Eco-physiological characterization of fluorescence in situ hybridization probe-targeted denitrifiers in activated sludge using culture-independent methods

AIMS: This study proposes the application of a culture-independent method fluorescence in situ hybridization (FISH)] and a bioreactor operation control strategy to characterize environmental micro-organisms according to their survival strategies in a mixed suspension culture. Eco-physiological characteristics of two 16S rRNA probe-targeted denitrifiers (DEN581 and DEN124) were investigated against the availability of two resources. METHODS AND RESULTS: Four sequencing batch reactors were operated with manipulation of the sludge retention times to enforce limited and excess availability of two nutrients, namely acetate and nitrite, to the biomass. DEN581 FISH probe-targeted denitrifiers demonstrated dominance when the ratio of either acetate or nitrite to biomass was in excess, while DEN124-targeted organisms dominated when the above were limited. CONCLUSIONS: The study demonstrated that microbial populations in mixed cultures can be selected by changing the substrate availability (Rs) to biomass (X) ratio. The manipulation of the specific resource availability (Rs/X) determined which one of the studied probe-targeted denitrifiers (DEN124 or DEN581) became dominant. Rs/X provides a basis to study the physiology of micro-organisms that cannot be isolated in pure culture from activated sludge. SIGNIFICANCE AND IMPACT OF THE STUDY: The eco-physiological characterization of micro-organisms responsible for biological nutrient removal is anticipated to assist process designers and operators to optimize a specific biological process, such as denitrification.