Lecture 6 _ Molecular Epidemiology

Question 1

What are phylogenetic trees

Answer 1

A graphical representation of the evolutionary relationships among a set of species or taxa. Can also be applies to populations, individuals, and even genes

Question 2

Phylogenetic trees are also called

Answer 2

Phylogenies or evolutionary trees & this field of study is known as phylogenetics

Question 3

Phylogenetic trees - nodes

Answer 3

Hypothetical ancestors of the children taxa

Question 4

Phylogenetic trees - root

Answer 4

Common ancestor of all species/individuals on chart. Sometimes you know the root, other times you don’t

Question 5

Phylogenetic trees - branches

Answer 5

Show who is related to who

Question 6

Phylogenetic trees - leaves/tips

Answer 6

Represent species, taxa, populations, individuals, or genes

Question 7

In an unrooted tree, what do you not know

Answer 7

Where the tree starts, who is common ancestor, and passage of time

Question 8

What is a cladogram vs phylogram?

Answer 8

Cladogram - length of branches is arbitrary & shows you clades (groups or species)
Phylogram - length of branches does mean something

Question 9

Phylogenetic trees - does axis that root is on have a meaning

Answer 9

No meaning

Question 10

Phylogenetic trees - does axis from root to branches have a meaning

Answer 10

Yes - shows passage of time

Question 11

QUESTION ON FINAL!!!! Look at slide 9 & 10 and figure out who shares a common ancestor

Answer 11

1. A, B, C, & D -
2. C&D also have a common ancestor
3. A & B have a common ancestor

Question 12

Haplotype network

Answer 12

A graphical representation of evolutionary relationships among haplotypes (sequence of a specific stretch of DNA/RNA) within a population or species
- could be a small stretch of DNA, big stretch of DNA, or could also be genome

Question 13

If you have a stretch of DNA/phenotypes you can build the

Answer 13

Haplotype

Question 14

In a Haplotype, the size of the circle shows

Answer 14

How abundant that sequence is

Question 15

Dashes in Haplotype represent

Answer 15

of differences between those 2 sequences (2 circles)

Question 16

With a Haplotype network, you can track

Answer 16

How sequences change over time in a population

Question 17

What is epidemiology

Answer 17

The study of the distribution (who, when, where) patterns, and determinants of disease in a defined population

Question 18

Examples of epi uses

Answer 18

Outbreak investigation, identifying cause of disease, disease transmission, identify risk factors, improve public health, and preventative health care

Question 19

Epidemiologic triangle

Answer 19

Agent
Host
Environment

With disease in the middle of the triangle

Question 20

Epidemiologic triangle - agent

Answer 20

Actual cause of the disease

Question 21

Epidemiologic triangle - host

Answer 21

Organisms that harbour the disease

Question 22

Epidemiologic triangle - environment

Answer 22

External factors that affect disease transmission (sanitation, temperature, etc)

Question 23

What is BRD

Answer 23

Bovine Respiratory Disease - A multifactorial disease caused by the interaction of pathogenic bacteria and viruses & causes a variety of clinical signs

Question 24

What’s a leading cause of death in dairy heifers

Answer 24

BRD

Question 25

Risk of BRD is reduced by

Answer 25

-Feeding calves with milk rather than milk replacers - feeding over 3.8L of milk per day to calves under 21d of age -frequent changing of maternity pen bedding -housing calves in all wood or plastic hutches rather than metal roofed hutches (very hot, leads to heat stress) -administration of BRD vaccines to dams before calving

Question 26

Molecular epidemiology can tell us

Answer 26

Cause of disease (agent) Source of infection (where did outbreak start - DNA) Routes of transmission Virulence - molecular tools used to find strain Host susceptibility - GWAS

Question 27

Molecular epidemiology tools for pathogen/species detection

Answer 27

PCR/qPCR DNA metabarcoding

Question 28

Molecular epidemiology tools - Within-species variation

Answer 28

Cutting up genome with restriction enzymes (looks for strain of different bacteria) Sequencing

Question 29

Criteria for selecting the most appropriate approach

Answer 29

Measure the construct of interest (PCR enough to test for presence/absence) - (need to sequence if want diff variants) Level of discrimination - sufficiently discriminatory in the population of interest Reliable, reproducible Requirements for specimen collection, storage, and handling Cost effective

Question 30

Cutting up of DNA uses

Answer 30

Restriction enzymes

Question 31

Restriction enzymes do what

Answer 31

Characterize differences between bacterial strains Variety is below species level

Question 32

What is difference between restriction enzymes vs restriction endonuclease

Answer 32

Restriction enzymes are one class of restriction endonucleases

Question 33

Restriction enzymes recognize a sequence motif then

Answer 33

Cut DNA at those points & cut the bacteria into pieces

Question 34

If restriction site is GAATTC, what is restriction enzyme cutting site

Answer 34

G - CUT - AATTC

Question 35

What is Pulses-Field Gel Electrophoresis (PFGE)?

Answer 35

Used in microbiology for typing bacteria (identifying bacteria strains) Grow bacteria colonies, then put restriction enzymes in, put through a gel, and bands show up where cuts are made (so where sequences were recognized)

Question 36

In PFGE, will bacteria that is genetically similar get cut at same spots?

Answer 36

Yes

Question 37

What is standard approach at CFIA and Public Health Agency of Canada (PHAC) for food borne and waterborne disease

Answer 37

PFGE

Question 38

E. coli are a diverse group of bacteria that normally live in

Answer 38

The intestines of humans and animals

Question 39

Most E Coli strains are harmless but

Answer 39

Some produce toxins that can cause disease such as severe and bloody diarrhea, and life-threatening kidney failure

Question 40

PFGE major drawbacks

Answer 40

Time consuming - 3:5 days Genome not always cuttable Identical bands do NOT equal identical genomes - only looking at few cut sites, not looking at all DNA (sequencing DNA is more accurate)

Question 41

For outbreak at goat farm, what were the methods for source tracing

Answer 41

Molecular sub typing to determine outbreak strains PFGE pattern = unique DNA fingerprint Isolates with same PFGE patterns = more likely to share common source Joint farm visit with Connecticut department of agriculture and local health director - conducted environmental and animal sampling

Question 42

Read case study in lecture slides on source tracing on goat farm !

Answer 42

Do it !

Question 43

In case study, they sampled for E. coli where

Answer 43

Kid barn, goats themselves, doe barn, & food samples stored Fecal samples & rectal swabs Cheeses & unpasteurized milk

Question 44

Conclusions of case study on goat farm outbreak

Answer 44

Epi & laboratory evidence for outbreak of E. coli Exposure to goats & widespread contamination in environment - farm had a lack of awareness of risks - no hand washing facilities - availability of hand sanitizer an ineffective measure

Question 45

What is Foot and Mouth Disease

Answer 45

A highly infection viral disease that causes fever, blister like sores on tongue, lips, mouth, teats, and between hooves. can also cause weight loss, lameness, and occasional abortions

Question 46

Which animals does foot and mouth disease affect

Answer 46

Cloven hoof animals (cattle, sheep, swine)

Question 47

FMD virus

Answer 47

Single strand, positive sense RNA with high mutation rate (lots of genetic variation)

Question 48

FMD virus - conventional molecular epidemiology

Answer 48

Sequencing of VP1 (8% of genome, which is not enough to resolve a single outbreak) - not enough detail

Question 49

Look at FMD case study slides 37-42

Answer 49

Do it <3

Question 50

How was FMD transmitted from farm to farm

Answer 50

No animal move,ent Animals form premises 1 & 2 were disinfected prior to infection of premise 5 -movement of person, object, and aerosols?