ch 5 control of microbial growth

approaches to control

control mechanisms are either physical or chemical

• may be combination of both

• physical methods

  • heat

  • irradiation

  • filtration

  • mechanical removal (wiping the surface of a table with disinfectant wipe)

• chemical methods

  • use antimicrobial chemicals like disinfectants and antiseptics

different types of treatments

• sterilization: destroys all microbes (including endospores and viruses)

• disinfection: removes most microbes, not all

  • disinfectants = used on inanimate objects and surfaces

  • antiseptics = used on living tissues, like mouthwash

• pasteurization: brief heating to destroy spoilage organisms and pathogens, in milk the process didn’t destroy everything but hopefully these things

• preservation: process to delay spoilage of perishable products (food, cosmetics)

situational considerations

microbial control methods depend upon situation and level of control required

daily life

routine control - to wash and scrub with soaps and detergents

• mechanically removes microbes

• beneficial skin microbiota not adversely affected by regular use

• hand washing with soap and water most important step in stopping spread of many infectious diseases

hospitals

minimizing microbial population is very important

• danger of healthcare-associated infections HAIs

• patients more susceptible to infection, keep hospitals free of microbes

• may undergo invasive procedures (surgery)

  • instruments sterilized to avoid introducing infection to deep tissues

  • operating room - cleaning critical

  • surfaces and air

• more pathogens found in hospital

  • most resistant to antimicrobial agents

microbiology laboratories

aseptic techniques used to prevent contamination of samples, self, laboratory

treatment of media both before and after use

• before: eliminates unwanted organisms

• after: prevent contamination of environment

washing hands, flaming inoculating loops

food and food production facilities

goal: retain quality of perishables longer (increase shelf-life)

• methods used to destroy, remove, or inhibit contaminating microbes

  • heat: most common

    • can alter flavor, appearance of products

  • irradiation: approved to treat certain foods

  • preservatives: chemical additives to prevent spoilage

water treatment facilities

ensure drinking water is free of pathogens

chlorine traditionally used to disinfect water

• some organisms resistant to chemical disinfectants like chlorine

  • cryptosporidium parvum (causes GI disease: diarrhea) - cyst very resistant to chemical disinfectants, including chlorine

  • filtering will remove the cysts from water and chlorination will remove bacteria (they’re still highly resistant)

selection of antimicrobial procedures

is complicated: ideal method does not exist

choice depends on numerous factors

• type and number of microbes

• environmental conditions

• risk of infection

• composition of infected item

selection of antimicrobial procedures - type of microorganism

most critical consideration, multiple highly resistant microbes

• bacterial endospores

• protozoan cysts and oocysts

• mycobacterium species

• pseudomonas species

• non-enveloped viruses

bacterial endospores

most resistant, only extreme heat or chemical treatment destroys them

protozoan cysts and oocysts

resistant to disinfectants, excreted in feces; if ingested causes diarrheal disease

mycobacterium species

resistant to many chemical treatments due to waxy cell walls

pseudomonas species

resistant to and can actually survive and multiply/grow in some disinfectants

• can grow into certain antiseptics

• iodine is generally effective against P but P can be unusually resistant to some chemical agents

non-enveloped viruses

protein coat with nucleic acid inside, lack lipid envelope; more resistant to disinfectants

• when the ones with the envelope are destroyed the virus is inactivated

• the ones that don’t have an envelope are harder to inactivate and more resistant

selection of antimicrobial procedures - number of microorganisms

can affect treatment time

• large population = more time needed

decimal reduction time (D value)

• time required to kill 90% of population under specific conditions

after 5 min (D) and theres 10,000 and 90% are killed → 1,000 and treat again for 5 min with 90% killed → 100 → 10 → 1

• not a good idea to stop here cuz it can just grow back so go to 0.1 with additional 5 minutes

selection of antimicrobial procedures - environmental conditions

pH, temp, and presence of organic materials

inc or dec effectiveness

• higher temps and lower pH increase effectiveness of treatment

• presence of organic molecules dec effectiveness

  • interfere with penetration of heat/antimicrobial agent

  • dirt, grease, bodily fluids

  • imp to thoroughly clean

• microorganisms in biofilm are more resistant - when they have a capsule or polysaccharide on the outside

  • slimy film on sink

  • like a protective layer and bacteria is under it so have to break it up before you treat the surface

selection of antimicrobial procedures - risk for infection

medical instrument categories according to risk for transmitting infectious agents

• critical items come in contact with body tissues

  • must be sterile

  • needles and scalpels

• semi-critical instruments contact mucous membranes but do not penetrate body tissues

  • must be free of pathogens

  • endoscopes are endotracheal tubes

• non-critical instruments contact unbroken skin only

  • low risk of transmission

  • countertops, stethoscopes, blood pressure cuffs

selection of antimicrobial procedures - composition of item

makes some sterilization and disinfection methods inappropriate

• heat sensitive items - heat treatment inappropriate

  • plastics, antibiotics (heat can damage/denature some proteins in some that have them)

• moisture-sensitive material

  • moist heat, liquid chemical disinfectants and steam and water cannot be used

using heat to destroy microorganisms and viruses

heat treatment - useful for microbial control

• reliable, safe, relatively fast, inexpensive, non-toxic

• can sterilize or disinfect

methods include

• moist heat

• dry heat

using heat to destroy microorganisms and viruses - moist heat/boiling

irreversibly denatures proteins, involves steam/water

• like frying an egg that destroys the proteins

• __ (100*C) - destroys most microorganisms and viruses

  • may not sterilize; endospores may survive

  • if you do it long enough it can destroy the endospores so can sterilize

using heat to destroy microorganisms and viruses - moist heat/pasteurization

uses quick heat to destroy pathogens and spoilage organisms/microbes in liquid food

• increase shelf life of food

• make sure ppl don’t get sick

• equivalent treatments for pasteurizing milk - NOTE FOR EXAM that as temp goes up time goes down

  • 63*C for 30 min: low temp long time LTLT

  • 72*C for 15 sec: high temp short time HTST

  • 140*C for 2-5 sec: ultra-high temp UHT

using heat to destroy microorganisms and viruses - moist heat/autoclaving

sterilizes using pressurized steam

• typical program: 121*C/15 psi/15 minutes

  • kills endospores and really all microbes except for prions

  • longer for larger volumes

• prions (infectious agent causes mad cow disease) thought to be destroyed at 132*C for 1 hr

  • this protein is very hard to destroy

  • so higher pressure and longer time has to be used

using heat to destroy microorganisms and viruses - moist heat effectiveness

moist heat has more contact - REASON for why its more effective

• water transfers heat more efficiently and causes faster protein denaturation

using heat to destroy microorganisms and viruses - dry heat

less effective than moist heat; require longer times, higher temps

• 200*C for 90 minutes vs 121*C for 15 minutes → d vs m

• hot air ovens: oxidize cell components, denature proteins killing all microbes

• incineration: oxidizes cell to ashes (sterilization)

  • burning of medical waste and animal carcasses

  • flaming laboratory inoculation loop

using other physical methods to remove or destroy microbes

heat sensitive materials - cant use heat cuz it’ll destroy them

methods of treatment:

• filtration

• irradiation

• high-pressure treatment

Filtration

removes microbes from fluids and air

• liquid filtration

  • membrane filters (0.2 nanometers pore size removes bacteria)

    • allows liquids to flow through

    • traps microbes on filter

• air filtration

  • uses high efficiency particulate air HEPA filters

    • filter has 0.3 nanometer pores to trap and remove nearly all microbes from air

Irradiation

Radiation

• shorter wavelength, higher energy, more antimicrobial

• can be ionizing or non-ionizing

Ionizing Radiation

able to strip electrons from atoms

• causes damage to dna and plasma membrane

• ex. gamma rays used to destroy microbes on heat-sensitive materials

  • medical equipment, surgical supplies, medications

  • food - FDA approved for spices/dried herbs, fruits, vegetables, grains, pork and other meats

• endospores can be resistant

Non-ionizing radiation (UV)

ultraviolet radiation (UV)

• damages DNA - formation of thymine dimers

• used to destroy microbes in the air, drinking water and surfaces

• limitation

  • poor penetrating power

    • thin films or covering can limit effect (glass or plastic)

microwaves - kills by heat

• microwave ovens heat food unevenly so cells can survive

using chemicals to destroy microorganisms and viruses

chemicals can disinfect or sterilize

• disinfectants

• antiseptics

react with vital cell sites

• proteins

• dna

• cell membrane

selecting the appropriate germicidal chemical

for destroying microorganisms and viruses

toxicity: benefits must be weighed against risk of use

compatibility with material being treated

• liquids cant be used on electrical equipment

cost and availability

environmental risk

• agent may need to be neutralized before disposal

potency required

potency of germicidal chemical formulations

sterilants destroy all microorganisms including spores

• heat sensitive critical instruments

high-level disinfectants destroy viruses, vegetative cells

• do not reliably kill endospores

• semi-critical instruments

intermediate level disinfectants destroy vegetative bacteria, mycobacteria, fungi, and most viruses

• disinfect non-critical instruments

low level disinfectants destroy fungi, vegetative bacteria (except mycobacteria), and enveloped viruses

• do not kill endospores, non-enveloped viruses

• disinfect furniture, floors, walls

classes of germicidal chemicals - alcohols

60-80% solutions of isopropyl or ethyl alcohol

• kill vegetative bacteria and fungi

• not effective against endospores, naked viruses

mode of action

• coagulation of proteins and enzymes

• damage to lipid membranes

used as antiseptic and disinfectant

limitations

• evaporates quickly, limiting contact time

• may damage material such as rubber and some plastics

classes of germicidal chemicals - aldehydes

kill bacteria/inactivate viruses by inactivating proteins and nucleic acids

• 2% glutaraldehyde solution - most widely used liquid sterilant

  • immersion for 10-12 hours to sterilize

  • toxic

• formaldehyde used as gas or as formalin (37% solution)

  • effective germicide

  • uses

    • to kill bacteria and inactivate viruses used as vaccines

    • to preserve specimens

classes of germicidal chemicals - biguanides

chlorhexidine

• most effective member, low toxicity

• found in antiseptics (skin creams, mouth rinse) and disinfectants

• destroys wide range of organisms

  • vegetative bacteria, fungi, some enveloped viruses

classes of germicidal chemicals - ethylene oxide

gaseous sterilant

mode of action: reacts with proteins

good penetration of material and useful in sterilizing heat or moisture-sensitive items

• fabrics, equipment, implantable devices (pacemakers, artificial hips)

• petri dishes, pipettes

limitations: toxic → mutagenic and potentially carcinogenic

classes of germicidal chemicals - halogens

common disinfectants

oxidizes proteins and other cell components

• chlorine

  • destroys all types of microbes

  • household bleach - 1:100 dilution

  • very low levels disinfect drinking water

    • Cryptosporidium oocysts, Giardia cysts survive

• iodine

  • kills vegetative cells, not endospores

  • found in tincture (Betadine) or iodophor used on skin iodine tablets for disinfecting drinking water

  • Pseudomonas species can survive

classes of germicidal chemicals - metal compounds

combine with enzymes and proteins (-SH group)

• interfering with function

high concentrations toxic to human tissue

low levels used as antimicrobial agent

• silver used as antiseptic: creams, bandages

  • silver nitrate eye drops given to newborn to prevent Neisseria gonorrhoeae infections

    • replaced by antibiotics

  • creams with silver sulfadiazine used to prevent secondary infections

classes of germicidal chemicals - ozone O3

unstable form of oxygen

• naturally occurring molecules made of 3 o atoms, quickly breaks down

• powerful oxidizing agent

  • strong oxidizer that destroys microorganisms by damaging cell walls, proteins, and genetic material

• used as alternative to chlorine

  • disinfectant for drinking water

    • eliminates bacteria, viruses, and protozoa

    • improves taste, odor, clarity

    • leaves no harmful chemical residue

  • wastewater disinfection

    • destroys pathogens and reduces odors

    • effective against a wide range of microorganisms

    • environmentally friendly; breaks down to oxygen

classes of germicidal chemicals - peroxygens

powerful oxidizers used as sterilants

• less toxic than ethylene oxide, glutaraldehyde

• hydrogen peroxide

  • more effective on inanimate object

  • vapor-phase can be used as sterilant

• peracetic acid: more potent than H2O2

  • effective on organic material

  • useful on wide range of material

classes of germicidal chemicals - phenolic compounds

kills most vegetative cells

• kills Mycobacterium at high concentrations

mode of action

• destroy plasma membrane

• denature proteins

phenol - one of the earliest disinfectants

• has unpleasant odor, irritates skin

triclosan and hexachlorophene : non-toxic on skin

• used in soaps and lotions

• triclosan now banned in house-hold products

classes of germicidal chemicals - quaternary ammonium compounds

cationic detergents

found in disinfectants

mode of action

• reduces surface tension

  • aids in removal of microbes, dirt and organic matter

• positive charge attracted to negative charge of cell and reacts with membrane

  • destroys vegetative bacteria and enveloped viruses

  • not effective on endospores, mycobacteria and naked viruses

preservation of perishable products - chemicals

use chemical preservatives

• extend shelf-life of products by preventing/slowing growth of microorganisms

chemical preservatives include

• weak organic acids (benzoic, sorbic, propionic, citric)

  • alter cell membrane function

  • control molds and bacteria in baked goods, cheeses, juices, and cosmetics

• nitrate and nitrite added to processed meats

  • inhibit germination of Clostridium botulinum endospores

    • also give meats pink color

  • shown to be carcinogenic forming nitrosamines

preservation of perishable products - low temp storage

refrigeration: inhibits growth of pathogens/spoilage organisms

• psychrotrophs, psychrophilic organisms may still grow

freezing: stops all microbial growth

• some cells killed by ice crystal formation, most still viable

preservation of perishable products - reducing available water

addition of salt, sugar

• increases environmental solutes → causes cellular plasmolysis

• some bacteria grow in high salt environments

  • Staphylococcus aureus

• drying often supplemented by salting

• lyophilization (freeze drying) foods

  • coffee, milk, meats, fruits, vegetables

• drying stops microbial growth but does not reliably kill