Effect of heat inactivation on the detection of the RNA of SARS-CoV-2 by RT-qPCR

SARS-CoV-Z treated at 56 degrees C for 30 min can be inactivated effectively. However, the effect of heat treatment on subsequent detection of the RNA of SARS-CoV-2 by real-time reverse transcription-quantitative polymerase Chain reaction (RT-qPCR) has not been reported. We filled this knowledge gap in present study. We used five SARS'CoV-Z-positive throat swabs. Each throat swab was divided into four parts and assigned to a group: control;56 degrees C for 30 min;56 degrees C for 45 min;56 degrees C for 60 min. After heat treatment, SARS-CoV-Z RNA was extracted and detected by RT-qPCR (absolute quantitation using a standard curve). We found that SARS- CoV-Z RNA was reduced by ~40% after treatment at 56 degrees C for 30 ' 60 min. There was no significant difference (P > 0.05 for all) in the test results between the treatment groups (56 degrees C for 30 min;56 degrees C for 45 min;56 degrees C for 60 min). Our study suggested that SARS-CoV-Z specimens could be inactivated at 56 degrees C for 30 min, before RNA extraction and RT-qPCR detection, which could protect the safety of personnel and the environment during testing. Heat inactivation had a limited effect upon RT-qPCR detection but it should be used with caution if the specimen result is near the critical value.

Profiling SARS-CoV-2 Infection by High-Throughput Shotgun Proteomics.

A comprehensive cartography of viral and host proteins expressed during the different stages of SARS-CoV-2 infection is key to decipher the molecular mechanisms of pathogenesis. For the most detailed analysis, proteins should be first purified and then proteolyzed with trypsin in the presence of detergents. The resulting peptide mixtures are resolved by reverse phase ultrahigh pressure liquid chromatography and then identified by a high-resolution tandem mass spectrometer. The thousands of spectra acquired for each fraction can then be assigned to peptide sequences using a relevant protein sequence database, comprising viral and host proteins and potential contaminants from the growth medium or from the operator. The peptides are evidencing proteins and their intensities are used to infer the abundance of their corresponding proteins. Data analysis allows for highlighting the viral and host proteins dynamics. Here, we describe the sample preparation method adapted to profile SARS-CoV-2 -infected cell models, the shotgun proteomics pipeline to record experimental data, and the workflow for data interpretation to analyze infection-induced proteomic changes in a time-resolved manner.

Optimization of Soybean Protein Extraction Using By-Products from NaCl Electrolysis as an Application of the Industrial Symbiosis Concept

Featured ApplicationThis study shows the use of a by-product from the manufacture of a novel antiseptic/disinfectant (HOCl) to obtain a protein isolate from defatted soybean flour (a co-product from the soybean oil industry);an optimization process was carried out to create an industrial symbiosis.Defatted soybean flour is generated during the oil extraction process of soybean, and it has a protein content of ~50%. On the other hand, an alkaline solution of NaOH is produced during the electrolysis process of NaCl in a novel method used to make a potent disinfectant/antiseptic (HOCl). In the present work, we suggest using these two products to produce soy protein isolate (SPI), aiming to create an industrial symbiosis. A Box–Behnken experimental design was executed, and a surface response analysis was performed to optimize temperature, alkaline solution, and time used for SPI extraction. The SPI produced at optimal conditions was then characterized. The experimental results fit well with a second-order polynomial equation that could predict 93.15% of the variability under a combination of 70 °C, alkaline solution 3 (pH 12.68), and 44.7 min of the process. The model predicts a 49.79% extraction yield, and when tested, we obtained 48.30% within the confidence interval (46.66–52.93%). The obtained SPI was comparable in content and structure with a commercial SPI by molecular weight and molecular spectroscopy characterization. Finally, the urease activity (UA) test was negative, indicating no activity for trypsin inhibitor. Based on the functional properties, the SPI is suitable for food applications.