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**Spoiler Alert** The world as we know it as fell when a virus called SHVID-1 spreads through mankind turning them into flesh eating zombies. SHVID-1 began from a shark bite so a team of scientist go to an underwater lab to find a cure as the world above them begins to crumble. However, deadly sharks and weak structure forces the remaining crew to flee to the world above forcing them to face those that have been changed by the virus. **Spoiler Alert**
A juvenile, male tiger shark (Galeocerdo cuvier) developed illness after capture in Florida waters and was euthanized. Gross lesions included mild skin abrasions, hepatic atrophy, and coelomic fluid. Histologically, gills contained multifocal lamellar epithelial cell necrosis and thromboses. Scattered gill and esophageal epithelial cells had large, basophilic, intracytoplasmic, and intranuclear inclusions. Ultrastructurally, lamellar epithelial cells contained arrays of intracytoplasmic viral particles and scattered intranuclear nucleocapsids. Capsulated virions were 148 11 nm with an 84 8 nm icosahedral nucleocapsid and an electron-dense core. Next-generation sequencing, quantitative polymerase chain reaction, and in situ hybridization performed on formalin-fixed tissue confirmed a herpes-like viral infection. The viral polymerase shared 24% to 31% protein homology with other alloherpesviruses of fish, indicating a divergent virus. This report documents the pathologic findings associated with a molecularly confirmed novel herpes-like virus in an elasmobranch.
FIGURE 4. Phylogenetic analysis of microbial and viral fractions from Shark Bay stromatolites for the Replication-associated protein (Rep) in CRESS viruses. Maximum likelihood phylogenetic tree of Rep protein sequences found in CRESS viruses in Shark Bay stromatolites. Protein sequence alignments were performed using MUSCLE and gaps in alignment were removed with UGENE. The tree was constructed with IQ-TREE v. 1.6.1 with 1000 bootstrap replicates and was visualized with iTOL. Number indicates bootstrap values, nodes with bootstrap values lower than 70 were not shown and represented by the collapsed branch. The collapsed branches in this figure represent reference sequences from Rosario et al. (2009a) for the replication (rep) protein that have lower than 70 bootstrap values.
FIGURE 5. Phylogenetic analysis of viral fractions from Shark Bay stromatolites for the major capsid protein (VP1) in Microviridae viruses. Maximum likelihood phylogenetic tree of VP1 protein sequences obtained from Microviridae viruses in Shark Bay stromatolites. Reference sequences were retrieved from the Uniprot database. Short sequences (
FIGURE 6. Phylogenetic analysis of viral fractions from Shark Bay stromatolites for phoH (of the pho regulon) in dsDNA viruses. Maximum-likelihood tree of phoH protein sequences obtained from dsDNA viruses. Reference sequences were retrieved from Uniprot database. Alignments were performed using MUSCLE and gaps in alignment were removed with UGENE. The tree was constructed with IQ-TREE v. 1.6.1 with 1000 bootstrap replicates and was visualized with iTOL. Number indicates bootstrap values, nodes with bootstrap values lower than 70 were not shown and represented by the collapsed branch. The collapsed branches in this figure represent reference sequences from Goldsmith et al. (2011) for the phosphate starvation inducible protein (phoH) that have lower than 70 bootstrap values.
The new VNARs will not be immediately available as a treatment in people, but they can help prepare for future coronavirus outbreaks. The shark VNARs were able to neutralize WIV1-CoV, a coronavirus that is capable of infecting human cells but currently circulates only in bats, where SARS-CoV-2, the virus that causes COVID-19, likely originated.
One VNAR, named 3B4, attached strongly to a groove on the viral spike protein near where the virus binds to human cells and appears to block this attachment process. This groove is very similar among genetically diverse coronaviruses, which even allows 3B4 to effectively neutralize the MERS virus, a distant cousin of the SARS viruses.
Future therapies would likely include a cocktail of multiple shark VNARs to maximize their effectiveness against diverse and mutating viruses. This new class of drug is cheaper and easier to manufacture than human antibodies, and can be delivered into the body through various routes, but has yet to be tested in humans. LeBeau is also studying the ability of shark VNARs to help in the treatment and diagnosis of cancers.
Digital on Demand: SHVID-1 is a shark-borne virus which has now transferred to humans, with lethal results. A group of researchers entrenched on the seafloor are racing to find a cure...but that's easier said than done when some of the researchers have the virus, and there are sharks both inside and outside the facility...
The facility is about to collapse and/or lose oxygen so everyone needs to escape. Gregory goes rogue, steals the cure and uses the only way out, an elevator, which gets eaten by the shark. The shark eats the communication cables too. The virus is making that shark a total douchebag.
The VNARs also showed promise as therapeutics for other known beta coronaviruses and future emergent diseases. They appear able to identify and bind to regions of amino acids that are the same among different coronaviruses.
Previous viral metagenomic studies have suggested that linking various viral genotypes to certain environments to establish viral biogeography is challenging. Often the same viral genotype is found in a variety of ecosystems suggesting that viruses have a cosmopolitan distribution (Breitbart and Rohwer, 2005). A viral metagenomic study that contrasted this concept of cosmopolitan viral biogeography suggested that viral ecotypes do exist in nature (Desnues et al., 2008). It was found that single-stranded DNA microphages from Highbourne Cay stromatolites were endemic and these specific viruses were not found among any other cross-examined ecosystem, including marine, freshwater, terrestrial or metazoan-associated systems. However, this is the only study to date that exists for viral communities among modern microbialites (Desnues et al., 2008).
Phylogenetic analysis of viral fractions from Shark Bay stromatolites for the major capsid protein (VP1) in Microviridae viruses. Maximum likelihood phylogenetic tree of VP1 protein sequences obtained from Microviridae viruses in Shark Bay stromatolites. Reference sequences were retrieved from the Uniprot database. Short sequences (
Single-domain antibodies derived from the unique New Antigen Receptor found in sharks have numerous potential applications, ranging from diagnostic reagents to therapeutics. Shark-derived single-domain antibodies possess the same characteristic ability to refold after heat denaturation found in single-domain antibodies derived from camelid heavy-chain-only antibodies. Recently, two shark derived single-domain antibodies specific for the nucleoprotein of Ebola virus were described. Our evaluation confirmed their high affinity for the nucleoprotein, but found their melting temperatures to be low relative to most single-domain antibodies. Our first approach towards improving their stability was grafting antigen-binding regions (complementarity determining regions) of one of these single-domain antibodies onto a high melting temperature shark single-domain antibody. This resulted in two variants: one that displayed excellent affinity with a low melting temperature, while the other had poor affinity but a higher melting temperature. These new proteins, however, differed in only 3 amino acids within the complementarity determining region 2 sequence. In shark single-domain antibodies, the complementarity determining region 2 is often referred to as hypervariable region 2, as this segment of the antibody domain is truncated compared to the sequence in camelid single-domain antibodies and conventional heavy chain variable domains. To elucidate which of the three amino acids or combinations thereof were responsible for the affinity and stability we made the 6 double and single point mutants that covered the intermediates between these two clones. We found a single amino acid change that achieved a 10C higher melting temperature while maintaining sub nM affinity. This research gives insights into the impact of the shark sdAb hypervariable 2 region on both stability and affinity.
Shark sdAb have been developed towards a number of targets relevant to human health; recently shark sdAb that recognize the nucleoprotein (NP) of the Ebola virus (EBOV; formerly the Zaire ebolavirus) have been described [19]. Although demonstrated in binding assays with live virus [19,20], neither the thermal stability of these sdAb nor their binding kinetics were evaluated. We prepared and evaluated these shark sdAb and found that while they had excellent binding affinity to a truncated recombinant NP construct, they showed melting temperatures of only 50C. In this work we focused on improving the stability of one of these shark-derived sdAb.
The DSTL096 and DSTL097 constructs are composed of the sdAb expressed as a fusion with a murine kappa light chain constant domain. An unfused construct containing only the sdAb portion of DSTL096 was prepared using its published protein sequence (Fig 1). The gene was synthesized and cloned into the pET22b(+) expression vector to produce the construct shark096. Other researchers had validated the ability of both the unfused shark anti-EBOV NP sdAb to bind live virus [20]. Similarly, our experiments confirmed that shark096 has essentially identical affinity and melting characteristics to the fusion construct with a KD = 0.3 nM (Fig 2) and melting temperature of 52C (Table 1). Using SPR we also verified the specificity of shark096, showing that it had no cross reactivity to other recombinant EBOV proteins (VP40, GP; data not shown). 781b155fdc
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