Production and Purification of LTB-RBD: A Potential Antigen for Mucosal Vaccine Development against SARS-CoV-2.

Most of the current SARS-CoV-2 vaccines are based on parenteral immunization targeting the S protein. Although protective, such vaccines could be optimized by inducing effective immune responses (neutralizing IgA responses) at the mucosal surfaces, allowing them to block the virus at the earliest stage of the infectious cycle. Herein a recombinant chimeric antigen called LTB-RBD is described, which comprises the B subunit of the heat-labile enterotoxin from E. coli and a segment of the RBD from SARS-CoV-2 (aa 439-504, carrying B and T cell epitopes) from the Wuhan sequence and the variant of concern (VOC)-delta. Since LTB is a mucosal adjuvant, targeting the GM1 receptor at the surface and facilitating antigen translocation to the submucosa, this candidate will help in designing mucosal vaccines (i.e., oral or intranasal formulations). LTB-RBD was produced in E. coli and purified to homogeneity by IMAC and IMAC-anionic exchange chromatography. The yields in terms of pure LTB-RBD were 1.2 mg per liter of culture for the Wuhan sequence and 3.5 mg per liter for the delta variant. The E. coli-made LTB-RBD induced seric IgG responses and IgA responses in the mouth and feces of mice when subcutaneously administered and intestinal and mouth IgA responses when administered nasally. The expression and purification protocols developed for LTB-RBD constitute a robust system to produce vaccine candidates against SARS-CoV-2 and its variants, offering a low-cost production system with no tags and with ease of adaptation to new variants. The E. coli-made LTB-RBD will be the basis for developing mucosal vaccine candidates capable of inducing sterilizing immunity against SARS-CoV-2.

A Comparative Study between the Three Waves of the Pandemic on the Prevalence of Oropharyngeal Dysphagia and Malnutrition among Hospitalized Patients with COVID-19.

BACKGROUND: The phenotype of patients affected by COVID-19 disease changed between the waves of the pandemic. We assessed the prevalence of oropharyngeal dysphagia (OD), malnutrition (MN), and mortality between the first three waves of COVID-19 patients in a general hospital. METHODS: a prospective observational study between April 2020-May 2021. Clinical assessment for OD was made with the volume-viscosity swallowing test; nutritional assessment was performed consistent with GLIM criteria. A multimodal intervention was implemented in the second and third wave, including (a) texturized diets-fork mashable (1900 kcal + 90 g protein) or pureed (1700 kcal + 75 g protein), (b) oral nutritional supplements (500-600 kcal + 25-30 g protein), and (c) fluid thickening (250 mPa·s or 800 mPa·s). RESULTS: We included 205 patients (69.3 ± 17.6 years) in the 1st, 200 (66.4 ± 17.5 years) in the 2nd, and 200 (72.0 ± 16.3 years;) in the 3rd wave (p = 0.004). On admission, prevalence of OD was 51.7%, 31.3% and 35.1%, and MN, 45.9%, 36.8% and 34.7%, respectively; mortality was 10.7%, 13.6% and 19.1%. OD was independently associated with age, delirium, and MN; MN, with age, OD, diarrhea and ICU admission; mortality, with age, OD and MN. (4) Conclusions: Prevalence of OD, MN and mortality was very high among COVID-19 patients. OD was independently associated with MN and mortality. An early and proactive multimodal nutritional intervention improved patients' nutritional status.

Nanoclays: Promising Materials for Vaccinology.

Clay materials and nanoclays have gained recent popularity in the vaccinology field, with biocompatibility, simple functionalization, low toxicity, and low-cost as their main attributes. As elements of nanovaccines, halloysite nanotubes (natural), layered double hydroxides and hectorite (synthetic) are the nanoclays that have advanced into the vaccinology field. Until now, only physisorption has been used to modify the surface of nanoclays with antigens, adjuvants, and/or ligands to create nanovaccines. Protocols to covalently attach these molecules have not been developed with nanoclays, only procedures to develop adsorbents based on nanoclays that could be extended to develop nanovaccine conjugates. In this review, we describe the approaches evaluated on different nanovaccine candidates reported in articles, the immunological results obtained with them and the most advanced approaches in the preclinical field, while describing the nanomaterial itself. In addition, complex systems that use nanoclays were included and described. The safety of nanoclays as carriers is an important key fact to determine their true potential as nanovaccine candidates in humans. Here, we present the evaluations reported in this field. Finally, we point out the perspectives in the development of vaccine prototypes using nanoclays as antigen carriers.

What Does Plant-Based Vaccine Technology Offer to the Fight against COVID-19?

The emergence of new pathogenic viral strains is a constant threat to global health, with the new coronavirus strain COVID-19 as the latest example. COVID-19, caused by the SARS-CoV-2 virus has quickly spread around the globe. This pandemic demands rapid development of drugs and vaccines. Plant-based vaccines are a technology with proven viability, which have led to promising results for candidates evaluated at the clinical level, meaning this technology could contribute towards the fight against COVID-19. Herein, a perspective in how plant-based vaccines can be developed against COVID-19 is presented. Injectable vaccines could be generated by using transient expression systems, which offer the highest protein yields and are already adopted at the industrial level to produce VLPs-vaccines and other biopharmaceuticals under GMPC-processes. Stably-transformed plants are another option, but this approach requires more time for the development of antigen-producing lines. Nonetheless, this approach offers the possibility of developing oral vaccines in which the plant cell could act as the antigen delivery agent. Therefore, this is the most attractive approach in terms of cost, easy delivery, and mucosal immunity induction. The development of multiepitope, rationally-designed vaccines is also discussed regarding the experience gained in expression of chimeric immunogenic proteins in plant systems.

Chapter 16 - Antivirals based on nanomaterials against SARS-CoV-2

In this chapter the use of nanomaterials (e.g., graphene oxide, quantum dots, silver, zinc oxide, and gold nanoparticles) to combat COVID-19 is presented. This chapter does not include nanomaterials used for the release of drugs or antiviral molecules where the nanomaterial is not a part of the antiviral effect. The nanomaterials presented somehow interact with the virus, therefore, having an antiviral effect per se. The chapter first reviews the updated efforts conducted evaluating nanomaterials against SARS-CoV-2. Later, the chapter reviews nanomaterials that have been evaluated against enveloped viruses (e.g., the feline coronavirus, influenza A virus, pseudorabies virus, herpes simplex virus, and respiratory syncytial virus), which could be tested against SARS-CoV-2. Most of the nanomaterials studied thus far are effective at inhibiting viruses when contacting them with the virus before infection. Incipient studies address the use of nanomaterials in therapeutic approaches. Similarly, most studies of nanomaterials against viruses have only been evaluated in vitro. Few studies have been conducted in vivo using mice. Virus inactivation is generally achieved by the interaction of the nanomaterial and the virus through electrostatic, hydrophobic, and affinity interactions. A combination of these interactions could arise depending on the properties of the nanomaterial. In few stances the nanomaterial studied responds to an external stimulation, such as near-infrared irradiation, to inactivate the virus. Finally, some works evaluate or envision the nanomaterials as antiviral agents of surfaces or components of personal protection equipment.

A Systematic and a Scoping Review on the Psychometrics and Clinical Utility of the Volume-Viscosity Swallow Test (V-VST) in the Clinical Screening and Assessment of Oropharyngeal Dysphagia.

(1) Background: The volume-viscosity swallow test (V-VST) is a clinical tool for screening and diagnosis of oropharyngeal dysphagia (OD). Our aims were to examine the clinical utility of the V-VST against videofluoroscopy (VFS) or fiberoptic endoscopic evaluation of swallow (FEES) and to map the V-VST usage with patients at risk of OD across the years since it was described for the first time, carrying a systematic and a scoping review. (2) Methods: We performed both a systematic review (SR) including studies that look at the diagnostic test accuracy, and a scoping review (ScR) with articles published from September 2008 to May 2020. Searches were done in different databases, including PubMed and EMBASE from September 2008 until May 2020, and no language restrictions were applied. A meta-analysis was done in the SR to assess the psychometric properties of the V-VST. Quality of studies was assessed by Dutch Cochrane, QUADAS, GRADE (SR), and STROBE (ScR) criteria. The SR protocol was registered on PROSPERO (registration: CRD42020136252). (3) Results: For the diagnostic accuracy SR: four studies were included. V-VST had a diagnostic sensitivity for OD of 93.17%, 81.39% specificity, and an inter-rater reliability Kappa = 0.77. Likelihood ratios (LHR) for OD were 0.08 (LHR-) and 5.01 (LHR+), and the diagnostic odds ratio for OD was 51.18. Quality of studies in SR was graded as high with low risk of bias. In the ScR: 34 studies were retrieved. They indicated that V-VST has been used internationally to assess OD's prevalence and complications. (4) Conclusions: The V-VST has strong psychometric properties and valid endpoints for OD in different phenotypes of patients. Our results support its utility in the screening and clinical diagnosis and management of OD.

COVID-19 is associated with oropharyngeal dysphagia and malnutrition in hospitalized patients during the spring 2020 wave of the pandemic.

BACKGROUND & AIMS: Prevalence and complications of oropharyngeal dysphagia (OD) and malnutrition (MN) in COVID-19 patients is unknown. Our aim was to assess the prevalence, risk factors and clinical outcomes of OD and MN in a general hospital during the first wave of the COVID-19 pandemic. METHODS: This was a prospective, observational study involving clinical assessment of OD (Volume-Viscosity Swallowing Test), and nutritional screening (NRS2002) and assessment (GLIM criteria) in COVID-19 patients hospitalized in general wards at the Consorci Sanitari del Maresme, Catalonia, Spain. The clinical characteristics and outcomes of patients were assessed at pre-admission, admission and discharge, and after 3 and 6-months follow-up. RESULTS: We included 205 consecutive patients (69.28 ± 17.52 years, Charlson 3.74 ± 2.62, mean hospital stay 16.8 ± 13.0 days). At admission, Barthel Index was 81.3 ± 30.3; BMI 28.5 ± 5.4 kg/m2; OD prevalence 51.7% (44.1% impaired safety of swallow); and 45.5% developed MN with a mean weight loss of 10.1 ± 5.0 kg during hospitalization. OD was an independent risk factor for MN during hospitalization (OR 3.96 [1.45-10.75]), and hospitalization was prolonged in patients with MN compared with those without (21.9 ± 14.8 vs 11.9 ± 8.9 days, respectively; p < 0.0001). OD was independently associated with comorbidities, neurological symptoms, and low functionality. At 6-month follow-up, prevalence of OD was still 23.3% and that of MN only 7.1%. Patients with OD at discharge showed reduced 6-month survival than those without OD at discharge (71.6% vs 92.9%, p < 0.001); in contrast, those with MN at discharge did not show 6-month survival differences compared to those without (85.4% vs 83.8%, p = 0.8). CONCLUSIONS: Prevalence and burden of OD and MN in patients hospitalized in COVID-19 wards is very high. Our results suggest that optimizing the management of MN might shorten the hospitalization period but optimizing the management of OD will likely impact the nutritional status of COVID-19 patients and improve their clinical outcomes and survival after hospital discharge. CLINICALTRIALS: gov Identifier: NCT04346212.

Synthesis and immunogenicity assessment of a gold nanoparticle conjugate for the delivery of a peptide from SARS-CoV-2.

The development of vaccines is a crucial response against the COVID-19 pandemic and innovative nanovaccines could increase the potential to address this remarkable challenge. In the present study a B cell epitope (S461-493) from the spike protein of SARS-CoV-2 was selected and its immunogenicity validated in sheep. This synthetic peptide was coupled to gold nanoparticles (AuNP) functionalized with SH-PEG-NH2 via glutaraldehyde-mediated coupling to obtain the AuNP-S461-493 candidate, which showed in s.c.-immunized mice a superior immunogenicity (IgG responses) when compared to soluble S461-493; and led to increased expression of relevant cytokines in splenocyte cultures. Interestingly, the response triggered by AuNP-S461-493 was similar in magnitude to that induced using a conventional strong adjuvant (Freund's adjuvant). This study provides a platform for the development of AuNP-based nanovaccines targeting specific SARS-CoV-2 epitopes.

The Potential of Algal Biotechnology to Produce Antiviral Compounds and Biopharmaceuticals.

The emergence of the Coronavirus Disease 2019 (COVID-19) caused by the SARS-CoV-2 virus has led to an unprecedented pandemic, which demands urgent development of antiviral drugs and antibodies; as well as prophylactic approaches, namely vaccines. Algae biotechnology has much to offer in this scenario given the diversity of such organisms, which are a valuable source of antiviral and anti-inflammatory compounds that can also be used to produce vaccines and antibodies. Antivirals with possible activity against SARS-CoV-2 are summarized, based on previously reported activity against Coronaviruses or other enveloped or respiratory viruses. Moreover, the potential of algae-derived anti-inflammatory compounds to treat severe cases of COVID-19 is contemplated. The scenario of producing biopharmaceuticals in recombinant algae is presented and the cases of algae-made vaccines targeting viral diseases is highlighted as valuable references for the development of anti-SARS-CoV-2 vaccines. Successful cases in the production of functional antibodies are described. Perspectives on how specific algae species and genetic engineering techniques can be applied for the production of anti-viral compounds antibodies and vaccines against SARS-CoV-2 are provided.

Can nanotechnology help in the fight against COVID-19?

INTRODUCTION: The current COVID-19 pandemic caused by the SARS-CoV-2 virus demands the development of strategies not only to detect or inactivate the virus, but to treat it (therapeutically and prophylactically). COVID-19 is not only a critical threat for the population with risk factors, but also generates a dramatic economic impact in terms of morbidity and the overall interruption of economic activities. AREAS COVERED: Advanced materials are the basis of several technologies that could diminish the impact of COVID-19: biosensors might allow early virus detection, nanosized vaccines are powerful agents that could prevent viral infections, and nanosystems with antiviral activity could bind the virus for inactivation or destruction upon application of an external stimulus. Herein all these methods are discussed under the light of cutting-edge technologies and the previously reported prototypes targeting enveloped viruses similar to SARS-CoV-2. This analysis was derived from an extensive scientific literature search (including pubmed) performed on April 2020. EXPERT OPINION: Perspectives on how biosensors, vaccines, and antiviral nanosystems can be implemented to fight COVID-19 are envisioned; identifying the approaches that can be implemented in the short term and those that deserve long term research to cope with respiratory viruses-related pandemics in the future.