Phage Out
Solving the Antibiotic Resistance Crisis
By Yeshin Kim, Ethan Yim and Ryan Lee
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Abstract
Bacteria are microscopic single-celled organisms that lack a nucleus and membrane-bound organelles. Most bacteria have a protective cell wall, a cell membrane, and a strand of DNA. Many also have flagella to help them move, and all reproduce by binary fission, splitting into two new identical cells. Bacteria are highly diverse - they live in all types of environments on Earth and come in various shapes like rods or spirals. Of the three domains used to classify life, two are bacteria: Archaea and Eubacteria. Archaea bacteria have unique genes enabling them to get energy from ammonia, methane, and hydrogen gas. Most bacteria fall into the Eubacteria domain. While some bacteria can cause illness, most perform important functions - gut bacteria help digest food, cyanobacteria produce oxygen through photosynthesis, and bacteria are used to make foods like yogurt and cheese. In fact, bacterial cells make up 90% of the cells in the human body.
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A virus is a microscopic infectious agent that consists of a strand of genetic material, either DNA or RNA, encased within a protein shell known as a capsid. Unlike living cells, viruses lack the cellular machinery necessary for self-replication - they require a host cell to infect and hijack in order to make copies of themselves. Viruses bind to specific receptors on the surface of host cells using proteins on their capsids, allowing them to gain entry. Once inside the cell, a virus will either inject its genetic material into the host cell cytoplasm or become fully engulfed into the cell. The viral genes then utilize the host cell's molecular mechanisms to produce numerous viral proteins and replicate the viral genome. These components self-assemble into new virus particles which accumulate inside the cell until the pressure causes the cell to burst, releasing a flood of viruses to infect surrounding cells. While some viruses are pathogenic and can cause disease, others have beneficial applications such as strengthening crop disease resistance or fighting cancer. Most remarkably, research suggests that ancient viruses infecting early bacteria may have provided the genetic material that led to the evolution of the nucleus within eukaryotic cells. In essence, the nucleus which defines all complex life may have originated from an ancient viral infection event.
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What is a Phage?
Definition and Classification
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Definition
Bacteriophages, commonly known as phages, are viruses that infect bacteria. The name comes from the Greek word “phagein” meaning to devour. Phages are composed of a protein capsid that encases their genetic material, which can be either DNA or RNA. The capsid attaches to a tail structure that allows the phage to recognize and bind to specific bacterial hosts.
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Categorization of Phages
Phages are categorized into several families based on their morphology. The most well-studied are the tailed phages belonging to the order Caudovirales. This order contains three main families: the Siphoviridae which have long, non-contractile tails; the Myoviridae which have contractile tails; and the Podoviridae which have short stubby tails. There are also filamentous and pleomorphic phages that do not fall into these tailed groups.
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Population
Phages are the most abundant biological entities on Earth. These microscopic predators play pivotal ecological roles in regulating bacterial populations and facilitating genetic diversity. Phages specific to disease-causing bacteria may offer solutions to growing antibiotic resistance. Engineered phages are emerging tools for modulating microbiomes and treating dysbiosis.
Abundance and Distribution
Phages are extremely abundant in nature and play a critical ecological role in regulating bacterial populations. There are an estimated 10^31 phage particles on Earth, making them the most numerous biological entities. Phages can be found in all environments populated by bacterial hosts, including soil, seawater, the ocean depths, and the gut microbiomes of animals.
Each gram of human gut bacteria contains over 1 billion phage particles. Phages are equally diverse, with an estimated 10^31 different phage species globally. Different phages infect specific bacterial hosts, though some have broader host ranges. This phage diversity allows them to infect the full bacterial diversity.
What is the Historical Context?
Bacteriophages were first discovered in 1915 by Frederick Twort and in 1917 by Félix d’Hérelle, who realized their potential for killing bacteria. After an initial heyday in the pre-antibiotic era, phages were largely disregarded in Western medicine due to the ease of using antibiotics. However, phage research and application continued in some countries like the Soviet Union, where phages were routinely isolated and used therapeutically against bacterial infections. Much early phage research focused on model phages like T4 that infected Escherichia coli. These model systems laid the foundation of molecular biology, with phages helping identify DNA as the genetic material and the 3 nucleotide codon system. Phages also enabled discovery of restriction enzymes. For decades, only a handful of phages were studied in depth.
The recent resurgence of interest in phage biology stems from realizing just how abundant and diverse phages are in bacterial environments, as revealed by microscopy and bacterial genome sequencing. This belatedly showed phages shape many aspects of bacterial ecology and evolution. The patchy knowledge of phage biology accumulated over the past century demonstrates phages play critical roles across the natural world. These insights have sparked renewed interest in phage research, including development of phage therapy approaches to combat antibiotic resistant bacterial infections. After decades of obscurity, bacteriophages are now recognized as central players in microbial ecosystems and have potential as living antibiotics.
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The structure of a phage is very important and different than other forms of life.
Capsid
All phages share a basic structure of a protein capsid containing the viral genome attached to some form of tail used for recognizing and infecting bacterial hosts. The capsid proteins assemble into a protective shell that encapsulates the viral genome, typically in the shape of an icosahedron though filamentous and pleomorphic shapes also exist.
The capsid protects the genome from damage and provides a stable package for the viral particles. Capsids are made from multiple copies of one or more types of capsid proteins organized into complex architectures. The capsid proteins encode the instructions for self-assembly.
Tail
Phage tails recognize and irreversibly bind to receptor sites on the bacterial surface to initiate infection. Siphoviridae like T5 have long, flexible non-contractile tails. Myoviridae like T4 have a contractile tail sheath that contracts upon attachment to drive the inner tube through the bacterial membrane. Podoviridae tails are very short and often end in tail fibers used to recognize specific host cells.
Baseplate and Tail Fibers
The baseplate is a multifaceted structure at the tail end that contains the receptor binding proteins (RBPs) which determine host specificity. RBPs are highly specific to receptors on the host cell surface like proteins, sugars, or lipopolysaccharides. Long tail fibers extending from the baseplate in some phages help attach to these bacterial receptors.
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The process of a bacteriophage infecting a bacterium encompasses specific attachment, genetic material injection, replication within the host, and eventual release of new phages.
Attachment and Penetration
The phage first attaches to a specific receptor on the bacterial cell surface, using its tail fibers for precise recognition. This attachment is highly specific and dictates the host range of the phage. Following attachment, the phage injects its genetic material (either DNA or RNA) into the bacterium. This is often accomplished through the contraction of the phage tail, which pierces the bacterial cell wall and membrane, creating a pathway for the transfer of the phage genome into the cell. The phage's protein coat does not enter the bacterium.
Biosynthesis and Assembly
Once the phage's genetic material is inside, it co-opts the bacterial machinery to replicate its genetic material and produce phage proteins. These components are necessary for assembling new phage particles. During this stage, the host's normal synthesis of DNA, RNA, and proteins is often disrupted or entirely hijacked for the production of new phage components.
Release
New phage particles are assembled within the bacterial cell. Once a significant number of new phages are formed, they typically cause the bacterial cell to lyse (break open), releasing the new phages to infect surrounding bacteria. Some phages, however, can initiate a lysogenic cycle where the phage DNA integrates into the host genome and replicates along with it, without immediately killing the host.
Importance
Introduction
Bacteriophages, or phages for short, are viruses that infect bacterial hosts. Discovered over a century ago, these microscopic viruses have recently regained prominence as promising alternatives to traditional antibiotics. This renewed interest comes as antibiotic resistance rises while new antibiotic development stagnates. The unique properties of phages may help overcome infections by highly drug-resistant superbugs.
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Ecological Significance
Phages are the most abundant biological entities, with an estimated 10^31 total viral particles on Earth. A single gram of human gut bacteria contains over 1 billion phages. They exist in all microbial habitats and exert strong evolutionary pressures on bacterial populations.
As predators of bacteria, phages help prevent overgrowth of strains and maintain balance in microbial communities through lysis. They transfer genes between bacteria via transduction, enhancing genetic diversity. Phages influence global nutrient cycles by releasing cell contents through lysis. Marine phages also help drive the "biological pump" sending carbon into the deep oceans.
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Therapeutic Potential
Interest in phage therapy declined with the rise of antibiotics but has reemerged given the antibiotic resistance crisis. Phages offer advantages over traditional antibiotics. They rapidly coevolve with bacteria, developing counter-resistance to bacterial mutations. Engineered phages can be tailored to target specific pathogens while sparing beneficial microbes.
Phage treatments can penetrate biofilms that protect bacteria from antibiotics. Used alongside antibiotics, phages improve bacterial clearance. They synergistically enhance antibiotic efficacy against resistant superbugs. Engineered phages may someday treat complex conditions like obesity by altering gut microbiomes.
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Biotechnology Applications
Beyond therapy, phages are widely used in biotechnology. Phage display utilizes phages to present peptides or proteins for the discovery of new drugs, vaccines, and diagnostics. Their specificity makes phages ideal for developing biosensors to detect pathogens. Phages are also workhorse model systems that enabled breakthroughs like identifying DNA as the genetic material.
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Infographic
Our inspiration for creating our phage discovery guide came from a Drexelian student, Bijaya, who discovered her own phage. We took inspiration from Bijaya to take matters into our own hands so more people could discover their own phages. With the rise of antibiotic-resistant bacteria, finding phages is of utmost importance to combat these so-called superbugs. Upon researching the phage discovery process, we only found complex instructions designed for college students or professors. At times, the steps were unclear, lengthy, and redundant. Thus, we dissected the convoluted web to create a simplified version on bacteriophage discovery. The process goes through isolating, purifying, and amplifying the phage lysate in a streamlined manner. This guide is meant for high school or middle school students who want to learn more about phages. Our steps in this infographic are straight to the point and eliminate the unnecessary wordiness of the current phage discovery information on the internet. We hope that this guide can be widely accessible across our global community. The World Economic Forum predicts that designer phages will be in the top ten emerging technologies in 2023 because of the pertinent antibiotic issue. Thus, it is necessary that our search for phages begins now. We can start with this infographic.
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Conclusion
After decades of obscurity, bacteriophages have returned to prominence as medicine confronts the antibiotic resistance crisis. Their unique biology and evolutionary capabilities lend phages to diverse applications. With deeper understanding, engineered phages may profoundly shape microbiomes and human health. The untapped potential of phages continues to drive new innovations.
Sources
https://www.intechopen.com/chapters/66740
https://www.youtube.com/watch?v=YI3tsmFsrOg&t=32s
https://www.ncbi.nlm.nih.gov/books/NBK493185/
https://seaphagesphagediscoveryguide.helpdocsonline.com/3-0-overview
https://intelligence.weforum.org/topics/a1G680000008gwGEAQ
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5899739/#:~:text=Phages%20are%20bacterial%20viruses%20and,receptor%20sites%20(Haq%20et%20al.
https://www.youtube.com/watch?v=SqLx1qNlvRA
https://www.youtube.com/watch?v=DeRk3BiqwtU