Plasmids face a paradox. 🔹 More copies = more mutational opportunities 📈 🔹 But random segregation = more drift, fewer fixations 📉 So, which force dominates as plasmid copy number (PCN) increases: more mutations or more losses? Let’s find out 🧬

We modelled plasmid inheritance using a Cannings model and validated it with computational simulations 🧫 Each plasmid copy is like an evolutionary unit, randomly split at division, then copied. Prediction: Mutation rate increases with PCN, but with diminishing returns: a logarithmic rise.

We conducted a mutation accumulation experiment to test our predictions using a plasmid with a tunable copy number 🫧 We evolved bacterial lines for 30 days under tight bottlenecks at three conditions: 🔹low (~1) 🔹medium (~10) 🔹high (~60 copies)

Experiments confirm it: plasmid mutation rate rises logarithmically with copy number! 🚀 At higher PCN, the boost in mutation supply outweighs the segregational drift 💥

What about natural plasmids? We analyzed 1,598 plasmids (NCBI) 💻 to detect mutations arising during culture growth by comparing sequencing reads to consensus assemblies. 💥 Result: High-copy plasmids exhibited significantly higher mutation rates, just like theory (and experiments!) predicted!

In summary, plasmids are not just gene delivery platforms. They significantly boost the availability of mutations, enabling rapid evolution of traits like resistance, metabolism, and virulence, highlighting their role as evolutionary powerhouses of bacterial cells. PCN is key to evolvability 🫧🤩