BISC 300 Lecture 17 & 18

fomites

inanimate objects that may carry microbial contamination

how are fomites treated

more aggressive control methods, for a longer time

to achieve lower levels of contamination

why cant we aggressively treat living tissue for contamination

living tissue is more fragile and either requires gentler methods

or we tolerate greater contamination levels

what methods of decontamination can you use for fomites (objects/surfaces)

disinfection

sanitization

sterilization

which decontamination method results in the lowest level of contamination

sterilizatoin

disinfection

reduces/destroys microbioal load of an inanimate item through heat or antimicrobial chemicals

sanitization

reduces microbioal load of inanimate object to safe public health levels through heat or antimicrobial chemicals

sterilization

completely eliminates all vegetative cells, endospores, and viruses from an inanimate object

what methods of decontamination can you use on living tissue

antisepsis

degerming

antisepsis

reduces microbioal load on skin or tissue by applying antimicrobial chemical

degerming

reduces microbial load on skin or tissue through gentle to firm scrubbing and the use of mild chemicals (soap)

aseptic technique

set of protocols that maintain sterility (asepsis), which prevents contamination of the patient with microbs

sepsis

systemic inflammatory response to a systemic infection

methods that kill microbes

end in -cide or -cidal

complete sterility

methods that inhibit growth

end in -stat or -static

allow immune system opportunity to clear infection

physical control methods

heat

radiation

refrigeration/freezing

pressure

chemical control methods

gas

liquid

mechanical removal control methods

filtration

biological control methods

virus

toxin

microbial death curve

semi-log plot

d-value on microbial death curve

decimal reduction time

time to kill 90% = 1 Log unit

types of heat control methods

boiling

dry-heat oven

incineration

what does boiling do to microbes

denatures proteins and alters membranes

what does dry-heat oven do to microbes

denatures proteins and alters membranes, dehydration, desiccation

what does incineration do to microbes

destroy by burning

how does clostridium botulinum respond to high temp

endospores

does boiling kill all microbes

no

some endospores survive >20 hours of boiling

less effective at higher altitudes

does incineration kill all microbes

yes at very high temps

autoclaving

denatures proteins and alters membranes

rely on moist-heat sterilization and raise temp above boiling point of water

most effective method

internal indicators to ensure autoclave sterilization

heat-sensitive autoclave tape

biological indicator spore test

biological indicator spore test

endospores of the thermophile geobacillis sterothermophilus to determine whether endospores were killed

pasteurization

denatures proteins and alters membranes

uses heat but does not render the food sterile

reduces # of spoilage-causing microbes

three levels of pasteurization

LTH

HTST
UHT

LTH

low temperature holding

63 degrees for 30 mins

HTST

high temp short temp

72 degrees for 15 seconds

lowers bacterial number and preserving quality

UHT

ultra high temp

138 degrees for 2 seconds

can be stores for a long time in sealed containers without refrigeration

refrigeration

inhibits metabolism

slows or arrests cell division

0 to 7 degrees

freezing

stops metabolism, may kill microbes

below 2 degrees

ultra-low temp

dry ice -70 degrees

liquid nitrogen tanks -196 degrees

for long-term storage or transport

high-pressure processing

100-800 MPa

denatures proteins and can cause cell lysis

hyperbaric oxygen therapy

air pressure 3x higher than normal

inhibits metabolism and growth of anaerobic microbes

how is pressure used in food

kills microbes while maintaining food quality and extending shelf life

how is pressure used clinically

hyperbaric oxygen therapy is used to treat infectoin

patient breathes pure o2

simple desiccation

drying

inhibits metabolism

desiccation reduce water activity

addition of salt or water

inhibits metabolism and can cause lysis

desiccation lyophilization

rapid freezing under vacuum- water is lost by sublimation

inhibits metabolism

water activity

water content

can be lowered by adding solutes

ionizing radiation

exposure to x rays or gamma rays or high energy electron beams

alters molecular structure, introduces double-stranded breaks into DNA

how does ionizing radiation interact with paper and plastic

penetrate paper and plastic to sterilize packaged materials

nonionizing radiation

ultraviolet (UV) light

introduces thymine dimers → mutation

less energetic/penetrating, used for surface disinfection

filtration

physical separation of microbes from air or liquid

high efficiency particulate air (HEPA)

filter have pores about 0.3 um

filter out bacteria, endospores, and many viruses

membrane filters

porous membranes with defines pore sizes

removed by physical screening

cellular microbes >0.2um

viruses > 0.1um

phenolic chemical disinfectants

denature proteins and disrupt membranes

metal chemical disinfectants

bind to proteins and inhibit enzyme activity

halogens chemical disinfectants

oxidation and destabilization of celluular macromolecules

alcohol chemical disinfectant

denature proteins and disrupt membranes

surfactant chemical disinfectant

lowers surface tension of water to help with washing away microbes and disrupt cell membranes

bisbiguanides chemical disinfectants

disruption of cell membranes

alkylating agents chemical disinfectants

inactivation of enzymes and nucleic acid

peroxygen chemical disinfectants

oxidation and destabilization of cellular macromolecules

supercritical gases as chemical disinfectants

penetrates cells, formed carbonic acid, lowers intracellular pH

chemical food preservatives as chemical disinfectants

decreases pH and inhibit enzymatic function

natural food preservatives as chemical disinfectants

inhibition of cell wall synthesis

disk-diffusion method for chemical agents

filter disks contain chemical placed on an agar plate inoculated with bacterium

compound causes zones of inhibition- size correlates with potency of compound

Kock’s postulates

•causative microbe must be present in diseased organisms, but not healthy organisms

•must be able to isolate the causative organism in pure culture

•must be able to infect a healthy organism with the isolated culture

•must be able to re-isolate microbe from experimentally infected organism

exceptions to koch’s postulates

•some pathogens are part of the normal microbiome

•some diseases involve multiple agents, or can be caused by different microbes

•some infections have varied symptoms (pleiotropic)

•some infectious agents cannot be cultured

primary pathogens

almost always cause disease

opportunistic pathogens

•normally exist outside of the host (e.g., commensals)

•cause infection under the right circumstances

•age, weakened immune system, injury

•Staphylococcus epidermidis

obligate pathogens

•cannot exist outside host in the natural environment

•(perhaps can be cultured in the lab)

•Chlamydia, Rickettsia, Mycobacteria

reservoir

natural population outside of the host

drinking water, soil, animals (zoonoses)

vectors

organisms that spread disease

mosquitos carrying malaria

extracellular pathogens

pathogens exist on or in host fluids and tissues, but do not enter host cells

may move through circulatory system or migrate through the matrix between host cells

can directly encounter elements of the immune system

examples of extracellular pathogens

E. coli

Staphylococcus aureus

Helicobacter pylori

Borrela burgdorferi

intracellular pathogens

microbes enter and multiply within host cells

allows them to evade many elements of host immune system

examples of intracellular pathogens

Listeria monocytogenes

Mycobacterium tuberculosis

Salmonella enterica

Legionella pneumophila

incubation period

pathogen entry, before symptoms

prodromal stage

first onset of symptoms

period of illness

disease is most severe, symptoms apparent

period of decline

body fights off infection

convalescence

symptoms resolve

infectious dose 50 (ID50)

number of pathogens that will infect 50% of hosts in a specified time

how does ID50 vary

•varies with pathogen, strain, etc.

•this is why antisepsis and disinfection are sufficient to reduce incidence of many infections

The dose of pathogens needed to bring on disease varies greatly – can be as few as 1

virulence

intensity of pathogenicity

pathogen lifestyle, opportunistic vs obligate

which type of pathogen is more likely to kill the host

opportunisitc

which type of pathogen is less likely to severely (or rapidly) harm the host

obligate

virulence factors

facilitate infection, tissue invasion, or farm

encoded by genes on chromosome or plasmid (HGT)

pathogenicity islands

where genes encoding virulence factors cluster on the chromosome

correlate with pathogenicity

absent in non-pathogenic strains

common phenotype of pathogenicity islands

•toxin secretion

•pilus, or other features for attachment to host

•iron uptake

•biofilm formation

adherence

once a pathogen has gained entry into host, it must adhere somewhere

recognizes specific host molecules

mediated by adhesins

adhesins in e coli

•diarrhea:  fimbriae bind sugars on intestinal epithelium

•hemolytic uremia: pili bind sugars on kidney cells

•urinary tract infection: pili bind sugars on urethral epithelium

invasiveness

ability to spread to adjacent tissues

exoenzymes

may aid in spreading by

break down extracellular matrix

degrade carbohydrate-protein matrix between cells

can also disrupt host cell surface

collagenase

breaks down collagen

hyaluronidase

breaks down hyaluronic acid

bacteremia

presence of bacteria in the blood

septicemia

pathogens of their toxins in the blood

endoxins

factors secreted by bacteria to cause damage to the host

•may induce tissue damage, aiding invasion

•may cause host cell lysis, releasing nutrients

channel-forming toxins

•self-assemble into pores in host-cell plasma membrane

•cause host cell lysis

AB toxins

•A and B components form a complex

•B component attaches to host cell, triggers endocytosis of AB complex

•A component is released, causes toxicity

•(many ways to cause cell death)