Control of microbial growth: Sterilization and disinfectants

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Control of microbial growthSterilization and
disinfectants

• Numerous terms are defined in the text be
  familiar with them. Sterile, disinfection,
  antiseptic, bacteriostatic, and bactericidal,
  plus others.
• Sterile devoid of life. Something is either
  sterile or not.
• Disinfect kill most microbes, especially harmful
  ones, but probably not spores which are
  resistant.
• Disinfectant use on inanimate objects like
  benchtops, etc.
• Antiseptic used to disinfect living tissue must
  be gentler.
• Bacteriostatic keeps bacteria from growing.
• Bactericidal kills them. Sometimes subtle
  differences between bacteriostatic and
  bactericidal.

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Kinetics of bacterial death

• Bacteria not only grow exponentially, but die
  that way too.
• Factors that affect the rate of death include
  temperature, pH, concentration of disinfectant,
  type of microbe, and presence of organic material.

The longer the treatment is applied, the more
that are killed. The more microbes there are, the
longer it will take.
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The Ideal Disinfectant

• Fast and effective, even in the presence of
  organic material (like blood, vomit, feces..)
• Effective, but non-toxic to humans.
• Penetrate materials without damaging them.
• Easy to prepare and stable over time.
• Inexpensive and easy to apply.
• Not stink!

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How do antimicrobial agents work?

• Attack proteins
• Oxidize, hydrolyze, or bind to proteins.
• Change 3-D structure, usually irreversibly ruin
  protein.
• Dissolve membranes or damages cell walls
• Leaky membranes means vitamins, metabolites,
  escape
• Proton gradient across membrane gone, little ATP
  made.
• Wall destroyed, loss of osmotic protection
• Damage to nucleic acids
• DNA denatured or broken, cell cant replicate
• RNA molecules in ribosomes affected

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Microbes vary in susceptibility

• Bacterial endospores hardest of all to kill
• Resist high heat, dessication, chemicals,
  radiation
• Cysts of protozoa
• Resting cells of eukaryotes, similar to
  endospores
• Mycobacteria waxy cell wall provides resistance
• Gram negative bacteria more resistant than G
• Viruses some hard to kill, other easier

Disinfectants vary in effectiveness Not all
disinfectants work equally well against all
microbes. These differences can be measured in
different ways.
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Disinfectants -1

• Halogens Cl, I, Br, Fl
• Chlorine Cl2 gas, hypochlorites (bleach), and
  chloramines (NH2Cl)
• I as tincture or as iodophors such as betadine.
• Phenols and phenolics
• O-phenyl phenol (Lysol), hexachlorophene,
  trichlosan, chlorhexidine
• Alcohols isopropanol, ethanol best at 70-95
• Good at removing lipids w/ attached bacteria from
  skin
• Weakly attacks proteins, cell membranes

phenol
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Disinfectants -2

• Hydrogen peroxide and ozone
• H2O2, stored at 30, used at 3
• new plasma gas sterilizers read article
• Soaps and detergents cationic (Quats), anionic
• Soaps are alkaline salts of fatty acids, weak
• Quaternary ammonium compounds weak but useful.
• Mostly wash away microbes or damage membranes.
• Heavy metals Hg, Ag, Se, Cu
• Used less toxic and corrosive. Hg and Ag
  historically

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Disinfectants -3

• Alkylating agents
• Formaldehyde, glutaraldehyde (effective at high
  pH)
• Ethylene oxide common, toxic, explosive
• Acids and alkalis often microbiostatic
• Acetic and lactic acids in foods
• Benzoate, sorbic acid and propionate
• Other stuff
• Sulfites (control regrowth in wine), nitrites
  (botulism or cancer?), various dyes (selective
  growth media)

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Physical methods

• Temperature
• Cold slows or prevents growth, may kill slowly
• Ultra-cold used to preserve bacteria long term
• Heat denatures enzymes, kills cells
• Moist heat
• Traditional pasteurization does not sterilize
• Standard protocols, time and temperature e.g.
  62.9/ 30 min
• flash pasteurization 71.6 / 15
• Newer UHT sterilizes 74-140-74 in 5

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More on temperature

• Sterilizing boiling, autoclave
• Boiling will not necessarily kill endospores
• Autoclave is steam heat under pressure, so above
  boiling
• 121C, 19-21 psi. Very effective.
• Compare moist heat and dry heat
• Dry heat 170 deg C, near 350 F, for 2 hours
• Water conducts heat much more effectively,
  sterilizes at lower temperature for shorter time.
• Dry heat not useful for liquids!
• Incineration (e.g. flaming loop) has its place
  too.

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Physical methods-2

• Drying cells need water. Remove it, and no
  growth.
• Freeze drying lyophilization also used to
  preserve cultures for long term storage.
• Osmotic pressure/high salt
• Sucks water out of cytoplasm salted meat,
  jellies, etc.
• Radiation
• UV used to sterilize air, surfaces in hospitals,
  etc.
• Ionizing radiation x-rays (electron beam) and
  gamma rays important treatment of plastics,
  various foods.
• Irradiation of meat important tool in food
  safety.
• Microwaves only boil ultrasonics used to break
  cells.

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Physical methods-3

• Filtration
• Membrane filtration thin plastic disks with
  holes of 0.22 or 0.45 micrometers, separate
  liquid from bacteria
• Used to collect bacteria or sterilize liquids
• Solutions of vitamins or proteins can be
  destroyed by heating
• Air filtration HEPA filters used in hospitals
  and also homes to help remove dust-borne
  bacteria, allergens.