scholarly journals The use of denaturing solution as collection and transport media to improve SARS-CoV-2 RNA detection and reduce infection of laboratory personnel

2021 ◽  
Author(s):  
Alex F. Carvalho ◽  
Raissa P. Rocha ◽  
Andreza P. Gonçalves ◽  
Thaís B. S. Silva ◽  
Hugo I. Sato ◽  
...  
Keyword(s):  
Rna Detection ◽  
Laboratory Personnel

scholarly journals THE USE OF DENATURING SOLUTION AS COLLECTION AND TRANSPORT MEDIA TO IMPROVE SARS-COV-2 RNA DETECTION AND REDUCE INFECTION OF LABORATORY PERSONNEL

2020 ◽  
Author(s):  
Alex Fiorini de Carvalho ◽  
Andreza Parreiras Gonçalves ◽  
Thaís Bárbara de Souza Silva ◽  
Hugo Itaru Sato ◽  
Larissa Vuitika ◽  
...  
Keyword(s):  
Diagnostic Tests ◽  
Diagnostic Procedures ◽  
Clinical Samples ◽  
Sample Collection ◽  
Rna Detection ◽  
Laboratory Personnel ◽  
Human Genes ◽  
Oropharyngeal Swab ◽  
The Uk ◽  
Infectious Potential

AbstractBackgroundSince the emergence of the COVID-19, health officials have struggled to devise strategies to counteract the speed of the pandemic’s spread across the globe. It became imperative to implement accurate diagnostic tests for the detection of SARS-CoV-2 RNA on respiratory samples. In many places, however, besides the limited availability of test reagents, laboratory personnel face the challenge of adapting their working routines to manipulate highly infective clinical samples. Here, we proposed the use of a virus-inactivating solution as part of a sample collection kit to decrease the infectious potential of the collected material without affecting the integrity of RNA samples used in diagnostic tests based on RT-qPCR.MethodsNasopharyngeal and oropharyngeal swab samples were collected from SARS-CoV-2-infected patients and from laboratory personnel using a commercially available viral transport solution (VTM) and the denaturing solution (DS) described here. RNA extracted from all samples was tested by RT-qPCR using probes for viral and human genes. Exposure of laboratory personnel to infective viruses was also accessed using ELISA tests.FindingsThe use of the DS did not interfere with the detection of viral genome or the endogenous human mRNA, since similar results were obtained from samples collected with VTM or DS. In addition, all tests of laboratory personnel for the presence of viral RNA and IgG antibodies against SARS-CoV-2 were negative.InterpretationThe methodology described here provides a strategy that allow high diagnostic accuracy as well as safe manipulation of clinical samples by those involved with diagnostic procedures.FundingCAPES, FAPEMIG, CNPq, MCTIC, FIOCRUZ and the UK Global Challenges Research Fund (GCRF).

Distinctive COVID-19 manifestations and re-exposure to SARS CoV 2 in clinical laboratory personnel under regular monitoring by RT-PCR and serology

2021 ◽  
Vol 2 (2) ◽  
Author(s):  
Fábio de Oliveira Martinez Alonso ◽  
◽  
Bruno Duarte Sabino ◽  
Marianna Tavares Venceslau ◽  
Rafael Brandão Varella ◽  
...  
Keyword(s):  
Clinical Laboratory ◽  
Rt Pcr ◽  
Rna Detection ◽  
Laboratory Personnel ◽  
Regular Monitoring ◽  
Antibody Levels ◽  
Lack Of Knowledge

Clinical laboratory personnel (CLP) present a unique opportunity to investigate distinct forms of COVID-19 as they are under constant testing for SARS CoV 2 infection. SARS CoV 2 RNA and antibodies were routinely investigated over a 5-month period in 26 professionals from a clinical laboratory in RJ, Brazil. Of them, three (11.5%) CLP presented the following peculiar COVID-19 manifestations: 2/26 (7.7%) had SARS CoV 2 antibodies without RNA detection during the follow-up, with a possible re-exposure in one case, and 1/26 (3.8%) a confirmed reinfection with RNA detection, and possibly a third re-exposure. Based on a long follow-up of SARS CoV 2 infection in CLP, this study showed that cases of COVID-19 without RNA detection are not common, but it does indicate the risk of re-exposure after the fall of antibody levels. Although scarcely reported, the investigation of less frequent forms of COVID-19 is relevant, given the lack of knowledge of its impact on the pandemic.

scholarly journals Seroprevalence of Anti-SARS-CoV-2 IgG and IgM among Adults over 65 Years Old in the South of Italy

Diagnostics ◽  
2021 ◽  
Vol 11 (3) ◽  
pp. 483
Author(s):  
Immacolata Polvere ◽  
Alfredina Parrella ◽  
Giovanna Casamassa ◽  
Silvia D’Andrea ◽  
Annamaria Tizzano ◽  
...  
Keyword(s):  
Antibody Response ◽  
Immunoglobulin M ◽  
Molecular Testing ◽  
Elisa Assay ◽  
The South ◽  
Serum Samples ◽  
Serological Screening ◽  
Rna Detection ◽  
In Vitro Diagnostic

SARS-CoV-2 is a zoonotic betacoronavirus associated with worldwide transmission of COVID-19 disease. By the beginning of March, WHO reported about 113,820,000 confirmed cases including more than 2,527,000 deaths all over the world. However, the true extent of virus circulation or its real infection/fatality ratio is not well-estimated due to the huge portion of asymptomatic infections. In this observational study, we have estimated the prevalence of specific immunoglobulin M and G directed towards SARS-CoV-2 antigen in a cohort of 1383 adult volunteers aged over 65 years old, living in the district of Benevento, in the South of Italy. Serological screening was carried out on capillary blood in September 2020, seven months after pandemic outbreak in Italy, to evaluate virus circulation and antibody response among elderly adults, in which severe symptoms due to viral infection are more common. The overall seroprevalence of anti-SARS-CoV-2 antibodies was 4.70% (CI 3.70%–5.95%) with no statistically significant differences between sexes. Among these, 69.69% (CI 55.61%–77.80%) tested positive to IgM, 23.08% (CI 14.51%–34.64%) to IgG and 9.23% (CI 4.30%–18.71%) was positive for both. All patients that were positive to IgM underwent molecular testing through RT-qPCR on oral-rhino pharyngeal swabs and only one specimen was positive for SARS-CoV-2 RNA detection. Instead, the presence of IgG from screened volunteers was confirmed by re-testing serum samples using both an ELISA assay validated for in vitro diagnostic use (IVD) and a recently published synthetic peptide-based ELISA assay. In conclusion, our report suggests that (1) early restrictions were successful in limiting COVID-19 diffusion in the district of Benevento; (2) rapid serological analysis is an ideal testing for both determining real seroprevalence and massive screening, whereas detection of viral RNA remains a gold standard for identification of infected patients; (3) even among people without COVID-19 related symptoms, the antibody response against SARS-CoV-2 antigens has individual features.

scholarly journals Superiority of MALDI-TOF Mass Spectrometry over Real-Time PCR for SARS-CoV-2 RNA Detection

Viruses ◽  
2021 ◽  
Vol 13 (5) ◽  
pp. 730
Author(s):  
Magda Rybicka ◽  
Ewa Miłosz ◽  
Krzysztof Piotr Bielawski
Keyword(s):  
Mass Spectrometry ◽  
Gold Standard ◽  
Viral Genome ◽  
False Negative ◽  
Rt Pcr ◽  
Rna Detection ◽  
Maldi Tof ◽  
Negative Results ◽  
False Negative Results ◽  
Pcr Test

At present, the RT-PCR test remains the gold standard for early diagnosis of SARS-CoV-2. Nevertheless, there is growing evidence demonstrating that this technique may generate false-negative results. Here, we aimed to compare the new mass spectrometry-based assay MassARRAY® SARS-CoV-2 Panel with the RT-PCR diagnostic test approved for clinical use. The study group consisted of 168 suspected patients with symptoms of a respiratory infection. After simultaneous analysis by RT-PCR and mass spectrometry methods, we obtained discordant results for 17 samples (10.12%). Within fifteen samples officially reported as presumptive positive, 13 were positive according to the MS-based assay. Moreover, four samples reported by the officially approved RT-PCR as negative were positive in at least one MS assay. We have successfully demonstrated superior sensitivity of the MS-based assay in SARS-CoV-2 detection, showing that MALDI-TOF MS seems to be ideal for the detection as well as discrimination of mutations within the viral genome.

scholarly journals Noncanonical crRNAs derived from host transcripts enable multiplexable RNA detection by Cas9

Science ◽  
2021 ◽  
pp. eabe7106
Author(s):  
Chunlei Jiao ◽  
Sahil Sharma ◽  
Gaurav Dugar ◽  
Natalia L. Peeck ◽  
Thorsten Bischler ◽  
...  
Keyword(s):  
Campylobacter Jejuni ◽  
Genetic Material ◽  
Simultaneous Detection ◽  
Type Ii ◽  
Rna Detection ◽  
Single Base ◽  
Dna Targeting ◽  
Single Base Resolution ◽  
Rna Complexes

CRISPR-Cas systems recognize foreign genetic material using CRISPR RNAs (crRNAs). In Type II systems, a trans-activating crRNA (tracrRNA) hybridizes to crRNAs to drive their processing and utilization by Cas9. While analyzing Cas9-RNA complexes from Campylobacter jejuni, we discovered tracrRNA hybridizing to cellular RNAs, leading to formation of “noncanonical” crRNAs capable of guiding DNA targeting by Cas9. Our discovery inspired the engineering of reprogrammed tracrRNAs that link the presence of any RNA-of-interest to DNA targeting with different Cas9 orthologs. This capability became the basis for a multiplexable diagnostic platform termed LEOPARD (Leveraging Engineered tracrRNAs and On-target DNAs for PArallel RNA Detection). LEOPARD allowed simultaneous detection of RNAs from different viruses in one test and distinguished SARS-CoV-2 and its D614G variant with single-base resolution in patient samples.

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