Initial Insights Into the Genetic Epidemiology of SARS-CoV-2 Isolates From Kerala Suggest Local Spread From Limited Introductions.

Coronavirus disease 2019 (COVID-19) rapidly spread from a city in China to almost every country in the world, affecting millions of individuals. The rapid increase in the COVID-19 cases in the state of Kerala in India has necessitated the understanding of SARS-CoV-2 genetic epidemiology. We sequenced 200 samples from patients in Kerala using COVIDSeq protocol amplicon-based sequencing. The analysis identified 166 high-quality single-nucleotide variants encompassing four novel variants and 89 new variants in the Indian isolated SARS-CoV-2. Phylogenetic and haplotype analysis revealed that the virus was dominated by three distinct introductions followed by local spread suggesting recent outbreaks and that it belongs to the A2a clade. Further analysis of the functional variants revealed that two variants in the S gene associated with increased infectivity and five variants mapped in primer binding sites affect the efficacy of RT-PCR. To the best of our knowledge, this is the first and most comprehensive report of SARS-CoV-2 genetic epidemiology from Kerala.

Rapid and accurate nucleobase detection using FnCas9 and its application in COVID-19 diagnosis.

Rapid detection of DNA/RNA pathogenic sequences or variants through point-of-care diagnostics is valuable for accelerated clinical prognosis, as witnessed during the recent COVID-19 outbreak. Traditional methods relying on qPCR or sequencing are tough to implement with limited resources, necessitating the development of accurate and robust alternative strategies. Here, we report FnCas9 Editor Linked Uniform Detection Assay (FELUDA) that utilizes a direct Cas9 based enzymatic readout for detecting nucleobase and nucleotide sequences without trans-cleavage of reporter molecules. We also demonstrate that FELUDA is 100% accurate in detecting single nucleotide variants (SNVs), including heterozygous carriers, and present a simple web-tool JATAYU to aid end-users. FELUDA is semi-quantitative, can adapt to multiple signal detection platforms, and deploy for versatile applications such as molecular diagnosis during infectious disease outbreaks like COVID-19. Employing a lateral flow readout, FELUDA shows 100% sensitivity and 97% specificity across all ranges of viral loads in clinical samples within 1hr. In combination with RT-RPA and a smartphone application True Outcome Predicted via Strip Evaluation (TOPSE), we present a prototype for FELUDA for CoV-2 detection closer to home.

A rapid and sensitive method to detect SARS-CoV-2 virus using targeted-mass spectrometry.

In the last few months, there has been a global catastrophic outbreak of severe acute respiratory syndrome disease caused by the novel coronavirus SARS-CoV-2 affecting millions of people worldwide. Early diagnosis and isolation are key to contain the rapid spread of the virus. Towards this goal, we report a simple, sensitive and rapid method to detect the virus using a targeted mass spectrometric approach, which can directly detect the presence of virus from naso-oropharyngeal swabs. Using a multiple reaction monitoring we can detect the presence of two peptides specific to SARS-CoV-2 in a 2.3 min gradient run with 100% specificity and 90.5% sensitivity when compared to RT-PCR. Importantly, we further show that these peptides could be detected even in the patients who have recovered from the symptoms and have tested negative for the virus by RT-PCR highlighting the sensitivity of the technique. This method has the translational potential of in terms of the rapid diagnostics of symptomatic and asymptomatic COVID-19 and can augment current methods available for diagnosis of SARS-CoV-2.