viral growth

8/8/2019 Viral Growth

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Viral Growth

Dr Paul [email protected]

BC10M: Introductory Biochemistry

Lecture 7


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Characteristics of Viruses

Non-living agents that infect all lifeforms (Bacteriophages, plant viruses, andanimal viruses)

One virus particle = virion (size: 10

500 nm)

Virus architecture Nucleic acid (DNA or RNA not both!) Protein coat = capsid ofvarious shapes

(isometric, helical, complex) Naked virions vs. enveloped viruses

Genome: ss-/ds-RNA, ss-/dsDNA; linearor circular


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Methods of Study

Much more expensive anddifficult to study animalviruses than bacteriophages

Cultivation in host cellsLiving animalEmbryonated chicken eggsCell or tissue culture (= in

vitro)


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Methods of Study cont:

QuantitationPlaque assay (useful for infective and lyticviruses)

Virion counting with EM

Quantal assay(ID

50or LD

50)

Haemagglutination(e.g.influenza virus)


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Culturing viruses requires

an appropriate host cell.

In this example a bacteriophage

is grown using bacterial cells

as host cells. Animal or plant

viruses can be grown in tissue

culture.


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Plaques in a lawn of

bacterial cells caused

by viruses lysing

bacterial cells.


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Replication Cycle -

OverviewObligate intracellularparasites using host cellmachinery

Very limited number of genes

encode proteins for Capsid formation Viral nucleic acid replication Movement of virus into and out of

cell

Kill or live in harmony withinthe host cell Outside thecell, viruses are inert

Size comparison

(see also Table 13.1)

Fig 13.


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Bacteriophage (Phage)

Obligate intracellular parasites that multiplyinside bacteria by making use of some or allof the host biosynthetic machinery

SignificanceModels for animal cell viruses Gene transfer in bacteria (transduction)Medical applications

Identification of bacteria - phage typing Treatment and prophylaxis???

Examples: T4 and Lambda ()


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Composition and Structure

CompositionNucleic acid

Genome sizeModified bases

ProteinProtectionInfection

Structure (T4)

SizeHead or capsid

Tail

Tail

Tail Fibers

Base Plate

Head/Capsid

Contractile

Sheath


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Infection of Host Cells

Irreversible attachment

Adsorption

LPS for T4

Sheath Contraction

Nucleic acid

injection


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Lytic Bacteriophage

Lytic or virulent phage: Phage that can onlymultiply within bacteria and kill the cell by

lysis. (e.g., T4)

O f


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One-step growth curve of a lytic

bacteriophage


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Lytic Phage Multiplication

Cycle

Eclipse Early genes

Phage DNA

synthesis Late genes

Intracellularaccumulation

Lysis and Release

Total

Phage

Extracellular

Phage

Eclipse

Intracellular

accumulation

phase

Time after Infection

Numbe

rofInfec

tiou

sParticles

Lysis


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Time course of events occurring during one-

step growth curve of bacteriophage T4.


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Lysogenic Bacteriophage

Lysogenic or temperate phage:Phage that can either multiply via thelytic cycle or enter a quiescent statein the bacterial cell. (e.g.,)Expression of most phage genes

repressed

ProphageLysogen


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Lysogenic Phage

DNA integrates into host chromosome Phage DNA = Prophage Infected bacterial cell = lysogenic cell or lysogen

Prophage state can be indefinite

Lysogenic conversion confers new propertiesonto host cells (e.g.: toxin production ofS. pyogenes scarlet fever)

Phage induction converts lysogenic to lyticstate


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Replication cycle in a lysogenic bacteriophage


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Events Leading to Lysogeny

Circularization of the phage chromosome Cohesive ends

Ligase

Closed Circle

Cohesive Ends

Linear Double Stranded Opened Circle


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Events Leading to Lysogeny

Site-specificrecombination

Phage coded

enzyme

Repression of the

phage genome Repressor protein Specific

Immunity tosuperinfection gal bio

gal bio

gal

bio


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Termination of Lysogeny

InductionAdverse conditions

Role of proteases

recA proteinDestruction ofrepressor

Gene expression

ExcisionLytic growth

gal

bio

gal bio

gal bio

gal bio


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Lytic vs Lysogenic Cycle?

Role of repressor

Role ofcro gene product

Role of proteases


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Significance of Lysogeny

Model for animal virus transformation

Lysogenic or phage conversion

Definition: A change in the phenotype ofa bacterial cell as a consequence oflysogenyModification ofSalmonella O antigen

Toxin production by Corynebacteriumdiphtheriae


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Host Range of Phages

Phage host cell interaction usually very specific

Limiting factors for host range

1. Phage has to bind to bacterial surfacereceptors Bacterial surface receptors mutate resistant cell

Lysogenic conversion changes surface receptors andprotects host

1. Restriction modification system of hostcell


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Medical Applications of Phage

I strongly believe phage could become an

effective antibacterial tool - Carl Merril, Chief of theLaboratory of Biochemical Genetics, National Institute of Mental

Health, NIH.

It might be another string on the bow, such thatwhen (conventional antibiotics) fail, heres

something that has a chance of working. But its

not going to be a panacea - Joshua Lederberg, SacklerFoundation Scholar at The Rockefeller University

Reassessment of Medicinal Phage Spurs Companies to Study Therapeutic Uses

American Society for Microbiology News 64:620-623, 1998

M di l A li ti f Ph


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Medical Applications of Phage

Exponential Biotherapies (Rockville, MD) Vancomycin resistant Enterococcus faecium and

Streptococcus pneumoniae

Phage Therapeutics (Bothell, WA) Staphylococcus aureus and Staphylococcus epidermidis

Intralytix, Inc. (Baltimore, MD) Salmonella in meat and poultry

Biopharm Ltd. (Tblisi, Georgia) Infections associated with burns

University of Idaho Escherichia coliO157:H7 in cattle

Reassessment of Medicinal Phage Spurs Companies to Study

Therapeutic Uses. American Society for Microbiology News 64:620-623,

1998.

Phages eyed as agents to protect against harmful E. coli. American

Society for Microbiology News 65:666-667, 1999.

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