Flow cytometric characterization of bacterial abundance and physiological status in a nitrifying-denitrifying activated sludge system treating landfill leachate

Abstract

Flow cytometry has recently been presented as a research tool in the assessment of the viability/activity of activated sludge from municipal wastewater treatment plants, but it has not put in practice for industrial biotreatments yet. In this study, for the first time ever, the reliability and significance of the multiparameter flow cytometry applied to the biological nitrification-denitrification treatment of leachate have been evaluated. Using a double staining procedure (cFDA/PI), the viable, damaged, and dead subpopulations were determined, and the results were compared to those obtained with conventional methods, such as nitrogen and oxygen uptake rates or plate counting. Flow cytometry showed that viable cells represented approximately 47% of the total population, whereas active cells accounted for 90%. For both sludge from nitrification and denitrification processes, with less than 1% of them being also culturable in plate. Either flow cytometry or uptake rates revealed that health status of sludge remained constant throughout the biotreatment, which is consistent with the high recirculation rates. Under anaerobic starvation conditions, physiological status of sludge remained constant as well as specific oxygen and denitrification rates. Nevertheless, both the culturability in plate and the nitrification rate significantly decreased. These findings proved that multiparameter flow cytometry is a useful tool for the assessment of the viability and activity of sludge from a nitrification-denitrification biotreatment process. These results gathered all the bacterial communities in the sludge, so the decay in minority populations, such as nitrifying bacteria, requires the use of a complementary technique to evaluate specific activities.

Prophage-Related Gene VpaChn25_0724 Contributes to Cell Membrane Integrity and Growth of Vibrio parahaemolyticus CHN25

“…Cell membrane damage was analyzed using the method described by Collado et al (Collado et al, 2017). The bacterial cell suspension was double dyed using propidium iodide (PI) (final concentration 10 mM) (Sangon, China) and 5(6)carboxydiacetate fluorescein succinimidyl ester (CFDA) (final concentration 10 mM) (Beijing Solarbio Science & Technology Co. Ltd., China).…”

Recombinant Rhodococcus erythropolis expressing HAO and AMO genes promotes nitrogen and organic matter removal efficiency in the treatment of landfill leachate

“…Nitrifying bacteria can increase the activity of AS and promote nitrogen metabolism by nitrification–denitrification biotreatment process in the wastewater (Collado et al, 2017). To identify the role of nitrifying bacteria, RH was isolated from aerobic reactor in the landfill leachate treatment system.…”

Multiplatform Physiologic and Metabolic Phenotyping Reveals Microbial Toxicity

“…Once the membrane is depolarized and loses membrane potential, DiBAC enters and binds to lipid-containing components. Another physiologic feature of bacteria is metabolic activity, which can be measured by fluorogenic esterase substrates, like carboxyfluorescein diacetate (CFDA) ( 12 , 59 , 60 ) and carboxyfluorescein diacetate succinimidyl ester (CFSE) ( 61 ). Fluorogenic esterase substrate is converted by intracellular esterase into fluorescein analogs, which are retained by cells.…”