Bio-SCAN: A CRISPR/dCas9-Based Lateral Flow Assay for Rapid, Specific, and Sensitive Detection of SARS-CoV-2.

Simple, rapid, specific, and sensitive point-of-care detection methods are needed to contain the spread of SARS-CoV-2. CRISPR/Cas9-based lateral flow assays are emerging as a powerful alternative for COVID-19 diagnostics. Here, we developed Bio-SCAN (biotin-coupled specific CRISPR-based assay for nucleic acid detection) as an accurate pathogen detection platform that requires no sophisticated equipment or technical expertise. Bio-SCAN detects the SARS-CoV-2 genome in less than 1 h from sample collection to result. In the first step, the target nucleic acid sequence is isothermally amplified in 15 min via recombinase polymerase amplification before being precisely detected by biotin-labeled nuclease-dead SpCas9 (dCas9) on commercially available lateral flow strips. The resulting readout is visible to the naked eye. Compared to other CRISPR-Cas-based pathogen detection assays, Bio-SCAN requires no additional reporters, probes, enhancers, reagents, or sophisticated devices to interpret the results. Bio-SCAN is highly sensitive and successfully detected a clinically relevant level (4 copies/µL) of synthetic SARS-CoV-2 RNA genome. Similarly, Bio-SCAN showed 100% negative and 96% positive predictive agreement with RT-qPCR results when using clinical samples (86 nasopharyngeal swab samples). Furthermore, incorporating variant-specific sgRNAs in the detection reaction allowed Bio-SCAN to efficiently distinguish between the α, ß, and δ SARS-CoV-2 variants. Also, our results confirmed that the Bio-SCAN reagents have a long shelf life and can be assembled locally in nonlaboratory and limited-resource settings. Furthermore, the Bio-SCAN platform is compatible with the nucleic acid quick extraction protocol. Our results highlight the potential of Bio-SCAN as a promising point-of-care diagnostic platform that can facilitate low-cost mass screening for SARS-CoV-2.

Rapid single-molecule detection of COVID-19 and MERS antigens via nanobody-functionalized organic electrochemical transistors.

The coronavirus disease 2019 (COVID-19) pandemic has highlighted the need for rapid and sensitive protein detection and quantification in simple and robust formats for widespread point-of-care applications. Here, we report on nanobody-functionalized organic electrochemical transistors with a modular architecture for the rapid quantification of single-molecule-to-nanomolar levels of specific antigens in complex bodily fluids. The sensors combine a solution-processable conjugated polymer in the transistor channel and high-density and orientation-controlled bioconjugation of nanobody-SpyCatcher fusion proteins on disposable gate electrodes. The devices provide results after 10 min of exposure to 5 µl of unprocessed samples, maintain high specificity and single-molecule sensitivity in human saliva and serum, and can be reprogrammed to detect any protein antigen if a corresponding specific nanobody is available. We used the sensors to detect green fluorescent protein, and severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) and Middle East respiratory syndrome coronavirus (MERS-CoV) spike proteins, and for the COVID-19 screening of unprocessed clinical nasopharyngeal swab and saliva samples with a wide range of viral loads.

SARS-CoV-2 genetic diversity and variants of concern in Saudi Arabia.

INTRODUCTION: In December 2019, a new severe acute respiratory syndrome coronavirus, SARS-CoV-2, emerged in China, causing coronavirus disease 2019. The present study investigated genetic profiles and variations of SARS-CoV-2 distributed in different regions of Saudi Arabia to begin to understand the pathogenesis and transmission of SARS-CoV-2 in this country and analyzed associations of these variations with host factors. METHODOLOGY: In total, 774 SARS-CoV-2 genomic sequences obtained and annotated by the Global Initiative on Sharing All Influenza Data (GISAID) were captured and analyzed. RESULTS: The most common SARS-CoV-2 clades in Saudi Arabia were GH followed by O, GR, G, and S. Statistically significant associations were detected between clades and patient outcome. Age, as a host factor, was significantly associated with many variables, including virus geographical location, clade, and patient outcome. The most common variants detected were the NSP12_P323L mutation 94.9%, followed by the D614G mutation (76%) and the NS3_Q57H mutation (71.4%). The concerned variants B.1.1.7, B.1.351, and P.1 were not detected in our population. D614G was associated with higher morbidities than the wild-type virus, including higher rates of death and hospitalization. The NS3_Q57H mutation was the only variant associated with better patient outcome than the wild type. Risk of death was highest with the NSP12_P323L mutation (OR = 1.84; 95% CI = 0.37-9.30) and lowest with the NS3_Q57H mutation (OR = 0.43; 95% CI = 0.25-0.727). CONCLUSIONS: SARS-CoV-2 has evolved uniquely and independently in Saudi Arabia. Our findings provide evidence to begin linking the evolutionary implications to host factors and their effects on the virus severity and transmission.

Human papillomaviruses: The cervical cancer saga in developing countries.

Cervical cancer is the fourth most common cancer affecting women, with worldwide annual incidence and mortality rates of 528,000 and 266,000, respectively, according to the World Health Organization. It is well established that cervical cancer is predominantly caused by a persistent human papillomavirus (HPV) infection of cervical cells. Increasing numbers of studies have investigated HPV and cervical cancer, contributing greatly to the global knowledge and unraveling some of the critical questions regarding HPV transmission, infection, and prevention. However, despite these studies, our knowledge is far from complete and much remains to be discovered. Although molecular detection and HPV prophylactic approaches have greatly advanced in recent years, approximately 85% of the global burden of mortality from cervical cancer still occurs in developing countries. Clinical and molecular epidemiological studies have demonstrated a need for developing countries in general to adopt cervical screening and vaccination programs. However, studies examining cervical cancer screening modalities and HPV prevalence as well as whether HPV vaccination programs should be implemented are lacking at the national level in some developing countries. Therefore, this review describes the current status of HPV in developing countries, presenting some of the existing challenges in implementing cervical screening and HPV vaccination programs.

Current studies on human papillomavirus in Saudi Arabia.

Human papillomavirus (HPV) infection is a significant etiological factor and an important prognosticator in cervical cancer. Indeed, researchers worldwide have confirmed these roles for high-risk HVPs in over 70% of cervical cancer cases. According to the World Health Organization, approximately 561,200 new cancer cases (5.2% of all new cancers) are attributed to HPV infection. Over 120 types of HPV are classified further as either low-risk HPV (LR-HPV) or high-risk HPV (HR-HPV) based on their oncological potential of transforming cells. The LR-HPV types cause benign hyperproliferative lesions (i.e. genital warts) while the HR-HPV types are strongly associated with premalignant and malignant cervical lesions. Data on the prevalence of HPV, survival of infected patients, and mortality rate are scarce in Saudi Arabia. The unsubstantiated assumption of a low prevalence of HPV in Saudi Arabia has contributed to limiting HPV research in this conservative country. Therefore, the goal of this review is to shed light on the current HPV research being conducted and the prevalence of HPV in Saudi Arabia.