Sulfur oxidizing bacteria have been widely exploited to remove thiosulfate with adverse effects on human health and environment. Since formation of elemental sulfur among other thiosulfate oxidation products is advantageous in industrial applications, we tend to statistically screen (fractional factor design) and optimize (response surface methodology) the operating conditions in this study so that the final thiosulfate removal (Y1) and the highest sulfur production (Y2) become maximum along with the appropriate final sulfate formation (Y3). Optimum conditions were then examined in a stirred bioreactor. Screening results showed the definite effectiveness of six factors on all responses however level modification and optimization were necessary due to significant P-value of curvature. Single response optimizations (maximization of each response) resulted in dissimilar sets of optimal conditions. Multiple response (Y-1 = 100 %, Y-2 and Y-3 = 50 %) optimization, using desirability function, was therefore used and resulted in agitation speed: 44 rpm, pH: 6.07, temperature: 24.7 degrees C, time: 2.64 day, inoculum ratio: 19.5 % and thiosulfate level: 2.87 mg l(-1) as optimal conditions. Bioreactor experiments (agitation speeds: 30-90 rpm) under optimal conditions led to the highest S-0 formation of 46 % and complete thiosulfate removal at 60 rpm and 40 h. Therefore, multiple response optimization followed by minor scale modifications can productively enhance industrial elemental sulfur formation from wastewater plants. (C) 2019 Institution of Chemical Engineers. Published by Elsevier B.V. All rights reserved.