**Summary: Transgenerational Inheritance of Learned Pathogen Avoidance in C. elegans**
The capacity to recognize and avoid harmful pathogens is crucial for animal survival. Some animals can detect specific molecules produced by pathogens, triggering immediate defensive responses. Others, however, learn to avoid pathogens through prior exposure—a process known as “learned avoidance.” A striking example of this phenomenon has been observed in the nematode worm, *Caenorhabditis elegans*, which can learn to steer clear of the pathogenic bacterium *Pseudomonas aeruginosa*, specifically the PA14 strain.
**Discovery of Transgenerational Avoidance**
In 2019, research led by Coleen Murphy at Princeton University found that *C. elegans*’s learned avoidance of PA14 could be passed down for up to four generations, even if the subsequent generations had no direct contact with the pathogen. This remarkable instance of transgenerational epigenetic inheritance means that the offspring benefit from the experiences of their ancestors, potentially improving their survival chances in environments where PA14 is present.
**Controversy Over Replicability**
However, the concept of transgenerational inheritance of learned avoidance has not been without contention. More recently, a team led by Craig Hunter at Harvard University revisited the phenomenon and reported different results. They confirmed that both the parent generation (P0) and the immediate offspring (F1) learned to avoid PA14, but they did not observe this avoidance behavior in the F2 generation. This raised questions about the robustness and reproducibility of the original findings.
The Murphy group responded, suggesting that modifications in experimental protocols by the Hunter group could explain the discrepancy. Now, a new study by Andres Vidal-Gadea and colleagues at Illinois State University has provided further support for the Murphy group’s results. By closely following the original protocol, they confirmed that learned avoidance of PA14 can indeed be transmitted to the F2 generation.
**Experimental Methods: The Avoidance Assay**
The studies employed a standard behavioral assay to assess avoidance. *C. elegans* were first grown on OP50, a benign strain of *E. coli* that serves as their standard laboratory food. For the avoidance test, worms were placed at the center of an agar plate with two spots at opposite ends: one containing PA14 and the other OP50. Sodium azide, a chemical that immobilizes worms, was added to both spots. After allowing the worms to roam, researchers counted how many worms were near each spot after an hour.
To induce learned avoidance, worms were exposed to PA14 for 24 hours—a process referred to as “training.” Following this exposure, a subset of worms was tested in the avoidance assay, and their eggs were collected for the next generation. Importantly, after the P0 generation, none of the subsequent generations encountered PA14 before testing, allowing researchers to measure inherited avoidance behavior. Avoidance in the F1 and F2 generations was compared