BIO 205 - Microbiology: Chapter 6 Study Guide

Chapter 6 Study Guide: Viruses, Viroids, Prions, and Gene Transfer

1. Structural Components of a Virus

All viruses share two fundamental components:

• Nucleic Acid: This is the genetic material of the virus, which can be DNA or RNA. It can be single-stranded (ss) or double-stranded (ds), and linear or circular.

• Capsid: A protein coat that surrounds and protects the nucleic acid. It is composed of protein subunits called capsomeres.

Components found in only some viruses:

• Envelope: A lipid bilayer membrane that surrounds the capsid. It is derived from the host cell's plasma membrane or internal membranes during viral budding. Viruses with an envelope are called enveloped viruses; those without are naked viruses.

• Spikes (Glycoproteins): Projections extending from the envelope (or sometimes the capsid of naked viruses). These are typically glycoproteins that aid in attachment to host cells.

• Enzymes: Some viruses carry their own enzymes within the capsid (e.g., reverse transcriptase in retroviruses, RNA-dependent RNA polymerase in some RNA viruses for replication).

2. Viral Capsid Shapes

Viral capsids are highly organized protein structures that determine the shape of the virion.

• Helical:

  • Description: Rod-shaped or filamentous. The capsomeres are arranged helically around the nucleic acid, forming a hollow tube. The length of the capsid is determined by the length of the nucleic acid.

  • Example: Tobacco mosaic virus, Ebola virus.

• Polyhedral (Icosahedral):

  • Description: Many-sided, often roughly spherical, with 20 triangular faces and 12 vertices. This is the most common polyhedral shape due to its structural stability and efficient packing. Capsomeres form the faces and edges.

  • Example: Adenovirus, poliovirus.

• Complex:

  • Description: These viruses have capsids that are neither purely helical nor purely polyhedral and may possess additional structures. They often have an irregular or intricate shape.

  • Example: Bacteriophages (e.g., T-even phages have an icosahedral head, a helical tail, and tail fibers), Poxviruses (large, brick-shaped).

3. Host Range of a Virus

• Host Range Definition: The specific range of host organisms (e.g., specific species or cell types) that a particular virus can infect.

• Determinants of Host Range:

  • Specific Host Cell Receptors: The primary determinant is the presence of specific, compatible receptor molecules on the surface of the host cell that the virus can bind to. Viral surface proteins (spikes, capsid proteins) must precisely match these receptors, much like a lock and key interaction.

  • Intracellular Factors: After attachment and entry, the host cell must possess the necessary cellular machinery and enzymes that the virus requires for replication and assembly. If these are absent, replication cannot proceed effectively.

4. Basic Steps in a Viral Life Cycle

All viral life cycles follow a generalized pattern, though specific mechanisms vary.

• a. Attachment (Adsorption):

  • The virion physically binds to specific receptors on the surface of the host cell. This binding is highly specific, dictating the virus's host range and tissue tropism.

  • For animal viruses, this typically involves viral glycoproteins (spikes) or capsid proteins interacting with host cell surface molecules (e.g., proteins, carbohydrates, lipids).

  • For bacteriophages, tail fibers often bind to receptors on the bacterial cell wall.

• b. Penetration (Entry):

  • The virus or its genetic material enters the host cell.

  • For bacteriophages: The viral nucleic acid is typically injected directly into the bacterial cytoplasm, leaving the capsid outside.

  • For animal viruses: Common mechanisms include:

    • Endocytosis: The host cell engulfs the entire virion (enveloped or naked) into a vesicle.

    • Fusion: For enveloped viruses, the viral envelope fuses directly with the host cell's plasma membrane, releasing the nucleocapsid into the cytoplasm.

  • Uncoating: For most animal viruses, the capsid is removed, either in the cytoplasm or nucleus, to release the viral nucleic acid. This step is generally absent in bacteriophages, which inject their genetic material directly.

• c. Biosynthesis (Replication and Synthesis):

  • The viral nucleic acid takes control of the host cell's metabolic machinery to replicate its genome and synthesize viral proteins.

  • Genome Replication: The viral genome is copied multiple times.

  • Transcription: Viral mRNA is produced from the viral genome.

  • Translation: Host ribosomes translate viral mRNA into viral proteins, including structural proteins (for capsids) and non-structural proteins (e.g., enzymes for replication).

  • The specific mechanism depends heavily on the type of viral nucleic acid (DNA, RNA, single-stranded, double-stranded, sense, anti-sense, retrovirus).

• d. Maturation (Assembly):

  • Newly synthesized viral nucleic acids and proteins spontaneously or enzymatically assemble into new virions (complete, infectious viral particles).

  • Capsomeres aggregate to form capsids, and the nucleic acid is packaged inside.

  • For enveloped viruses, this process often occurs near a host cell membrane where viral envelope proteins have been inserted.

• e. Release:

  • Newly formed virions exit the host cell.

  • For naked viruses and bacteriophages: Typically by lysis of the host cell, which often kills the cell.

  • For enveloped animal viruses: Usually by budding, where the virion acquires its envelope from the host cell membrane as it exits, often without immediately killing the cell (though it eventually does).

5. Lytic Versus Lysogenic Cycles of Bacteriophages

These describe two distinct pathways temperate bacteriophages can take upon infecting a bacterial cell.

The Lytic Cycle

• Description: The lytic cycle is a virulent pathway that directly leads to the production of new virions and the lysis (destruction) of the host bacterial cell.

• Steps:

  1. Attachment: Phage attaches to specific receptors on the bacterial cell wall.

  2. Penetration: Phage DNA is injected into the bacterial cytoplasm.

  3. Biosynthesis: The phage DNA immediately takes over the host's machinery. It directs the synthesis of phage proteins and replicates the phage genome. Host DNA degradation often occurs simultaneously.

  4. Maturation: Phage components (DNA and proteins) assemble into complete virions.

  5. Release: Newly assembled phages produce enzymes (e.g., lysozyme) that lyse the bacterial cell wall, releasing hundreds of new phage particles. The host cell dies.

The Lysogenic Cycle

• Description: The lysogenic cycle is a temperate pathway where the phage genome integrates into the host bacterial chromosome, becoming a prophage. The host cell survives and replicates the prophage along with its own DNA.

• Steps:

  1. Attachment and Penetration: Same as the lytic cycle, phage DNA enters the host cell.

  2. Integration: Instead of immediately replicating, the phage DNA (now called a prophage) integrates itself into the host bacterial chromosome. This integration is mediated by specific enzymes (integrases).

  3. Prophage State: The prophage remains largely inactive. Most phage genes are repressed by a repressor protein, but one or a few genes may be expressed (e.g., for phage conversion).

  4. Replication: When the host bacterium replicates its chromosome, the prophage DNA is replicated along with it, effectively propagating the phage genome to daughter cells without producing new virions.

  5. Induction: Under certain environmental stresses (e.g., UV radiation, chemicals), the prophage can excise itself from the host chromosome. This event triggers the phage to enter the lytic cycle, leading to biosynthesis, maturation, and lysis of the host cell.

6. Phage Conversion

• Definition: Phage conversion (also known as lysogenic conversion) is a phenomenon where a bacterial cell gains new characteristics or virulence factors when it carries a prophage genome.

• How it Occurs: The prophage, while integrated into the host chromosome, expresses certain genes that alter the phenotype of the bacterial cell. These genes are often responsible for producing toxins, enzymes, or surface antigens.

• Significance: Many medically important bacterial pathogens acquire their virulence by phage conversion. For example:

  • Corynebacterium diphtheriae becomes pathogenic (produces diphtheria toxin) only when infected by a specific lysogenic phage.

  • Vibrio cholerae (causes cholera) produces cholera toxin due to genes carried by a lysogenic phage.

  • Clostridium botulinum (causes botulism) produces botulinum toxin due to a prophage.

7. Consequences of the Lysogenic Cycle for an Infected Bacterial Cell

The lysogenic state carries three significant consequences for the infected bacterial cell:

1. Immunity to Superinfection: A lysogenized bacterium (one carrying a prophage) becomes immune to subsequent infection by the same type of phage (or closely related phages). This is because the repressor protein produced by the prophage prevents the expression of genes in any new, incoming phage of the same type, thus blocking their lytic cycle.

2. Phage Conversion: As described above, the host bacterium may exhibit new properties, often virulence factors, due to the expression of prophage genes.

3. Specialized Transduction: When a prophage excises imperfectly from the bacterial chromosome, it can sometimes carry a small piece of adjacent bacterial DNA along with its own genome. This hybrid phage particle can then transfer this specific (specialized) bacterial gene to a new host cell upon subsequent infection.

8. Contrasting Bacteriophage and Animal Virus Life Cycles

While both follow the general steps of attachment, penetration, biosynthesis, maturation, and release, there are key differences due to the differing structures of bacterial and animal cells.

9. Types of Penetration (Animal Viruses): Fusion and Endocytosis

These are two common mechanisms by which animal viruses enter host cells.

• Endocytosis:

  • Process: The host cell's plasma membrane invaginates, engulfing the entire virus particle (either naked or enveloped) into a vesicle (endosome). This is a general cellular process used for nutrient uptake. The virion is then uncoated within the endosome or after its release into the cytoplasm, often triggered by changes in pH.

  • Viruses: Both naked and enveloped viruses can enter via endocytosis.

• Fusion:

  • Process: The viral envelope directly fuses with the host cell's plasma membrane. This releases the viral nucleocapsid directly into the host cell's cytoplasm, bypassing the need for endosomal vesicles.

  • Required Structure: A virus must have an envelope to undergo fusion. The viral envelope proteins mediate this fusion process.

10. Viral Replication in RNA Viruses

RNA viruses exhibit diverse replication strategies based on the polarity and structure of their RNA genome. They often rely on RNA-dependent RNA polymerase (RdRp), an enzyme not found in host cells.

• a. Single-stranded + sense RNA (ssRNA+):

  • Nature: The genomic RNA acts directly as messenger RNA (mRNA). It can be immediately translated by host ribosomes upon entry into the cell.

  • Replication:

    1. The incoming genomic ssRNA+ is translated to produce viral proteins, including the RdRp.

    2. The RdRp then uses the ssRNA+ genome as a template to synthesize a complementary negative-sense RNA strand (ssRNA-) intermediate.

    3. This ssRNA- intermediate then serves as a template to synthesize new genomic ssRNA+ strands, which can either be packaged into new virions or serve as mRNA for further protein synthesis.

  • Examples: Poliovirus, Hepatitis A virus, Zika virus.

• b. Single-stranded - sense RNA (ssRNA-):

  • Nature: The genomic RNA cannot be directly translated into protein. It is complementary to mRNA.

  • Replication:

    1. The virus must carry its own RdRp within the virion because host cells do not have an enzyme to synthesize RNA from an RNA template.

    2. Upon entry, the viral RdRp uses the ssRNA- genome as a template to synthesize complementary positive-sense mRNA strands (ssRNA+).

    3. These ssRNA+ strands are then translated by host ribosomes to produce viral proteins.

    4. The RdRp also uses the ssRNA- genome as a template to synthesize additional ssRNA+ intermediates, which then serve as templates for the synthesis of new genomic ssRNA- strands.

    5. These new ssRNA- genomes are packaged into new virions.

  • Examples: Influenza virus, Measles virus, Rabies virus, Ebola virus.

• c. Double-stranded RNA (dsRNA):

  • Nature: The genomic RNA is double-stranded.

  • Replication:

    1. The virus carries its own RdRp within the virion.

    2. Upon entry, the dsRNA genome is transcribed by the viral RdRp to produce positive-sense mRNA strands (ssRNA+). This usually occurs within the partially uncoated capsid.

    3. These ssRNA+ strands serve as mRNA for protein synthesis.

    4. They also serve as templates for the synthesis of new complementary negative-sense RNA strands by the RdRp, forming new dsRNA genomes.

    5. The new dsRNA genomes are packaged into new virions.

  • Examples: Rotavirus.

11. Retroviruses

• Distinction from other RNA viruses: Retroviruses are unique among RNA viruses because they replicate through a DNA intermediate. Unlike other RNA viruses that synthesize RNA directly from an RNA template, retroviruses convert their RNA genome into DNA, which then integrates into the host cell's chromosome.

• Enzyme for RNA to DNA conversion: The enzyme that allows retroviruses to convert RNA to DNA is reverse transcriptase (also known as RNA-dependent DNA polymerase).

• Why reverse transcriptase is special: It's special because it defies the central dogma of molecular biology, which states that genetic information flows from DNA to RNA to protein. Reverse transcriptase reverses the flow by synthesizing DNA from an RNA template. This enzyme is crucial for the retroviral life cycle and is a prime target for antiviral drugs.

12. Major Virus Families Discussed in Lecture and Their Key Traits

(Note: As the specific lecture content is not provided, I will outline common major families and their general characteristics that would typically be covered.)

• Herpesviridae:

  • Genome: dsDNA, linear.

  • Envelope: Enveloped.

  • Key Trait: Known for causing latent infections, where the virus can remain dormant in host cells (e.g., neurons) and reactivate later, causing recurrent disease. Wide host range within humans, causing cold sores (HSV-1), genital herpes (HSV-2), chickenpox/shingles (VZV), mononucleosis (EBV), and cytomegalovirus infections (CMV).

• Orthomyxoviridae:

  • Genome: ssRNA-, segmented (usually 8 segments).

  • Envelope: Enveloped.

  • Key Trait: Causes influenza (flu). The segmented genome allows for antigenic shift (major changes) through reassortment, leading to new pandemic strains. Possesses hemagglutinin (HA) and neuraminidase (NA) spikes crucial for attachment and release.

• Retroviridae:

  • Genome: ssRNA+ (diploid - two identical copies).

  • Envelope: Enveloped.

  • Key Traits: Unique for using reverse transcriptase to convert its RNA genome into dsDNA, which then integrates into the host cell's chromosome as a provirus. The Integrated DNA can remain latent for long periods. HIV (Human Immunodeficiency Virus) is the most well-known member, causing AIDS.

• Adenoviridae:

  • Genome: dsDNA, linear.

  • Envelope: Naked (non-enveloped).

  • Key Trait: Commonly causes respiratory infections (e.g., common cold), conjunctivitis (pinkeye), and gastroenteritis. Often used in gene therapy vectors due to its ability to transfer DNA efficiently.

• Poxviridae:

  • Genome: dsDNA, linear (large genome).

  • Envelope: Enveloped; complex morphology.

  • Key Trait: Largest and most complex animal viruses. Replicates entirely in the cytoplasm. Causes smallpox (eradicated) and molluscum contagiosum. Vaccinia virus used in smallpox vaccine.

• Picornaviridae:

  • Genome: ssRNA+.

  • Envelope: Naked.

  • Key Trait: Very small viruses. Includes poliovirus (causes polio), rhinovirus (causes common cold), and hepatitis A virus.

13. Roles of Retrovirus Enzymes

Retroviruses, notably HIV, package several key enzymes within their virions to facilitate their complex life cycle:

• Reverse Transcriptase (RT):

  • Role: Catalyzes the synthesis of double-stranded DNA from the viral single-stranded RNA genome. This enzyme has three activities:

    1. RNA-dependent DNA polymerase: Synthesizes a DNA strand complementary to the viral RNA genome.

    2. RNase H activity: Degrades the RNA template strand once the complementary DNA strand is synthesized.

    3. DNA-dependent DNA polymerase: Synthesizes the second DNA strand, creating a double-stranded viral DNA molecule.

• Integrase (IN):

  • Role: Facilitates the integration of the newly synthesized viral double-stranded DNA copy into the host cell's chromosome. This integrated viral DNA is called a provirus.

  • Mechanism: Integrase cleaves the host DNA and ligates the viral DNA into the gap, becoming a permanent part of the host genome. This step is essential for persistent infection and replication of the viral genome alongside host DNA.

• Protease (PR):

  • Role: After viral genes are transcribed and translated into long polypeptide chains (polyproteins), protease is responsible for cleaving these large precursor proteins into individual, functional viral proteins (e.g., structural proteins like capsid proteins, and other enzymes like reverse transcriptase and integrase).

  • Significance: This cleavage is crucial for the maturation of infectious virions. Without functional protease, viruses assemble incorrectly and are non-infectious.

14. Antiviral Drugs: Targets and Interference Stages

(Note: Specific drugs were not listed in the prompt, so I will describe general categories of antiviral drug targets and the life cycle stages they interfere with. Actual lecture content would include names like Acyclovir, Zidovudine, Ritonavir, Oseltamivir, etc.)

Antiviral drugs aim to inhibit specific viral processes without significantly harming the host cell. They target unique viral enzymes or mechanisms.

• Fusion/Entry Inhibitors:

  • Target: Viral proteins involved in binding to host cell receptors or in the fusion of the viral envelope with the host membrane.

  • Interference Stage: Attachment and Penetration. Prevents the virus from entering the host cell.

  • Example (HIV): Maraviroc (targets host CCR5 receptor), Enfuvirtide (targets gp41 protein, preventing fusion).

• Uncoating Inhibitors:

  • Target: Viral proteins or cell processes required for capsid disassembly and release of the viral genome into the cytoplasm.

  • Interference Stage: Penetration/Uncoating. Prevents the viral genetic material from becoming accessible for replication.

  • Example (Influenza A): Amantadine and Rimantadine (inhibit M2 ion channel, preventing uncoating).

• Nucleic Acid Synthesis Inhibitors (Nucleoside/Nucleotide Analogs):

  • Target: Viral polymerases (DNA polymerase, RNA polymerase, reverse transcriptase).

  • Interference Stage: Biosynthesis (Genome Replication). These drugs are structural analogs of normal dNTPs or NTPs. They get incorporated into the replicating viral DNA/RNA, leading to chain termination or faulty replication.

  • Examples:

    • Acyclovir: (HSV) Targets viral DNA polymerase.

    • Zidovudine (AZT): (HIV) Nucleoside reverse transcriptase inhibitor (NRTI). Lacks a 3'-OH group, causing chain termination during DNA synthesis by reverse transcriptase.

    • Sofosbuvir: (HCV) Nucleotide analog for RNA polymerase.

• Non-Nucleoside/Nucleotide Polymerase Inhibitors:

  • Target: Allosteric sites on viral polymerases, altering their conformation and inhibiting their activity.

  • Interference Stage: Biosynthesis (Genome Replication).

  • Example (HIV): Efavirenz (NNRTI - Non-nucleoside reverse transcriptase inhibitor).

• Integrase Inhibitors:

  • Target: The viral integrase enzyme.

  • Interference Stage: Biosynthesis (Integration). Prevents the viral DNA from integrating into the host chromosome.

  • Example (HIV): Raltegravir.

• Protease Inhibitors:

  • Target: The viral protease enzyme.

  • Interference Stage: Maturation. Prevents the cleavage of viral polyproteins into functional proteins, leading to the assembly of immature, non-infectious virions.

  • Example (HIV): Ritonavir, Saquinavir.

• Neuraminidase Inhibitors:

  • Target: Neuraminidase enzyme on the surface of influenza virus.

  • Interference Stage: Release. Neuraminidase is essential for cleaving sialic acid residues on host cells, allowing newly formed virions to detach and spread. Inhibition prevents efficient release of new virions.

  • Example (Influenza): Oseltamivir (Tamiflu), Zanamivir (Relenza).

15. Structure of Viruses, Viroids, Virusoids, and Prions

These are all infectious agents, but their structures and compositions differ significantly:

• Viruses:

  • Structure: Consist of genetic material (DNA or RNA, single or double-stranded) enclosed within a protein coat called a capsid. Some viruses also have an outer lipid envelope derived from the host cell membrane.

  • Key Characteristic: Obligate intracellular parasites, requiring a host cell's machinery for replication.

• Viroids:

  • Structure: Small, circular, single-stranded RNA molecules that are naked (not enclosed in a capsid) and do not encode any proteins.

  • Key Characteristic: Known pathogens of plants, causing diseases by interfering with gene expression in the host. Replicate using host cell enzymes.

• Virusoids (Satellite RNA):

  • Structure: Similar to viroids – small, circular, single-stranded RNA molecules that are naked and do not encode proteins.

  • Key Characteristic: Unlike viroids, virusoids require a helper virus (a legitimate virus) to replicate. They are encapsulated within the capsid of their helper virus and are replicated by the helper virus's polymerase enzymes. Often plant pathogens (e.g., Hepatitis D virus is a human virusoid-like agent).

• Prions:

  • Structure: Infectious protein particles that lack nucleic acid (DNA or RNA).

  • Key Characteristic: Consist of abnormally folded versions of a normal cellular protein (PrPC, cellular prion protein). The infectious, misfolded form is designated PrPSc (scrapie prion protein). They cause transmissible spongiform encephalopathies (TSEs) in animals and humans (e.g., Mad Cow disease, Creutzfeldt-Jakob disease).

16. How Prions Propagate

Prions propagate through a unique, non-nucleic acid-based mechanism:

• Mechanism: Prions propagate by converting normally folded cellular prion proteins (PrPC) into the abnormally folded, infectious form (PrPSc).

  1. An exogenous (from infection) or endogenously produced (from mutation) PrPSc molecule comes into contact with a normally folded PrPC molecule.

  2. The PrPSc acts as a template or catalyst, inducing PrPC to misfold into the PrPSc conformation.

  3. This newly formed PrPSc then goes on to convert other PrPC molecules, creating a chain reaction that amplifies the amount of PrPSc protein.

  4. PrPSc molecules tend to aggregate, forming amyloid plaques in neural tissue, which leads to neuronal damage and the characteristic spongiform changes observed in TSEs.

17. Vertical Versus Horizontal Gene Transfer

These terms describe how genetic material is passed between organisms.

• Vertical Gene Transfer (VGT):

  • Definition: The transfer of genetic material from parent to offspring. This is the primary mode of gene transfer in multicellular organisms and occurs during reproduction.

  • Mechanism: In eukaryotes, it involves the inheritance of chromosomes during cell division (mitosis and meiosis). In bacteria, it's the replication of the bacterial chromosome and distribution to daughter cells during binary fission.

• Horizontal Gene Transfer (HGT) / Lateral Gene Transfer (LGT):

  • Definition: The transfer of genetic material between organisms that are not parent and offspring (i.e., between different individuals of the same generation or even different species).

  • Significance: HGT is a major driver of bacterial evolution, contributing to antibiotic resistance, virulence, and adaptation to new environments. It allows for rapid acquisition of new traits.

18. Three Types of Horizontal Gene Transfer (HGT) Conducted by Bacteria

Bacteria utilize three main mechanisms for horizontal gene transfer:

• a. Conjugation:

  • Description: Direct transfer of genetic material from one bacterial cell to another through physical contact. This usually involves the transfer of a plasmid (e.g., F plasmid) but can also transfer parts of the bacterial chromosome.

  • Mechanism:

    1. A donor cell (F+ cell, containing the F plasmid) forms a specialized pilus (sex pilus or conjugation pilus) that attaches to a recipient cell (F- cell, lacking the F plasmid).

    2. The pilus retracts, bringing the cells into close contact.

    3. One strand of the F plasmid is transferred from the F+ cell to the F- cell. Replication occurs concurrently in both cells to synthesize the complementary strand.

    4. The F- cell becomes F+ and can then act as a donor.

    5. If the F plasmid integrates into the bacterial chromosome, the cell becomes an Hfr (High frequency of recombination) cell, which can transfer parts of its chromosome to an F- cell.

• b. Transformation:

  • Description: The uptake of naked DNA (DNA released from a dead or lysed bacterial cell) from the environment by a competent recipient bacterial cell.

  • Mechanism:

    1. A donor cell disintegrates, releasing its DNA fragments into the surrounding environment.

    2. A recipient bacterial cell, which must be in a state of competence (physiological state allowing DNA uptake, often induced by stress or growth conditions), binds these free DNA fragments to its surface.

    3. One strand of the DNA fragment is transported across the cell membrane into the cytoplasm.

    4. The incoming DNA can then be integrated into the recipient cell's chromosome via homologous recombination, if the sequences are sufficiently similar.

  • Discovery: Demonstrated by Frederick Griffith in his experiments with Streptococcus pneumoniae.

• c. Transduction:

  • Description: The transfer of bacterial DNA from one bacterium to another via a bacteriophage (a virus that infects bacteria).

  • Mechanism: Occurs in two main forms:

    1. Generalized Transduction:

       • Occurs during the lytic cycle of a bacteriophage.

       • During phage assembly, a fragment of host bacterial DNA is mistakenly packaged into a phage capsid instead of phage DNA. This results in a transducing particle.

       • When this transducing particle infects a new bacterial cell, it injects the donor bacterial DNA. This DNA can then recombine with the recipient's chromosome, transferring bacterial genes.

       • Any gene from the donor chromosome can potentially be transferred.

    2. Specialized Transduction:

       • Occurs during the lysogenic cycle of a temperate bacteriophage.

       • When a prophage excises imperfectly from the host chromosome, it sometimes takes a small piece of bacterial DNA from an adjacent region with it, leaving some phage DNA behind. This creates a phage genome containing both phage and bacterial genes.

       • This