Summary The coronavirus disease 2019 (Covid‐19) pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2), is an international public health crisis with devastating effects. In particular, this pandemic has further exacerbated the burden in tropical and subtropical regions of the world, where dengue fever, caused by dengue virus (DENV), is already endemic to the population. The similar clinical manifestations shared by Covid‐19 and dengue fever have raised concerns, especially in dengue‐endemic countries with limited resources, leading to diagnostic challenges. In addition, cross‐reactivity of the immune responses in these infections is an emerging concern, as pre‐existing DENV‐antibodies might potentially affect Covid‐19 through antibody‐dependent enhancement. In this review article, we aimed to raise the issue of Covid‐19 and dengue fever misdiagnosis, not only in a clinical setting but also with regards to cross‐reactivity between SARS‐CoV‐2 and DENV antibodies. We also have discussed the potential consequences of overlapping immunological cascades between dengue and Covid‐19 on disease severity and vaccine development.

Bioinspired self-sorting and self-assembling systems using engineered versions of natural protein cages are being developed for biocatalysis and therapeutic delivery. The packaging and intracellular delivery of guest proteins is of particular interest for both in vitro and in vivo cell engineering. However, there is a lack of bionanotechnology platforms that combine programmable guest protein encapsidation with efficient intracellular uptake. We report a minimal peptide anchor for in vivo self-sorting of cargo-linked capsomeres of murine polyomavirus (MPyV) that enables controlled encapsidation of guest proteins by in vitro self-assembly. Using Förster resonance energy transfer, we demonstrate the flexibility in this system to support coencapsidation of multiple proteins. Complementing these ensemble measurements with single-particle analysis by super-resolution microscopy shows that the stochastic nature of coencapsidation is an overriding principle. This has implications for the design and deployment of both native and engineered self-sorting encapsulation systems and for the assembly of infectious virions. Taking advantage of the encoded affinity for sialic acids ubiquitously displayed on the surface of mammalian cells, we demonstrate the ability of self-assembled MPyV virus-like particles to mediate efficient delivery of guest proteins to the cytosol of primary human cells. This platform for programmable coencapsidation and efficient cytosolic delivery of complementary biomolecules therefore has enormous potential in cell engineering.

Metabolic pathways are commonly organized by sequestration into discrete cellular compartments. Compartments prevent unfavorable interactions with other pathways and provide local environments conducive to the activity of encapsulated enzymes. Such compartments are also useful synthetic biology tools for examining enzyme/pathway behavior and for metabolic engineering. Here, we expand the intracellular compartmentalization toolbox for budding yeast ( Saccharomyces cerevisiae ) with Murine polyomavirus virus-like particles (MPyV VLPs). The MPyV system has two components: VP1 which self-assembles into the compartment shell and a short anchor, VP2C, which mediates cargo protein encapsulation via binding to the inner surface of the VP1 shell. Destabilized green fluorescent protein (GFP) fused to VP2C was specifically sorted into VLPs and thereby protected from host-mediated degradation. An engineered VP1 variant displayed improved cargo capture properties and differential subcellular localization compared to wild-type VP1. To demonstrate their ability to function as a metabolic compartment, MPyV VLPs were used to encapsulate myo-inositol oxygenase (MIOX), an unstable and rate-limiting enzyme in d -glucaric acid biosynthesis. Strains with encapsulated MIOX produced ∼20% more d -glucaric acid compared to controls expressing “free” MIOX—despite accumulating dramatically less expressed protein—and also grew to higher cell densities. This is the first demonstration in yeast of an artificial biocatalytic compartment that can participate in a metabolic pathway and establishes the MPyV platform as a promising synthetic biology tool for yeast engineering.

Coronavirus disease 2019 (COVID-19) pandemic continues to constitute a public health emergency of international concern. Multiple vaccine candidates for COVID-19, which is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), have entered clinical trials. However, some evidence suggests that patients who have recovered from COVID-19 can be reinfected. For example, in China, two discharged COVID-19 patients who had recovered and fulfilled the discharge criteria for COVID-19 were retested positive to a reverse transcription polymerase chain reaction (RT-PCR) assay for the virus. This finding is critical and could hamper COVID-19 vaccine development. This review offers literature-based evidence of reinfection with SARS-CoV-2, provides explanation for the possibility of SARS-CoV-2 reinfection both from the agent and host points of view, and discusses its implication for COVID-19 vaccine development.

The Polyomavirus coat protein, VP1 has been developed as an epitope presentation system able to provoke humoral immunity against a variety of pathogens, such as Influenza and Group A Streptococcus. The ability of the system to carry cytotoxic T cell epitopes on a surface-exposed loop and the impact on protein solubility has not been examined. Four variations of three selected epitopes were cloned into surface-exposed loops of VP1, and expressed in Escherichia coli. VP1 pentamers, also known as capsomeres, were purified via a glutathione-S-transferase tag. Size exclusion chromatography indicated severe aggregation of the recombinant VP1 during enzymatic tag removal resulting from the introduction the hydrophobic epitopes. Inserts were modified to possess double aspartic acid residues at each end of the hydrophobic epitopes and a highthroughput buffer condition screen was implemented with protein aggregation monitored during tag removal by spectrophotometry and dynamic light scattering. These analyses showed that the insertion of charged residues at the extremities of epitopes could improve solubility of capsomeres and revealed multiple windows of opportunity for further condition optimization. A combination of epitope design, pH optimization, and the additive L-arginine permitted the recovery of soluble VP1 pentamers presenting hydrophobic epitopes and their subsequent assembly into virus-like particles.