Lecture 6 _ Molecular Epidemiology Flashcards
What are phylogenetic trees
A graphical representation of the evolutionary relationships among a set of species or taxa. Can also be applies to populations, individuals, and even genes
Phylogenetic trees are also called
Phylogenies or evolutionary trees & this field of study is known as phylogenetics
Phylogenetic trees - nodes
Hypothetical ancestors of the children taxa
Phylogenetic trees - root
Common ancestor of all species/individuals on chart. Sometimes you know the root, other times you don’t
Phylogenetic trees - branches
Show who is related to who
Phylogenetic trees - leaves/tips
Represent species, taxa, populations, individuals, or genes
In an unrooted tree, what do you not know
Where the tree starts, who is common ancestor, and passage of time
What is a cladogram vs phylogram?
Cladogram - length of branches is arbitrary & shows you clades (groups or species)
Phylogram - length of branches does mean something
Phylogenetic trees - does axis that root is on have a meaning
No meaning
Phylogenetic trees - does axis from root to branches have a meaning
Yes - shows passage of time
QUESTION ON FINAL!!!! Look at slide 9 & 10 and figure out who shares a common ancestor
- A, B, C, & D -
- C&D also have a common ancestor
- A & B have a common ancestor
Haplotype network
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
If you have a stretch of DNA/phenotypes you can build the
Haplotype
In a Haplotype, the size of the circle shows
How abundant that sequence is
Dashes in Haplotype represent
of differences between those 2 sequences (2 circles)
With a Haplotype network, you can track
How sequences change over time in a population
What is epidemiology
The study of the distribution (who, when, where) patterns, and determinants of disease in a defined population
Examples of epi uses
Outbreak investigation, identifying cause of disease, disease transmission, identify risk factors, improve public health, and preventative health care
Epidemiologic triangle
Agent
Host
Environment
With disease in the middle of the triangle
Epidemiologic triangle - agent
Actual cause of the disease
Epidemiologic triangle - host
Organisms that harbour the disease
Epidemiologic triangle - environment
External factors that affect disease transmission (sanitation, temperature, etc)
What is BRD
Bovine Respiratory Disease - A multifactorial disease caused by the interaction of pathogenic bacteria and viruses & causes a variety of clinical signs
What’s a leading cause of death in dairy heifers
BRD
Risk of BRD is reduced by
-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
Molecular epidemiology can tell us
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
Molecular epidemiology tools for pathogen/species detection
PCR/qPCR
DNA metabarcoding
Molecular epidemiology tools - Within-species variation
Cutting up genome with restriction enzymes (looks for strain of different bacteria)
Sequencing
Criteria for selecting the most appropriate approach
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
Cutting up of DNA uses
Restriction enzymes
Restriction enzymes do what
Characterize differences between bacterial strains
Variety is below species level
What is difference between restriction enzymes vs restriction endonuclease
Restriction enzymes are one class of restriction endonucleases
Restriction enzymes recognize a sequence motif then
Cut DNA at those points & cut the bacteria into pieces
If restriction site is GAATTC, what is restriction enzyme cutting site
G - CUT - AATTC
What is Pulses-Field Gel Electrophoresis (PFGE)?
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)
In PFGE, will bacteria that is genetically similar get cut at same spots?
Yes
What is standard approach at CFIA and Public Health Agency of Canada (PHAC) for food borne and waterborne disease
PFGE
E. coli are a diverse group of bacteria that normally live in
The intestines of humans and animals
Most E Coli strains are harmless but
Some produce toxins that can cause disease such as severe and bloody diarrhea, and life-threatening kidney failure
PFGE major drawbacks
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)
For outbreak at goat farm, what were the methods for source tracing
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
Read case study in lecture slides on source tracing on goat farm !
Do it !
In case study, they sampled for E. coli where
Kid barn, goats themselves, doe barn, & food samples stored
Fecal samples & rectal swabs
Cheeses & unpasteurized milk
Conclusions of case study on goat farm outbreak
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
What is Foot and Mouth Disease
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
Which animals does foot and mouth disease affect
Cloven hoof animals (cattle, sheep, swine)
FMD virus
Single strand, positive sense RNA with high mutation rate (lots of genetic variation)
FMD virus - conventional molecular epidemiology
Sequencing of VP1 (8% of genome, which is not enough to resolve a single outbreak) - not enough detail
Look at FMD case study slides 37-42
Do it <3
How was FMD transmitted from farm to farm
No animal move,ent
Animals form premises 1 & 2 were disinfected prior to infection of premise 5
-movement of person, object, and aerosols?
