Microwaves: a Novel Approach to SARS-CoV-2 Genome Extraction (preprint)

On this study an innovative approach to the heat extraction has been tested: the use of microwaves, which can decrease dramatically the time needed to do the genome extraction. The method can obtain the virus with enough quality to assure the identification by q(RT)-PCR and minimize procedures and contaminations.

Comparison of fourteen Rapid Point-of-Care Antigen Tests for SARS-CoV-2: Use & Sensitivity (preprint)

Fast sensitive techniques are advisable for SARS-CoV-2 detection. Various rapid SARS-CoV-2 antigen detection tests have been developed, but type and quality of the sample, stage of the disease and viral load can all have an impact on their sensitivity. For this study, a total of 486 swabs were processed and checked with various commercially available tests, and then compared with q(RT)-PCR (the gold-standard method). Total sensitivity varied considerably, for example, 42.10% (nal von minden and Tody Laboratories), 68.42% (Cahnos) and 84.78% (PCL). Sensitivity reached 100% when cycle threshold (Ct) was lower than 22 in almost all tests, although this dropped considerably when Ct was higher above 30, where only three tests identified 40% or more positive samples and in 5 cases it was 0%. What is more, only two cases were 100% accurate when viral load was higher than 5 log/103 cells and accuracy was 0% in 12 cases where viral load was lower than 4 log/103 cells. These results, particularly taking into consideration the fact they used normalized viral load, suggest that antigen detection tests have their role in the fast triage of positive patients, but that considerable care should be taken with negative results, which is even more important if they are used for massive screening.

C4COVID-19 Human: A Rapid SARS-Cov-2 Salivary Test Compared to the Gold Standard – Clinical Results (preprint)

The SARS-CoV-2 detection campaign is a key element in the viral pandemic response. Here we present a monocentric study to evaluate a simplified molecular amplification technology alternative to classical RT-qPCR platforms. The test, C4Covid-19 HumanTM makes use of saliva as a starting sample allowing for self, painless collection, avoiding risk of viral transmission for the operator who performs the nasopharyngeal (NP) procedure. Total time to result is 30 minutes. The principle relies on an extraction-free detection of two targets within the viral genomic RNA, RdRp and N genes, by real-time reverse transcription loop mediated isothermal amplification (RT-LAMP). Saliva and NP swabs were collected simultaneously from 1491 symptomatic or contact cases and analyzed in parallel by RT-LAMP and RT-qPCR respectively. The 249 positives RT-qPCR samples had a Ct <37. Sensitivity and specificity of RT-LAMP were 85.7% (95% CI 80.4 to 89.7) and 97.3% (95% CI 96.1 to 98.1) respectively. Among the total cohort, the 56.9% of asymptomatic subjects showed higher sensitivity (90.6% with a 95% CI of 81.0 to 95.6) while specificity remained unchanged (97.4% with a 95% CI of 95.6 to 98.4). C4Covid-19 HumanTM is a rapid, simple and reliable SARS-CoV-2 detection test for population screening.

The East African Community (EAC) mobile laboratory networks in Kenya, Burundi, Tanzania, Rwanda, Uganda, and South Sudan-from project implementation to outbreak response against Dengue, Ebola, COVID-19, and epidemic-prone diseases.

BACKGROUND: East Africa is home to 170 million people and prone to frequent outbreaks of viral haemorrhagic fevers and various bacterial diseases. A major challenge is that epidemics mostly happen in remote areas, where infrastructure for Biosecurity Level (BSL) 3/4 laboratory capacity is not available. As samples have to be transported from the outbreak area to the National Public Health Laboratories (NPHL) in the capitals or even flown to international reference centres, diagnosis is significantly delayed and epidemics emerge. MAIN TEXT: The East African Community (EAC), an intergovernmental body of Burundi, Rwanda, Tanzania, Kenya, Uganda, and South Sudan, received 10 million € funding from the German Development Bank (KfW) to establish BSL3/4 capacity in the region. Between 2017 and 2020, the EAC in collaboration with the Bernhard-Nocht-Institute for Tropical Medicine (Germany) and the Partner Countries' Ministries of Health and their respective NPHLs, established a regional network of nine mobile BSL3/4 laboratories. These rapidly deployable laboratories allowed the region to reduce sample turn-around-time (from days to an average of 8h) at the centre of the outbreak and rapidly respond to epidemics. In the present article, the approach for implementing such a regional project is outlined and five major aspects (including recommendations) are described: (i) the overall project coordination activities through the EAC Secretariat and the Partner States, (ii) procurement of equipment, (iii) the established laboratory setup and diagnostic panels, (iv) regional training activities and capacity building of various stakeholders and (v) completed and ongoing field missions. The latter includes an EAC/WHO field simulation exercise that was conducted on the border between Tanzania and Kenya in June 2019, the support in molecular diagnosis during the Tanzanian Dengue outbreak in 2019, the participation in the Ugandan National Ebola response activities in Kisoro district along the Uganda/DRC border in Oct/Nov 2019 and the deployments of the laboratories to assist in SARS-CoV-2 diagnostics throughout the region since early 2020. CONCLUSIONS: The established EAC mobile laboratory network allows accurate and timely diagnosis of BSL3/4 pathogens in all East African countries, important for individual patient management and to effectively contain the spread of epidemic-prone diseases.

Comparison of in-house SARS-CoV-2 genome extraction procedures. A need for COVID-19 pandemic (preprint)

Purpose: Among the methods used to diagnose COVID-19, those based on genomic detection by q(RT)-PCR are the most sensitive. To perform these assays, a previous genome extraction of the sample is required. The dramatic increase in the number of SARS-CoV-2 detection assays has increased the demand for extraction reagents hindering the supply of commercial reagents. Homemade reagents and procedures could be an alternative. Methods: Nasopharyngeal samples were extracted by seven different methods as well as the automatic method MagNaPure96, to detect SARS-CoV-2. Results: All protocols show sensitivity higher than 87%, in comparison with reference method, for detecting SARS-CoV-2 as well as human β- globin. Conclusions: Our results support that these procedures, using common and cheap reagents, are effective to extract RNA (from SARS-CoV-2) or DNA (from human β-globin) genome from nasopharyngeal swabs. Furthermore, these procedures could be easily adopted by routine diagnostic laboratories to implement detection methods to help to fight against COVID-19 pandemic.

SARS-CoV-2 analysis on environmental surfaces collected in an intensive care unit. Keeping Ernest Shackleton´s spirit. (preprint)

Background: Intensive care unit workers are at high risk of acquiring COVID-19 infection, especially when performing invasive techniques and certain procedures that generate aerosols (<5 µm). Therefore, one of the objectives of the health systems should implement safety practices to minimize the risk of contagion among these health professionals. Monitoring environmental contamination of SARS-CoV-2 may help to determine the potential of the environment as a transmission medium in an area highly exposed to SARS-CoV-2, such as an intensive care unit. The objective of the study was to analyze the environmental contamination by SARS-CoV-2 on surfaces collected in an intensive care unit, which is dedicated exclusively to the care of patients with COVID-19 and equipped with negative pressure of -10 pascals and an air change rate of 20 cycles per hour. Furthermore, all ICU workers were tested for COVID-19 by quantitative RT-PCR and ELISA methods. Results: A total of 102 samples (72 collected with pre-moistened swabs used for collection of nasopharyngeal exudates and 30 with moistened wipes used in the environmental microbiological control of the food industry) were obtained from ventilators, monitors, perfusion pumps, bed rails, lab benches, containers of personal protective equipment, computer keyboards and mice, telephones, workers' shoes, floor and other areas of close contact with COVID-19 patients and healthcare professionals who cared for them. The analysis by quantitative RT-PCR showed no detection of SARS-CoV-2 genome in environmental samples collected by any of the two methods described. Furthermore, none of the ICU workers was infected by the virus. Conclusions: Presence of SARS-CoV-2 on the ICU surfaces could not be determined supporting that a strict cleaning protocol with sodium hypochlorite, a high air change rate and a negative pressure in the ICU are effective in preventing environmental contamination. These facts together with the protection measures used could also explain the absence of contagion among staff inside ICUs.