Microbio Exam 4

Culture-based identification of microorganisms

Use selective/differential media to determine physiological properties

Ex. antibiotic sensitivities, biochemical properties, gram-reaction, cell shape

Culture-based identification of microorganisms is best and worst for

Best for bacterial pathogens

Not good for viruses

Dichotomous keys

Minimize number of tests/resources needed to ID an organism based on physiological characteristics

Microscopy is useful for

Identifying pathogens with unusual morphology and for determining Gram reaction

Culture-independent methods

Next-gen sequencing

PCR

RT-PCR

Serology

Immunoassays

Serology

Study of blood serum and bodily fluids

Mainly looks for antibodies

Immunoassays

Tests using antibodies/antigens to measure presence/concentration of molecules in a solution

Antibodies

Proteins made by the body that recognize antigens

Antigens

Recognized as foreign by the body

Agglutination

Antibodies crosslink antigenic particles to form clumps

Visible clumps on some wells of microtiter plate

Immunofluorescence

Antibody (primary or secondary) bound with a fluorochrome binds to antigen to diagnose specific antigens in tissues

Direct or indirect

Direct immunofluorescence

Fixing specimen to a slide and staining it with antibody-linked fluorophores

Indirect immunofluorescence

Detect antibodies in blood serum of patient after exposure to pathogen

ELISA

Antibody attached to an enzyme binds to an antigen

Examples of immunoassays

Agglutination

Immunofluorescence

ELISA

Why are microbes frenemies

1000+ species that are mainly good, but some pathogenic

Make up 3 lbs of weight

Infection

Multiplication of a microbe in a host and competition for supremacy

Disease

Change in the body away from good health

5 types of human pathogens

Viruses

Helminths

Protozoa

Fungi

Bacteria

What type of pathogen causes most human infections

>65% of human infections are from RNA viruses

Are all pathogens equally virulent

No, some more than others

B. anthracis is highly virulent

Lactobacillus is non-virulent

Polymicrobial diseases

Caused by more than one microbe acting together or in succession

Primary vs secondary infection

Primary in healthy body

Secondary in individual weakened by primary infection

Public Health

Science of protecting populations from disease and improving health through education, lifestyle change, and prevention

Why do public health agencies map outbreak to geographical locations

Help identify the source, routes of transmission, and identify at-risk populations

5 factors to break chain of transmission

Agent

Virulence

Exposure

Dose

Susceptibility

Why have so many diseases appeared

Changes in land use

Changes in human demographics

Poor population health

Pathogen evolution

Contamination

Global travel

Failure of public health
Global trade

Climate change

Morbidity

Incidence/population

Mortality

Deaths/incidence

Common source epidemic vs propagated epidemic

Common source from non-communicable disease

Propagated from communicable disease

Herd Immunity

Resistance of population to infection because enough people in population are immune to the infection

Not effective against non-communicable diseases

2 ways to control infections

Reduce/eliminate source/reservoir

Reduce susceptible population

Passive vs active immunity

Passive is immunity from someone/thing else

Active is immunity from self

Epidemiologists identify what 3 things

Reservoir

Mode of transmission

Presence among susceptible hosts

Reservoir

Source of microbe

AKA carriers

Mode of transmission for disease

Communicability?

Direct or indirect contact?

Aerosol transmission?

Vehicle

Inanimate object that spreads disease

Water, food, fomite

Vectors

Animate objects that spread disease

Arthropods, mosquitos

Nosocomial infections

Infections uniquely present in healthcare settings

Nosocomial infections depend on what 3 things

Compromised host

Hospital pathogens

Chain of transmission

Innate immunity characteristics

Non-specific

Doesn’t change throughout life

Rapid response (hours)

Barriers, cells, cytokines, inflammation, etc

Adaptive immunity characteristics

Specific

Evolves over lifetime

Slow response (days/years)

Memory

Antibodies, T-cells, B-cells

What barriers are found in the innate immunity

Skin

Mucous

Normal microbiota

How do the barriers in the innate immune system fight off pathogens

Flushing

Lysozyme/antimicrobials

pH

Sebaceous microenvironments on skin

Forehead, behind ear, back, sides of nostrils

Propionibacterium dominant

Moist areas on skin

Staphylococci occupy this

Corynebacteria

What does staphylococci use as a N source

Urea

Corynebacteria characteristic

Hard to study in vitro but apparent big role

Dry areas of skin characteristics

Most diverse (all 4 dominant phyla)

Lots of gram- bacteria

Are microbes on the skin constant on the skin

No, they vary by site and person

Lysozyme characteristics

Present in tears, saliva, mucus

Cleaves PG, resulting in osmotic lysis of bacterial cells

Female vs male genital barriers

Female: urethra colonized, vagina colonized (mainly LAB), dysbiosis can lead to vaginosis

Male: only distal end of urethra colonized

Types of leukocytes

Phagocytes (neutrophils, macrophages)

Lymphocytes (B and T cells)

Where are the phagocytes in the body

Neutrophils in blood

Macrophages in tissue

What do leukocytes recognize

MAMPs

Microbe associated molecular patterns

Role of primary organs in lymphatic system

Produce lymphocytes

Thymus for T cells

Bone marrow for B cells

Hematopeoesis

Formation of blood cells
Occurs throughout life

How do leukocytes recognize MAMPs

Through pattern recognition molecules (PRMs) or toll-like receptors (TLRs)

Phagocyte role

Engulf prey → internal lysozymes, oxidizers, and hydrolases digest it

Phagocytosis

Deadly blow of ROS, RNS, acid and/or other antimicrobial chemicals

What happens when MAMP bind TLRs

Cytokines synthesized

Cytokines

Low-molecular weight proteins that signal immune cells

Immune “hormones”

Cytokines regulate

Innate resistance

Hematopoiesis

Adaptive immunity

Opsonization

Coating pathogen with chemicals to increase phagocytosis

Complement system

~30 inactive proteins in the bloodstream that get cleaved to activate

Complement are the first opsonins to be produced in response to infection

Inflammation on a cellular level

Vasodilation and increased vascular permeability allow protein, inflammatory cells and fluid to enter tissue

Infected tissue becomes inflamed

Neutrophils travel to infected tissue where they engulf and kill bacteria

Neutrophils die in tissue and are processed by macrophages

Adaptive immunity contains what 2 responses

Humoral

Cell-mediated

Humoral response includes

B-cells

Antibodies

Targets extracellular antigens

Cell-mediated immune response includes

T-cells

Targets abnormal cells

How do B and T cells mature and go through clonal selection

Rapid cell division

Cells that recognize self are eliminated

Naive lymphocytes ready to differentiate

Lymphocytes encounter antigens

Antigens activate clones that carry matching receptors to trigger proliferation and differentiation

When does the antigen-independent period take place

In a fetus

When does the antigen-dependent period occur

After birth and throughout life

What are present on the surface of B cells

Antibodies

How many types of B cells in a person

10^13

Each specific for a single epitope of an antigen

How are B cells activated

When the antibody binds an antigen

Antibody

Proteins that react with epitopes in ECF

Variable region and conserved region

How is antibody diversity generated

Combinatorial joining (rearrangement of antibody exons)

Generation of different codons during antibody gene splicing

Variable regions are mutation hot spots

B cells all have different genotypes

6 ways antibodies work

Viral inhibition

Neutralization

Opsonization

Agglutination

Precipitation

Phagocytosis

2 Types of Antibody Response

Primary (IgM first then IgG much later, lower total antibody concentration)

Secondary (IgM first then IgG shortly after, higher total antibody concentration

What do B cells differentiate into

Plasma cells (antibody machine guns)

Memory B cells (can produce more effector cells)

What does B cell activation cause

A T-cell response → Cytokine production → facilitate B cell differentiation

Macrophage role

Present antigens from EC sources on their surface in MHC-II receptors

MHC-I characteristics

On all nucleated cells of the body to ID as self, varies person to person

Binds antigens from cytoplasm

CD8+ (cytotoxic) recognize MHC-I

MHC-II characteristics

Produced by phagocytes

Bind processed antigens from EC pathogens taken up from phagocytosis

CD4+ (helper) recognize MHC-II

Tolerance of Self

T cells recognize self are destroyed through apoptosis

Leads to autoimmune disease

Minimal self damage

Immune response must be strong enough to eliminate pathogen without damaging host

Overreaction leads to anaphylactic shock (1918 Spanish Flu)

Superantigens

Crosslink MHC-II to T-cells without an antigen

Overstimulates T cell → huge amount of cytokines

Cytokines stimulate vasodilation and inflammation (cytokine storm)

Can lead to tissue damage and possible multi-organ failure

What molecules make bad antigens

Central metabolites

DNA

Lipids

Highly conserved proteins

Intracellular proteins

Animal virion shapes

Coronavirus

Retrovirus

Paramyxovirus

RNA vs DNA viruses

RNA: own replication enzymes, error prone, replicates in cytoplasm

DNA: uses cell DNA replication apparatus, replicates in nucleus

Do viral pathogens target the same sites in the body

No, have many different sites

Respiratory system, GI system, CV system, etc

How does measles, coronavirus, influenza, and HIV transmit

Respiratory droplets: measles, coronavirus, influenza

Contact of bodily fluids: HIV

Measles characteristics

Respiratory virus

Symptoms include cough, fever, headache, conjunctivitis

Outbreaks increasing due to lack of vaccination

Measles virus characteristics

(+) RNA virus

16 kb genome

Low mutation rate

H and F spikes bind to many distributed receptors

Enters lungs, fuses with macrophage membrane, then transported into lymph

How does measles infect and leave B-cells

Infects B-cells using RdRp to replicate and leaves by budding

Measles complications

Subacute sclerosing panencephalitis (SSPE, neurological deterioration, 100% fatal, vaccination is 100% effective in prevention, rare)

Immune amnesia (resets immune memory, lasts months to years after infection)

What coronaviruses have crossed species barrier since 2000

SARS-CoV-1 in 2002 (11% fatal)

MERS-CoV in 2012 (35% fatal)

SARS-CoV-2 in 2019 (0.2-4.9% fatal)

SARS-CoV-2 virus characteristics

(+) ssRNA

30 kb genome (large)

RdRp has some proofreading capabilities, but source of mutations

Spike, E, and M proteins can be targets for immune system

Where does SARS-CoV-2 attach, and how does it lead to symptoms

To ACE-2 receptor, which is in different quantities in different people, leading to varying symptoms

How does SARS-CoV-2 replicate and what is the immune response

Spike protein binds to ACE2 receptor

+RNA is translated by host

RdRp makes genome copies

Immune cells release chemokines to attack infected cells