In situ microbial community analysis of container soilless substrates

Microbial communities are important components of soilless substrates, especially those amended with composts, that benefit plant health through nutrient mobilization and disease suppression. These communities are affected by plant age, substrate composition, environmental factors (temperature, humidity, etc.), and substrate management (fertilizer, irrigation, etc.). Their characterization usually requires destructive sampling. The hypothesis of this research is that microbial communities in soilless substrates can be characterized in situ using leachates collected from the containers. To test this, a container mix consisting of manure compost, peat moss, and perlite (20:65:15 v/v) was added to 1.33 L containers (n=4) and sown with sunflower (Helianthus annus L. ‘Choco Sun’). Samples were collected using two different methods at 0, 3, and 6 weeks after planting. The first, leachate (LH) was a non-destructive method whereby each pot was watered to saturation. After 40 min, 250 mL of additional water was added, and the resulting leachate was collected. The leachate was centrifuged for 30 min at 5000 rpm. The resulting pellets were resuspended and used for DNA extraction. The second method, composite (CP) involved sampling the growth media by first removing the entire root ball, detaching the sunflower shoots and roots; and sampling the remaining soilless substrate after thorough mixing. A representative 10 g sample of the media was used for DNA extraction. DNA was extracted and purified using a DNeasy PowerSoil Kit for both methods. PCR was using universal bacterial 16s rRNA primers 515F and 806R. From the amplified sequences, exact amplicon sequence variants (ASVs) were inferred, and taxonomy was assigned using the SILVA database with a dada2-based pipeline in R 4.0. Results showed minimal differences between bacterial communities identified using the two methods by phyla and genera abundance, PCA analysis of amplified sequences, and Shannon diversity indices. Both exhibited similar relative abundances and ribotype distributions. Significant differences, however, were found in communities over the three sampling dates. These results demonstrate that a simple non-destructive method can be used to collect nucleic acids for the in situ characterization of bacterial communities in container media.