S06 - Session O8 - In situ microbial community analysis in container soilless substrates.

Authors: Silvia Valles *, James Altland, Frederick Michel

Microbial communities in soilless substrates are dynamic and can vary based on age, substrate components and environmental (temperature, humidity, etc.) and management (fertilizer, irrigation, etc.) factors. Analysis of these communities usually requires destructive sampling. The objective of this research was to investigate whether microbial communities in soilless substrates could be analyzed in situ . To test this, a mix consisting of manure compost, peatmoss, and perlite (20:65:15 v/v) was filled in 15-cm pots (n=4) and sown with sunflower ( Helianthus annus L. ' Choco Sun'). Samples were collected 0, 3 and 6 weeks after planting using two different methods. One was a non-destructive, pour-through method whereby each pot was watered until saturated, then after 40 minutes, 250 mL of tap water was poured through the container and the leachate was collected. The leachate was transferred into four 50-mL falcon tubes and centrifuged for 30 min at 5000 rpm. The resulting pellet was resuspended and used for DNA collection. For the second method, the entire root ball was removed from the container, the sunflower shoots and roots detached, and the remaining soilless substrate mixed thoroughly and homogenously, and a representative 10 g sample used for DNA collection. For both, DNA was extracted and purified using a DNeasy PowerSoil Kit. A 25 µL aliquot of the DNA solution at 5 mg·mL -1 was PCR amplified using universal bacterial primers, 515F and 806R. The amplified sequences were identified by comparison to those in the Greengenes 16S gene database using R 4.0. software. Results showed no significant differences between bacterial communities using the two methods. Both exhibited similar relative abundances and types of ribotypes. Significant differences were found over the three sampling dates. These results demonstrate that a simple non-destructive method can be used to collect nucleic acids for in situ characterization of microbial communities in container media.