Environmental Monitoring & Assessment Flashcards

When testing environmental water samples by qPCR, humic substances most commonly cause which problem?

Answer: PCR inhibition causing delayed Ct values or false negatives

Humic and fulvic acids inhibit polymerase activity and quench fluorescence, so Ct values come later or the target is missed.

Which control best detects PCR inhibition in an individual environmental sample extract?

Answer: An internal amplification control (IAC) spiked into each reaction

An IAC amplified in the sample's own reaction shows whether that matrix suppresses amplification.

A soil extract gives Ct 34 undiluted and Ct 31 when diluted 1:10. What does this indicate?

Answer: Inhibitors are present and dilution relieved them

A lower Ct after dilution is a classic sign of inhibition being diluted away; without inhibition a 1:10 dilution would raise Ct by about 3.3.

In environmental DNA (eDNA) surveys, a field blank (filtered sterile water carried to the site) is used to detect:

Answer: Contamination introduced during field sampling and handling

Field blanks go through the same handling as real samples to show contamination added in the field.

Why are eDNA water samples typically filtered and frozen or preserved soon after collection?

Answer: To limit microbial and enzymatic degradation of extracellular DNA

Extracellular DNA breaks down quickly in water, so fast filtration and preservation keep it detectable.

Which sample type is generally most challenging for PCR due to co-extracted inhibitors?

Answer: Organic-rich soil and sediment

Soils and sediments contain humic acids, polysaccharides, and metal ions that co-purify with DNA.

Wastewater surveillance by RT-qPCR often normalizes target concentration using a fecal marker such as PMMoV. The purpose is to:

Answer: Account for variation in fecal content and flow between samples

Fecal-strength markers adjust for dilution and population input, so samples can be compared.