Thermal stability of diagnostic targets plasmodium falciparum histidine rich protein II and plasmodium lactate dehydrogenase in rapid detection.

During the COVID-19 epidemic, blood samples are usually processed at 56 to attenuate the virus before pathogen detection. 71 blood samples of malaria patients reported by Shanghai Center for Disease Control and Prevention in 2017-2019 were collected, including 38 with Plasmodium falciparum infection, 8 P. malariae, 11 P. ovale and 14 P. vivax. The effect of inactivation on the thermal stability of P. falciparum histidine rich protein II (PfHRPII) and Plasmodium lactate dehydrogenase (pLDH) in blood samples was assessed before and after incubation at 56 for 30 min using the rapid diagnostic test (RDT) kit. The results showed that among the 38 P. falciparum T1-positive (PfHRPII) blood samples before heat treatment, 35 samples remained to be T1-positive (92.11%, 35/38, chi2=3.123, P>0.05) after heat treatment;while 54 blood samples (26 P. falciparum, 6 P. vivax, 10 P. ovale and 12 P. vivax) that were T2-positive (pLDH) before heat treatment turned to be T2-negative (positive rate 0, 0/54, chi2=87.755, P<0.01) after heat treatment. It was demonstrated that PfHRPII is stable during incubation at 56 for 30 min, while pLDH is unstable and degraded or inactivated during the heating. Therefore, the detection results of P. falciparum will not be affected by RDT, but diagnosis of the parasites other than P. falciparum in blood samples may be missed.Copyright © 2021, National Institute of Parasitic Diseases. All rights reserved.

Molecularly imprinted polymer-based nanodiagnostics for clinically pertinent bacteria and virus detection for future pandemics

There seems to be a growing curiosity for utilizing MIPs to recognize molecules that can be applied in numerous fields, such as biomimetic antibodies, detection of viruses and bacteria, the broad range of sensing devices, etc., owing to its scalability and economic viability. MIPs have higher thermal and chemical stability, delivering a promising potential for recognizing bacteria and viruses. The bacteria and virus imprinted polymer exhibit elongated product life-time, reproducible fabrication, robustness, reusability, sensitivity, and high target selectivity. Moreover, the MIPs could give consistent screening along with negligible false positive/negative outcomes, which is vital for the control and prevention of viral and bacterial infections. In the viral and bacterial imprinting process, critical aspects, such as compositional complexity, fragility, solubility, and target size, should be systematically evaluated and analytically considered. Although, the application of MIPs has a number of drawbacks and challenges that require solving to develop sensing platforms with high specificity and sensitivity for clinical application. In the present review, current progress and advancement regarding the reasoning and applications of MIPs as recognition molecules in various biosensors for detecting bacteria and viruses and its existing noteworthy challenges along with future perspectives are also reflected.

Recent advances of nanotechnology in COVID 19: A critical review and future perspective

The global anxiety and economic crisis causes the deadly pandemic coronavirus disease of 2019 (COVID 19) affect millions of people right now. Subsequently, this life threatened viral disease is caused due to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). However, morbidity and mortality of infected patients are due to cytokines storm syndrome associated with lung injury and multiorgan failure caused by COVID 19. Thereafter, several methodological advances have been approved by WHO and US-FDA for the detection, diagnosis and control of this wide spreadable communicable disease but still facing multi-challenges to control. Herein, we majorly emphasize the current trends and future perspectives of nano-medicinal based approaches for the delivery of anti-COVID 19 therapeutic moieties. Interestingly, Nanoparticles (NPs) loaded with drug molecules or vaccines resemble morphological features of SARS-CoV-2 in their size (60–140 nm) and shape (circular or spherical) that particularly mimics the virus facilitating strong interaction between them. Indeed, the delivery of anti-COVID 19 cargos via a nanoparticle such as Lipidic nanoparticles, Polymeric nanoparticles, Metallic nanoparticles, and Multi-functionalized nanoparticles to overcome the drawbacks of conventional approaches, specifying the site-specific targeting with reduced drug loading and toxicities, exhibit their immense potential. Additionally, nano-technological based drug delivery with their peculiar characteristics of having low immunogenicity, tunable drug release, multidrug delivery, higher selectivity and specificity, higher efficacy and tolerability switch on the novel pathway for the prevention and treatment of COVID 19. © 2022 The Author(s)

Microwave treatment to optimize physicochemical properties of modified Busil (Xanthosoma sagittifolium) starch

COVID-19 pandemic encourages the utilization of local food sources to ensure food availability. Busil (Xanthosoma sagittifolium) was readily available and affordable in Banjarnegara Regency in the Province of Central Java in Indonesia. However, the busil starch utilization was still rare due to the low functional properties of the native busil starch. The objective of this study was to explore busil starch physicochemical characterization enhancement after microwave irradiation treatment, especially on the stability of heat processing. This research was conducted in two steps. First, microwave treatment (with a variation of energy and irradiation time) of native busil starch (NBS), and the second was modified busil starch (MBS) physicochemical characterization. A rise in amylose was observed on MBS. SEM analysis was shown MBS granules are breakdown. Through viscosity, final viscosity, setback viscosity, peak time, and the pasting temperature of MBS generally were increased. Meanwhile, peak viscosity and breakdown viscosity of MBS was decreased. Thermal properties of MBS like onset (To), peak (Tp), and conclusion (Tc) temperatures were also increased. The degree of whiteness index (DW) of MBS was decreased. FTIR analysis has shown that microwave treatment did not cause functional group alteration. XRD analysis has also demonstrated no change in the diffraction pattern but a slight change in the crystallinity index. Generally, microwave treatment leads to MBS thermal stability and potentially broaden MBS utilization on food processing product.

Design, production and characterization of antiviral proteins targeting SARS-CoV-2

The virus responsible for the current COVID -19 pandemic is SARS-CoV-2, which has caused >400 million infections and >5 million deaths (as of February 2022). Despite vaccination efforts, there is still an urgent need to develop strategies to control infection and treat patients. One of the proteins bound to the viral membrane is the spike (S) protein, which consists of two subunits: S1, which contains a receptor-binding domain (RBD) responsible for binding to the host cell receptor, and S2, which facilitates membrane fusion between the viral and host cell membranes, previously published in: Jackson CB et al. (2018) Nat Rev Mol Cell Biol 23, 3-20. Thus, this protein is primarily responsible for the ability of the virus to enter host cells, making it one of the most promising therapeutic targets of coronavirus, previously published in: Cao L et al. (2020) Science 6515, 426- 431. The aim of this work was to design and produce antiviral proteins that could prevent the interaction between the two proteins and thus block infection by binding to the RBD region and blocking its interaction with the host receptor, angiotensin converting enzyme-2 (ACE2) protein. First, several antiviral proteins were computationally designed using the Rosetta program based on the interactions between ACE2 and the RBD. Next, six molecular dynamics simulations (MD) of 1 ls of three candidates were performed to test their interaction with the RBD. This was followed by experimental validation after expression and purification of the three candidates. The secondary structure and thermostability of these proteins were tested by far-UV circular dichroism spectropolarimetry. Surface plasmon resonance was used to evaluate the affinity of each candidate for RBD. Neutralization assays were performed to investigate the neutralization ability of the proteins. The experimental results show that one of the developed proteins is a promising therapeutic approach that will be further improved in the future.

Thermostable vaccines: an innovative concept in vaccine development.

INTRODUCTION: Vaccines represent he most common and safer ways of combating infectious diseases. Loss of potency owing to thermal denaturation or degradation of almost all the vaccines necessitates their storage, transportation, and final dissemination under refrigerated conditions. However, maintenance of a continuous cold chain raises the costs of vaccines significantly. A large number of life-saving vaccines are discarded before their application owing to exposure to sub-optimum temperatures. Therefore, there is a pressing need for the development of a thermostable vaccine with a long shelf life at ambient temperature. AREAS COVERED: A literature search was performed to compile a list of different vaccines, and their storage and handling conditions. Similarly, a separate list was prepared for different coronavirus vaccines. A literature survey was also performed to look at different approaches undertaken globally to address the issue of the cold-chain problem. We emphasized the importance of yeast cells in the development of thermostable vaccines. In the end, we discussed why thermostable vaccines are required, not only in resource-poor countries but also for resource-rich countries . EXPERT OPINION: Temperature change can severely impact the stability of various life-saving vaccines. Therefore, there is a pressing need for the development of thermostable vaccines with a long shelf lives.

A flexible, thermostable nanostructured lipid carrier platform for RNA vaccine delivery.

Current RNA vaccines against severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) are limited by instability of both the RNA and the lipid nanoparticle delivery system, requiring storage at -20°C or -70°C and compromising universally accessible vaccine distribution. This study demonstrates the thermostability and adaptability of a nanostructured lipid carrier (NLC) delivery system for RNA vaccines that has the potential to address these concerns. Liquid NLC alone is stable at refrigerated temperatures for ≥1 year, enabling stockpiling and rapid deployment by point-of-care mixing with any vaccine RNA. Alternatively, NLC complexed with RNA may be readily lyophilized and stored at room temperature for ≥8 months or refrigerated temperature for ≥21 months while still retaining the ability to express protein in vivo. The thermostability of this NLC/RNA vaccine delivery platform could significantly improve distribution of current and future pandemic response vaccines, particularly in low-resource settings.

Thermostable designed ankyrin repeat proteins (DARPins) as building blocks for innovative drugs.

Designed ankyrin repeat proteins (DARPins) are antibody mimetics with high and mostly unexplored potential in drug development. By using in silico analysis and a rationally guided Ala scanning, we identified position 17 of the N-terminal capping repeat to play a key role in overall protein thermostability. The melting temperature of a DARPin domain with a single full-consensus internal repeat was increased by 8 °C to 10 °C when Asp17 was replaced by Leu, Val, Ile, Met, Ala, or Thr. We then transferred the Asp17Leu mutation to various backgrounds, including clinically validated DARPin domains, such as the vascular endothelial growth factor-binding domain of the DARPin abicipar pegol. In all cases, these proteins showed improvements in the thermostability on the order of 8 °C to 16 °C, suggesting the replacement of Asp17 could be generically applicable to this drug class. Molecular dynamics simulations showed that the Asp17Leu mutation reduces electrostatic repulsion and improves van-der-Waals packing, rendering the DARPin domain less flexible and more stable. Interestingly, this beneficial Asp17Leu mutation is present in the N-terminal caps of three of the five DARPin domains of ensovibep, a SARS-CoV-2 entry inhibitor currently in clinical development, indicating this mutation could be partly responsible for the very high melting temperature (>90 °C) of this promising anti-COVID-19 drug. Overall, such N-terminal capping repeats with increased thermostability seem to be beneficial for the development of innovative drugs based on DARPins.

Next-Generation COVID-19 Vaccines Should Take Efficiency of Distribution into Consideration.

The dire need for safe and effective coronavirus disease (COVID-19) vaccines is met with many vaccine candidates being evaluated in pre-clinical and clinical studies. The COVID-19 vaccine candidates currently in phase 3 or phase 2/3 clinical trials as well as those that recently received emergency use authorization (EUA) from the United States Food and Drug Administration (FDA) and/or other regulatory agencies worldwide require either cold (i.e., 2-8°C) or even freezing temperatures as low as -70°C for storage and distribution. Thus, existing cold chain will struggle to support both the standard national immunization programs and COVID-19 vaccination. The requirement for cold chain is now a major challenge towards worldwide rapid mass vaccination against COVID-19. In this commentary, we stress that thermostabilizing technologies are available to enable cold chain-free vaccine storage and distribution, as well as potential needle-free vaccination. Significant efforts on thermostabilizing technologies must now be applied on next-generation COVID-19 vaccines for more cost-effective worldwide mass vaccination and COVID-19 eradication.

Never let a crisis go to waste: Repurposing independent research projects to enhance students' critical thinking skills.

The recent lockdown of laboratories in the COVID-19 era has negatively impacted many lab-based courses for undergraduate students; however, research project classes have been hardest hit. Unlike the lab-based lecture courses, where many of the lab instructors have resorted to virtual experiments and videos to substitute for actual lab experiments, senior year independent research courses, which are supposed to involve real lab-based research, are struggling to find an appropriate solution. A possible solution to cases where the coronavirus pandemic lockdown has led to no or little student-generated lab data are to shift the focus of these research project courses such that they are taught as mentor-guided critical thinking exercises, using the primary literature. As an example, the lack of complete understanding for the molecular basis of protein thermostability can be used to promote such higher-level thinking skills.