Optimization of single plate-serial dilution spotting (SP-SDS) with sample anchoring as an assured method for bacterial and yeast cfu enumeration and single colony isolation from diverse samples

We propose a simple technique for bacterial and yeast cfu estimations from diverse samples with no prior idea of viable counts, designated as single plate-serial dilution spotting (SP-SDS) with the prime recommendation of sample anchoring (100 stocks). For pure cultures, serial dilutions were prepared from 0.1 OD (100) stock and 20 μl aliquots of six dilutions (101-106) were applied as 10-15 micro-drops in six sectors over agar-gelled medium in 9-cm plates. For liquid samples 100-105 dilutions, and for colloidal suspensions and solid samples (10% w/v), 101-106 dilutions were used. Following incubation, at least one dilution level yielded 6-60 cfu per sector comparable to the standard method involving 100 μl samples. Tested on diverse bacteria, composite samples and Saccharomyces cerevisiae, SP-SDS offered wider applicability over alternative methods like drop-plating and track-dilution for cfu estimation, single colony isolation and culture purity testing, particularly suiting low resource settings.