You are trying to identify if a specific foodborne pathogen, Clostridium perfringens, is present in food. It is important for you to get results in an hour and know where on the food the pathogen exists. Which method of DNA detection would be best?

Question

You are trying to identify if a specific foodborne pathogen, Clostridium perfringens, is present in food. It is important for you to get results in an hour and know where on the food the pathogen exists.  Which method of DNA detection would be best?

BlackTom AI · Accepted Answer

FISH (fluorescence in situ hybridization) targets ribosomal RNA/DNA in intact cells, so it gives species-specific nucleic acid detection while preserving spatial information about where organisms sit on or within a food sample.

Relevant knowledge points:
- FISH uses short, fluorescently labeled oligonucleotide probes complementary to species- or genus-specific rRNA sequences (often 16S rRNA), giving high specificity.
- Because probes hybridize inside intact cells, fluorescence microscopy shows the physical location of target cells on the food matrix.
- rRNA is abundant in live or metabolically active bacteria, increasing signal and sensitivity compared with single-copy DNA targets.
- Time-to-result for optimized FISH protocols can be on the order of an hour; culture methods take many hours to days, and PCR requires extraction that removes spatial context.

Step-by-step reasoning and typical timing:
1. Sample collection and small-area smear or thin sectioning of the food surface (5–10 min).
2. Fixation (e.g., paraformaldehyde or ethanol) to preserve cell morphology and permeabilize membranes (10–15 min).
3. Hybridization with fluorescent probe (species-specific Clostridium perfringens probe) under controlled temperature/formamide conditions (15–25 min for rapid protocols).
4. Brief washes to remove unbound probe (5–10 min).
5. Fluorescence microscopy to detect and localize labeled cells (immediate).
Total optimized time ≈ 45–60 minutes.

Why FISH is correct:
- It provides both DNA/RNA-based specificity and in situ localization (where on the food the pathogen is).
- Probes can be designed to uniquely identify C. perfringens, and rRNA abundance yields visible signal in an hour-scale workflow.

Why other methods are less suitable:
- PCR/qPCR: fast and sensitive for presence/quantification but requires nucleic acid extraction, losing spatial information about where cells are on the food surface.
- Culture: provides localization only if plating but requires many hours to days for growth.
- Immunoassays/ELISA: may detect antigen but often lack species-level nucleic acid specificity and may not reliably localize cells in situ; antibody availability and cross-reactivity can be limiting.

Conclusion: FISH is the best method for rapid (≈1 hour), species-specific DNA/RNA detection while showing the exact location of Clostridium perfringens on the food sample, confirming the provided answer: FISH.

You are trying to identify if a specific foodborne pathogen, Clostridium perfringens, is present in food. It is important for you to get results in an hour and know where on the food the pathogen exists. Which method of DNA detection would be best?单项选择题

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