Viral reactivations and co-infections in COVID-19 patients: a systematic review.

BACKGROUND: Viral reactivations and co-infections have been reported among COVID-19 patients. However, studies on the clinical outcomes of different viral reactivations and co-infections are currently in limit. Thus, the primary purpose of this review is to perform an overarching investigation on the cases of latent virus reactivation and co-infection in COVID-19 patients to build collective evidence contributing to improving patient health. The aim of the study was to conduct a literature review to compare the patient characteristics and outcomes of reactivations and co-infections of different viruses. METHODS: Our population of interest included confirmed COVID-19 patients who were diagnosed with a viral infection either concurrently or following their COVID-19 diagnosis. We extracted the relevant literature through a systematic search using the key terms in the online databases including the EMBASE, MEDLINE, Latin American Caribbean Health Sciences Literature (LILACS), from inception onwards up to June 2022. The authors independently extracted data from eligible studies and assessed the risk of bias using the Consensus-based Clinical Case Reporting (CARE) guidelines and the Newcastle-Ottawa Scale (NOS). Main patient characteristics, frequency of each manifestation, and diagnostic criteria used in studies were summarized in tables. RESULTS: In total, 53 articles were included in this review. We identified 40 reactivation studies, 8 coinfection studies, and 5 studies where concomitant infection in COVID-19 patients was not distinguished as either reactivation or coinfection. Data were extracted for 12 viruses including IAV, IBV, EBV, CMV, VZV, HHV-1, HHV-2, HHV-6, HHV-7, HHV-8, HBV, and Parvovirus B19. EBV, HHV-1, and CMV were most frequently observed within the reactivation cohort, whereas IAV and EBV within the coinfection cohort. In both reactivation and coinfection groups, patients reported cardiovascular disease, diabetes, and immunosuppression as comorbidities, acute kidney injury as complication, and lymphopenia and elevated D-dimer and CRP levels from blood tests. Common pharmaceutical interventions in two groups included steroids and antivirals. CONCLUSION: Overall, these findings expand our knowledge on the characteristics of COVID-19 patients with viral reactivations and co-infections. Our experience with current review indicates a need for further investigations on virus reactivation and coinfection among COVID-19 patients.

Implementation of a COVID-19 Genomic Surveillance Regional Network for Latin America and Caribbean region.

The timely release of SARS-CoV-2 first genomic sequences allowed the identification of the etiologic agent and development of diagnostic protocols. Genomic sequencing was a crucial step in generating data for driving laboratory response and detections of SARS-CoV-2 since the start of the COVID-19 pandemic. Because of all the progression and achievements that timely release of genetic sequence data represents in the public health response, the Pan American Health Organization (PAHO) in collaboration with countries' public health laboratories, started implementation of a network for strengthening the Latin America and Caribbean (LAC) region on timely generation of SARS-CoV-2 genomic data. Here we describe the implementation of the COVID-19 Genomic Surveillance Regional Network in the Americas region during the beginning of the pandemic. The establishment of this network has strengthened laboratory response capacity at the country level, as well as facilitated timely release of SARS-CoV-2 genomic information to be used to complement the multiple response strategies for COVID-19 pandemic mitigation. As genomic epidemiology is useful for guiding public health decisions on outbreak and response, we also analysed the first SARS-CoV-2 genomic sequence data from countries of the Latin America and Caribbean Region.

Technical viability of the YF MAC-HD ELISA kit for use in yellow fever-endemic regions.

Yellow fever (YF), an arboviral disease, affects an estimated 200,000 people and causes 30,000 deaths per year and recently has caused major epidemics in Africa and South America. Timely and accurate diagnosis of YF is critical for managing outbreaks and implementing vaccination campaigns. A YF immunoglobulin M (IgM) antibody-capture (MAC) enzyme-linked immunosorbent assay (ELISA) kit, the YF MAC-HD, was successfully introduced starting in 2018 to laboratories in Africa and South America. The YF MAC-HD kit can be performed in 3.5 hours, test up to 24 samples, and includes all reagents necessary to perform the test, except for water used to dilute wash buffer. In 2018 and 2019, a total of 56 laboratory personnel from 39 countries in Africa and South America were trained to use the kit during workshops, followed by take-home YF IgM proficiency testing (PT) exercises. Participants received either a 10- or 20-sample YF PT panel and performed testing using the YF MAC-HD kit. All countries obtained 90% or higher correct results. These results verified the technical viability and transferability of YF MAC-HD kit use for laboratories in YF-endemic countries.

Standardization and validation of an in house RT-LAMP molecular test for the diagnosis of SARS-CoV-2. / Estandarización y validación de una prueba molecular RT-LAMP in house para el diagnóstico de SARS-CoV-2.

OBJECTIVES: To standardize and validate an in-house RT-LAMP test for the detection of SARS-CoV-2, based on laboratory and field assays using samples from COVID-19 suspected patients. MATERIALS AND METHODS: An in-house SARS-CoV-2 RT-LAMP molecular test was standardized, establishing the detection limit with Vero cells of isolated Peruvian strains of SARS-CoV-2, and the robustness to various concentrations of primers. The laboratory validation was performed with 384 nasal and pharyngeal swab samples (UFH) obtained between March and July 2020. The field validation was performed with 383 UFH obtained from COVID-19 suspected symptomatic cases. All samples were tested by RT-LAMP and RT-qPCR. The RT-qPCR was considered as the reference standard test. The concordance measures and diagnostic performance were calculated. RESULTS: The detection limit was consistent in cases with Ct <30 in both tests, showing efficiency to detect up to 1000 copies/µL of the target gene. Robustness was evidenced with half of the primer concentrations and 20 µL of final volume. Absence of amplification was identified for other HCoVs. Concordance showed a kappa index of 0.88 (95% CI: 0.83-0.93) and 0.89 (95% CI: 0.84 - 0.94) in laboratory and field settings, respectively. The sensitivity value in the laboratory was 87.4% (95% CI: 80.8 - 92.4) and 88.1% in the field (95% CI: 81.6 - 92.9). The specificity value in both settings was 98.8% (95% CI: 96.4-99.7). CONCLUSIONS: The in-house SARS-CoV-2 RT-LAMP test was successfully validated based on its adequate robustness, no cross-reactions, good concordance, and diagnostic performance compared to RT-qPCR.

OBJETIVOS: Estandarizar una prueba RT-LAMP in house para la detección de SARS-CoV-2 y validarla con muestras de laboratorio y de campo en pacientes con sospecha clínica de COVID-19. MATERIALES Y MÉTODOS: Se estandarizó una prueba molecular RT-LAMP in house para la detección de SARS-CoV-2 estableciéndose el límite de detección con células Vero de cepas peruanas aisladas de SARS-CoV-2. Se validó la prueba en laboratorio con 384 muestras de hisopado nasal y faríngeo (HNF) obtenidas entre marzo y julio de 2020. Para la validación de campo se obtuvieron muestras de HNF de 383 casos sintomáticos sospechosos de COVID-19. Todas las muestras fueron evaluadas por RT-LAMP y RT-qPCR. Para la validación de laboratorio y de campo se consideró como estándar de referencia al RT-qPCR, se calcularon medidas de concordancia y rendimiento diagnóstico. RESULTADOS: El límite de detección fue consistente en los casos con umbral de ciclo (Ct) Ct < 30 en ambas pruebas, mostrando eficiencia para detectar hasta 1000 copias/µL del gen diana. Se evidenció robustez con la mitad de las concentraciones de cebadores y 20 µL de volumen final. Se identificó ausencia de amplificación para otros coronavirus humanos. La concordancia en laboratorio obtuvo un Kappa de 0,88 (IC 95%: 0,83-0,93) y en campo fue de 0,89 (IC 95%: 0,84−0,94); la sensibilidad en laboratorio fue de 87,4% (IC 95%: 80,8−92,4) y en campo fue de 88,1% (IC 95%: 81,6−92,9), la especificidad en ambos escenarios fue de 98,8% (IC 95%: 96,4−99,7). CONCLUSIONES: La prueba RT-LAMP in house fue validada por presentar una adecuada robustez, sin reacciones cruzadas, buena concordancia y rendimiento diagnóstico comparado con el RT-qPCR.

COVID-19 laboratory preparedness and response in the Americas Region: Lessons learned.

By the time the etiologic agent of the COVID-19 was identified as a novel coronavirus, no country in the Americas Region had laboratory capacity for detecting this new virus. A strategic multilevel approach with specific reagent purchase and delivery, regional trainings, in-country missions, and the provision of technical support was established for timely preparedness of national reference laboratories for SARS-CoV-2 detection. All countries should be prepared to timely detect any potential pandemic emerging agent. The rapid SARS-CoV-2 molecular detection implementation throughout the Americas showed the importance of an efficient and coordinated laboratory response for preparedness. Here we present how in 25 days the Americas Region went from no SARS-CoV-2 diagnostic capacity, to molecular detection fully implemented in 28 Member States, under the coordinated strategy of the Pan American Health Organization and collaborative work at regional and country level with national authorities and public health laboratories.

A faster and less costly alternative for RNA extraction of SARS-CoV-2 using proteinase k treatment followed by thermal shock.

One of the biggest challenges during the pandemic has been obtaining and maintaining critical material to conduct the increasing demand for molecular tests. Sometimes, the lack of suppliers and the global shortage of these reagents, a consequence of the high demand, make it difficult to detect and diagnose patients with suspected SARS-CoV-2 infection, negatively impacting the control of virus spread. Many alternatives have enabled the continuous processing of samples and have presented a decrease in time and cost. These measures thus allow broad testing of the population and should be ideal for controlling the disease. In this sense, we compared the SARS-CoV-2 molecular detection effectiveness by Real time RT-PCR using two different protocols for RNA extraction. The experiments were conducted in the National Institute of Health (INS) from Peru. We compared Ct values average (experimental triplicate) results from two different targets, a viral and internal control. All samples were extracted in parallel using a commercial kit and our alternative protocol-samples submitted to proteinase K treatment (3 µg/µL, 56°C for 10 minutes) followed by thermal shock (98°C for 5 minutes followed by 4°C for 2 minutes); the agreement between results was 100% in the samples tested. In addition, we compared the COVID-19 positivity between six epidemiological weeks: the initial two in that the Real time RT-PCR reactions were conducted using RNA extracted by commercial kit, followed by two other using RNA obtained by our kit-free method, and the last two using kit once again; they did not differ significantly. We concluded that our in-house method is an easy, fast, and cost-effective alternative method for extracting RNA and conducing molecular diagnosis of COVID-19.