The Viral Infidelity
Viruses are pseudo-living organisms existing since the dawn of the earth. They are versatile creatures which are well-equipped in attacking living cells and directing them to synthesize multiple copies of viruses. All viruses don’t infect all cells. There is a specified host-range for each virus which it infects. One is never less astonished by the structural specificity and complexity of the assembly and infectious cycle of viruses. One can only appreciate the beauty with which these little creatures roam around ruling the reigns.
Viruses are a subject of research since they have been discovered. Years of continued research in elucidating the infectious processes of viruses have resulted in many fascinating conclusions. This knowledge has been further exploited to develop applications including phage therapy, novel anti-viral targets for drug discovery, vaccination, drug delivery, diagnostics and engineering genetic circuits for use as anti-bacterials with reduced immune response and higher therapeutic efficacy.
Patients are treated with a combination of the engineered phage + the conventional antibiotic. The results supported the fact that the combination reduced the antibiotic-resistance upto some extent. Bacteriophages are also used in diagnostics wherein the specific interactions between the expressed peptide and the antigen are selected from a pool of antigens thereby confirming the presence of the pathogenic antigen.
This being said, there is considerable research going-on for constructing synthetic bacteriophages. In-fact, the Massachusetts Institute of Technology patented a synthetic bacteriophage in 2018. They constructed the genome via synthetic biology and was aimed to be used as an anti-bacterial with experimentally proved reduced side-effects and immune response.
Research in Virology has been mainstreamed into engineering bacteriophages given their ability to infect bacteria and their malleable nature allowing the scientific community to juggle with the system which is also simplified by the short size of the genome and comparatively less complexity.
There are 2 widely used approaches for engineering viruses (bacteriophages):
1. Homologous Recombination
This method exploits the traditional recombination process to incorporate foreign DNA into the phage genome. A bacterial strain having high recombination efficiency is transformed with phage DNA and the desired DNA in a vector. This result in a modified phage DNA which is further amplified in the phage’s native host. This engineered phage will have the desired gene/phenotype required to perform the desired function. A drawback of this traditional method is the tedious selection step after transformation. To overcome this difficulty, CRISPR-Cas systems are deployed which is based on a counter-selection method, wherein, the wild-type phage genome is cleaved by nulceases resulting in their destruction and on lysis only engineered phages are obtained.
2. Rebooting Genomes
This approach involves cell-free systems like the Gibbson Assembly to bring together pieces of DNA to complete the viral genome or the tk-tl (transcription-translation) like cell free protein synthesis systems for producing engineered phages. For large DNA fragments, the genome is modified via transformation assisted recombination (TAR) in yeasts by yeast artificial chromosomes. These below methods are widely used given their non-requirement for selection of transformed phages
a. In-Vitro Gibson Assembly.
b. Transformation assisted recombination in yeasts.
Now, if engineering of genomes itself hasn’t got you startled enough, we further arrive at interactions between viruses. A general perception regarding viral infections among common people being that the infection is a result of a single virus strain. Wherein, the fact being that the infection is a result of complex interactions between different viral strains present in the micro-environment. The outcome of a viral infection is governed by a myriad of biological and ecological factors. The literature on virology suggests the presence of four important factors namely, host ecology, host taxonomy, host defense mechanisms and virus-virus interactions.
The virus-virus interactions (VVI) form the most important factor of the viral infection. These VVI are classified into 3 types in the literature as given below
1. Direct Interaction of viral gene products.
2. Indirect environmental Interactions.
3. Immunological Interactions.
These are further sub-classified into 15 different types of interactions. This gives us an idea about the vastness of the interactions and the myriad effects these interactions can have on the host and the outcome of infection.
I am willing to discuss some specific points in direct interactions of viral gene products between viruses. Direct interactions involve
1. Pseudo-typed viruses.
These are viruses which on co-infection with viruses of related species wear the envelope of the later with the internal genomic content being its own. Pseudo-type viruses have found applications in study of virus entry mechanisms, as vectors for gene therapy and as a safe option for studying biosafety 3 and 4 level viruses. This behavior can be put to good use for tackling viral infections. A specific example is given below.
The Vesicular Stomatitis Virus belongs to the family Rhabdoviridae and is capable of Infecting a broad range of hosts including horses, cattle and swine. Literature on pseudo-type viruses has established that Vesicular Stomatitis Virus (VSV) is capable of transforming into a pseudo-typed virus Inferring its use in combating viral infections. Moreover, VSV doesn’t infect humans therefore there is a minimum chance of infection due to VSV. The mechanism of hacking the envelope is not clear yet, but it can be inferred that VSV acquires not only the peripheral glycoproteins which mediate interaction with the receptors but the envelope as a whole.
In view of my Insufficient knowledge about cell systems, I wish to postulate a strategy for deploying engineered viruses harboring CRISPR spacers from the Japanese Encephalitis virus (JEV) into VSV. The CRISPR system deployed could be used for treating cells co-infected with JEV and VSV. However this method can be successful given that multiplicity of infection (MOI) of VSV is greater than JEV, not only JEV but any virus having MOI less than VSV can be hypothesized to mitigate the infection if not completely eradicate it. There might be many more factors at play for the cas nucleases to show its effect.
There is always a possibility of CRISPR escape mutants (CEM) being generated. A minor change can be made: the spacer sequences can be the gene sequence for the envelope receptor protein. Therefore, even if the virus (JEV or any other virus) tried to evade CRISPR cleavage, there should be a good chance of a CEM being generated where the mutation in the receptor distorts its structure and therefore prevents the entry of the virus into its host cell. (How about that smarty pants, lmao).
2. Heterologous transactivation.
It refers to trans-activation of a gene of one virus species by gene products of a heterologous virus. This method is well documented in literature and the enticing point for its application stems from the overproduction of the protein (mainly for the increase in virus titer) which trans-activates a gene from another co-infected virus in a cell. Again applying the same engineering principles to generate a recombinant virus wherein a repressor is overproduced instead of a promoter or transcriptional factors which would eventually lead to downregulation of the former virus thereby lowering the infection. I don’t foresee this method as a sole anti-viral treatment but can be supplemented with traditional anti-viral drugs and therapies.
Having reflected my thoughts, it would be great to ponder over some more fascinating biological processes so that the quarantine just passes by like a second. However, there’s something called as academics and no matter how relevant your academics is to your passion (in my case its biological science, you probably might be knowing that by now) the kind of ecstasy that is perceived after one follows their curiosity and Intel to pursue answers to petty questions, thoughts and doubts about their passion through the Internet (only approved scientific evidence is looked at) is just inexhaustible.
In times of pandemic, there is an immediate need for novel methods to control and contest these tiny morons. The basic concept of virus-virus Interactions can lead to a novel method of targeting viruses. Not a long time ago, the discovery of phage therapy had and still continues to pull us out from reaching deadlocks in biological sciences. This is the main reason I am so affected with basic science. Nonetheless, I am pretty sure, if not to be too confident, that these virus-virus Interactions might lead to a halt in viral Infections for some time, if not for an indefinite period. However, it is not to be dismissed that we are subservient to mother nature and she has her own mystical ways to surprise us. Therefore, unless we find a way to stop the random mutations in nucleic acids, we are doomed to face antibiotic resistance, viral mutants, and autoimmune disorders.
The two methods mentioned above are a result of pure hypothetical speculation done by the author. The author is unknown of any reported counter-evidence against these methods. It is to be noted, these strategies can be useful only for a small niche of viruses or might not be useful at all (obviously, without research and documented evidence this cannot be predicted). The only Intention the author had in mind was to present his hypothesis and to gather constructive criticism from the scientific community which will aid in bolstering his knowledge about this mysterious cell community.
Hope you find something insightful in here. Would be happy to hear from you about the developments in biological sciences and new speculations, theories and questions one might have because I would be more than happy to ponder over such beautiful conversations. Thanks for reading ;)
In case one would like to know more about the information referred in the text above, the references are mentioned below.
References:
1. Bacteriophages as potential new therapeutics to replace or supplement antibiotics, Mzia Kutateladze and Revaz Adamia.
2. Engineering Bacteriophages as Versatile Biologics, Samuel Kilcher, and Martin J. Loessner.
3. Genetically Engineered Phages: a Review of Advances over the Last Decade, Diana P. Pires, Sara Cleto, Sanna Sillankorva, Joana Azeredo, Timothy K. Lua.
4. Phage Display: An Overview in Context to Drug Discovery, Selena Mimmi, Domenico Maisano, Ileana Quinto and Enrico Iaccino.
5. A systematic approach to virus–virus interactions, T. DaPalma, B.P. Doonan, N.M. Trager, L.M. Kasman Dept. of Microbiology and Immunology, Medical University of South Carolina, United States.
6. Development and applications of VSV vectors based on cell tropism, HidekiTani, Shigeru Morikawa and Yoshiharu Matsuura.
7. Viral coinfection is shaped by host ecology and virus–virus interactions across diverse microbial taxa and environments, Samuel L. Dıaz-Munoz
