospectivos , Riesgo , /aislamiento & purificación , Saliva/virología , Esputo/virología , Adulto Joven
2.
Dan Med J ; 68(5)2021 Apr 07.
Artículo en Inglés | MEDLINE | ID: mdl-33832565

RESUMEN

INTRODUCTION: The reference test to evaluate patients with suspected respiratory virus infection is a real-time reverse transcription-polymerase chain reaction (RT-PCR) from a nasopharyngeal swab (NPS). However, other specimen collection methods such as an oropharyngeal swab (OPS) or saliva specimen are also used for SARS-CoV-2 testing during the ongoing COVID-19 pandemic. However, it remains unclear if rates of SARS-CoV-2 detection differ between sampling methods. This study will compare the rates of SARS-CoV-2 detection by saliva, OPS, and NPS sampling in a public setting. METHODS: Individuals referred for outpatient SARS-CoV-2 testing will be invited to participate in a prospective clinical study. They will have saliva, OPS and NPS specimens collected that will be analysed separately for SARS-CoV-2 RNA by RT-PCR. The rate of SARS-CoV-2 detection in saliva, OPS and NPS will be compared using a logistic regression mixed-effect model analysis. A sample of 19,110 participants is required at an expected 1.5% test-positive rate in order to detect a 25.6% difference. The total sample size will be adjusted as the test-positive rate changes. CONCLUSIONS: This study will provide evidence for the optimal site of specimen collection to detect SARS-CoV-2. The results may help guide the health authorities. FUNDING: This is an investigator-initiated trial based on an unrestricted grant from the Novo Nordisk Foundation and the Aage og Johanne Louis-Hansens Fond. The foundations have had no say in the decisions on study design or reporting. TRIAL REGISTRATION: ClinicalTrials.gov (ID: NCT04715607).


Asunto(s)
/diagnóstico , Nasofaringe/virología , Orofaringe/virología , Saliva/virología , Humanos , Modelos Logísticos , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Manejo de Especímenes
3.
Sci Rep ; 11(1): 6264, 2021 03 17.
Artículo en Inglés | MEDLINE | ID: mdl-33731722

RESUMEN

Many educational institutions have partially or fully closed all operations to cope with the challenges of the ongoing COVID-19 pandemic. In this paper, we explore strategies that such institutions can adopt to conduct safe reopening and resume operations during the pandemic. The research is motivated by the University of Illinois at Urbana-Champaign's (UIUC's) SHIELD program, which is a set of policies and strategies, including rapid saliva-based COVID-19 screening, for ensuring safety of students, faculty and staff to conduct in-person operations, at least partially. Specifically, we study how rapid bulk testing, contact tracing and preventative measures such as mask wearing, sanitization, and enforcement of social distancing can allow institutions to manage the epidemic spread. This work combines the power of analytical epidemic modeling, data analysis and agent-based simulations to derive policy insights. We develop an analytical model that takes into account the asymptomatic transmission of COVID-19, the effect of isolation via testing (both in bulk and through contact tracing) and the rate of contacts among people within and outside the institution. Next, we use data from the UIUC SHIELD program and 85 other universities to estimate parameters that describe the analytical model. Using the estimated parameters, we finally conduct agent-based simulations with various model parameters to evaluate testing and reopening strategies. The parameter estimates from UIUC and other universities show similar trends. For example, infection rates at various institutions grow rapidly in certain months and this growth correlates positively with infection rates in counties where the universities are located. Infection rates are also shown to be negatively correlated with testing rates at the institutions. Through agent-based simulations, we demonstrate that the key to designing an effective reopening strategy is a combination of rapid bulk testing and effective preventative measures such as mask wearing and social distancing. Multiple other factors help to reduce infection load, such as efficient contact tracing, reduced delay between testing and result revelation, tests with less false negatives and targeted testing of high-risk class among others. This paper contributes to the nascent literature on combating the COVID-19 pandemic and is especially relevant for educational institutions and similarly large organizations. We contribute by providing an analytical model that can be used to estimate key parameters from data, which in turn can be used to simulate the effect of different strategies for reopening. We quantify the relative effect of different strategies such as bulk testing, contact tracing, reduced infectivity and contact rates in the context of educational institutions. Specifically, we show that for the estimated average base infectivity of 0.025 ([Formula: see text]), a daily number of tests to population ratio T/N of 0.2, i.e., once a week testing for all individuals, is a good indicative threshold. However, this test to population ratio is sensitive to external infectivities, internal and external mobilities, delay in getting results after testing, and measures related to mask wearing and sanitization, which affect the base infection rate.


Asunto(s)
/prevención & control , Pandemias/prevención & control , Instituciones Académicas/normas , Universidades/normas , Enfermedades Asintomáticas , Simulación por Computador , Trazado de Contacto/métodos , Humanos , Saliva/virología
4.
Sci Rep ; 11(1): 5448, 2021 03 09.
Artículo en Inglés | MEDLINE | ID: mdl-33750853

RESUMEN

To safely re-open economies and prevent future outbreaks, rapid, frequent, point-of-need, SARS-CoV-2 diagnostic testing is necessary. However, existing field-deployable COVID-19 testing methods require the use of uncomfortable swabs and trained providers in PPE, while saliva-based methods must be transported to high complexity laboratories for testing. Here, we report the development and clinical validation of High-Performance Loop-mediated isothermal Amplification (HP-LAMP), a rapid, saliva-based, SARS-CoV-2 test with a limit of detection of 1.4 copies of virus per µl of saliva and a sensitivity and specificity with clinical samples of > 96%, on par with traditional RT-PCR based methods using swabs, but can deliver results using only a single fluid transfer step and simple heat block. Testing of 120 patient samples in 40 pools comprised of 5 patient samples each with either all negative or a single positive patient sample was 100% accurate. Thus, HP-LAMP may enable rapid and accurate results in the field using saliva, without need of a high-complexity laboratory.


Asunto(s)
/diagnóstico , Saliva/virología , /virología , Humanos , Límite de Detección , Técnicas de Diagnóstico Molecular , Nasofaringe/virología , Técnicas de Amplificación de Ácido Nucleico , ARN Viral/metabolismo , Sensibilidad y Especificidad , Temperatura
5.
Emerg Infect Dis ; 27(4)2021 04.
Artículo en Inglés | MEDLINE | ID: mdl-33755009

RESUMEN

We analyzed feasibility of pooling saliva samples for severe acute respiratory syndrome coronavirus 2 testing and found that sensitivity decreased according to pool size: 5 samples/pool, 7.4% reduction; 10 samples/pool, 11.1%; and 20 samples/pool, 14.8%. When virus prevalence is >2.6%, pools of 5 require fewer tests; when <0.6%, pools of 20 support screening strategies.


Asunto(s)
/métodos , Saliva/virología , Manejo de Especímenes/métodos , /diagnóstico , Creación de Capacidad/métodos , Asignación de Recursos para la Atención de Salud , Humanos , Límite de Detección , Asignación de Recursos/métodos , Sensibilidad y Especificidad , Estados Unidos
6.
Biosens Bioelectron ; 180: 113111, 2021 May 15.
Artículo en Inglés | MEDLINE | ID: mdl-33743492

RESUMEN

Significant barriers to the diagnosis of latent and acute SARS-CoV-2 infection continue to hamper population-based screening efforts required to contain the COVID-19 pandemic in the absence of widely available antiviral therapeutics or vaccines. We report an aptamer-based SARS-CoV-2 salivary antigen assay employing only low-cost reagents ($3.20/test) and an off-the-shelf glucometer. The test was engineered around a glucometer as it is quantitative, easy to use, and the most prevalent piece of diagnostic equipment globally, making the test highly scalable with an infrastructure that is already in place. Furthermore, many glucometers connect to smartphones, providing an opportunity to integrate with contact tracing apps, medical providers, and electronic health records. In clinical testing, the developed assay detected SARS-CoV-2 infection in patient saliva across a range of viral loads - as benchmarked by RT-qPCR - within 1 h, with 100% sensitivity (positive percent agreement) and distinguished infected specimens from off-target antigens in uninfected controls with 100% specificity (negative percent agreement). We propose that this approach provides an inexpensive, rapid, and accurate diagnostic for distributed screening of SARS-CoV-2 infection at scale.


Asunto(s)
Antígenos Virales/análisis , Técnicas Biosensibles/métodos , /diagnóstico , Pruebas en el Punto de Atención , Saliva/virología , Adulto , Femenino , Humanos , Masculino , Fosfoproteínas/análisis , Técnica SELEX de Producción de Aptámeros , Sensibilidad y Especificidad , Glicoproteína de la Espiga del Coronavirus/análisis
7.
Nat Commun ; 12(1): 1931, 2021 03 26.
Artículo en Inglés | MEDLINE | ID: mdl-33771993

RESUMEN

The COVID-19 pandemic continues to have an unprecedented impact on societies and economies worldwide. There remains an ongoing need for high-performance SARS-CoV-2 tests which may be broadly deployed for infection monitoring. Here we report a highly sensitive single molecule array (Simoa) immunoassay in development for detection of SARS-CoV-2 nucleocapsid protein (N-protein) in venous and capillary blood and saliva. In all matrices in the studies conducted to date we observe >98% negative percent agreement and >90% positive percent agreement with molecular testing for days 1-7 in symptomatic, asymptomatic, and pre-symptomatic PCR+ individuals. N-protein load decreases as anti-SARS-CoV-2 spike-IgG increases, and N-protein levels correlate with RT-PCR Ct-values in saliva, and between matched saliva and capillary blood samples. This Simoa SARS-CoV-2 N-protein assay effectively detects SARS-CoV-2 infection via measurement of antigen levels in blood or saliva, using non-invasive, swab-independent collection methods, offering potential for at home and point of care sample collection.


Asunto(s)
/métodos , /sangre , Saliva/virología , /epidemiología , /genética , Epidemias , Servicios de Atención de Salud a Domicilio , Humanos , Sistemas de Atención de Punto , Curva ROC , /fisiología , Manejo de Especímenes/métodos
8.
Emerg Infect Dis ; 27(4): 1146-1150, 2021 04.
Artículo en Inglés | MEDLINE | ID: mdl-33754989

RESUMEN

The expense of saliva collection devices designed to stabilize severe acute respiratory syndrome coronavirus 2 RNA is prohibitive to mass testing. However, virus RNA in nonsupplemented saliva is stable for extended periods and at elevated temperatures. Simple plastic tubes for saliva collection will make large-scale testing and continued surveillance easier.


Asunto(s)
/métodos , ARN Viral , Saliva/virología , /diagnóstico , /virología , Creación de Capacidad/métodos , Humanos , Estabilidad del ARN , ARN Viral/aislamiento & purificación , ARN Viral/fisiología , Reproducibilidad de los Resultados , Asignación de Recursos , /aislamiento & purificación , Manejo de Especímenes/economía , Manejo de Especímenes/instrumentación , Manejo de Especímenes/métodos
10.
Elife ; 102021 03 29.
Artículo en Inglés | MEDLINE | ID: mdl-33779548

RESUMEN

Here, we develop a simple molecular test for SARS-CoV-2 in saliva based on reverse transcription loop-mediated isothermal amplification. The test has two steps: (1) heat saliva with a stabilization solution and (2) detect virus by incubating with a primer/enzyme mix. After incubation, saliva samples containing the SARS-CoV-2 genome turn bright yellow. Because this test is pH dependent, it can react falsely to some naturally acidic saliva samples. We report unique saliva stabilization protocols that rendered 295 healthy saliva samples compatible with the test, producing zero false positives. We also evaluated the test on 278 saliva samples from individuals who were infected with SARS-CoV-2 but had no symptoms at the time of saliva collection, and from 54 matched pairs of saliva and anterior nasal samples from infected individuals. The Saliva TwoStep test described herein identified infections with 94% sensitivity and >99% specificity in individuals with sub-clinical (asymptomatic or pre-symptomatic) infections.


Asunto(s)
/diagnóstico , Portador Sano/diagnóstico , Portador Sano/virología , Saliva/virología , /metabolismo , Humanos , Técnicas de Diagnóstico Molecular/métodos , Técnicas de Amplificación de Ácido Nucleico/métodos , ARN Viral/genética , Sensibilidad y Especificidad , Manejo de Especímenes/métodos
11.
Allergol Immunopathol (Madr) ; 49(1): 159-164, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-33528945

RESUMEN

Coronavirus disease 2019 (COVID-19) is a disease caused by a new strain of coronavirus named as severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). Globally, since the outbreak, more than seven million confirmed cases of COVID-19 have been reported. The rapid spread and increase in the number of new cases is due to person-to-person transmission. To further control its transmission, early laboratory diagnosis of both asymptomatic and symptomatic patients is crucial. Presently, the COVID-19 diagnosis of infected individuals is dependent on computed tomography scanning and real-time polymerase chain reaction (PCR). The latter is considered more sensitive and efficient for early diagnosis. In this review, general comparisons are made (cases, fatality rate, incubation period, clinical features, and reservoirs) and diagnostic laboratory procedures (specimens, extraction methods, and positive rates by real-time PCR) are compared between SARS, Middle East Respiratory Syndrome, and SARS-2. In total, 8982 SARS-2 suspected patients specimen data were retrieved, in which 40.9% (n = 3678) were detected as positive by real-time PCR. The specimen-wise high detection rate was observed from bronchoalveolar lavage, followed by saliva, nasal swabs, and sputum. As the COVID-19 cases are persistently increasing, the selection of appropriate specimens and laboratory assay would help in rapid and timely diagnosis.


Asunto(s)
/diagnóstico , Reacción en Cadena en Tiempo Real de la Polimerasa , /aislamiento & purificación , Lavado Broncoalveolar , /virología , Infecciones por Coronavirus/diagnóstico , Infecciones por Coronavirus/virología , Humanos , Nasofaringe/virología , Saliva/virología , Síndrome Respiratorio Agudo Grave/diagnóstico , Síndrome Respiratorio Agudo Grave/virología , Esputo/virología
12.
Viruses ; 13(2)2021 02 06.
Artículo en Inglés | MEDLINE | ID: mdl-33562073

RESUMEN

The contemporary surge in metagenomic sequencing has transformed knowledge of viral diversity in wildlife. However, evaluating which newly discovered viruses pose sufficient risk of infecting humans to merit detailed laboratory characterization and surveillance remains largely speculative. Machine learning algorithms have been developed to address this imbalance by ranking the relative likelihood of human infection based on viral genome sequences, but are not yet routinely applied to viruses at the time of their discovery. Here, we characterized viral genomes detected through metagenomic sequencing of feces and saliva from common vampire bats (Desmodus rotundus) and used these data as a case study in evaluating zoonotic potential using molecular sequencing data. Of 58 detected viral families, including 17 which infect mammals, the only known zoonosis detected was rabies virus; however, additional genomes were detected from the families Hepeviridae, Coronaviridae, Reoviridae, Astroviridae and Picornaviridae, all of which contain human-infecting species. In phylogenetic analyses, novel vampire bat viruses most frequently grouped with other bat viruses that are not currently known to infect humans. In agreement, machine learning models built from only phylogenetic information ranked all novel viruses similarly, yielding little insight into zoonotic potential. In contrast, genome composition-based machine learning models estimated different levels of zoonotic potential, even for closely related viruses, categorizing one out of four detected hepeviruses and two out of three picornaviruses as having high priority for further research. We highlight the value of evaluating zoonotic potential beyond ad hoc consideration of phylogeny and provide surveillance recommendations for novel viruses in a wildlife host which has frequent contact with humans and domestic animals.


Asunto(s)
Quirópteros/virología , Virus/aislamiento & purificación , Zoonosis/virología , Animales , Reservorios de Enfermedades/veterinaria , Reservorios de Enfermedades/virología , Heces/virología , Genoma Viral/genética , Humanos , Aprendizaje Automático , Metagenómica , Filogenia , Virus de la Rabia/clasificación , Virus de la Rabia/genética , Virus de la Rabia/aislamiento & purificación , Saliva/virología , Virus/clasificación , Virus/genética
13.
Expert Rev Mol Diagn ; 21(1): 31-42, 2021 01.
Artículo en Inglés | MEDLINE | ID: mdl-33523770

RESUMEN

Introduction: The unprecedented outbreaks of corona virus disease of 2019 (COVID-19) have highlighted the necessity of readily available, reliable, precise, and faster techniques for its detection. Nasopharyngeal swab has been the gold standard for the diagnosis of COVID-19. However, it is not an ideal screening procedure for massive screening as it implicates the patient's stay in the hospital or at home until diagnosis, thus causing crowding of the specimen at the diagnostic centers. Present study deal with the exploration of potential application of different body fluids using certain highly objective techniques (Optical and e-Nose) for faster detection of molecular markers thereby diagnosing viral infections.Areas covered: This report presents an evaluation of different body fluids, and their advantages for the rapid detection of COVID-19, coupled with highly sensitive optical techniques for the detection of molecular biomarkers.Expert opinion: Tears, saliva, and breath samples can provide valuable information about viral infections. Our brief review strongly recommends the application of saliva/tears and exhaled breath as clinical samples using technics such as high-performance liquid chromatography-laser-induced fluorescence, photoacoustic spectroscopy, and e-Nose, respectively, for the fast diagnosis of viral infections.


Asunto(s)
/diagnóstico , /aislamiento & purificación , Biomarcadores/metabolismo , Líquidos Corporales/virología , Pruebas Respiratorias , Cromatografía Liquida , Espiración , Humanos , Rayos Láser , Tamizaje Masivo/métodos , Nanotecnología , Técnicas Fotoacústicas , Saliva/virología , Sensibilidad y Especificidad , Lágrimas/virología
14.
J Clin Virol ; 136: 104760, 2021 03.
Artículo en Inglés | MEDLINE | ID: mdl-33610926

RESUMEN

The new coronavirus infection (COVID-19) is a major public health concern, with a high burden and risk for infection among patients and healthcare workers. Saliva droplets containing SARS-COV-2 are a major vector for COVID-19 infection, making saliva a promising alternative for COVID-19 testing using nasopharyngeal swab samples. To diagnose COVID-19 patients in the field, a point-of-care test (POCT) using saliva was conceptualized. We have developed a simple method for extracting RNA from saliva samples using semi-alkaline proteinase, a sputum homogenizer typically used for preparing samples for tuberculosis testing, and a subsequent simple heating step with no need for centrifugation or RNA extraction. Further, we newly developed a triplex reverse transcription loop-mediated isothermal amplification approach (RT-LAMP) which utilizes colorimetric readout using a heat block, with results evaluated with the unaided eye. In 44 clinical patients suspected of having COVID-19 infection, the test took 45 min, and resulted in a diagnostic sensitivity of 82.6% (19/23) and diagnostic specificity of 100% (21/21), compared to the reference standard. The limit of detection was 250 copies/reaction (25,000 copies/mL). Our newly developed POCT approach achieved simple RNA extraction and constant RT-LAMP detection. This POCT has the potential to be used for simple inspection stations in a field setting, helping reduce the risk of infection by simplifying and accelerating testing for COVID-19.


Asunto(s)
/métodos , Pruebas en el Punto de Atención , ARN Viral/análisis , Saliva/virología , Humanos , Límite de Detección , Técnicas de Diagnóstico Molecular/métodos , Técnicas de Amplificación de Ácido Nucleico/métodos , /aislamiento & purificación
15.
Sci Rep ; 11(1): 4500, 2021 02 24.
Artículo en Inglés | MEDLINE | ID: mdl-33627730

RESUMEN

Emerging evidences have shown the utility of saliva for the detection of SARS-CoV-2 by PCR as alternative to nasopharyngeal swab (NPS). However, conflicting results have been reported regarding viral loads between NPS and saliva. We conducted a study to compare the viral loads between NPS and saliva in 42 COVID-19 patients. Viral loads were estimated by the cycle threshold (Ct) values. SARS-CoV-2 was detected in 34 (81%) using NPS with median Ct value of 27.4, and 38 (90%) using saliva with median Ct value of 28.9 (P = 0.79). Kendall's W was 0.82, showing a high degree of agreement, indicating equivalent viral loads in NPS and saliva. After symptom onset, the Ct values of both NPS and saliva continued to increase over time, with no substantial difference. Self-collected saliva has a detection sensitivity comparable to that of NPS and is a useful diagnostic tool with mitigating uncomfortable process and the risk of aerosol transmission to healthcare workers.


Asunto(s)
/virología , /genética , Adulto , /métodos , Pruebas Diagnósticas de Rutina/métodos , Femenino , Humanos , Masculino , Persona de Mediana Edad , Nasofaringe/virología , Reacción en Cadena de la Polimerasa/métodos , ARN Viral/genética , Saliva/virología , Manejo de Especímenes/métodos , Carga Viral/métodos
16.
BMJ Open Respir Res ; 8(1)2021 02.
Artículo en Inglés | MEDLINE | ID: mdl-33627333

RESUMEN

BACKGROUND: An outbreak of novel coronavirus (SARS-CoV-2)-associated respiratory infectious diseases (COVID-19) emerged in 2019 and has spread rapidly in humans around the world. The demonstration of in vitro infectiousness of respiratory specimens is an informative surrogate for SARS-CoV-2 transmission from patients with COVID-19; accordingly, viral isolation assays in cell culture are an important aspect of laboratory diagnostics for COVID-19. METHODS: We developed a simple and rapid protocol for isolating SARS-CoV-2 from respiratory specimens using VeroE6/TMPRSS2 cells, a cell line that is highly susceptible to the virus. We also investigated a correlation between isolation of SARS-CoV-2 and viral load detected by real-time RT-PCR (rRT-PCR) using N2 primer/probe set that has been developed for testing of COVID-19 in Japan. RESULTS: The SARS-CoV-2 isolation protocol did not require blind passage of inoculated cells and yielded the results of viral isolation within 7 days after inoculation. Specimens with cycle threshold (Ct) values of <20.2, determined by rRT-PCR, were predicted to be isolation-positive. On the other hand, 6.9% of specimens with Ct values >35 were virus isolation-positive, indicating that low viral loads (high Ct values) in upper respiratory specimens do not always indicate no risk of containing transmissible virus. CONCLUSION: In combination with rRT-PCR, the SARS-CoV-2 isolation protocol provides a means for assessing the potential risk of transmissible virus in upper respiratory specimens.


Asunto(s)
/transmisión , /patogenicidad , Animales , Línea Celular , Chlorocebus aethiops , Efecto Citopatogénico Viral , Humanos , Cavidad Nasal/virología , Nasofaringe/virología , Saliva/virología , Serina Endopeptidasas/genética , Manejo de Especímenes , Células Vero
17.
J Med Virol ; 93(5): 3268-3272, 2021 05.
Artículo en Inglés | MEDLINE | ID: mdl-33527375

RESUMEN

Current diagnostic standards involve severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) detection in nasopharyngeal swabs (NPS), but saliva is an attractive and noninvasive option for diagnosis. The objectives were to determine the performance of saliva in comparison with NPS for detecting SARS-CoV-2 and to compare the optimized home brew reverse-transcription polymerase chain reaction (RT-PCR) with a commercial RT-PCR. Paired NPS and saliva specimens were prospectively collected and tested by RT-PCR from patients presenting at an emergency room with signs and symptoms compatible with coronavirus disease-2019. A total of 348 samples from 174 patients were tested by RT-PCR assays. Among 174 patients with symptoms, 63 (36%) were SARS-CoV-2 positive in NPS using the optimized home-brew PCR. Of these 63 patients, 61 (98%) were also positive in saliva. An additional positive SARS-CoV-2 saliva was detected in a patient with pneumonia. Kappa Cohen's coefficient agreement between NPS and saliva was 0.96 (95% confidence interval [CI], 0.90-0.99). Median Ct values in NPS versus saliva were 18.88 (interquartile range [IQR], 15.60-23.58; range, 11.97-38.10) versus 26.10 (IQR, 22.75-30.06; range, 13.78-39.22), respectively (p < .0001). The optimized home-brew RT-PCR demonstrated higher analytical and clinical sensitivity compared with the commercial RT-PCR assay. A high sensitivity (98%) and agreement (kappa 0.96) in saliva samples compared to NPS was demonstrated when using an optimized home-brew PCR even when the viral load in saliva was lower than in NPS. This noninvasive sample is easy to collect, requires less consumable and avoids discomfort to patients. Importantly, self-collection of saliva can diminish exposure to healthcare personnel.


Asunto(s)
/diagnóstico , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa/métodos , Saliva/virología , Manejo de Especímenes/métodos , Adulto , Anciano , Servicio de Urgencia en Hospital , Femenino , Humanos , Masculino , Persona de Mediana Edad , Estudios Prospectivos
18.
Diagn Microbiol Infect Dis ; 100(1): 115324, 2021 May.
Artículo en Inglés | MEDLINE | ID: mdl-33529938

RESUMEN

With surging global demand for SARS-CoV-2 testing capacity, laboratories seek automated, high-throughput molecular solutions, particularly for specimens not requiring specialized collection devices or viral transport media. Saliva specimens submitted from patients under investigation for COVID-19 from March to July 2020 were processed in the laboratory with sterile phosphate-buffered saline in a 1:2 dilution and tested using manual extraction and a commercial assay for detection of the SARS-CoV-2 E gene (LightMix®) in comparison to the Roche cobas® SARS-CoV-2 Test on the cobas® 6800 instrument. 34.4% (22/64) of saliva samples were positive for SARS-CoV-2. Positive and negative concordance between the LightMix® and cobas® assays were 100%. The overall invalid rate for saliva on the cobas® 6800 (1/128, 0.78%) was similar to the baseline invalid rate observed for nasopharyngeal swabs/viral transport media. Saliva is a feasible specimen type for SARS-CoV-2 testing on the cobas® 6800 platform, with potential to improve turnaround time and enhance testing capacity.


Asunto(s)
/métodos , Saliva/virología , Automatización de Laboratorios , Humanos , Técnicas de Diagnóstico Molecular , Manejo de Especímenes/métodos
19.
J Med Microbiol ; 70(3)2021 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-33629949

RESUMEN

This study tests the release of SARS-CoV-2 RNA into the air during normal breathing, without any sign of possible risk of contagion such as coughing, sneezing or talking. Five patients underwent oropharyngeal, nasopharyngeal and salivary swabs for real-time reverse transcriptase PCR (RT-PCR) detection of SARS-CoV-2 RNA. Direct SARS-CoV-2 release during normal breathing was also investigated by RT-PCR in air samples collected using a microbiological sampler. Viral RNA was detected in air at 1 cm from the mouth of patients whose oropharyngeal, nasopharyngeal and salivary swabs tested positive for SARS-CoV-2 RNA. In contrast, the viral RNA was not identified in the exhaled air from patients with oropharyngeal, nasopharyngeal and salivary swabs that tested negative. Contagion of SARS-CoV-2 is possible by being very close to the mouth of someone who is infected, asymptomatic and simply breathing.


Asunto(s)
Microbiología del Aire , /aislamiento & purificación , Aerosoles/análisis , Anciano , Infección Hospitalaria/diagnóstico , Infección Hospitalaria/virología , Hospitales , Humanos , Italia/epidemiología , Nasofaringe/virología , Orofaringe/virología , Aisladores de Pacientes , Saliva/virología
20.
Pathol Res Pract ; 220: 153381, 2021 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-33640711

RESUMEN

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a single-stranded RNA virus that causes coronavirus disease 2019, which spread worldwide immediately after the first patient infected with this virus was discovered in Wuhan, China, in December 2019. Currently, polymerase chain reaction (PCR) specimens for the detection of SARS-CoV-2 include saliva, nasopharyngeal swabs, and lower respiratory tract-derived materials such as sputum. Initially, nasopharyngeal swab specimens were applied mainly to the PCR detection of SARS-CoV-2. There was a risk of infection to healthcare workers due to coughing or sneezing by the subjects at the time of sample collection. In contrast, saliva specimens have a low risk of droplet infection and are easy to collect, and their application to PCR testing has been promoted. In this study, we have determined the detection limit of SARS-CoV-2 in saliva samples and examined the effects of storage temperature and storage time of saliva samples on the PCR detection results. As a result, 5 × 103 copies of SARS-CoV-2 could be detected in 1 mL phosphate-buffered saline, whereas 5 × 104 copies of SARS-CoV-2 were needed in 1 mL saliva to detect the virus by real-time one-step PCR. Interestingly, SARS-CoV-2 (5 × 103 copies/mL) could be detected in saliva supplemented with an RNase inhibitor. Concerning the saliva samples supplemented with an RNase inhibitor, the optimal temperature for sample storage was -20 °C, and PCR detection was maintained within 48 h without problems under these conditions. These finding suggest that RNase in the saliva can affect the detection of SARS-CoV-2 by PCR using saliva samples.


Asunto(s)
/métodos , Reacción en Cadena en Tiempo Real de la Polimerasa/métodos , Ribonucleasas , Saliva/virología , Humanos , Límite de Detección , ARN Viral/análisis , Saliva/enzimología , Manejo de Especímenes/métodos
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