Antimicrobial Agents

• Antimicrobial agents (AMA) prevent the spread of diseases.

Control of Microbial Growth Terminology

• Sterilization completely removes or kills all vegetative cells, endospores, and viruses from a targeted item or environment.
• Aseptic technique maintains sterility in clinical or microbiology lab procedures through combined protocols.
• Commercial sterilization destroys pathogens responsible for food poisoning.
• Disinfection inactivates most microbes on inanimate or non-living surfaces.
• Disinfectants achieve disinfection.
• Antisepsis inactivates most microbes on living tissues.
• Antiseptics achieve antisepsis.
• Sanitization cleanses inanimate objects to remove enough microbes for public health safety.
• Degerming reduces microbial numbers significantly by gently scrubbing living tissue, like washing hands.
• Microbial control that kills has the suffix "-cide" like virucide.
• Microbial control that only inhibits microbial growth has the suffix "-static."
• Removing a microbe from a "-static" microbial control allows it to resume growth, like bacteriostatic.

Factors Affecting Antimicrobial Agent Effectiveness

• Effectiveness is affected by the amount of time the agent needs to be in contact with the item.
• Effectiveness is affected by the concentration of the agent needed.
• Effectiveness is affected by other conditions that limit contact, such as organic matter.
• Regularly cleaned floors require less contact time and more diluted disinfectant.

Modes of Action of Antimicrobial Agents

• Physical methods non-specifically kill cells by disrupting membranes or damaging cellular proteins/nucleic acids.
• Chemical methods can work non-specifically or specifically to kill cells.
• Chemical safety is critical when used with humans and the environment.

Efficiency of Anti-Microbial Agents

• A disc-diffusion assay assesses the relative effectiveness of a chemical-based antimicrobial agent.
• An effective chemical agent exhibits a larger zone of inhibition in a disc-diffusion assay.

Sterilization Situations and Methods

• Sterilization is required in microbiology labs for equipment, glassware, media, etc.
• Sterilization is required for hospital items that contact susceptible people.
• Moist heat (autoclave) combines heat and steam, useful in labs for sterilizing anything wettable like media, agar, glassware, and lab waste.
• Dry heat (direct flaming or oven) can sterilize anything heated, like an inoculation loop.
• Slow-freezing forms ice crystals that can damage proteins, useful to control eukaryotic parasites.
• Ionizing radiation (gamma or X-rays) has limited use due to toxic effects on humans, but can penetrate tissue, paper, and plastic.
• Ionizing radiation is used for tissues for transplantation, materials made of paper/plastic, and food.
• Non-ionizing radiation (UV rays) only sterilizes the surface, and is used to purify water and in germicidal lamps.
• Filtration passes a liquid or gas through a filter with pores to physically remove microbes.
• Filtration is used for solutions that cannot be heated, like those with vitamins or antibiotics, and for air and water filters.
• Alkylating agents are the only chemical capable of sterilization but is carcinogenic.
• Two types of Alkylating agents are 2% glutaraldehyde (liquid) and ethylene oxide (gas), used for surgical equipment and hospital surfaces.

Disinfection (disinfectants and antiseptics)

• Disinfection cleaning is required for disease prevention after an outbreak, in hospital settings, and for deeper household cleaning.
• Filtration can sterilize media containing vitamins prior to inoculation
• 2% glutaraldehyde or ethylene oxide sterilize a hospital mattress, although gas can sterilize the inside of the mattress

Chemical Methods of Disinfection

• Phenol and phenolics are effective against gram-positive bacteria, fungi, and technically yeast.
• Phenol and phenolics are not effective against gram-negative bacteria, viruses, and endospores.
• Lysol is used for house cleaning disinfectant.
• Triclosan is used in toothpaste and anti-bacterial soaps.
• Heavy metals are effective against all cells except for endospores, but toxic to humans, and found in products embedded with silver, zinc, or copper.
• Mouthwash contains silver nitrate.
• Halogens (iodophor or modified iodine) are effective against all cells, excluding endospores.
• Halogens are used for water purification disinfectant.
• Halogens are used for hand scrubs before surgery or cleaning skin wounds (betadine) antiseptic.
• Halogens (chlorine and chlorine derivatives) are effective against all cells except endospores, with one exception.
• Halogens are used in water purification (e.g., swimming pools, drinking water) and surface cleaning disinfectant.
• Bleach is capable of removing endospores.
• Alcohol (in a solution with water) is effective against bacteria, fungi, enveloped viruses, but not non-enveloped viruses, and induces endospore formation.
• 70% ethanol is used to clean surfaces and skin surfaces.
• Surfactants (QUATS) are effective against bacteria, fungi, and enveloped viruses, but not non-enveloped viruses and endospores.
• Lysol (for housecleaning) and Mouthwash are Surfactants.
• Peroxygens are effective against all cells except endospores.
• Higher concentrations of peroxygens are needed against bacteria that can produce catalase.
• Hydrogen peroxide (in liquid and gas forms) is a disinfectant used to clean hospitals.
• Hydrogen peroxide is an antiseptic found in acne medication and toothpaste.

Anti-Microbial effectiveness by type

• Bleach is most effective for cleaning a daycare after a Norwalk viral outbreak.
• QUAT is appropriate for cleaning a toilet every two weeks at home because it is a sterilizing agent unnessecary

Sanitization Situations and Methods

• Situations requiring this level of cleaning include general, daily cleaning
• Pasteurization (mild heating) increases the shelf life of food items like dairy products, alcohol, and juices
• Refrigeration (bacteriostatic only) provides short-term storage of food items or bacterial cultures
• Lyophilization (bacteriostatic only) - no ice crystals, is a freeze-drying (rapid freezing combined with desiccation) for long-term storage Applications
• Osmotic pressure (high concentrations of sugar or salt) is used in salted foods, honeys, and jams
• Surfactants - soaps cause regular mechanical removal of microbes through emulsification of oils and microbes into small drops, and then rinsed away

Antimicrobial drugs

• Antimicrobial drugs (AMD) are chemicals used to treat diseases
• Narrow spectrum antibiotics target only specific subset of pathogens
  • Use if the causative pathogen has been identified
  • Minimizes damage to normal flora (resident microbiota)
  • Only kills pathogens and doesnt kill other bacteria
• Broad spectrum antibiotics target a wide variety of pathogens
  • Use if needs to target a wide variety of pathogens quickly
  • Treatment needs to start immediately
  • Mixed infection with multiple causative pathogens
  • Prevention of infection of invasive surgical procedures
  • Failure in treatment by a narrow spectrum antibiotic

Antimicrobial drug risks

• Risks include:
  • Normal flora can be targeted which can increase the risk of developing a opportunistic infection (e.g., yeast infection)
  • Contributes to antibiotic resistance

Kirby-Bauer Technique

• Used to determine the effectiveness of an antibiotic; similar to the disc diffusion assay where a more effective antibiotic would have a larger zone of inhibition
• Unlike for antiseptics/disinfectants, the effectiveness of an antibiotic must reach a specific threshold (results are analyzed using antibiotic susceptibility tables)

Major Modes of Action for Bacterial Antibiotics

• Cell wall synthesis inhibitors specifically target the peptidoglycan layer
  • Prevents the formation of cell wall, making it more susceptible to cell lysis
  • Less side effects for humans
  • Example: penicillin
• Membrane function inhibitors have detergent-like properties, disrupting outer and plasma membranes
  • Can be toxic to humans if ingested, so used only in topical creams
  • Example: polymyxin
• Protein biosynthesis inhibitors target ribosomes, preventing protein synthesis
  • Less side effects as prokaryotic ribosomes are different from eukaryotic ribosomes
  • Example: Tetracycline (broad spectrum antibiotic)
• Nucleic acid synthesis inhibitors interfere with DNA replication or block RNA polymerase
  • Typically have more side effects, so used with other antibiotics
  • Example: Rifampin blocks RNA polymerase
• Metabolic pathway inhibitors act as antimetabolites and competes with regular substrates for the same enzyme, leading to loss of function
  • Side effects depends on the actual antibiotic
  • Example: sulfonamides block the synthesis of folic acid

Antimicrobial drug resistance overview

• Genetic variation will lead to some cells being resistant to drugs
• Susceptible cells will die through antimicrobial drugs
• Resistant cells will survive through antimicrobial drugs
• Over time, the proportion of drug-resistant cells will increase

Mechanisms through which bacteria can resist drugs

• Efflux: Bacterium contains pumps that can transport the drug out of the cell
  • Found in both gram-negative and gram-positive bacteria
• Drug modification or inactivation: Chemically modify or degrade the antibiotic, making the antibiotic inactive
  • Found in both gram-negative and gram-positive bacteria
• Prevention of cellular uptake: Smaller pores of gram-negative bacteria prevent the antibiotic from entering the cell
• Target modification: the drug targets change over time due to antibiotic selection
  • Example: the ribosomes in tetracycline-resistant bacteria have been modified through mutations over time to remain function, but not affected by tetracycline(antibiotic)
  • Found in both gram-negative and gram-positive bacteria

Anti-viral drugs

• Viruses are obligate intracellular pathogens that requires host cell machinery to replicate; targeting viruses would also target our own cells ; use hosts resources (e.g. Ddrp, DNA, Polymerase, ribosomes)

Viral drugs

• Neuraminidase inhibitors are used to target influenza viruses and prevents release of the virus from the host cell
• HIV drugs:
  • Reverse transcriptase inhibitors
  • Integrase inhibitors prevent the formation of the provirus
  • RdRp inhibitors for RNA viruses

Phage therapy

• Phage therapy involves the collection and growth of bacteriophages that target the pathogen
• Can be administered intravenously

Phage therapy advantages

- Will only target bacteria
- Specificity of bacteriophages means that normal flora is unaffected
- Bacteriophages are also capable of evolution/ butation along with bacteria