Extended Data Fig. 1: PA14 sRNAs induce maternal PA14 avoidance and increased daf-7 expression in the ASI neurons.

a, Worms exposed to a PA14 bacterial lawn for 24 h learn to avoid PA14 in a choice assay, but PA14 DNA exposure alone does not induce maternal avoidance of PA14. b, Training with large RNA (>200 nt) isolated from bacterial lawns of PA14 is not sufficient for maternal PA14 avoidance. c, Bioanalyzer results of isolated PA14 total RNA and fractionated small and large RNAs. RNA levels were normalized for worm training. d, ΔlasR sRNA exposure does not induce PA14 learned avoidance. e, Development of progeny of PA14 sRNA-trained mothers was not delayed compared to progeny of OP50-trained mothers. n = 6 plates per condition with 23–142 progeny per plate, mean ± s.e.m. f, pmk-1(km25) worms learn to avoid PA14 when exposed to PA14 bacteria lawns. g, pmk-1(km25) is not required for PA14 sRNA-induced pathogenic learning. h, irg-1p::gfp expression is induced by PA14 bacterial lawn exposure, but not by PA14 sRNAs alone. i, GFP intensity from h was quantified, n = 33, 24, 43 and 35 worms, left to right. j, Model of PA14 bacteria lawn and sRNA learning. k, Location of ASI and ASJ neurons. l, The 24-h exposure of worms to a PA14 lawn or PA14 sRNA increases daf-7p::gfp expression in the ASI neurons. Two-way ANOVA, Tukey’s multiple comparison test. n = 35, 41, 41 and 49 neurons, left to right. m, ΔlasR sRNA exposure does not induce changes in daf-7p::gfp ASI expression. One-way ANOVA, Tukey’s multiple comparison test. n = 38, 27 and 29 neurons, left to right. Biological replicates: 2 (a), 3(b–d, f–i, m), 6 (l). For choice assays, each dot represents an individual choice assay plate (average of 115 worms per plate) with data shown from all replicates. For box plots, centre line is the median, box extends from the 25th to the 75th percentile; whiskers denote minimum–maximum values. One-way (d, m) and Two-way ANOVA (a, b, e–g, i, l), Tukey’s multiple comparison test. *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P < 0.0001, n.s., not significant. Mean differences are shown using Cumming estimation plots⁴⁷, with each graphed as a bootstrap sampling distribution. Mean differences are depicted as dots; 95% confidence intervals are indicated by the ends of the vertical bars. See Supplementary Table 4 for exact sample sizes (n) and P values.