Control of Growth

Lister

• “ Saved more lives by introduction of his system than all the wars of the 19th century together had sacrificed”

• Revolutionized surgery

Definitions

• Sterilization

• Disinfection

• Cide vs stasis

• Sepsis

• Asepsis

Rate of Microbial Death

• Factors that influence effectiveness– # of microbes– Presence of organic material interferes with

action of disinfectant– Temperature– Time of exposure– Characteristic of organism

Potential Risk of Infection

• Critical items-direct contact with body tissues

• Semi critical- contact with mucous membranes but do not penetrate body tissue

• Non critical-contact with intact skin

Actions of Agents

• Damage to membrane-leakage

• Damage to proteins & nucleic acids– Enzymes– Nucleic acids -lethal

Daily Life

• Soaps and detergents

• Hand cleansing

Physical Methods of Control

• Heat– Denatures enzymes

• Moist heat coagulates proteins– Boiling kills vegetative forms of pathogens

Autoclave

• Uses temperature above boiling water• Steam under pressure• Preferred method unless material is damaged• Higher the pressure, higher the temperature• Need direct contact with steam• All air is evacuated from chamber• 15 psi at 121 C for 15 mins

Autoclave

• Prions- protein only– 134 C for 18 mins– Soak in 1N NaOH for 1 hour

• Flash sterilization-at 134 C for 3min– Used for individual instruments

• Packaging

• Use of indicators– Biological indicators-endospores

Physical Methods of Control

• Pasteurization for heat sensitive products– Mild heating to kill pathogens

• Dry heat– Kills by oxidation– Flaming or hot air

Filtration

• Passage of liquid through screen device

• Pores small enough to retain microbes

• Sterilize heat sensitive materials

• Negative-uses vacuum

• Positive uses pressure

• HEPA hoods & TB rooms

High Pressure

• Can alter structure of proteins

• Kill vegetative forms

• Endospores relatively resistant

• Osmotic pressure reduces availability of water

Radiation

• Ionizing radiation-less than 1nm– Gamma rays-cobalt or electron beams– X rays

• Principle-ionization of water

Radiation

• Nonionizng radiation– UV light– Causes pyrimidine dimers

• T-T, T-C

Chemicals

• Considered pesticides• Regulated by EPA• Sterilants-sterilize in 6-10 hrs• High level disinfection do not kill all endospores

• Intermediate level kill bacteria and some viruses• Low level – general purpose

Microbial Characteristics

• Gram negatives more resistant– External layer of lipopolysaccharide– Porins –holes in cell wall, selective

• Mycobacterium-lipids in cell wall

• Viruses without a lipid layer more resistant

Antimicrobial Chemotherapy

History

• Paul Ehrlich- magic bullets• Chemotherapeutic agent

• Antimicrobial• Antibiotic• Synthetic drugs produced in lab

History

• Sulfa drugs- wide spread use in 1935• Fleming-penicillin in 1928

• Waksman- bacteria from soil- Streptomyces griseus-streptomycin

Selective Toxicity

• Harm microbe without causing significant damage to host

• Penicillin

• Streptomycin to tx TB

Drug Administration

• Effective concentration of abx to site of infection-route

• Topical-infection at body surface– Athlete’s feet

• Systemic-IV fastest but need hospitalization• IM intra muscular- peak levels in BS within 15

mins

Drug Administration

• PO- enters GI then into BS– Painless, but slower and not as effective– Only fraction enters BS– Must follow directions– Great for outpts

Drug Distribution

• Barriers to fast distribution: cell membrane & proteins– Prevent entry or bind drug

• CM of cells around BS can prevent free passage into tissues from blood vessels

Drug Distribution

• Drug binding proteins

• Penicillin: about 65% reversible bound to albumin

• Race between distribution of drug and elimination

Elimination of Drugs

• Converted metabolically to another compound: liver

• Excreted

• Penicillin excreted rapidly via kidneys

• Within 90 minutes, most in urine

• Take regularly and often or

• Resistance can develop

Drug Resistance

• Natural or acquired resistance• Species lacks target: penicillin attacks

peptidoglycan in cell wall

• Many can’t enter gram negative cell wall

Spectrum of Activity

• Range of microbes the agent acts against

• Broad –gram positive & negative

• Narrow –small number of microbes

• Prevents destruction of normal flora

Acquired Resistance

• Broad spectrum antibiotic changes normal flora:

• Mutations and genetic exchange among bacteria

• Antibiotics favor resistant strains but do not cause genetic changes

Mechanisms

• Alteration of target of drug action

• Alteration of membrane permeability

• Efflux or pumps out the drug

Mechanism

• Development of enzymes: most common– Destroy or inactivate drug– B-lactamase-several

Genetics

• Mutations on chromosome• Develop gradually over years of antibiotic

usage• Need higher & higher concentrations of

drug• Eventually becomes useless

Plasmid Borne Genes

• Occurs almost immediate via genetic recombination

• R plasmids or factors

• Penicillin resistance from beta- lactamase is an example

Slowing Use of Antibiotics

• 3/4 of all hospital pts receive several courses of abxs-over kill

• 50 million of 150 million outpatient antibiotics are unneeded

• Maintain high levels in body long enough to kill all pathogens

Slow Resistance

• Limit use of antibiotics in animals

• Transmission of drug resistant organisms from animals to humans

• Some countries drugs sold over the counter

Drug Dosage

• Depends upon route of administration, drug distribution, drug elimination

• Effective concentration depends upon susceptibility of organism to drug

• Kirby-Bauer method– MIC: minimum inhibitory concentration– Microbial sensitivities

Kirby Bauer Method

• Disk diffusion method-standardized

• Agent spread over MH plate

• Disks impregnated with antibiotic

• Agent diffuse out, lower MW faster

• Zones of inhibition-clear areas around disk

• Standardized for S, R I

Targets of Antimicrobial Drugs

• Bactericidal-kills

• Bacteriostatic-inhibits growth– Phagocytosis & antibodies kill organisms

• Inhibition of cell wall synthesis

Cell Membrane

• Membrane of bacteria & fungi differ somewhat from animal cell membranes– Some selective toxicity but not ideal

Inhibition of Protein Synthesis

• Selective toxicity depends upon difference between ribosomes

• Bacteria-70S (30S & 50S) & animals -80S– Human 40S & 60S but mitochondria have 70S– Side effects do occur

Inhibition of Nucleic Acid Synthesis

• Rifampin binds to bacterial RNA polymerase– inhibits RNA synthesis- TB– Colors urine and contacts orange