key: cord-0701821-4i4kdsed authors: López de la Iglesia, Jaime; Fernández-Villa, Tania; Rivero, Ana; Carvajal, Ana; Simon, Estefanía Bay; Martínez, Miriam Martínez; Argüello, Héctor; Puente, Héctor; Vázquez, Jose Pedro Fernández title: Predictive factors of Covid-19 in patients with negative RT-qPCR date: 2020-06-23 journal: Semergen DOI: 10.1016/j.semerg.2020.06.010 sha: 09114cea3143abccf3e462f38de7bc05590921b5 doc_id: 701821 cord_uid: 4i4kdsed Abstract Objective: To evaluate the factors associated with false negatives in RT-qPCR in patients with mild-moderate symptoms of COVID-19. Materials and methods: This was a cross-sectional study that used a random sample of non-hospitalized patients from the primary care management division of the Healthcare Area of Leon (58 RT-qPCR-positive cases and 52 RT-qPCR-negative cases). Information regarding symptoms was collected and all patients were simultaneously tested using two rapid diagnostic test - RDTs (Combined - cRDT and Differentiated - dRDT). The association between symptoms and SARS-CoV-2 infection was evaluated by non-conditional logistic regression, with estimation of Odds Ratio. Results: A total of 110 subjects were studied, 52% of whom were women (mean age: 48.2 ± 11.0 years). 42.3% of negative RT-qPCRs were positive in some RDTs. Fever over 38ºC (present in 35.5% of cases) and anosmia (present in 41.8%) were the symptoms most associated with SARS-CoV-2 infection, a relationship that remained statistically significant in patients with negative RT-qPCR and some positive RDT (aOR=6.64; 95%CI=1.33-33.13 and aOR=19.38; 95% CI=3.69-101.89 respectively Conclusions: RT-qPCR is the technique of choice in the diagnosis of SARS-CoV-2 infection, but it is not exempt from false negatives. Our results show that those patients who present mild or moderate symptoms with negative RT-qPCR, but with fever and/or anosmia, should be considered suspicious cases and should be evaluated with other diagnostic methods. J o u r n a l P r e -p r o o f Objetivo: Evaluar los factores asociados con falsos negativos a RT-qPCR negativa y sintomatología leve o moderada de COVID-19. Materiales y métodos: Estudio transversal. Se utilizó una muestra aleatoria de pacientes no hospitalizados de la Gerencia de Atención Primaria del Área de Salud de León (58 con RT-qPCR positiva y 52 con RT-qPCR negativa). Se recogió información sobre síntomas y a todos se les realizó simultáneamente dos pruebas de diagnóstico rápido -PDR (Combinada: PRD-C y Diferenciada: PRD-D). La asociación de los síntomas con la infección por SARS-CoV-2 se evaluó mediante regresión logística no condicional, con el cálculo de Odds Ratio. One of the main strategies for the prevention and control of the COVID-19 pandemic is the detection and isolation of the infection sources 1 . The Reverse transcription quantitative polymerase chain reaction (RT-qPCR) is the technique of choice for detecting infection sources due to its high sensitivity and specificity 2 . Also, this is a well-known and widespread technique in the clinical laboratories of our hospitals 3 . This method, however, is not free of false negatives, being the most frequent causes of them the inadequate sample collection, delays in transport, labelling errors and the poor virus elimination in the patient 4, 5 . For these reasons, in the face of clinical suspicion, a negative RT-qPCR result must be contrasted with other diagnostics test 4 . The aim of this article is to understand the factors associated with false negatives in RT-qPCR in patients with mild-moderate symptoms of COVID-19. A cross-sectional study was carried out. A random selection was made of 58 non-hospitalized patients with positive RT-qPCR and 52 negative RT-qPCR. Patients were selected from the register of confirmed or suspected COVID-19 cases from primary care management of the Healthcare Area of Leon. In all of them more than 14 days had passed since the beginning of the symptoms. Participation in the study was voluntary. The invitation to participate in the study was made by a telephone call, in which the participants were cited for the collection of a biological sample and information. During the collection of information and samples, all protection regulations were followed and the project was approved by the Ethics Committee of the health area of León and the Bierzo (reference: 2073). After signing an informed consent, each participant completed a brief ad hoc questionnaire that collected information on socio-demographic data, symptoms, date of onset and end of symptoms, date of RT-qPCR. All patients were tested simultaneously with two RDTs 6 : Both tests were performed on a fingerstick whole blood sample, collected by two nurses. The first test jointly determines the presence of IgM and IgG, while the second test makes a differentiated measurement of both antibody subtypes. The result of the tests was read 10-15 minutes after they were carried out. We considered as a case of COVID19 those patients with a positive result to at least one of the RDTs or RT-qPCR. Central and dispersion measures were calculated in quantitative variables (mean and standard deviation (SD)) and frequencies with their 95% confidence intervals in qualitative variables. Using non-conditional logistic regression models, adjusted at least by age and sex, we obtained the Odds Ratio (aOR) to be considered COVID-19 case, performing stratified analysis according to the results of the RDTs and the RT-qPCR. The characteristics of those patients with negative RT-qPCR and positive PDR were examined. All analyses were performed with the STATA 15 statistical package 7 . A total of 110 subjects were studied, 51.8% of whom were women. The age range was between Table 1 shows the distribution of symptoms and their association with being a COVID-19. A statistically significant association with fever, anosmia, ageusia, myalgia and anorexia were observed in the model adjusted for age and sex. Adjusting for age, sex and the signs and symptoms statistically associated with COVID-19, the significant association with anosmia (aOR=20.39; 95%CI=4.74-87.73) and fever (aOR=4.33; 95%CI=1.24-15.11) was maintained. Figure 2 shows that all the patients with fever and anosmia have been positive to some test and in the case of patients with negative RT-qPCR but positive to some RDT, more than 80% (18/22) have either fever or anosmia or both. Table 2 shows how anosmia (aOR=19.38) and the presence of fever (aOR=6.64) are strong and significantly associated with having COVID-19 in subjects previously tested negative by RT-qPCR. * aOR: adjusted odds ratio by sex and age. The results of our study reflect that 42.3% of respondents with negative RT-qPCR were positive for some RDT. The symptoms most commonly associated with SARS-CoV-2 infection were fever over 38ºC (present in 35.5% of cases) and anosmia (present in 41.8%), a relationship that remained significant in individuals with negative RT-qPCR and some positive RDT (aOR=6.64; 95%CI=1.33-33.13 and aOR=19.38; 95% CI=3.69-101.89 respectively). RT-qPCR is the most widely used technique in the diagnosis of SARS-CoV-2 infection, given its high reliability 3 . However, it is not exempt from false negatives, which may be due to tests with fewer genes detected, mild symptoms or very early stage of the disease, upper respiratory tract sample, denaturation of the materials used, delay in transport or processing of the sample 4,5 . Our data show that 42.3% of the subjects analyzed were classified as healthy according to the With regard to symptoms associated with SARS-CoV-2 infection, fever and anosmia were significant in the false negatives. These symptoms associated with the infection were not surprising, since the beginning of the pandemic, fever, along with coughing and difficulty in breathing were the alarm symptoms. According to data published in a recent review, fever is present in 83-98% of cases, but up to 17% of patients may have afebrile illness 5 . The Centers for Disease Control and Prevention (CDC) has expanded the list of symptoms associated with the disease to include chills, muscle pain, sore throat, and recent loss of taste or smell 10-12 . This last symptom, anosmia, is emerging strongly among the cases detected worldwide, having been described by several studies with a prevalence of between 33-86% 13, 14 . Despite not being present in all cases, these are very characteristic symptoms that can arise in the early stage of the disease, so it should be taken into account as clinical suspicion. The results obtained in this study must be interpreted with caution, being the main limitations the descriptive nature of the study and the low sample size. However, it is one of the first research studies in Spain to highlight the factors associated with false negatives in the choice test in the diagnosis of COVID-19. This research has not received specific support from public sector agencies, the commercial sector or non-profit organizations. Información científico -técnica Recent progress on the diagnosis of 2019 Novel Coronavirus Técnicas y sistemas de diagnóstico para COVID-19 Recomendaciones institucionales. Documento de posicionamiento de la SEIMC sobre el diagnóstico microbiológico del COVID-19 SARS CoV-2, manifestaciones clínicas y consideraciones en el abordaje diagnóstico de COVID-19. Revista Médica de Costa Rica Informe sobre estrategia de diagnóstico microbiológico del COVID-19 (Actualización) Madrid 19 de abril de 2020 StataCorp. 2017. Stata: Release 15. Statistical Software. College Station, TX: StataCorp LLC Evaluating the accuracy of different respiratory specimens in the laboratory diagnosis and monitoring the viral shedding of 2019-nCoV infections Nota de prensa: Faltan estudios bien diseñados que apoyen la eficacia de los tratamientos antivirales y antiinflamatorios en la COVID-19 Síntomas de la enfermedad del coronavirus Evolution of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) as coronavirus disease 2019 (COVID-19) pandemic: A global health emergency Olfactory Dysfunction in COVID-19: Diagnosis and Management Self-reported Olfactory and Taste Disorders in Patients With Severe Acute Respiratory Coronavirus 2 Infection: A Crosssectional Study Olfactory and gustatory dysfunctions as a clinical presentation of mild-to-moderate forms of the coronavirus disease (COVID-19): a multicenter32. European study The authors would like to thank all patients for their participation in this study who have voluntarily agreed to collaborate in the evaluation of diagnostic techniques against COVID-19.Thank you to the primary care management division of the Healthcare Area of Leon and the University of Leon also, because their collaboration has been crucial in developing this study. The rapid spread of the COVID-19 pandemic requires the rapid generation of knowledge to combat this disease. Our results reflect how those patients with fever and anosmia and negative RT-qPCR, should be considered as suspected COVID-19 cases in clinical practice, given the percentage of false negatives that this test of choice is presenting. Furthermore, it highlights the need to perform a combination of tests to ensure the diagnosis of the disease, such as the use of RDTs. The authors declare no conflict of interest.