Bacterial Quantification in Different Plating Methodologies

Determining the number of bacterial cells in a product is a fundamental procedure in the inoculant industry, as it ensures its quality. The current legislation in Brazil recommends the spreading technique as the standard procedure to count viable cells per gram or milliliters (mL). However, in order to carry out these analyses, there is a considerable material demand. Hence, the objective of this study was to test an alternative methodology, the microdrop technique, and determine whether it mitigates costs and material demands, with the added benefit of being faster and more accurate for operators. To carry out the experiment, 8 treatments were made in both saline solution (SS) and saline solution (0.9%) + Tween 80 (0.1%), at different concentrations, using a homemade inoculant as inoculum. After plating, bacterial growth was conducted in a growth chamber for 4–6 days. A parameter of 30–300 colonies per plate was used to count colony-forming units (CFU). In addition, contaminant tests were performed for Tryptic Soy Agar (TSA) bacteria and saprophytic fungi (Sabouraud). Dilution samples (105) and inoculum from commercial packages were evaluated by Propidium monoazide quantitative polymerase chain reaction (PMA-qPCR). Despite the microdroplet presenting higher CFU values when compared to the spreading technique, the dilutions did not alter its order of magnitude (107 CFU.mL-1). The use of the microdrop method, when combined with Tween 80 (0.1%), increased the unitary CFU value, providing a cost-effective method for counting viable Azospirillum brasilense cell numbers compared to standard culture media.