Bacteriophage, Optimum Growth Temp, Human Microbiome

• Viruses that infect bacteria are called

  bacteriophages
• lytic phage


  takes over metabolism of the host cell completely (makes a copy of itself) resulting in the lysis of the cell
• steps of lytic cycle

  attachment, entry (injection), replication, assembly, release
• full steps of lytic cycle


  1. attachment of the bacterial cell wall

  2. injection of viral DNA into bacterial cytoplasm

  3. Viral genome then commands cell to produce viral proteins - used fo assembly of phages

  4. once assembly is complete -> cell lyses and releases phages which attack other bacterial cells and begin replication cycle anew
• An example of a lytic phage is

  T3 (infects E coli)
• Temperate (latent) phages / Lysogenic


  live in harmony with the host cell and multiply as the host cell multiplies
• Lysogenic steps of infection


  1. Attachment 2. Injection

  3. Viral nucleic acid becomes integrated into the host cells chromosome. Takeover of cellular processes is not immediate

  4. Replication of phage is delayed

  - can remain like this for long times

  5. Under stressful condition -> lysogenic prophage switches to the lytic mode to try and escape from the cell before cell death
• Lysogeny


  a state in which phage DNA is incorporated into the host cell without lysis

  - results in a prophage that can remain dormant for long periods of time
• An example of a lysogenic phage


  Lambda (infects E coli)
• Examples of stimuli that induce switch from lysogenic to lytic phage


  exposure to environmental stress factors like radiation & toxic chemicals
• A difference between lytic and lysogenic

  Lytic- viruses force the host cell to produce more viruses and lysis of host

  Lysogenic- viral DNA inserts itself into the host cells DNA (prophage) and is replicated during mitosis
• Plaques


  lysis of bacterial cells growing on an agar plate produces a clearing that can be viewed with the naked eye
• Plaque assay


  uses plaque formation as a means to calculate phage concentration in a given sample
• phage titer (density)


  concentration of infectious phage particles per milliliter of growth medium
• plaque assay similar to standard plate count

  employs serial dilution to produce countable plates needed for later calculations

  At suitable dilutions of phage particles -> each plaque that forms represents the point at which a phage particle was deposited

  similar to plate count of bacteria where each colony formed represents a single bacterium in the initial inoculum
• Process

  1. Diluted phage added to small amount of broth culture

  2. This phage host mix is added to soft agar tube and then poured onto nutrient agar = agar overlay

  3. Consistency of solidified soft agar -> immobilize bacteria while allowing smaller bacteriophages to diffuse short distances and infect surrounding cells,

  4. During incubation -> phage host produces lawn of growth on plate in which plaques appear where contingous cells have been lysed by the virus

  5. At suitable dilutions of phage particles -> each plaque that forms represents the point at which a single phage particle was deposited

  -> pfu/ml (phage particles per ml / plaque forming units)
• Enumeration


  bacteriophage counts made by counting # of plaques formed on host bacteria grown on agar
• Pfu formula


  (# of plaques) / (vol plated in ml)*(dilution plated)
• Can we grow bacteriophages on NA plates? Why or why not?

  We can not grow bacteriophage on NA plates because a virus requires a host cell in order to replicate.
• Adult is colonized by

  trilliions of microorganisms
• There are _ microbial cells for every _ human cell

  3 / 1
• All these microorganisms add up to _____ in healthy adult

  1.5 -2.7 kilos
• human microflora/microbiome
• bacteria are most numerous in


  large intestine (the colon)
• bacteria heavily colonize


  skin, oral cavity, scalp, and nostrils
• human body contains many environmental niches


  "niche" refers to specific ecological roles or habitats within the human body where environmental conditions can vary. It indicates the various places or environments within the body that have different conditions such as water availability, salinity, temperature, pH levels, and the presence of fats and oils.
• Great diversity of bacteria associated with a healthy human

  around 1000 diff species
• Staphylococcus


  found on normal skin (external surfaces and in hair follicles), facultative anaerobes, chemoorganotropic, colonies are opaque and pearly white, may have yellow gold pigmentation
• Pathogen most often encountered in Staphylococcus is


  S aureus (carbuncles, osteomyelitis, bacterial pneumonia, wound infection, food poisoning, toxic shock syndrome)
• Staphylococcus epidermidis

  common commensal of health skin, can produce opportunistic pathogen
• Staphylococcus saprophyticus


  often found in the normal flora of female genital tract and perineum; produce UTI (urinary tract infections) in females
• bacterial cells produce ___ during aerobic respiration

  H2O2
• if hydrogen peroxide accumalates


  cell becomes toxic
• Aerobic and facultative anaerobic bacteria possess an enzyme called


  Catalase
• catalase


  breaks down hydrogen peroxide to water and oxygen
• Catalase Test(differential & selective?)

  (differential) to determine whether or not the bacterium produces the enzyme catalase
• Catalase + bacteria (bubbles = production of O2)


  Staphylococcus (epidermis G+) and Micrococcus
• Catalase - bacteria

  streptococcus & enterococcus (faecalis G+)
• Staphylococcus

  Gram positive cocci in grape-like clusters
• Streptococcus

  mutansNeisseria genusfound on teeth, gram positive

  gram negative