Lab Detection of Drug Resistance

Question 1

Types of ressistance

Answer 1

• genotypic
• phenotypic
• clinical

Question 2

Problems with molecular testing

Answer 2

• genotype-phenotype relationship incompletely understood
• geographic variability in mutation distribution
• heteroresistance and mixed infections

Question 3

Problems with culture-based testing

Answer 3

• slow
• complex for some drugs
• controversy over critical concentrations
• biosafety

Question 4

3 approaches to culture-based DST

Answer 4

• absolute concentration
• resistance ratio
• proportion (reference standard)

Question 5

Testing genotypic rif resistance

Answer 5

RIF binds to B-subunit of DNA-dependent RNA polymerase

- prevents initiation of transcription

Question 6

Steps in line probe assays (Genotype MTBDRplus)

Answer 6

• DNA isolation
• PCR amplification
• Detection
• Reverse hydridisation and signal detection

Question 7

Steps in Gene Xpert

Answer 7

• sputum liquefaction and inactivation
• sample filtered and washed
• ultrasonic lysis of filter capture organisms to release DNA
• DNA molecucles mixed with dry PCR reagents
• semi-nested real-time amplification and detection in integrated reaction tube

Question 8

Gaps in Xpert-driven algorithm

Answer 8

• no routine INH susceptibility testing
• complexity of algorithms
• delay for 2nd line DST

Question 9

Advantage of sequencing

Answer 9

• broad detection
• missense vs silent mutations
• may detect mixed allelic variants

Question 10

Disadvantages of sequencing

Answer 10

• well trained staff
• costly equipment
• bioinformatics
• centralized model