Dielectric Spectroscopy: Revealing the True Colors of Biological Matter

Accurate characterization of biological matter, for example, in tissue, cells, and biological fluids, is of high importance. For example, early and correct detection of abnormalities, such as cancer, is essential as it enables early and effective type-specific treatment, which is crucial for mortality reduction [1]. Moreover, it is imperative to investigate the effectiveness and toxicity of pharmaceutical treatments before administration in clinical practice [2]. However, biological matter characterization still faces many challenges. State-of-the-art imaging and characterization methods have drawbacks, such as the requirement to attach difficult-to-find and costly labels to the biological target (e.g., COVID-19 rapid tests), expensive equipment (e.g., magnetic resonance imaging), low accuracy (e.g., ultrasound), use of ionizing radiation (e.g., X-rays), and invasiveness [3]. The characterization of biological matter using microwave (μW), millimeter-wave (mmW), and terahertz (THz) spectroscopy is a promising alternative: it is label-free, does not require ionizing radiation, and can be noninvasive. Moreover, there is a significant difference in how different biological materials absorb, reflect, and transmit electromagnetic (EM) waves [4] that is due to the difference in their dielectric properties. The dielectric properties are described by the frequency-dependent material parameter called the complex permittivity f, which expresses how the material responds to an external oscillating electric field. The complex permittivity of a material determines how the material absorbs, reflects, and transmits EM waves at different frequencies (Figure 1). Since each biological material's permittivity spectrum is different, it acts as an EM fingerprint. A material's complex permittivity can be calculated from the reflection and transmission of EM waves through the material, described by the S-parameters, which can be measured using a vector network analyzer (VNA) transmitting and receiving EM waves over a range of frequencies. The amplitude and phase of the transmitted and reflected EM waves at different frequencies are influenced by different underlying biological effects at different scales. That causes the entire spectrum to provide information from the supracellular to the molecular and even atomic scale. © 2000-2012 IEEE.

Stability of SARS-CoV-2 in Biological Fluids of Animals.

Since its first emergence in 2019, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has continued to evolve genetically, jump species barriers, and expand its host range. There is growing evidence of interspecies transmission including infection of domestic animals and widespread circulation in wildlife. However, knowledge of SARS-CoV-2 stability in animal biological fluids and their role in transmission is still limited as previous studies focused on human biological fluids. Therefore, this study aimed to determine the SARS-CoV-2 stability in biological fluids from three animal species, cats, sheep and white-tailed deer (WTD). Saliva, feces, 10% fecal suspensions, and urine of cats, sheep, and WTD were mixed with a known concentration of virus and incubated under indoor and three different climatic conditions. Our results show that the virus was stable for up to 1 day in the saliva of cats, sheep, and WTD regardless of the environmental conditions. The virus remained infectious for up to 6 days in feces and 15 days in fecal suspension of WTD, whereas the virus was rather unstable in cat and sheep feces and fecal suspensions. We found the longest survival of SARS-CoV-2 in the urine of cats, sheep, and WTD. Furthermore, side-by-side comparison with different SARS-CoV-2 strains showed that the Alpha, Delta, and Omicron variants of concern were less stable than the ancestral Wuhan-like strain in WTD fecal suspension. The results of our study provide valuable information for assessing the potential role of various animal biological fluids in SARS-CoV-2 transmission.

Ancestral Lineage of SARS-CoV-2 Is More Stable in Human Biological Fluids than Alpha, Beta, and Omicron Variants of Concern.

SARS-CoV-2 is a zoonotic virus first identified in 2019, and has quickly spread worldwide. The virus is primarily transmitted through respiratory droplets from infected persons; however, the virus-laden excretions can contaminate surfaces which can serve as a potential source of infection. Since the beginning of the pandemic, SARS-CoV-2 has continued to evolve and accumulate mutations throughout its genome leading to the emergence of variants of concern (VOCs) which exhibit increased fitness, transmissibility, and/or virulence. However, the stability of SARS-CoV-2 VOCs in biological fluids has not been thoroughly investigated. The aim of this study was to determine and compare the stability of different SARS-CoV-2 strains in human biological fluids. Here, we demonstrate that the ancestral strain of the Wuhan-like lineage A was more stable than the Alpha VOC B.1.1.7, and the Beta VOC B.1.351 strains in human liquid nasal mucus and sputum. In contrast, there was no difference in stability among the three strains in dried biological fluids. Furthermore, we also show that the Omicron VOC B.1.1.529 strain was less stable than the ancestral Wuhan-like strain in liquid nasal mucus. These studies provide insight into the effect of the molecular evolution of SARS-CoV-2 on environmental virus stability, which is important information for the development of countermeasures against SARS-CoV-2. IMPORTANCE Genetic evolution of SARS-CoV-2 leads to the continuous emergence of novel virus variants, posing a significant concern to global public health. Five of these variants have been classified to date into variants of concern (VOCs); Alpha, Beta, Gamma, Delta, and Omicron. Previous studies investigated the stability of SARS-CoV-2 under various conditions, but there is a gap of knowledge on the survival of SARS-CoV-2 VOCs in human biological fluids which are clinically relevant. Here, we present evidence that Alpha, Beta, and Omicron VOCs were less stable than the ancestral Wuhan-like strain in human biological fluids. Our findings highlight the potential risk of contaminated human biological fluids in SARS-CoV-2 transmission and contribute to the development of countermeasures against SARS-CoV-2.

Quantitative analysis of favipiravir and hydroxychloroquine as FDA-approved drugs for treatment of COVID-19 using synchronous spectrofluorimetry: application to pharmaceutical formulations and biological fluids.

Coronavirus disease 2019 (COVID-19) is a contagious viral infection caused by coronavirus 2 (SARS-CoV-2) that causes severe acute respiratory syndrome. It has ravaged several countries and burdened many healthcare systems. As the process of authorizing a novel treatment for human use is extensive and involves multiple phases to obtain safety information and identify potential concerns. Therefore, the fastest and easiest choice was to use United States Food and Drug Administration (US FDA)-approved drugs such as favipiravir and hydroxychloroquine. For the simultaneous estimation of both medications, a simple synchronous spectrofluorimetric approach was established in which both drugs were measured at 372 and 323 nm, respectively in the presence of each other without interference at Δλ 60 nm. The effect of various experimental conditions on synchronous fluorescence intensities were thoroughly investigated and optimized. The maximum synchronous fluorescence intensities were obtained at pH 5.4 using acetate buffer (0.2 M, 0.5 ml) and ethanol as a diluent. Excellent linearity ranges were obtained using 1.0-18.0 ng/ml and 10.0-120.0 ng/ml for favipiravir and hydroxychloroquine, respectively. The approach exhibited high sensitivity with detection limits down to 0.25 ng/ml and 1.52 ng/ml and quantitation limits down to 0.77 ng/ml and 4.62 ng/ml, respectively. Spiking human plasma samples with the studied drugs yielded high % recoveries, allowing a significant bioanalytical application. Moreover, the method was validated according to International Conference on Harmonization guidelines and further applied to commercial pharmaceutical preparations with good results.

Highly Sensitive and Selective Sensing Probe for Determination of Anti-Covid-19 Remdesvir: Application to Pharmaceutical Dosage Form and Biological Fluids

We reported the first investigational electrochemical study for Remdesvir (REM). REM is a promising antiviral agent used recently for the treatment of the most dangerous pandemic disease nowadays (COVID-19). Anionic surfactant, silica nanoparticles, and multiwall carbon nanotubes modified carbon paste (SDS/SiO2/MWCNT/CPE) sensor was designed to introduce our approach. The results revealed irreversible diffusion oxidative reaction of REM with two well-defined peaks (E1/V = 1.19, E2/V = 1.35) in 0.1 M phosphate buffer of pH 6 using differential pulse (DP) voltammetry. A linear relationship between the peak current and the drug concentration was established over the concentration range of 1.66 × 10-7-3.52 × 10-6 M (100-200 ng ml-1) with a limit of detection (LOD) of 4.80 × 10-8 M and limit of quantitation (LOQ) of 8.0 × 10-8 M and mean % recovery ± % RSD of 99.05 ± 1.94. The proposed method succeeded in the determination of the drug in its pharmaceutical dosage form, in human plasma with and human urine samples. Finally, the method was validated according to ICH guidelines and FDA guidance for the determination of the drug in biological fluids. The developed data was found to be in good agreement with a validated reported method. © 2022 The Electrochemical Society ("ECS").

Electroanalytical Methods for Determination of Antiviral Drugs in Pharmaceutical Formulation and Biological Fluids: A Review

Viruses can infect all kinds of life forms, from humans, animals and plants to microorganisms, including bacteria and archaea, and can cause serious illness, including a pandemic. Antivirals act as a blocking mechanism against the viral replication cycle at various stages to treat viral infections. Currently, antiviral therapy is available for a limited number of infections and is mainly used to minimize symptoms, further spread, and shorten the duration of the illness. For this reason, it is very important to develop fast, precise and reliable methods to analyze antiviral drugs from dosage forms or biological samples. Electroanalytical methods have been used frequently in the analysis of antiviral drugs comprising of different structures. Electrochemical methods have an important place among analytical methods with their unique features such as specificity, high sensitivity, reliability, economical, easy sample preparation, and no pre-analysis process. The purpose of this review article is to compile published electrochemical methods for the determination of antiviral drugs in pharmaceutical preparations and biological fluids. In addition to these applied methods, the types of electrodes used, and the materials used in the modification and designation of these electrodes are classified and briefly explained with their limitations.

Beyond liquid biopsy: Toward non-invasive assays for distanced cancer diagnostics in pandemics.

Liquid biopsy technologies have seen a significant improvement in the last decade, offering the possibility of reliable analysis and diagnosis from several biological fluids. The use of these technologies can overcome the limits of standard clinical methods, related to invasiveness and poor patient compliance. Along with this there are now mature examples of lab-on-chips (LOC) which are available and could be an emerging and breakthrough technology for the present and near-future clinical demands that provide sample treatment, reagent addition and analysis in a sample-in/answer-out approach. The possibility of combining non-invasive liquid biopsy and LOC technologies could greatly assist in the current need for minimizing exposure and transmission risks. The recent and ongoing pandemic outbreak of SARS-CoV-2, indeed, has heavily influenced all aspects of life worldwide. Ordinary tasks have been forced to switch from "in presence" to "distanced", limiting the possibilities for a large number of activities in all fields of life outside of the home. Unfortunately, one of the settings in which physical distancing has assumed noteworthy consequences is the screening, diagnosis and follow-up of diseases. In this review, we analyse biological fluids that are easily collected without the intervention of specialized personnel and the possibility that they may be used -or not-for innovative diagnostic assays. We consider their advantages and limitations, mainly due to stability and storage and their integration into Point-of-Care diagnostics, demonstrating that technologies in some cases are mature enough to meet current clinical needs.

A Review on Analytical Strategies for the Assessment of Recently Approved Direct Acting Antiviral Drugs.

Human beings are in dire need of developing an efficient treatment against fierce viruses like hepatitis C virus (HCV) and Coronavirus (COVID-19). These viruses have already caused the death of over two million people all over the world. Therefore, over the last years, many direct-acting antiviral drugs (DAADs) were developed targeting nonstructural proteins of these two viruses. Among these DAADs, several drugs were found more effective and safer than the others as sofosbuvir, ledipasvir, grazoprevir, glecaprevir, voxilaprevir, velpatasvir, elbasvir, pibrentasvir and remdesivir. The last one is indicated for COVID-19, while the rest are indicated for HCV treatment. Due to the valuable impact of these DAADs, larger number of analytical methods were required to meet the needs of the clinical studies. Therefore, this review will highlight the current approaches, published in the period between 2017 to present, dealing with the determination of these drugs in two different matrices: pharmaceuticals and biological fluids with the challenges of analyzing these drugs either alone, with other drugs, in presence of interferences (pharmaceutical excipients or endogenous plasma components) or in presence of matrix impurities, degradation products and metabolites. These approaches include spectroscopic, chromatographic, capillary electrophoretic, voltametric and nuclear magnetic resonance methods that have been reported during this period. Moreover, the analytical instrumentation and methods used in determination of these DAADs will be illustrated in tabulated forms.

Monitoring COVID-19 Transmission Risks by Quantitative Real-Time PCR Tracing of Droplets in Hospital and Living Environments.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) environmental contamination occurs through droplets and biological fluids released in the surroundings from patients or asymptomatic carriers. Surfaces and objects contaminated by saliva or nose secretions represent a risk for indirect transmission of coronavirus disease 2019 (COVID-19). We assayed surfaces from hospital and living spaces to identify the presence of viral RNA and the spread of fomites in the environment. Anthropic contamination by droplets and biological fluids was monitored by detecting the microbiota signature using multiplex quantitative real-time PCR (qPCR) on selected species and massive sequencing on 16S amplicons. A total of 92 samples (flocked swabs) were collected from critical areas during the pandemic, including indoor (three hospitals and three public buildings) and outdoor surfaces exposed to anthropic contamination (handles and handrails, playgrounds). Traces of biological fluids were frequently detected in spaces open to the public and on objects that are touched with the hands (>80%). However, viral RNA was not detected in hospital wards or other indoor and outdoor surfaces either in the air system of a COVID hospital but only in the surroundings of an infected patient, in consistent association with droplet traces and fomites. Handled objects accumulated the highest level of multiple contaminations by saliva, nose secretions, and fecal traces, further supporting the priority role of handwashing in prevention. In conclusion, anthropic contamination by droplets and biological fluids is widespread in spaces open to the public and can be traced by qPCR. Monitoring fomites can support evaluation of indirect transmission risks for coronavirus or other flu-like viruses in the environment.IMPORTANCE Several studies have evaluated the presence of SARS-CoV-2 in the environment. Saliva and nasopharyngeal droplets can land on objects and surfaces, creating fomites. A suitable indicator would allow the detection of droplets or biofluids carrying the virus. Therefore, we searched for viral RNA and droplets and fomites on at risk surfaces. We monitored by qPCR or next generation sequencing (NGS) droplets through their microbiota. Although the study was performed during the pandemic, SARS-CoV-2 was not significantly found on surfaces, with the only exception of environmental areas near infectious patients. Conversely, anthropic contamination was frequent, suggesting a role for biofluids as putative markers of indirect transmission and risk assessment. Moreover, all SARS-CoV-2-contaminated surfaces showed droplets' microbiota. Fomite monitoring by qPCR may have an impact on public health strategies, supporting prevention of indirect transmission similarly to what is done for other communicable diseases (e.g., influenza and influenza-like infections).