Monday, March 1, 2010
Sunday, February 28, 2010
Friday, February 19, 2010
Tuesday, February 16, 2010
ENERGY PATHWAYS FOR EXERCISE
We've learned that carbohydrates and fats being the source of energy for physical activities. In the same token we know that the main source of energy is ATP either from aerobic or anaerobic metabolism.
What exactly goes within the cells during exercise is explained in this article. Go for it now.
energy pathways during exercise
What exactly goes within the cells during exercise is explained in this article. Go for it now.
energy pathways during exercise
Tuesday, February 9, 2010
Monday, January 25, 2010
Wednesday, January 20, 2010
slides on gm negative bacteria
hey guys! you missed class the last time. check out the slides on gm negative bacteria on January 2008 posting and be ready for quiz...concentrate on N. meningitidis, spirochaete and leptospirosis...nice reading
Sunday, January 10, 2010
HIV lec slide
• A - Acquired - because it's a condition one must
acquire or get infected with
• I - Immune - because it affects the body's
immune system
• D - Deficiency - immune system because it
makes the immune system deficient
• S - Syndrome - because someone with AIDS
may experience a wide range of different
diseases and opportunistic infections
What is HIV?
• H - Human - because this virus can only infect
human beings.
• I - Immuno-deficiency - a failure to work
properly within the body's immune system
• V - Virus - because this organism is a virus,
incapable of reproducing by itself.
It reproduces by taking over the
machinery of the human
• HIV Disease" cover the entire HIV spectrum
• from initial infection to full-blown AIDS
• stages are representative of the experience of many people with HIV
• The time is variable
• For most people---the process of HIV disease is fairly slow
taking several years from infection to the development of severe immunodeficiency
• InfectionHIV enters the bloodstream
• begins to take up residence in the cells
• A person with HIV is infectious at all times.
• Maybe asymptomatic
• only way to find out if a person is infected is by taking an HIV antibody test.
Primary Infection (or Acute Infection)
• the first stage of HIV disease
• when the virus first establishes itself in the body.
• period of time between ---first infected with HIV and when antibodies are produced
• usually 6- 12 weeks
• 70% of people experience some "flu-like" symptoms.
fevers, chills, night sweats and rashes
• The remaining percentage of people maybe asymptomatic
acute HIV infection
• the virus makes its way to the lymph nodes
• HIV actively reproduces and releases new virus particles into the bloodstream.(2 mos.)
people with acute HIV infection usually will have negative HIV antibodies test
• takes the body approximately one to three months to produce antibodies against HIV.
Seroconversion
• time when the body produce antibodies to the virus.
• 95% within three months after infection.
• If first result is negative a second test done three months later.
Immune System Decline
• The virus appears to slowly damage the immune system for a number of years after infection
• a faster decline of the immune system occurs at some point
• the virus rapidly replicates
• This damage can be seen in blood tests such as lowered T-cell counts, before any actual symptoms are experienced.
Mild, Non-Specific Symptoms
• Once the immune system is damaged
• skin rashes
• fatigue
• slight weight loss
• night sweats
• thrush in the mouth, etc.
• it takes the average person five to seven years to experience their first mild symptom.
More Severe Symptoms; Opportunistic Infections and Diseases
• immune system damage is more severe
• may experience opportunistic infections
• Centers for Disease Control’s definition of full-blown "AIDS."
• Does everyone who has HIV
eventually get sick? =majority of untreated people do eventually
become ill from HIV
Some long-term survivors may do so well because
of their unique body chemistry,
or access to a combination of medical, emotional
and spiritual support,or something yet unknown to us
Treatment
• Antiviral Therapy
• The goal of antiviral therapy
• disable HIV replication
• reverse transcriptase-- a protein the virus must use in order to reproduce.
Protease inhibitors----interfere with HIV's use of the protease enzyme, another essential component to its reproduction
• Immune Boosting Therapy
• attempt to bolster the body's ability to fight HIV
• "therapeutic" vaccines+anti-viral therapy
• may improve the body's response to HIV.
• Other treatments boost CD4 count (T-cells), though this approach is not believed to be useful unless it is combined with an antiviral treatment.
Tests That Monitor The Immune System
• HIV viral load.
• testing measures the amount of HIV in blood plasma.
• CD4 count
• measures the number of CD4 cells in a blood sample.
• The CD4 count is one indicator of how much damage HIV has caused to the immune system.
• CD8 count
• CD8s are a different subset of T-cells that include suppressor T-cells and "killer" T-cells.
At this stage, several things can happen
• The new virus ("provirus") can remain inactive for a long time without triggering the reproduction of virus,
• divide into two proviruses- mitosis
• or it can start producing new virus
• budding off from the T-cell wall-- eventually destroying the T-cell.
• In the process of viral reproduction
• the virus destroys increasing numbers of T-cells
• leaving the body open to
opportunistic infections
How HIV Is Spread
• Requirements For Transmission to Occur
• 1. HIV must be present
• 2. In sufficient quantity
• 3. And it must get into the bloodstream.
Where is HIV Found in the Body?
• HIV can be transmitted from an infected person to another through:
Blood (including menstrual blood)
Semen
Vaginal secretions
Breast milk
• Possibly infectious "bodily fluids"
Pre-seminal fluid (pre-cum)
• Non-infectious "bodily fluids"
Saliva
Tears
Sweat
Feces
Urine
Activities That Allow HIV Transmission
• there are three primary ways in which this can happen:
• 1. Unprotected sexual contact
• 2. Direct blood contact
including injection drug needles, blood transfusions, accidents in health care settings or certain blood products
• 3. Mother to baby
before or during birth, or through breast milk
Sexual Routes Of Transmission
• Sexual intercourse (vaginal and anal)
• Oral sex (mouth-penis, mouth-vagina)
• Heterosexual transmission studies
Non-Sexual Routes Of Transmission
• Sharing injection needles
• Needle sticks
• Blood transfusions
• Hemophilia treatments
pooled blood of many donors
• Other blood products
• Mother to Child
HIV Is NOT Transmitted By
• Insect bites
• Casual contact
• sharing of dishes/foods
• Donating blood
• Swimming pools/bath tubs
• Pets
• Contact with saliva, tears,sweat,urine,feces
• Rape/sexual assault?
• Transmission Through Tattooing, Piercing, Acupuncture, Electrolysis, and Shaving
• What is the risk?
• Universal precautions
How the Immune System Fights Disease
Aids interfere with normal body response
acquire or get infected with
• I - Immune - because it affects the body's
immune system
• D - Deficiency - immune system because it
makes the immune system deficient
• S - Syndrome - because someone with AIDS
may experience a wide range of different
diseases and opportunistic infections
What is HIV?
• H - Human - because this virus can only infect
human beings.
• I - Immuno-deficiency - a failure to work
properly within the body's immune system
• V - Virus - because this organism is a virus,
incapable of reproducing by itself.
It reproduces by taking over the
machinery of the human
• HIV Disease" cover the entire HIV spectrum
• from initial infection to full-blown AIDS
• stages are representative of the experience of many people with HIV
• The time is variable
• For most people---the process of HIV disease is fairly slow
taking several years from infection to the development of severe immunodeficiency
• InfectionHIV enters the bloodstream
• begins to take up residence in the cells
• A person with HIV is infectious at all times.
• Maybe asymptomatic
• only way to find out if a person is infected is by taking an HIV antibody test.
Primary Infection (or Acute Infection)
• the first stage of HIV disease
• when the virus first establishes itself in the body.
• period of time between ---first infected with HIV and when antibodies are produced
• usually 6- 12 weeks
• 70% of people experience some "flu-like" symptoms.
fevers, chills, night sweats and rashes
• The remaining percentage of people maybe asymptomatic
acute HIV infection
• the virus makes its way to the lymph nodes
• HIV actively reproduces and releases new virus particles into the bloodstream.(2 mos.)
people with acute HIV infection usually will have negative HIV antibodies test
• takes the body approximately one to three months to produce antibodies against HIV.
Seroconversion
• time when the body produce antibodies to the virus.
• 95% within three months after infection.
• If first result is negative a second test done three months later.
Immune System Decline
• The virus appears to slowly damage the immune system for a number of years after infection
• a faster decline of the immune system occurs at some point
• the virus rapidly replicates
• This damage can be seen in blood tests such as lowered T-cell counts, before any actual symptoms are experienced.
Mild, Non-Specific Symptoms
• Once the immune system is damaged
• skin rashes
• fatigue
• slight weight loss
• night sweats
• thrush in the mouth, etc.
• it takes the average person five to seven years to experience their first mild symptom.
More Severe Symptoms; Opportunistic Infections and Diseases
• immune system damage is more severe
• may experience opportunistic infections
• Centers for Disease Control’s definition of full-blown "AIDS."
• Does everyone who has HIV
eventually get sick? =majority of untreated people do eventually
become ill from HIV
Some long-term survivors may do so well because
of their unique body chemistry,
or access to a combination of medical, emotional
and spiritual support,or something yet unknown to us
Treatment
• Antiviral Therapy
• The goal of antiviral therapy
• disable HIV replication
• reverse transcriptase-- a protein the virus must use in order to reproduce.
Protease inhibitors----interfere with HIV's use of the protease enzyme, another essential component to its reproduction
• Immune Boosting Therapy
• attempt to bolster the body's ability to fight HIV
• "therapeutic" vaccines+anti-viral therapy
• may improve the body's response to HIV.
• Other treatments boost CD4 count (T-cells), though this approach is not believed to be useful unless it is combined with an antiviral treatment.
Tests That Monitor The Immune System
• HIV viral load.
• testing measures the amount of HIV in blood plasma.
• CD4 count
• measures the number of CD4 cells in a blood sample.
• The CD4 count is one indicator of how much damage HIV has caused to the immune system.
• CD8 count
• CD8s are a different subset of T-cells that include suppressor T-cells and "killer" T-cells.
At this stage, several things can happen
• The new virus ("provirus") can remain inactive for a long time without triggering the reproduction of virus,
• divide into two proviruses- mitosis
• or it can start producing new virus
• budding off from the T-cell wall-- eventually destroying the T-cell.
• In the process of viral reproduction
• the virus destroys increasing numbers of T-cells
• leaving the body open to
opportunistic infections
How HIV Is Spread
• Requirements For Transmission to Occur
• 1. HIV must be present
• 2. In sufficient quantity
• 3. And it must get into the bloodstream.
Where is HIV Found in the Body?
• HIV can be transmitted from an infected person to another through:
Blood (including menstrual blood)
Semen
Vaginal secretions
Breast milk
• Possibly infectious "bodily fluids"
Pre-seminal fluid (pre-cum)
• Non-infectious "bodily fluids"
Saliva
Tears
Sweat
Feces
Urine
Activities That Allow HIV Transmission
• there are three primary ways in which this can happen:
• 1. Unprotected sexual contact
• 2. Direct blood contact
including injection drug needles, blood transfusions, accidents in health care settings or certain blood products
• 3. Mother to baby
before or during birth, or through breast milk
Sexual Routes Of Transmission
• Sexual intercourse (vaginal and anal)
• Oral sex (mouth-penis, mouth-vagina)
• Heterosexual transmission studies
Non-Sexual Routes Of Transmission
• Sharing injection needles
• Needle sticks
• Blood transfusions
• Hemophilia treatments
pooled blood of many donors
• Other blood products
• Mother to Child
HIV Is NOT Transmitted By
• Insect bites
• Casual contact
• sharing of dishes/foods
• Donating blood
• Swimming pools/bath tubs
• Pets
• Contact with saliva, tears,sweat,urine,feces
• Rape/sexual assault?
• Transmission Through Tattooing, Piercing, Acupuncture, Electrolysis, and Shaving
• What is the risk?
• Universal precautions
How the Immune System Fights Disease
Aids interfere with normal body response
Friday, December 25, 2009
Friday, December 18, 2009
Saturday, December 12, 2009
bacterial cell lec slide
Prokaryotes (Bacteria)
• Eubacter "True" bacteria
– human pathogens
– clinical or environmental
– one kingdom
• Archaea
– Environmental organisms
– second kingdom
Eukaryotes
• Other cell-based life e.g.
– plants
– animals
– fungi
Prokaryotic Cell (versus Eukaryotic Cell
• Not compartmentalized
• Cell membranes lack sterols (e.g. cholesterol)
• Single circular chromosome
• Ribosomal are 70S
Bacteria
• Plasmids
• Extra-chromosomal DNA
• multiple copy number
• coding pathogenesis and antibiotic
resistance factors
• bacterial replication
The Cell Envelope
Oxidative phosphorylation occurs at cell membrane
(since there are no mitochondria
The ceined: caAyer or glycocalyx
• usually polysaccharide
• often lost on in vitro culture
• protective in vivo
• Many bacterial cells secrete some extracellular material in the form of a capsule or a slime layer
• A slime layer is loosely associated with the bacterium and can be easily washed off
• whereas a capsule is attached tightly to the bacterium and has definite boundaries
Endospores (spores
• Dormant cell
• Produced when starved
• Resistant to adverse conditions
- high temperatures
- organic solvents
• Bacillus and Clostridium
Culture and identification of infectious agents
Taxonomy
• Defines common traits among strains for a bacterial species
• Usually genetic
• Allows development of diagnostic kits
TAXONOMY
Classification, Nomenclature, Laboratory Identification
Identification of infectious agents
in the diagnostic laboratory
• Aids treatment
• Helps antibiotic selection
• General hospital laboratory
– physiological tests
• Reference laboratories
– Genetic tests
Steps in isolation and identification
• Step 1. Streaking culture plates
�Ayer or glycocalyx
• usually polysaccharide
• often lost on in vitro culture
• protective in vivo
• Many bacterial cells secrete some extracellular material in the form of a capsule or a slime layer
• A slime layer is loosely associated with the bacterium and can be easily washed off
• whereas a capsule is attached tightly to the bacterium and has definite boundaries
Endospores (spores
• Dormant cell
• Produced when starved
• Resistant to adverse conditions
- high temperatures
- organic solvents
• Bacillus and Clostridium
Culture and identification of infectious agents
Taxonomy
• Defines common traits among strains for a bacterial species
• Usually genetic
• Allows development of diagnostic kits
TAXONOMY
Classification, Nomenclature, Laboratory Identification
Identification of infectious agents
in the diagnostic laboratory
• Aids treatment
• Helps antibiotic selection
• General hospital laboratory
– physiological tests
• Reference laboratories
– Genetic tests
Steps in isolation and identification
• Step 1. Streaking culture plates
– colonies on incubation (e.g 24 hr)
– size, texture, color, hemolysis
– oxygen requirement
Samples of body fliuds (blood,urine,csf)
• Step 2. Colonies Gram stained
– cells observed microscopically
Gram stain morphology
• Shape
– cocci (round)
– bacilli (rods)
– spiral or curved (e.g. spirochetes)
• Single or multiple cells
– clusters (e.g. staphylococci)
– chains (e.g. streptococci)
• Gram positive or negative
Step 3. Isolated
bacteria are speciated
• Generally using physiological tests
Step 4.
Antibiotic susceptibility testing
Molecular differentiation
• Genomics
• Gene characterization
– Sequencing
– PCR
• Hybridization
• % guanine + cytosine
16S rRNA Sequencing
• Differentiates bacterial species
• Development of clinical tests based on sequence (e.g. PCR)
Rapid diagnosis without culture
• WHEN AND WHY?
• grow poorly
• can not be cultured
Microscopy
• spinal fluids (meningitis)
• sputum (tuberculosis)
• sensitivity poor
Serologic identification
• antibody response to the infecting agent
• several weeks after an infection has occurred
Nutrition, Growth and Metabolism
Bacterial requirements for growth
• oxygen (or absence)
• energy
• nutrients
• optimal temperature
• optimal pH
Respiration
• Respiration is the oxidation of a source of energy by removal of electrons and donation to an inorganic terminal electron accepter.
Fermentation
• Fermentation is defined as an energy yielding process whereby organic molecules serve as both electron donors and electron accepters
• The molecule being metabolized does not have all its potential energy extracted from it. In other words, it is not completely oxidized
Oxygen Requirements
Obligate aerobes must grow in the presence of oxygen they cannot carry out fermentation
Obligate anaerobes do not carry out oxidative phosphorylation they are killed by oxygen
Aerotolerant anaerobes are bacteria that respire anaerobically but can survive in the presence of oxygen
Facultative anaerobes can perform both fermentation and aerobic respiration
Microaerophilic bacteria grow well in low concentrations of oxygen,but are killed by higher concentrations.
Obligate aerobes
• grow in presence of oxygen
• no fermentation
• oxidative phosphorylation
Obligate anaerobes
• no oxidative phosphorylation
• fermentation
• killed by oxygen
• lack certain enzymes
superoxide dismutase
O2-+2H+ H2O2
catalase
H2O2 H20 + O2
peroxidase
H2O2 H20 /NAD NADH
Facultative anaerobes
• fermentation
• aerobic respiration
• survive in oxygen
Microaerophilic bacteria
• grow
– low oxygen
• killed
– high oxygen
Optimal growth temperature
• Mesophiles:
– human body temperature
* pathogens
* opportunists
• pyschrophile
– close to freezing
• thermophile
– close to boiling
pH
• Many grow best at neutral pH
• Some can survive/grow
- acid
- alkali
Nutrient Requirements
• Carbon
• Nitrogen
• Phosphorus
• Sulfur
• Metal ions (e.g. iron)
Siderophores (S)
Small molecule secreted by bacteria that bind iron
Measuring bacterial mass in liquid cultures of bacteria
Common methods include:
a) turbidity (the cloudiness of a liquid culture of bacteria –
a measure of total bacteria [live and dead]
b) the number of viable bacteria in a culture –
usually assessed by counting the number of colonies
that grow after streaking a known volume on a plate
Measuring bacterial mass (live + dead) in liquid culture
Measured with spectrophotometer
Measuring viable bacteria
Colony forming units
SUGAR CATABOLISM
• Glycolysis
– Embden Meyerhof Parnas Pathway
– most bacteria
– also animals and plants
STERILIZATION
• All killed
• non-selective
• autoclaving
• 121oC (heat/pressure)
* Heat resistant materials
• ethylene oxide
• Non heat resistant
– usually equipment
• ultra-violet light
– surfaces (e.g operating rooms)
not totally effective
• gamma radiation
– food
– some mail
Disinfection
• Liquids that kill bacteria
– e.g. phenol based
– too toxic for skin surfaces
Antiseptics
• Topical (e.g. skin)
– e.g. iodine or 70% alcohol
– “reduce” bacterial load
ANTIBIOTICS
• Selectively toxic for bacteria
– bactericidal (killing)
– bacteriostatic (growth inhibition)
• no harm to patient
• destroy structures
• present in bacteria
• not present in host
Antibiotics work together
with immune system
Minimal inhibitory concentration
• lowest level stopping growth
• e. g. zone of inhibition around a disk impregnated with antibiotic
• Antibiotics that inhibit cell wall
biosynthesis are bactericidal
• Without cell wall, osmotic pressure
causes bacteria to burst
Antibiotics: Protein Synthesis, Nucleic Acid Synthesis and Metabolism
Principles and Definitions
• Selectivity
– Selectivty8 toxicity9
• Therapeutic index
– Toxic dose/ Effective dose
• Categories of antibiotics
– Bactericidal
• Usually antibiotic of choice
– Bacteriostatic
• Duration of treatment sufficient for host defenses
• Categories of antibiotics
– Use of bacteriostatic vs bactericidal antibiotic
• Therapeutic index better for bacteriostatic antibiotic
• Resistance to bactericidal antibiotic
• Protein toxin mediates disease – use bacteriostatic protein synthesis inhibitor
• Antibiotic susceptibility testing (in vitro)
– Minimum inhibitory concentration (MIC)
• Lowest concentration that results in inhibition of visible growth
– Minimum bactericidal concentration (MBC)
• Lowest concentration that kills 99.9% of the original inoculum
• Combination therapy
– Prevent emergence of resistant strains
– Temporary treatment until diagnosis is made
– Antibiotic synergism
• Penicillins and aminoglycosides
• CAUTION: Antibiotic antagonism
• Penicillins and bacteriostatic antibiotics
Antibiotics that Inhibit Protein Synthesis
Protein Synthesis Inhibitors
• Mostly bacteriostatic
• Selectivity due to differences in prokaryotic and eukaryotic ribosomes
• Some toxicity - eukaryotic 70S ribosomes
Inhibitors of Nucleic Acid Synthesis
• Rifampin, Rifamycin, Rifampicin, Rifabutin (bactericidal
• Quinolones (bactericidal)
nalidixic acid, ciprofloxacin, ofloxacin, norfloxacin, levofloxacin, lomefloxacin, sparfloxacin
Antimetabolite Antimicrobials
• Inhibitors of Folic Acid Synthesis
• Sulfonamides, Sulfones (bacteriostatic)
• Trimethoprim, Methotrexate, Pyrimethamine (bacteriostatic)
Antimicrobial Drug Resistance
Principles and Definitions
• Clinical resistance
• Resistance can arise by mutation or by gene transfer (e.g. acquisition of a plasmid)
• Resistance provides a selective advantage
• Resistance can result from single or multiple steps
• Cross resistance vs multiple resistance
– Cross resistance -- Single mechanism-- closely related antibiotics
– Multiple resistance -- Multiple mechanisms -- unrelated antibiotics
Antimicrobial Drug Resistance Mechanisms
• Altered permeability
– Altered influx
• Gram negative bacteria
– Altered efflux
• tetracycline
• Inactivation
– $-lactamse
– Chloramphenicol acetyl transferase
• Altered target site
– Penicillin binding proteins (penicillins)
– RNA polymerase (rifampin)
– 30S ribosome (streptomycin)
• Replacement of a sensitive pathway
– Acquisition of a resistant enzyme (sulfonamides, trimethoprim)
Pathogenicity
• virulence factors
• number of initial organisms
• immune status
Koch's postulates
• isolated
– diseased not healthy people
• growth
– pure culture
• induce disease
– susceptible animals
• re-isolated
– susceptible animals
Opportunistic infections
• compromised people
– normal flora
– environment
Opportunists - normal flora
• Skin
– Staphylococcus aureus,
– S. epidermidis
– Propionibacterium acnes
• Intestine
– Bacteroides
* high numbers
– Enterobacteriaceae
* low number
Opportunists - environment
» air
• water
• soil
• food
• Opportunists in hospital- nosocomial
Transmission
• airborne droplets
• food
• water
• sexual contact
Host defenses
• Gut
– peristalsis
– defecation
• respiratory tract
– ciliary action
– coughing
– sneezing
• urogenital tract
– urination
COMMENSALS CAUSE DISEASE
WHEN HOSTS DEFENCES ARE LOW
PATHOGENS POSSESS VIRULENCE FACTORS
THAT ENABLE THEM TO CAUSE DISEASE IN
THE PRESENCE OF NORMAL DEFENCES
DETERMINANTS OF BACTERIAL PATHOGENISIS
TRANSMISSION
INFECTING DOSE-INOCULUM
PORTAL OF ENTRY
BACTERIAL VIRULENCE FACTORS
CAPSULES
INVASIVENESS
ADHESINS
EXOENZYMES
TOXINS
CAPSULES
LOCATED EXTERNAL TO THE CELLWALL
COMMON VIRULENCE FACTOR
POLYSACCHARIDES
AVOID OR SURVIVE PHAGOCYTOSIS
ADHESINS
ALLOW BACTERIA TO STICK ON MUCOSAL SURFACE
ADHESINS ARE SURFACE FACTORS
PILI / FIMBRIAE
LPS LIPOPOLYSACCARIDE SIDECHAINS
M PROTEIN
INVASIVENESS
ABILITY TO INVADE HOST CELL
MULTIFACTORIAL COMPLEX PROCESS
MAY INCLUDE ADHESION AND ENZYMES FACTORS
EXOENZYMES
PRODUCE AND SECRETED BY BACTERIA
ENZYMES THAT BREAKDOWN COLLAGEN AND FIBRIN
COLLAGENASES / HYALURONIDASES / FIBRINOLYSINS
ENZYMES THAT BREAKDOWN CELLULAR MATERIALS
PROTEASES / LECITHINASES
ENZYMES THAT DEACTIVATE AND MODIFY ANTIBIOTICS
BETA LACTAMASE
ENZYMES THAT DESTROY NEUTROPHILS
AND MACROPHAGES
LEUKOCIDINE
ENZYMES THAT ACCELERATE FIBRIN CLOT
COAGULASE
TOXINS
2 CLASSIFICATIONS
EXOTOXINS – PROTEINS PRODUCED AND
SECRETED BY BACTERIA
ENDOTOXINS – PART OF THE BACTERIA ITSELF
EXOTOXIN
NEUROTOXINS- AFFECT NERVOUS TISSUE
TETANUS TOXIN
ENTEROTOXINS – AFFECT GASTROINTESTINAL TRACT
CHOLERA TOXIN
CYTOTOXINS – AFFECT CELLS OF VARIOUS TISSUES
DIPHTERIA TOXIN
ENDOTOXINS
ALL GM NEGATIVE BATERIA HAVE ENDOTOXINS
IN OUTER MEMBRANE
RELEASED AS BACTERIA ARE LYSED
RESPONSIBLE FOR SEPSIS AND
SEPTIC SHOCK - FATAL
• Eubacter "True" bacteria
– human pathogens
– clinical or environmental
– one kingdom
• Archaea
– Environmental organisms
– second kingdom
Eukaryotes
• Other cell-based life e.g.
– plants
– animals
– fungi
Prokaryotic Cell (versus Eukaryotic Cell
• Not compartmentalized
• Cell membranes lack sterols (e.g. cholesterol)
• Single circular chromosome
• Ribosomal are 70S
Bacteria
• Plasmids
• Extra-chromosomal DNA
• multiple copy number
• coding pathogenesis and antibiotic
resistance factors
• bacterial replication
The Cell Envelope
Oxidative phosphorylation occurs at cell membrane
(since there are no mitochondria
The ceined: caAyer or glycocalyx
• usually polysaccharide
• often lost on in vitro culture
• protective in vivo
• Many bacterial cells secrete some extracellular material in the form of a capsule or a slime layer
• A slime layer is loosely associated with the bacterium and can be easily washed off
• whereas a capsule is attached tightly to the bacterium and has definite boundaries
Endospores (spores
• Dormant cell
• Produced when starved
• Resistant to adverse conditions
- high temperatures
- organic solvents
• Bacillus and Clostridium
Culture and identification of infectious agents
Taxonomy
• Defines common traits among strains for a bacterial species
• Usually genetic
• Allows development of diagnostic kits
TAXONOMY
Classification, Nomenclature, Laboratory Identification
Identification of infectious agents
in the diagnostic laboratory
• Aids treatment
• Helps antibiotic selection
• General hospital laboratory
– physiological tests
• Reference laboratories
– Genetic tests
Steps in isolation and identification
• Step 1. Streaking culture plates
�Ayer or glycocalyx
• usually polysaccharide
• often lost on in vitro culture
• protective in vivo
• Many bacterial cells secrete some extracellular material in the form of a capsule or a slime layer
• A slime layer is loosely associated with the bacterium and can be easily washed off
• whereas a capsule is attached tightly to the bacterium and has definite boundaries
Endospores (spores
• Dormant cell
• Produced when starved
• Resistant to adverse conditions
- high temperatures
- organic solvents
• Bacillus and Clostridium
Culture and identification of infectious agents
Taxonomy
• Defines common traits among strains for a bacterial species
• Usually genetic
• Allows development of diagnostic kits
TAXONOMY
Classification, Nomenclature, Laboratory Identification
Identification of infectious agents
in the diagnostic laboratory
• Aids treatment
• Helps antibiotic selection
• General hospital laboratory
– physiological tests
• Reference laboratories
– Genetic tests
Steps in isolation and identification
• Step 1. Streaking culture plates
– colonies on incubation (e.g 24 hr)
– size, texture, color, hemolysis
– oxygen requirement
Samples of body fliuds (blood,urine,csf)
• Step 2. Colonies Gram stained
– cells observed microscopically
Gram stain morphology
• Shape
– cocci (round)
– bacilli (rods)
– spiral or curved (e.g. spirochetes)
• Single or multiple cells
– clusters (e.g. staphylococci)
– chains (e.g. streptococci)
• Gram positive or negative
Step 3. Isolated
bacteria are speciated
• Generally using physiological tests
Step 4.
Antibiotic susceptibility testing
Molecular differentiation
• Genomics
• Gene characterization
– Sequencing
– PCR
• Hybridization
• % guanine + cytosine
16S rRNA Sequencing
• Differentiates bacterial species
• Development of clinical tests based on sequence (e.g. PCR)
Rapid diagnosis without culture
• WHEN AND WHY?
• grow poorly
• can not be cultured
Microscopy
• spinal fluids (meningitis)
• sputum (tuberculosis)
• sensitivity poor
Serologic identification
• antibody response to the infecting agent
• several weeks after an infection has occurred
Nutrition, Growth and Metabolism
Bacterial requirements for growth
• oxygen (or absence)
• energy
• nutrients
• optimal temperature
• optimal pH
Respiration
• Respiration is the oxidation of a source of energy by removal of electrons and donation to an inorganic terminal electron accepter.
Fermentation
• Fermentation is defined as an energy yielding process whereby organic molecules serve as both electron donors and electron accepters
• The molecule being metabolized does not have all its potential energy extracted from it. In other words, it is not completely oxidized
Oxygen Requirements
Obligate aerobes must grow in the presence of oxygen they cannot carry out fermentation
Obligate anaerobes do not carry out oxidative phosphorylation they are killed by oxygen
Aerotolerant anaerobes are bacteria that respire anaerobically but can survive in the presence of oxygen
Facultative anaerobes can perform both fermentation and aerobic respiration
Microaerophilic bacteria grow well in low concentrations of oxygen,but are killed by higher concentrations.
Obligate aerobes
• grow in presence of oxygen
• no fermentation
• oxidative phosphorylation
Obligate anaerobes
• no oxidative phosphorylation
• fermentation
• killed by oxygen
• lack certain enzymes
superoxide dismutase
O2-+2H+ H2O2
catalase
H2O2 H20 + O2
peroxidase
H2O2 H20 /NAD NADH
Facultative anaerobes
• fermentation
• aerobic respiration
• survive in oxygen
Microaerophilic bacteria
• grow
– low oxygen
• killed
– high oxygen
Optimal growth temperature
• Mesophiles:
– human body temperature
* pathogens
* opportunists
• pyschrophile
– close to freezing
• thermophile
– close to boiling
pH
• Many grow best at neutral pH
• Some can survive/grow
- acid
- alkali
Nutrient Requirements
• Carbon
• Nitrogen
• Phosphorus
• Sulfur
• Metal ions (e.g. iron)
Siderophores (S)
Small molecule secreted by bacteria that bind iron
Measuring bacterial mass in liquid cultures of bacteria
Common methods include:
a) turbidity (the cloudiness of a liquid culture of bacteria –
a measure of total bacteria [live and dead]
b) the number of viable bacteria in a culture –
usually assessed by counting the number of colonies
that grow after streaking a known volume on a plate
Measuring bacterial mass (live + dead) in liquid culture
Measured with spectrophotometer
Measuring viable bacteria
Colony forming units
SUGAR CATABOLISM
• Glycolysis
– Embden Meyerhof Parnas Pathway
– most bacteria
– also animals and plants
STERILIZATION
• All killed
• non-selective
• autoclaving
• 121oC (heat/pressure)
* Heat resistant materials
• ethylene oxide
• Non heat resistant
– usually equipment
• ultra-violet light
– surfaces (e.g operating rooms)
not totally effective
• gamma radiation
– food
– some mail
Disinfection
• Liquids that kill bacteria
– e.g. phenol based
– too toxic for skin surfaces
Antiseptics
• Topical (e.g. skin)
– e.g. iodine or 70% alcohol
– “reduce” bacterial load
ANTIBIOTICS
• Selectively toxic for bacteria
– bactericidal (killing)
– bacteriostatic (growth inhibition)
• no harm to patient
• destroy structures
• present in bacteria
• not present in host
Antibiotics work together
with immune system
Minimal inhibitory concentration
• lowest level stopping growth
• e. g. zone of inhibition around a disk impregnated with antibiotic
• Antibiotics that inhibit cell wall
biosynthesis are bactericidal
• Without cell wall, osmotic pressure
causes bacteria to burst
Antibiotics: Protein Synthesis, Nucleic Acid Synthesis and Metabolism
Principles and Definitions
• Selectivity
– Selectivty8 toxicity9
• Therapeutic index
– Toxic dose/ Effective dose
• Categories of antibiotics
– Bactericidal
• Usually antibiotic of choice
– Bacteriostatic
• Duration of treatment sufficient for host defenses
• Categories of antibiotics
– Use of bacteriostatic vs bactericidal antibiotic
• Therapeutic index better for bacteriostatic antibiotic
• Resistance to bactericidal antibiotic
• Protein toxin mediates disease – use bacteriostatic protein synthesis inhibitor
• Antibiotic susceptibility testing (in vitro)
– Minimum inhibitory concentration (MIC)
• Lowest concentration that results in inhibition of visible growth
– Minimum bactericidal concentration (MBC)
• Lowest concentration that kills 99.9% of the original inoculum
• Combination therapy
– Prevent emergence of resistant strains
– Temporary treatment until diagnosis is made
– Antibiotic synergism
• Penicillins and aminoglycosides
• CAUTION: Antibiotic antagonism
• Penicillins and bacteriostatic antibiotics
Antibiotics that Inhibit Protein Synthesis
Protein Synthesis Inhibitors
• Mostly bacteriostatic
• Selectivity due to differences in prokaryotic and eukaryotic ribosomes
• Some toxicity - eukaryotic 70S ribosomes
Inhibitors of Nucleic Acid Synthesis
• Rifampin, Rifamycin, Rifampicin, Rifabutin (bactericidal
• Quinolones (bactericidal)
nalidixic acid, ciprofloxacin, ofloxacin, norfloxacin, levofloxacin, lomefloxacin, sparfloxacin
Antimetabolite Antimicrobials
• Inhibitors of Folic Acid Synthesis
• Sulfonamides, Sulfones (bacteriostatic)
• Trimethoprim, Methotrexate, Pyrimethamine (bacteriostatic)
Antimicrobial Drug Resistance
Principles and Definitions
• Clinical resistance
• Resistance can arise by mutation or by gene transfer (e.g. acquisition of a plasmid)
• Resistance provides a selective advantage
• Resistance can result from single or multiple steps
• Cross resistance vs multiple resistance
– Cross resistance -- Single mechanism-- closely related antibiotics
– Multiple resistance -- Multiple mechanisms -- unrelated antibiotics
Antimicrobial Drug Resistance Mechanisms
• Altered permeability
– Altered influx
• Gram negative bacteria
– Altered efflux
• tetracycline
• Inactivation
– $-lactamse
– Chloramphenicol acetyl transferase
• Altered target site
– Penicillin binding proteins (penicillins)
– RNA polymerase (rifampin)
– 30S ribosome (streptomycin)
• Replacement of a sensitive pathway
– Acquisition of a resistant enzyme (sulfonamides, trimethoprim)
Pathogenicity
• virulence factors
• number of initial organisms
• immune status
Koch's postulates
• isolated
– diseased not healthy people
• growth
– pure culture
• induce disease
– susceptible animals
• re-isolated
– susceptible animals
Opportunistic infections
• compromised people
– normal flora
– environment
Opportunists - normal flora
• Skin
– Staphylococcus aureus,
– S. epidermidis
– Propionibacterium acnes
• Intestine
– Bacteroides
* high numbers
– Enterobacteriaceae
* low number
Opportunists - environment
» air
• water
• soil
• food
• Opportunists in hospital- nosocomial
Transmission
• airborne droplets
• food
• water
• sexual contact
Host defenses
• Gut
– peristalsis
– defecation
• respiratory tract
– ciliary action
– coughing
– sneezing
• urogenital tract
– urination
COMMENSALS CAUSE DISEASE
WHEN HOSTS DEFENCES ARE LOW
PATHOGENS POSSESS VIRULENCE FACTORS
THAT ENABLE THEM TO CAUSE DISEASE IN
THE PRESENCE OF NORMAL DEFENCES
DETERMINANTS OF BACTERIAL PATHOGENISIS
TRANSMISSION
INFECTING DOSE-INOCULUM
PORTAL OF ENTRY
BACTERIAL VIRULENCE FACTORS
CAPSULES
INVASIVENESS
ADHESINS
EXOENZYMES
TOXINS
CAPSULES
LOCATED EXTERNAL TO THE CELLWALL
COMMON VIRULENCE FACTOR
POLYSACCHARIDES
AVOID OR SURVIVE PHAGOCYTOSIS
ADHESINS
ALLOW BACTERIA TO STICK ON MUCOSAL SURFACE
ADHESINS ARE SURFACE FACTORS
PILI / FIMBRIAE
LPS LIPOPOLYSACCARIDE SIDECHAINS
M PROTEIN
INVASIVENESS
ABILITY TO INVADE HOST CELL
MULTIFACTORIAL COMPLEX PROCESS
MAY INCLUDE ADHESION AND ENZYMES FACTORS
EXOENZYMES
PRODUCE AND SECRETED BY BACTERIA
ENZYMES THAT BREAKDOWN COLLAGEN AND FIBRIN
COLLAGENASES / HYALURONIDASES / FIBRINOLYSINS
ENZYMES THAT BREAKDOWN CELLULAR MATERIALS
PROTEASES / LECITHINASES
ENZYMES THAT DEACTIVATE AND MODIFY ANTIBIOTICS
BETA LACTAMASE
ENZYMES THAT DESTROY NEUTROPHILS
AND MACROPHAGES
LEUKOCIDINE
ENZYMES THAT ACCELERATE FIBRIN CLOT
COAGULASE
TOXINS
2 CLASSIFICATIONS
EXOTOXINS – PROTEINS PRODUCED AND
SECRETED BY BACTERIA
ENDOTOXINS – PART OF THE BACTERIA ITSELF
EXOTOXIN
NEUROTOXINS- AFFECT NERVOUS TISSUE
TETANUS TOXIN
ENTEROTOXINS – AFFECT GASTROINTESTINAL TRACT
CHOLERA TOXIN
CYTOTOXINS – AFFECT CELLS OF VARIOUS TISSUES
DIPHTERIA TOXIN
ENDOTOXINS
ALL GM NEGATIVE BATERIA HAVE ENDOTOXINS
IN OUTER MEMBRANE
RELEASED AS BACTERIA ARE LYSED
RESPONSIBLE FOR SEPSIS AND
SEPTIC SHOCK - FATAL
Friday, December 4, 2009
ectron transport chain of proteins
synthesis of ATP is a series of reduction-oxidation(redox) reaction..
watch and learn by reviewing the questions
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