The monitoring tasks will allow to compare the purification yields of the different planted species
The PCR technique will serve to identify the denitrifying activity of the microbial communities that grow in the cells of the wetland
The new horizontal flow wetland built in Flores de Ávila as a pilot for the Aquamundam project is operational and its monitoring has begun. The objective of this pilot is to compare whether the passage of water through five cells with the same common species of reed (Phragmites australis) or through five cells with five different species has the same purification effect.
In addition to taking samples to measure water parameters at the inlet and outlet of the water flow, BOD, COD, nitrogen charge and others were carried out by the CHD. CARTIF has begun to carry out the treatment of samples from the different cells with the aim of studying the microbial diversity they present. These have been taken by the ICTHIOS team that, together with the CHD technical team, are in charge of doing the field work in the wetland.
One of the ways to remove pollutants in these wetlands, such as excess nitrogen from wastewater, is through the action of microorganisms that grow in the cells. The objective is to study the microbial communities that proliferate in cells populated by different plant species and to compare their effect on water purification. For this, in CARTIF the PCR technique will be used to identify the different species that present denitrifying activity, among others.
After receiving the material from the different wetlands (mixture of water and soil), the solid part was separated from the liquid by centrifugation and the solid part was frozen to subsequently extract the total genomic DNA (gDNA) from the microbiota present in the soil.
The extraction of the gDNA has been carried out following a protocol for the extraction of DNA from the soil and, after completing the process, its concentration and integrity have been verified by means of agarose gel electrophoresis. These techniques are based on breaking the bacterial cell wall and extracting DNA from them. Next, it is checked whether it has been extracted by staining this DNA and observing it with UV light on an agarose gel. This gel is subjected to an electric current and taking advantage of the fact that the DNA is negatively charged –which causes it to move through the gel towards the positive pole– a band of DNA is observed.
During the next few weeks, the identification analyses of the metabolic activities of the microbial communities will be carried out using quantitative PCR (qPCR). This technique is based on identifying genes that are characteristic of the bacteria that carry out these activities and the result of the qPCR allows us to compare whether these genes exist and how much they are expressed, that is, whether the bacteria present in the sample actually carry out these decontamination activities. of wastewater.
Among the main metabolic activities, those related to nitrogen and sulfur metabolism will be studied. The expression of various genes involved in these pathways, which are commonly used as detection markers for nitrifying and denitrifying bacteria present in wetlands, will be measured. The results obtained will contribute to the comparison of the purification efficiency of the two parallel lines (control and experimental) of the wetland.