SELECTION OF INDIGENOUS DENITRIFYING MICROORGANISMS FOR THE CONSTRUCTION OF A SYNTHETIC CONSORTIUM FOR BIOAUGMENTATION OF BIOLOGICAL DENITRIFICATION REACTORS TREATING GROUNDWATER FOR HUMAN CONSUMPTION.

MASSICOT, AGUSTINA; CRISTIÁN DOTTO; MACCHIAROLI, NATALIA; KAMENETZKY, LAURA; FIGUEROLA, EVA

Chapadmalal

Congreso; XVIII Congreso de la Sociedad Argentina de Microbiología General; 2023

SAMIGE

Elevated nitrate levels in groundwater are a global concern attributed to human activities, such as agriculture, and poor municipal and industrial waste management. The presence of nitrate in drinking water is harmful to human health causing methemoglobinemia and potentially nitrosamine formation in the digestive tract. Biological denitrification (BD) is a well-established alternative for water treatment utilized in several countries. This process allows the transformation of nitrate into gaseous nitrogen with low operating costs and high water recovery. BD is mainly carried out in fixed-bed reactors with an inert material to support the denitrifying biofilm, composed of a self-assembled consortium of indigenous microorganisms, naturally selected by the process conditions. We determined, In a previous stage, the optimal conditions for the establishment of denitrifying bacterial mixed communities at the laboratory-scale, and subsequently set up a 10X scaling test and a pilot plant in Llavallol, Buenos Aires, at a groundwater extraction well affected by high levels of nitrate. At all scale levels, an acclimatization period of approximately 100 hydraulic retention times (HRT) was observed. We hypothesize that bioaugmentation with a synthetic consortium of indigenous microorganisms will shorten the time to reach the steady state. To advance in this direction, the first step was to obtain, characterize and identify isolates of autochthonous bacteria. Isolated colonies were obtained from the grains of sand of the laboratory-scale bioreactors by a streaking method on agar plates with groundwater plus nutrients as a culture medium. These cultures were analyzed to study their capacity for gas generation and nitrate accumulation. MALDI-TOF spectroscopy was carried out to discard redundant isolates. Denaturalization gradient polyacrylamide gel electrophoresis (DGGE) was performed on the 16S rRNA gene amplicons from non-redundant isolates and the metagenome of the bioreactors, to detect dominant strains and discard redundant ones that skip the MALDI-TOF analysis as well. Third Generation Sequencing and genome analysis were applied to selected isolates identifying target genes of the denitrifying pathway. From the 66 strains isolated, 32 were classified to gender level and 7 to species level. We discover a set of strains with complementary denitrification genes that could be promising for the assembly of the denitrifying consortia.