1. 260 summer 2016
pathogenesis

2. Clicker questions about viral replication
Which viruses can use the host’s replication enzymes to make new genomes
Which viruses have a genome that can be directly used as an mRNA
Which viruses require a RNA-dep RNA pol
Which viruses need to carry the RNA-dep RNA pol in the capsid
Reverse transcriptase uses a ____ template to make ____
A = DNA viruses B = all RNA viruses C = all RNA viruses except not retroviruses D=retroviruses only
A = DNA  DNA B = DNA  RNA C = RNA  DNA C = RNA  RNA
A = dsDNA virus B = dsRNA virus C = “+”sense RNA D = “-”sense RNA E= retrovirus

3. About PCR Compare replication to PCR
1. list the things you need (reagents, enzymes) for replication and in what order
2. list the things you put into your PCR tube (reagents, enzymes).
3. compare – for the things in replication that are missing in the PCR tube, what did you add to fill that missing role?

4. The study of disease Pathogenesis – the study of disease progression
Next class – epidemiology we’ll study disease again but from a public health perspective

5. Pathogenesis (chapter 16)
Principles of disease aka
The Chapter of Many Definitions

6. Why do care? What specific agent causes disease
How does the agent cause disease in the body
How infectious agents are spread

7. A brief history
Diseases and plagues attributed to supernatural forces, incurable and terrifying
Observation: disease seems to be communicable
1600s there are microbes
1800s Microbes cause disease
Robert Koch (German physician ) – systematic proof of causative agent

8. Koch’s Postulates
The same pathogen must be present in every case of the disease
The pathogen must be isolated from the diseased host and grown in pure culture
The pathogen from the pure culture must cause the disease when it is inoculated into a healthy, susceptible lab animal
The pathogen must be isolated from the inoculated animal and must be shown to be the original organism

9. Koch’s Postulates Obtain pure culture of the organism
Pure culture used to infect healthy lab animal causes the disease
Same organism is present
in every case of the disease
Figure 14.3

10. Koch’s Postulates
Organism cultured from infected animal is identical to original infection
Figure 14.3

11. Koch’s Postulates
Koch's postulates can be used to prove the cause of an infectious disease..
But can Koch’s postulates be used to identify the agent/microbe responsible for all diseases?
Some pathogens can cause several disease conditions
Some pathogens cause disease only in humans
Some pathogens are not easily cultivated

12. Pathology, Infection, and Disease
Pathology: The study of disease
Etiology: The study of the cause of a disease
Pathogenesis: The development of disease
Infection: Colonization of the body by pathogens
Disease: An abnormal state in which the body is not functioning normally

13. Normal Microbiota and the Host
Transient microbiota may be present for days, weeks, or months then disappear
Normal microbiota permanently colonize the host
Symbiosis is the relationship between normal microbiota and the host

14. Representative Normal Microbiota
Figure 14.1

15. Is this bad news?

16. Symbiosis
In commensalism, one organism benefits, and the other is unaffected
In mutualism, both organisms benefit
In parasitism, one organism benefits at the expense of the other
Some normal microbiota are opportunistic pathogens such as?

17. Normal Microbiota on the Human Body
Table 14.1

18. What is the role of normal microbiota?

19. Normal Microbiota and the Host
Microbial antagonism is a competition between microbes.
Normal microbiota protect the host by
Occupying niches that pathogens might occupy
Producing waste eg. acids
Producing bacteriocins
Probiotics: Live microbes applied to or ingested into the body, intended to exert a beneficial effect

20. Effect of antibiotics
Why are you at risk of a yeast infection or bacterial GI illness after taking antibiotics?

21. Principles of disease
Symptom: A subjective change in body function that is felt by a patient as a result of disease
Sign: An objective change in a body that can be measured or observed as a result of disease
Syndrome: A specific group of signs and symptoms that accompany a disease

22. For example You catch a respiratory virus
You feel tired, have a headache, your sinuses itch, you have a sore throat
The doctor finds swollen lymph nodes, notes an elevated body temperature and views redness in your throat and nasal passages
What are the symptoms, signs
How are the signs and symptoms related to the concept of a syndrome

23. The Stages of a Disease
Figure 14.5

24. Development of disease
Incubation period – from transmission to first symptoms
Prodromal period – early mild symptoms
Period of illness – period of severe symptoms
Period of decline – number of pathogens declines (may overlap with severe symptoms)
Period of convalescence (numbers continue to drop; mild or no symptoms)

25. For example 10/8 – your partner sneezes on you 10/10 – sore throat
10/12 – liquid goo phase, sneezing, fever
10/17 – back to salsa dancing!
Give the dates of the incubation period, prodromal period and period of illness

26. Severity or Duration of a Disease
Acute disease: Symptoms develop rapidly
Chronic disease: Disease develops slowly
Subacute disease: Symptoms in between acute and chronic
Subclinical disease: infection without noticeable symptoms
Latent disease: Disease with a period of no symptoms when the causative agent is inactive

27. Order of disease
Primary infection: Acute infection that causes the initial illness
Secondary infection: Opportunistic infection after a primary (predisposing) infection

28. For example
1. a few weeks after the salsa dancing success, you find you have a bacterial sinus infection
2. a patient with HIV develops thrush

29. Types of infection
Local infection: Pathogens are limited to a small area of the body
Systemic infection: Microbe or toxins spread throughout the body by the blood/lymph
Focal infection: Infection that began as a local infection spreads via the blood/lymph

30. Spread of disease – inside the body
Bacteremia: Bacteria present in the blood
Toxemia: Toxins present in the blood
Viremia: Viruses present in the blood
Septicemia: “blood poisoning” - systemic infection arising from multiplication of pathogens in the blood

31. Question
What’s the difference between bacteremia and bacterial septicemia
What’s the difference between bacteremia and toxemia

32. Spread of disease – in the population
Communicable disease: A disease that is spread from one host to another
Contagious disease: A disease that is easily spread from one host to another
Noncommunicable disease: A disease that is not transmitted from one host to another – SUCH AS??

33. For example
Determine whether these are communicable or not and if so, are they contagious
HIV
Influenza
Botulism

34. Mechanisms of Pathogenicity
Pathogenicity: Ability to cause disease
Are all microbes pathogens?
Virulence: The extent of pathogenicity
ID50: Dose that causes infection in 50% of population
LD50: Dose of toxin that is lethal for 50% of test population

35. Questions
For a given microbe, is the ID50 the same in every situation?
Which is less likely to cause disease – a microbe with a higher ID50or lower ID50?

36. Toxins Which organism has the most potent toxin?? Toxin LD50 Botulinum
0.03 ng/kg
Shiga toxin
250 ng/kg
Staphylococcal enterotoxin
1350 ng/kg
Which organism has the most potent toxin??

37. reservoirs What do you think? Human Animal (zoonotic) Environment
Carrier – human without symptoms

38. For example What is the reservoir for HIV Rabies
Mycobacterium tuberculosis
Clostridium botulinum

39. Modes of transmission Direct contact transmission – touch
Indirect contact transmission - fomite
Vehicle transmission – a vector brings the pathogen
Mechanical vector (rides the roof)
Biological vector (rides inside)

40. question What is the reservoir for malaria
What kind of transmission is it?
Direct contact, indirect contact, zoonotic, mechanical vector, biological vector
More than one right answer

41. Portals of Entry Preferred route of entry
Some have multiple routes (e.g. Yersina pestis causes plague; B. anthracis causes anthrax)
Other routes may not cause disease (e.g. Salmonella typhi on skin; Streptococci pneumoniae that are ingested)

42. Portals of Entry How do microbes get into your body? Mucous membranes
GI tract, respiratory tract most common
Genitourinary tract, conjunctiva (membrane of eyes)
Skin
Major barrier to entry, mostly inaccessible to pathogens
Some openings: hair follicles, sweat gland ducts
Hookwork larvae can bore through skin; some fungi grow on keratin
Parenteral route
Penetration of skin /mucous membranes (punctures, cuts, bites, etc.)

43. Bacillus anthracis Portal of Entry ID50 Skin 10–50 endospores
Inhalation
10,000–20,000 endospores
Ingestion
250,000–1,000,000 endospores
Which portal is most likely to cause infection??

44. So for example Plasmodium (malaria) – what route?
C botulinum (botulism) – what route?
Influenzavirus – what route?
HIV (AIDS) – what route?
Bacillus anthracis (anthrax) – what route?
Why are these routes different?

46. biofilms
~65% of infections
Dental plaque
Implants
Contact lenses

47. Adherence

48. Alveolar macrophage attacking E. coli; RBC at top left
capsules
Can prevent phagocytosis – make cells “invisible”
Alveolar macrophage attacking E. coli; RBC at top left

49. Alveolar macrophage attacking E. coli; RBC at top left
Capsules
Capsules prevent phagocytosis
Streptococcus pneumoniae; Haemophilus influenzae; Bacillus anthracis
Often avirulent without capsule since phagocytosed (eg S. pneumoniae)
Note - many nonpathogenic bacteria also produce capsules (capsulation alone is not sufficient to designate a pathogen)
Alveolar macrophage attacking E. coli; RBC at top left

50. Cell Wall Components M protein resists phagocytosis
Surface of fimbriae
Streptococcus pyogenes (strep throat, scarlet fever, TSS)
Opa protein inhibits T helper cells
fimbriae
Neisseria gonorrhoeae
Mycolic acid resists digestion
Waxy lipid where?
Mycobacterium tuberculosis

51. Hijacking host Cytoskeleton
Cytoskeleton filament - actin
Microbes (e.g. Salmonella; E. coli) produce invasins that rearrange actin filaments of cytoskeleton  membrane ruffling, penetration of cell
Some species use actin to propel themselves through and between cells (e.g. Shigella; Listeria; Ricksettia)
BEGIN HERE

52. Membrane Ruffling
Invasins: alters host actin to enter host cell eg.Salmonella
Figure 15.2

53. Comet tails Some bacteria organize actin behind them for propulsion
eg.Listeria, Shigella, Ricksettia

54. Damage to Host Cells Consumption of host’s nutrients
Direct damage in vicinity of invasion
TOXINS  blood/lymph  distant sites

55. Consumption of host nutrients
Lack of iron limits bacterial growth
Secrete siderophores to capture
Take up from blood; some lyse cells to release

56. Human Herpes Virus 6 budding off
Direct damage
Disrupt host cell function
Release wastes
Rupture  release more viruses/bacteria
Human Herpes Virus 6 budding off

57. Toxins Toxin: Substance that contributes to pathogenicity
Toxigenicity: Ability to produce a toxin
Toxemia: Presence of toxin in the host's blood
Toxoid: Inactivated toxin used in a vaccine
Antitoxin: Antibodies against a specific toxin

58. All about toxins May be transported by blood or lymph
what is the important implication?
May  fever, diarrhea, shock, cardiovascular and nervous system disruption
~220 known types; ~40% damage cell membranes
Two general types:
Exotoxins
Endotoxins
What do you think the difference is?

59. Exotoxins and Endotoxins
Lipid A of lipopolysaccharides (LPS) of outer membrane of Gram negatives
Proteins produced by bacteria and secreted or released following lysis
Figure 15.4

60. Exotoxins Specific for a structure or function in host cell
Figure 15.4a

61. Naming exotoxins By type of cell affected By associated disease
(e.g., neurotoxins, cardiotoxins, hepatotoxins, leukotoxins, enterotoxins, cytotoxins)
What does each affect?
By associated disease
(e.g., diphtheria toxin, tetanus toxin)
What disease is associated with each toxin?
By bacterium producing
(e.g., botulinum toxin, Vibrio enterotoxin
What organism produces each of these toxins?

62. What are exotoxins, really?
Proteins, usually enzymes  even small amounts harmful
1 mg botulinum exotoxin can kill 1 million guinea pigs
Genes usually carried on ?
Soluble in host fluids  transported widely
plasmids, phage – WHY??
START HERE

63. What are exotoxins, really?
Proteins, usually enzymes  even small amounts harmful
1 mg botulinum exotoxin can kill 1 million guinea pigs
Genes usually carried on ?
Soluble in host fluids  transported widely
Mode of action– 3 main modes
Inhibit metabolism (usually protein synthesis)
Destroy parts of host cells
overactivate immune system
plasmids, phage
START HERE

64. What are exotoxins, really?
Proteins, usually enzymes  even small amounts harmful
1 mg botulinum exotoxin can kill 1 million guinea pigs
Genes usually carried on ?
Soluble in host fluids  transported widely
Mode of action
Inhibit metabolism (usually protein synthesis)
Destroy parts of host cells
overactivate immune system
Types
A-B
membrane-disrupting
superantigen
plasmids, phage
START HERE

65. Exotoxin type: A-B Exotoxin
eg. diptheria,
botulinum,
tetanus,
vibrio (cholera)
Two parts – A and B, proteins
B (binding) – binds host receptor
A-B enter
A and B separate
A – enzyme,
usually inhibits
protein synthesis
Figure 15.5

66. Membrane-Disrupting exotoxin
Lyse host’s cells in one of two ways
Making protein channels in the plasma membrane
Leukocidins
Hemolysins
Streptolysins (b-hemolysis Streptococcus)
Disrupting phospholipid bilayer
Phospholipases remove polar head –gas gangrene (C. perfringens)
Why is this bad?

67. Exotoxin type: Superantigens
Evoke intense immune response
Proliferation T cells (lymphocytes)  excess cytokines
Symptoms: fever, nausea, vomiting, diarrhea, shock, death
Eg. Staphylococcus (food poisoning, toxic shock syndrome)

68. Last words about exotoxins
Exotoxins are disease specific
Due to exotoxin, not bacterial infection
foodborne intoxication
botulism, staphylococcal food poisoning
Fighting exotoxins
Body makes antibodies (antitoxins)  provide immunity
Inactivate (heat, HCHO, iodine)  toxoids
Toxoids used as vaccine to stimulate antibody production
tetanus, diphtheria
START HERE

69. Endotoxins – part of the cell
Part of outer membrane of Gram-negatives
Lipid portion of LPS, called lipid A
Death and lysis of bacteria; antibiotics can worsen
Stimulate macrophages -release high level cytokines
Fever, nausea, vomiting, diarrhea, sometimes shock, death (similar to but not as potent as superantigen exotoxins…)
Remember this?

70. Endotoxins and the Pyrogenic Response
Figure 15.6

71. Damage to Host Cells

72. Exotoxin Source Gram? Relation to microbe ?made/part of Chemistry
What kind of molecule?
Neutralized by antitoxin?
Y/N?
LD50
Need a little or a lot?
Figure 15.4a

73. Exotoxin Source Mostly Gram + Relation to microbe Made by growing cell
Chemistry
Protein
Neutralized by antitoxin?
Yes
LD50
Small
Figure 15.4a

74. Endotoxins Source Gram? Relation to Microbe Made/part? Chemistry
Molecule?
Neutralized by Antitoxin?
Y/N?
LD50
How much you need?
Figure 15.4b

75. Endotoxins Source Gram Relation to Microbe Outer membrane Chemistry
Lipid A
Neutralized by Antitoxin? No
LD50
Relatively large
Figure 15.4b

76. Horizontal transmission
Where do bacteria get their toxin genes?

77. Horizontal transmission
Transformation
Conjugation
Transduction

78. A word about plasmids Circular small DNA containing 4-5 genes
What did we just transform into E. coli in lab?
What genes were on that plasmid?
DIAGRAM WHITE BOARD
Many antibiotic resistance genes are transmitted between species by sharing plasmids
Transformation and conjugation

79. Plasmids can code for Antibiotic resistance genes Metabolic enzymes
An example from lab?
Metabolic enzymes
Examples from this class?
AND toxins
Tetanus neurotoxin
Staphylococcal enterotoxin
Dextransucrase (Streptococcus mutans  tooth decay)
Adhesins and coagulase (Staphylococcus aureus)
Fimbria (Enteropathogenic E. coli)

80. Other ways to get toxins: transduction
Lysogenic conversion: prophage in chromosome
Shiga toxin of E. coli O157
Virbrio enterotoxin of Vibrio cholerae (cholera)
Botulinum neurotoxin
Capsule of Streptococcus pneumoniae (virulence factor)
Diphtheria toxin

81. Exotoxins due to Lysogenic Conversion
Corynebacterium diphtheriae
A-B toxin
Streptococcus pyogenes
Membrane-disrupting erythrogenic toxin
Clostridium botulinum
A-B toxin; neurotoxin
C. tetani
Vibrio cholerae
A-B toxin; enterotoxin
Staphylococcus aureus
Superantigen

82. Pathogenicity of Viruses
Gain entry via attachment sites for cell receptors
Resulting damage termed cytopathic effects
Production of virus particles (inclusion bodies)
Inhibition of biosynthesis (DNA, RNA, protein synthesis)
Host cell lysis
Inclusion body in brain tissue of rabies victim

83. Syncytium (giant cell) caused by measles virus
Cytopathic effects
Cause lysosomes to release enzymes
Cause cells to fuse into syncytium
Changes in host cell function
Production of interferon
Chromosomal damage
Loss of contact inhibition
Syncytium (giant cell) caused by measles virus

84. Pathogenic Properties of Fungi
May cause disease, but usually lack toxins
Fungal waste products may cause symptoms
Chronic infections provoke an allergic response
Proteases
Candida
Capsule prevents phagocytosis
Cryptococcus (meningitis)

85. Exceptions - Fungal toxins
Ergot toxin - hallucinations
Claviceps purpurea
Aflatoxin (peanut butter)
Aspergillus
Mycotoxins (mushrooms)
Neurotoxins: Phalloidin, amanitin
Amanita phalloides

86. Pathogenic Properties of Protozoa
Presence of protozoa
Protozoan waste products may cause symptoms
Avoid host defenses by
Growing in phagocytes
Antigenic variation

87. Plasmodium
What disease?
reproduction in host cells

88. Giardia
Giardia lamblia (diarrhea) – digest cells and tissue fluids

89. Trypanosomes African trypanosomiasis – sleeping sickness
Chagas disease (US)
reproduction inside host

90. Helminths Use host tissue and resources Interfere with host function
Waste can cause symptoms

91. Algae A few species produce toxins
Paralytic shellfish poisoning – “red tide”
Dinoflagellates
Saxitoxin

92. Portals of Exit
Why do we care how the pathogen gets out?

93. Portals of Exit Important in spread of disease (epidemiology)
Respiratory tract
Coughing and sneezing
Gastrointestinal tract
Feces and saliva; “oral-fecal”
Genitourinary tract
STDs, urine
Skin
Wound infections; drainage
Blood vectors important
Biting arthropods and needles or syringes

94. Mechanisms of Pathogenicity
Figure 15.9