Adaptation and population dynamics of Azotobacter vinelandii during aerobic biological treatment of olive-mill wastewater

Olive-mill wastewater (OMW) has a high organic and polyphenol content and is resistant to biodegradation. Its disposal leads to a major environmental pollution problem in the Mediterranean basin. The detoxification of OMW following inoculation with Azotobacter vinelandii (strain A) was performed for two successive 5-day-period cycles in an aerobic, biowheel-type reactor, under non-sterile conditions. The phytotoxicity of the processed product was reduced by over 90% at the end of both cycles. To exclusively monitor the A. vinelandii population in the reactor a most probable number-PCR approach was employed and applied daily to serial dilutions of total DNA extracted from reactor samples. PCR sensitivity was independent of the presence of OMW or non-target DNA. The A. vinelandii population dynamics were successfully monitored, showing an initial adaptation period, followed by a sharp population maximum on the fourth day of both cycles (1.6×10 8 and 9.6×10 7 cells ml −1 respectively), after a major phytotoxicity decline. N 2 fixation rates were estimated using the acetylene reduction assay and reached a peak during the first 1–2 days of each cycle (36 and 29 nmol C 2H 2 ml −1 h −1 respectively). The data are consistent with an initial physiological adaptation phase, where the presence of phenolic compounds limits A. vinelandii growth but stimulates N 2 fixation, followed by a rapid growth phase as phytotoxicity declines.