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Microbes and You > Bacteriophage L5 > Flashcards

Bacteriophage L5 Flashcards

1
Q

what is bacteriophage

A

obligate intracellular bacterial parasites – strictly reliant on a host to complete its life cycle and reproduce new phage particles

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2
Q

what is ubiquitous environmental distribution

A

wherever there are bacterial hosts there will be be phage which will infect them

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3
Q

how many bacteriophage are present

A

Most abundant biological entity on Earth

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4
Q

what are tailed phages

A

Caudovirales

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5
Q

what are examples of caudovirales

A

Myoviridae
Podoviridae
Siphoviridae

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6
Q

what are Podoviridae

A

short tailw

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7
Q

what are Siphoviridae

A

non-contractile tail

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8
Q

what are Myoviridae

A

long contractile tail

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9
Q

what are the two groups of phage in terms of life cycles

A
  1. Temperate phage

2. Lytic phage

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10
Q

examples of temperate phage

A

Podoviridae P22

Siphoviridae λ

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11
Q

examples of lytic phage

A

Myoviridae T4

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12
Q

what happens to myoviridae when contact with bacterial host

A

conformational change in tail proteins – contraction allows injection of DNA into bacterial host

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13
Q

what happens in phage lifecycles - temperate phage

A

integrates in host chromosome (stable genetic element-prophage)

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14
Q

what does phage lifecycle enhance

A

Enhancement of host virulence (lysogenic conversion)

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15
Q

what can lifecycle control

A

Can control host gene expression (e.g. serotype conversion)

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16
Q

what is the driver of bacterial genome evolution

A

Phage lifecycles – temperate phage and lysogeny

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17
Q

what happens in DNA replication of phage lifecycles

A

Every time divides the phage is passed onto next generation

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18
Q

what happens if there is DNA damage in phage lifecycles

A

phage will sense that and actively excise itself and start producing more phage particles and get out of that cell

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19
Q

what happens in λ phage integration

A

phage integrase mediates recombination between attP and attB
Phage DNA integrates inside attB in bacterial chromosome
Phage needs to bring about certain DNA sequences that are recognised as integration sites in attL and attR
circular DNA gets linearized and integrated in
Integrated prophage with attBP and attPB sites either sides

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20
Q

what happens in λ phage excision

A

Integrated prophage + bacterial lysogen
Phage uses excisionase, promotes recombination between attL and attR, allows prophage genome to excise
Once excised recircularises and starts DNA replication, phage protein production and new phage particles are created

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21
Q

what is Int

A

phage integrase

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22
Q

what is attB

A

bacteria attachment site

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23
Q

what is attP

A

phage attachment site

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24
Q

what is the problem with lysogens

A

Acquisition of virulence factors can be a consequence of lysogeny

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25
Q

what happens in the adsorption process of lytic pathway

A

initial adsorption process, phage attaches to bacterial cell surface, triggers conformational change

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26
Q

what happens in the injection stage of lytic pathways

A

inject DNA, degrades hosts DNA so is a pool of nucleotides to create phage genomic DNA

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27
Q

what happens in the synthesis protein production stage of lytic stage

A

DNA synthesis, gene transcription and protein synthesis (produce phage structural proteins which make up the head and tail)

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28
Q

what happens in the assembly stage of lytic cycle

A

Assembly of phage particles

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29
Q

what happens in the lysis stage of lytic cycle

A

Lysis of bacterial cell using endolysins - multiple phage burst out, release 1-> 200 particles
repeat process as long as host persists

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30
Q

what do phage packaging signals do

A

Some phage have packaging signals, will only package a certain amount of DNA into head based of sequence signal recognition in the phage head

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31
Q

what happens to phage packaging if there are no signals present

A

Other phage will keep packing DNA till head is full

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32
Q

what happens when phage degrades hosts genomic DNA

A

phage degrades hosts genomic DNA there may be fragments which are packaged in head, creates a phage which can attach to a new host, and transfers bacterial genes (rather than infecting with its own DNA)

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33
Q

what is used in generalised transduction

A

any sequence, can be large (some transducing phages >200 kb)

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34
Q

what is specialised transduction

A

imprecise excision of prophages from bacterial chromosome
Will use excisionase to cut self out, may take a gene or two that’s flanking either side of phage chromosome, e.g. a resistance gene

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35
Q

what does phage transduction contribute to

A

Major contributors of horizontal gene transfer

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36
Q

what is phage transduction useful for

A

Very useful genetic tools for bacterial studies

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37
Q

what is a capsid head made of

A

Icosahedral head structure, made up of multiple repeats of major capsid protein

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38
Q

what does capsid head associate with

A

Gp24
Hoc
Soc

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39
Q

what is Gp24

A

minor capsid

40
Q

what is Hoc

A

highly-antigenic outer capsid

41
Q

what is Soc

A

small outer capsid

42
Q

what are Hoc and Soc utilised for

A

phage display

43
Q

what do capsid heads do

A

Encapsidates phage DNA, keeps it protected from environment

44
Q

what is the neck protein

A

portal protein (Gp20)

45
Q

what proteins seal the portal vertex

A

Gp13 and Gp14

46
Q

what is the necks critical role

A
  • head assembly
  • genome packaging
  • head-neck-tail connection
  • genome ejection
47
Q

what do some necks of phage have

A

‘whisker’-type structures

48
Q

what are Gp15

A

Tail terminator protein attaches head-to-tail

49
Q

what are Gp3

A

Tail tube terminator protein

50
Q

what happens when reach point where phage binds to its bacterial host

A

if all conformational requirements met, see contraction of proteins in neck, creates pressure in head and forces out DNA

51
Q

what does the tail tube consist of

A

repeating units of tail sheath protein (Gp19) surrounding tube protein (Gp18)

52
Q

what does the baseplate & tail terminate with

A

Terminates with baseplate structure

53
Q

what does the phage bind to in bacterial host

A

Phage binding to bacterial hosts involves fibres, spikes, and adhesins

54
Q

what is baseplate & tail involved in

A

host recognition and adsorption

55
Q

what do baseplate and tail degrade

A

surface structures e.g. host LPS and peptidoglycan

56
Q

what do baseplate and tail allow access to

A

cognate receptor – fibre-positioning

57
Q

what does conformational change in baseplate trigger

A

DNA ejection

58
Q

what are tail proteins involved in

A

host-adsorption

59
Q

what can tail proteins do

A

Capable of degrading diverse targets (LPS, PG, and lipids)

60
Q

what are needed for catalytic activity

A

Conserved catalytic residues Glu-359, Asp-392, and Asp-395 needed for catalytic activity

61
Q

what does trimeric structure have potential to do

A

agglutinate bacteria

62
Q

what has potential antimicrobial applications

A

Recombinant tail spikes, needles, and neck-appendages

63
Q

what can be used as antimicrobial proteins

A

Phage tails target LPS can use them as antimicrobial proteins

64
Q

when can a cell no longer be infected

A

Can no longer infected as the LPS has been modified by phage that integrated into bacterial chromosome

65
Q

what are examples of bacterial resistance mechanisms

A
  • phase variation
  • restriction/modification
  • CRISPR systems
66
Q

what is phage variation bacterial resistance mechanism

A

switch off the gene that the phage targets

67
Q

what is restriction bacterial resistance mechanism

A

if phage DNA does not have correct methylation pattern will be cut by restriction enzymes

68
Q

what is CRISPR

A

Clustered regularly interspaced short palindromic repeats

69
Q

what does CRISPR provide

A

defence against mobile genetic elements e.g. phages or plasmid
Prokaryotic adaptive immunity

70
Q

what happens in CRISPR-mediated immunity

A

Phage infect its bacterial host
CRISPR invoke adaptation process
if have a similar phage with the same DNA sequence as spacer = expression of CRISPR RNA
CRISPR RNA bind to bacteriophage DNA
Cas9 endonuclease recognises it has a CRISPR RNA attached to it, will cut the DNA up
Degradation of bacteriophage DNA and bacterial cell stopped infection

71
Q

what is the CRISPR adaptation process

A

chop up bits of phage DNA and incorporate into CRISPR array

72
Q

what are the Cas1 DNA endonuclease and Cas2 RNA endonuclease involved in

A

acquisition of CRISPR spacers

73
Q

how many types of CRISPR-mediated immunity

A

Multiple types of CRISPR systems that all lead to adaptive immunity

74
Q

how are phage studied in the laboratory

A

classical experiments:

  • adsorption assays
  • one-step growth curves
  • host-ranges
  • pulsed-field gel electrophoresis
  • transduction assays
75
Q

what is PFGE

A

pulsed-field gel electrophoresis

76
Q

how is the morphology of phage found

A

scanning/transmission electron microscopy

77
Q

what does whole genome sequencing provide

A

useful information regarding genes and proteins

78
Q

what is eclipse in phage graph

A

no phage amplification

79
Q

what are therapeutic applications

A

targeting pathogenic microorganisms with phages

80
Q

what are the stages in the phage graph

A

Eclipse
Latent period – nothing then fast increasing
Rise period – when start to plateau
Top of graph is burst then it plateaus

81
Q

what does T4 DNA ligase do

A

if have a linear piece of DNA want to circularise it or ligate it together

82
Q

what is the central dogma of molecular biology

A

DNA > RNA > Protein

83
Q

what helped establish central dogma of molecular biology

A

phages

84
Q

what is phage associated to

A

Historical association with molecular biology and genetics

85
Q

which were the first ever sequences genomes

A

ɸχ174 and MS2

86
Q

what are phages a source of

A

a wide-range of commonly used enzymes e.g. T4 DNA ligase, Phi29 DNA pol

87
Q

what is a threat to modern civilisation

A

Antimicrobial resistance

88
Q

what are the multidrug resistant strains

A
  • Staphylococcus aureus (MRSA)
  • Enterobacteriaceae
  • Pseudomonas aeruginosa
  • Acinetobacter baumanii
89
Q

what are phages an alternative to

A

antibiotics

90
Q

what helps the bacteria burst out of cell

A

endolysins

91
Q

is there chance of phage resistance developing

A

instantaneous is very little chance of resistance developing

92
Q

what are phage lysins effective against

A

Gram Positives - Catalytic domain and cell binding domain as soon as comes in contact with bacterial cell see disruption of cell membrane and pops

93
Q

why are phage lysins not effective on gram-negative bacteria

A

Outer membrane and periplasm inner membrane etc in gram-negative phage lysins doesn’t work on

94
Q

what do endolysin based products treat

A

eczema, rosacea, acne, and skin-irritation

95
Q

what are advantages of clinical development issues

A
  • Highly-specific
  • target MDR pathogens
  • Selective decontamination
  • Self-amplification at site of infection
  • potential to disrupt biofilms
  • encode vast array of proteins with antibacterial properties
  • evolve to overcome bacterial resistance
96
Q

what are the disadvantages of clinical development issues

A
  • need knowledge of pathogen
  • possible lysis-induced toxin release
  • polymicrobial infections
  • no existing regulatory framework in place
  • genetic heterogeneity as a natural consequence of replication

Microbes and You (14 decks)

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