Optimizing RNAi-Target by Nicotiana benthamiana-Soybean Mosaic Virus System Drives Broad Resistance to Soybean Mosaic Virus in Soybean

Soybean mosaic virus (SMV) is a prevalent pathogen of soybean (Glycine max). Pyramiding multiple SMV-resistance genes into one individual is tedious and difficult, and even if successful, the obtained multiple resistance might be broken by pathogen mutation, while targeting viral genome via host-induced gene silencing (HIGS) has potential to explore broad-spectrum resistance (BSR) to SMV. We identified five conserved target fragments (CTFs) from S1 to S5 using multiple sequence alignment of 30 SMV genome sequences and assembled the corresponding target-inverted-repeat constructs (TIRs) from S1-TIR to S5-TIR. Since the inefficiency of soybean genetic transformation hinders the function verification of batch TIRs in SMV-resistance, the Nicotiana benthamiana-chimeric-SMV and N. benthamiana-pSMV-GUS pathosystems combined with Agrobacterium-mediated transient expression assays were invented and used to test the efficacy of these TIRs. From that, S1-TIR assembled from 462 bp CTF-S1 with 92% conservation rate performed its best on inhibiting SMV multiplication. Accordingly, S1-TIR was transformed into SMV-susceptible soybean NN1138-2, the resistant-healthy transgenic T1-plants were then picked out via detached-leaf inoculation assay with the stock-plants continued for progeny reproduction (T1 dual-utilization). All the four T3 transgenic progenies showed immunity to all the inoculated 11 SMV strains under individual or mixed inoculation, achieving a strong BSR. Thus, optimizing target for HIGS via transient N. benthamiana-chimeric-SMV and N. benthamiana-pSMV-GUS assays is crucial to drive robust resistance to SMV in soybean and the transgenic S1-TIR-lines will be a potential breeding source for SMV control in field.

Immune lag is a major cost of prokaryotic adaptive immunity during viral outbreaks

Clustered regularly interspaced short palindromic repeat (CRISPR)-Cas adaptive immune systems enable bacteria and archaea to efficiently respond to viral pathogens by creating a genomic record of previous encounters. These systems are broadly distributed across prokaryotic taxa, yet are surprisingly absent in a majority of organisms, suggesting that the benefits of adaptive immunity frequently do not outweigh the costs. Here, combining experiments and models, we show that a delayed immune response which allows viruses to transiently redirect cellular resources to reproduction, which we call ‘immune lag’, is extremely costly during viral outbreaks, even to completely immune hosts. Critically, the costs of lag are only revealed by examining the early, transient dynamics of a host–virus system occurring immediately after viral challenge. Lag is a basic parameter of microbial defence, relevant to all intracellular, post-infection antiviral defence systems, that has to-date been largely ignored by theoretical and experimental treatments of host-phage systems.

Human embryonic stem cell-derived cardiomyocyte platform screens inhibitors of SARS-CoV-2 infection

Patients with cardiovascular comorbidities are more susceptible to severe infection with SARS-CoV-2, known to directly cause pathological damage to cardiovascular tissue. We outline a screening platform using human embryonic stem cell-derived cardiomyocytes, confirmed to express the protein machinery critical for SARS-CoV-2 infection, and a SARS-CoV-2 spike-pseudotyped virus system. The method has allowed us to identify benztropine and DX600 as novel inhibitors of SARS-CoV-2 infection in a clinically relevant stem cell-derived cardiomyocyte line. Discovery of new medicines will be critical for protecting the heart in patients with SARS-CoV-2, and for individuals where vaccination is contraindicated.

Screening and Identification of Lujo Virus Inhibitors Using a Recombinant Reporter Virus Platform

Lujo virus (LUJV), a highly pathogenic arenavirus, was first identified in 2008 in Zambia. To aid the identification of effective therapeutics for LUJV, we developed a recombinant reporter virus system, confirming reporter LUJV comparability with wild-type virus and its utility in high-throughput antiviral screening assays. Using this system, we evaluated compounds with known and unknown efficacy against related arenaviruses, with the aim of identifying LUJV-specific and potential new pan-arenavirus antivirals. We identified six compounds demonstrating robust anti-LUJV activity, including several compounds with previously reported activity against other arenaviruses. These data provide critical evidence for developing broad-spectrum antivirals against high-consequence arenaviruses.

Potent neutralization antibodies induced by a recombinant trimeric Spike protein vaccine candidate containing PIKA adjuvant for COVID-19

Neutralizing antibodies are critical to prevent corona virus infection. The structures of immunogens to elicit most potent neutralization antibodies are still under investigation. Here we tested the immunogenicity of the trimeric, full length Spike protein with 2 proline mutations to preserve its prefusion conformation. Recombinant trimeric Spike protein expressed by CHO cells was used with polyI:C (PIKA) adjuvant to immunize mice by 0-7-14 day schedule. The results showed that Spike-specific antibody was induced at day 21 with titer of more than 50,000 in average as measured by direct binding to Spike protein. The titer of neutralization reached more than 1000 in average when tested by a pseudo-virus system, using monoclonal antibodies (40592-MM57 and 40591-MM43) with neutralizing IC50 at 1 μg/ml as standards. Protein/peptide array showed that the antibodies induced by trimeric S protein vaccine bind similarly to natural infection with the receptor binding domain (RBD) as major immunodominant region. No linear epitopes were found in RBD, although several linear epitopes were found in C-terminal domain right after RBD, and heptad repeat regions. Our study supports the efficacy of recombinant trimeric Spike protein vaccine candidate for COVID-19, with excellent safety and readiness for storage and distribution in developing countries.

Human embryonic stem cell-derived cardiomyocytes express SARS-CoV-2 host entry proteins: screen to identify inhibitors of infection

Patients with cardiovascular comorbidities are more susceptible to severe infection with SARS-CoV-2, known to directly cause pathological damage to cardiovascular tissue. We outline a screening platform using human embryonic stem cell-derived cardiomyocytes, confirmed to express the protein machinery critical for SARS-CoV-2 infection, and a pseudotyped virus system. The method has allowed us to identify benztropine and DX600 as novel inhibitors of SARS-CoV-2 infection.

An Account on Mycoviruses and Their Applications

Mycoviruses are obligate parasites of fungi and can infect the majority of the fungal groups. They remain mysterious to various communities throughout the globe. Mycoviruses are responsible for certain changes in fungal hyphae, which could be asymptomatic and may cause a reduction or elimination of the virulence capacity of fungal hosts by the process called hypovirulence. Such fungal-virus system could be valuable for the development of novel biocontrol approaches against fungal pathogens for the development of a sustainable environment. There are adequate reports where mycovirus has been employed as a biocontrol approach against the pathogenic fungi in the fields of agriculture and other allied sciences. The prime focus of this review is to emphasize naturally available mycoviruses and strategies to adopt the mycovirus therapy which could serve as an excellent alternative strategy against chemical prophylactic and therapeutic approaches.