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Optimized and scalable synthesis of magnetic nanoparticles for RNA extraction in response to developing countries' needs in the detection and control of SARS-CoV-2.

ABSTRACT: Ecuador is one of the most affected countries, with the coronavirus disease 2019 (COVID-19) infection, in Latin America derived from an ongoing economic crisis. One of the most important methods for COVID-19 detection is the use of techniques such as real time RT-PCR based on a previous extraction/purification of RNA procedure from nasopharyngeal cells using functionalized magnetic nanoparticles (MNP). This technique allows the processing of?~?10,000 tests per day in private companies and around hundreds per day at local Universities guaranteeing to reach a wide range of the population. However, the main drawback of this method is the need for specialized MNP with a strong negative charge for the viral RNA extraction to detect the existence of the SARS-CoV-2 virus. Here we present a simplified low cost method to produce 10 g of nanoparticles in 100 mL of solution that was scaled to one litter by parallelizing the process 10 times in just two days and allowing for the possibility of making?~?50,000 COVID-19 tests. This communication helps in reducing the cost of acquiring MNP for diverse biomolecular applications supporting developing country budgets constraints and chemical availability specially during the COVID-19 International Health Emergency.

SUBMITTER: Chacon-Torres JC

PROVIDER: S-EPMC7642403 | biostudies-literature | 2020 Nov

REPOSITORIES: biostudies-literature

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Optimized and scalable synthesis of magnetic nanoparticles for RNA extraction in response to developing countries' needs in the detection and control of SARS-CoV-2.

Chacón-Torres Julio C JC Reinoso C C Navas-León Daniela G DG Briceño Sarah S González Gema G

Scientific reports 20201104 1

Ecuador is one of the most affected countries, with the coronavirus disease 2019 (COVID-19) infection, in Latin America derived from an ongoing economic crisis. One of the most important methods for COVID-19 detection is the use of techniques such as real time RT-PCR based on a previous extraction/purification of RNA procedure from nasopharyngeal cells using functionalized magnetic nanoparticles (MNP). This technique allows the processing of ~ 10,000 tests per day in private companies and around ...[more]

PMID: 33149153

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Project description:The recent outbreak of the novel corona virus disease 2019 (COVID-19) has been made a serious global impact due to its high infectivity and severe symptoms. The Severe Acute Respiratory Syndrome (SARS-CoV-2) RNA extraction is considered as one of the most important steps in COVID-19 detection. Several commercially available kits and techniques are currently being used for specific extraction of SARS-CoV-2 RNA. However, such methods are time consuming and expensive due to the requirement of trained labors, and several chemical reagents. To overcome the mentioned limitations, magnetic RNA adsorption methodology of glycine functionalized iron oxide nanoparticles (GNPs) was established. It showed an efficient potential in SARS-CoV-2 RNA extraction due to pH responsive nature of GNPs. The highly magnetic pH responsive GNPs were synthesized by one-pot co-precipitation method. Random morphology and average 20 nm size of GNPs were denoted by Transmission Electron Microscopy (TEM). X-ray diffractometer (XRD) showed the crystalline magnetite nature. Fourier transform infrared spectroscopy (FT-IR) and UV-visible spectrometry confirmed the presence of glycine on the surface of magnetic nanoparticles. Furthermore, the magnetic nature and thermal properties of GNPs were examined by vibrating sample magnetometer (VSM) and thermo-gravimetric analysis (TGA), respectively. In this study, glycine performed the role of RNA adsorbent. The adsorption of RNA onto the surface of GNPs was achieved in acidic medium (pH 6). In contrary, the elution of RNA from the surface of GNPs was achieved in basic medium (pH 8). The purity of obtained RNA was analyzed by UV-visible spectrometry. Further, the obtained RNA was examined for the presence of SARS-CoV-2 specific Envelope (E), RNA dependent RNA polymerase (RDRP) and Nucleocapsid (N) genes using an RT-PCR analysis. It showed the sudden rise in amount of these genes after 25 cycles of RT-PCR and hence indicated the efficient RNA extraction by GNPs. Agarose gel electrophoresis was used for validation of the quantity and quality of RNA extracted from SARS-CoV-2 patient's sample. The reusability studies of GNPs were performed by monitoring the repeated use of GNPs for SARS-CoV-2 RNA extraction. This method possesses potential role in the field of disease diagnosis. The extraction results of RNA from SARS-CoV-2 patient's sample indicated that the GNPs have an outstanding property over the current existing extraction protocols. It leads to the new advancements in extraction and detection of RNA.Graphical abstractGraphical abstract of the pH responsive SARS-CoV-2 RNA extraction by using glycine functionalized magnetic iron oxide nanoparticles (GNPs) which were prepared by modified cost effective one pot chemical synthesis method. Prepared GNPs were characterized by XRD, FT-IR and UV-Visible spectrometry, Scanning electron microscopy (SEM) and Transmission electron microscopy (TEM). Glycine present on the surface of nanoparticles (NPs) played an important role in pH responsive RNA extraction procedure. When nanoparticles added in acidic (pH < 7) medium, glycine gained positive surface charge hence overall surface charge of NPs became positive. Thereby SARS-CoV-2 RNA adsorption/binding occurred on the surface of GNP. Later, the RNA-GNP complex was separated by an external magnet. Separated complex was added in basic (pH > 7) medium to elute RNA from GNP. This phenomenon occurred due to surface negative charge of glycine that caused charge repulsion with RNA. Eluted RNA was examined qualitatively and quantitatively by RT-PCR, nanodrop technique and agarose gel electrophoresis. Results were compared with kit based extracted RNA.Supplementary informationThe online version contains supplementary material available at 10.1007/s10853-022-07464-6.

Project description:Wastewater-based epidemiology (WBE) has emerged as a valuable approach to forecast disease outbreaks in developed countries with a centralized sewage infrastructure. On the other hand, due to the absence of well-defined and systematic sewage networks, WBE is challenging to implement in developing countries like Bangladesh where most people live in rural areas. Identification of appropriate locations for rural Hotspot Based Sampling (HBS) and urban Drain Based Sampling (DBS) are critical to enable WBE based monitoring system. We investigated the best sampling locations from both urban and rural areas after evaluating Bangladesh's sanitation infrastructure for forecasting COVID-19 prevalence. A total of 168 wastewater samples were collected from 14 districts of Bangladesh during each of two peak pandemic seasons. RT-qPCR commercial kits were used to target ORF1ab and N genes. The presence of SARS-CoV-2 genetic materials was found in 98% (165/168) of samples in the first round and 95% (160/168) of samples in the second round, respectively. Although wastewater effluents from both the marketplace and isolation center drains were found with the highest amount of genetic materials according to the mixed model, quantifiable SARS-CoV-2 RNAs were also identified in other four sampling sites. Hence, wastewater samples of the marketplace in rural areas and isolation centers in urban areas can be considered as appropriate sampling sites to detect contagion hotspots. This is the first complete study to detect SARS-CoV-2 genetic components in wastewater samples collected from rural and urban areas for monitoring the COVID-19 pandemic. The results based on the study revealed correlation between viral copy numbers in wastewater samples and SARS-CoV-2 positive cases reported by Directorate General of Health Services (DGHS) as part of the national surveillance program for COVID-19 prevention. The findings of this study will help in setting strategies and guidelines for the selection of appropriate sampling sites, which will facilitate in developing comprehensive wastewater-based epidemiological systems for surveillance of rural and urban areas of low-income countries with inadequate sewage infrastructure.

Dataset Information

Optimized and scalable synthesis of magnetic nanoparticles for RNA extraction in response to developing countries' needs in the detection and control of SARS-CoV-2.

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Optimized and scalable synthesis of magnetic nanoparticles for RNA extraction in response to developing countries' needs in the detection and control of SARS-CoV-2.

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