Sample collection and pathogen identification week 4

Question 1

Intended learning outcomes

Answer 1

• Describe methods for detection an monitoring of pathogens in marine mammals
• How can samples be collected and what can they be used for?
• How can the data we obtain be used to say something about distribution, abundance and circulation of pathogens?

Question 2

Pathogen abundance studies rely on…

Answer 2

• Antibodies
• Indicates the presence/absence of a pathogen
• The animal has been infected
• Serum samples –> are rarely collected (requires the blood of a live of very fresh animal)
• Minus: Does not tell anything about the origin of the pathogen
• DNA/RNA
• Possibility of tracing the origin of the pathogen
• The animal still have the pathogen in its system
• Swabs and tissue samples
• Minus: Low chance of finding positive samples

Question 3

Genetic identification of a viral pathogen

Answer 3

Sample collection –> DNA/RNA extractions –> Quality check (gel electrophoresis) –> Samples of good quality –> RT-PCR or RT-qPCR –> Gelectrophoresis of PCR products –> Sequencing

Question 4

Live animals - Sample collection

Answer 4

Behaviour
* Satellite tagging
* Consumption: faecal samples

Population genetics
* Biopsies

Pathogens and microbiomes
* Swabs
* FTA cards
* Blood

Question 5

Necropsy - Sample collection

Answer 5

Population genetics
* Muscle tissue

Pathogens and microbiomes
* Viruses:
- Tissue: Brain, spleen, lung
- Blood
- Swabs: trachea, conjunctival, rectal

• Bacteria:
• Tissue: Brain, spleen, lung
• Intestine (faeces)
• Swabs: trachea, rectal

Question 6

Environmental specimen banks

Answer 6

National sample collection of different tissues:
* Muscle
* Brain
* Spleen
* Lung
* Kidney
* Liver
* Intestine
* Stomach etc.

Question 7

Targeted Pathogen screenings

Answer 7

• Detection of a selected pathogen
• Plus: Less expensive, rapid answer to presence/identification
• Minus: Information is restricted to the targeted pathogen

Question 8

Non-targeted pathogen screenings

Answer 8

• Detection of a broad range of pathogens
• Metabarcoding (e.g. 16S for bacteria)
• Shotgun sequencing (everything)
• Plus: Detection of a broad range of pathogens
• Minus: more expensive, potential bioinformatic errors resulting in false positive hits

Question 9

PCR definition

Answer 9

Polymerase Chain Reaction (aka amplification of a specified piece of genetic material)

Question 10

Species identification?

Answer 10

BLAST
* Genetic sequence –> Alignment: Comparison of DNA/RNA sequences to an online data base
* Sometimes leads to rapid species identification

Question 11

Phylogenetic trees

Answer 11

• Diagram describing the ancestral relationships among genetic sequences (individuals, populations, species etc.)
• Each split represent the last time two branches shared a common ancestor.

Question 12

Time calibrated phylogenetic trees

Answer 12

• Bayesian phylogenetic analyses.
• More complex setup.
• Incorporation of collection dates.
• Ex) used to monitor the evolution and origin of viral diseases.