Abstract
Nitritation, the first stage of ammonia removal process is known to be
limiting for total process performance. Ammonia oxidizing bacteria (AOB)
which perform this process are obligatory activated sludge habitants, a
mixture consisting of Bacteria, Protozoa and Metazoa used for biological
wastewater treatment. Due to this fact they are an interesting bacterial
group, from both the technological and ecological point of view. AOB
changeability and biodiversity analyses both in wastewater treatment
plants and lab-scale reactors are performed on the basis of 16S rRNA gene
sequences using PCR-DGGE (Polymerase Chain Reaction – Denaturing Gradient
Gel Electrophoresis) as a molecular biology tool. AOB researches are
usually led with nested PCR. Because the application of nested PCR is
laborious and time consuming, we have attempted to check the possibility
of using only first PCR round to obtain DGGE fingerprinting of microbial
communities. In this work we are comparing the nested and non-nested
PCR-DGGE monitoring of an AOB community and presenting advantages and
disadvantages of both methods used. The experiment revealed that PCR
technique is a very sensitive tool for the amplification of even a minute
amount of DNA sample. But in the case of nested-PCR, the sensitivity is
higher and the template amount could be even smaller. The nested PCR-DGGE
seems to be a better tool for AOB community monitoring and complexity
research in activated sludge, despite shorter fragments of DNA
amplification which seems to be a disadvantage in the case of bacteria
identification. It is recommended that the sort of analysis approach
should be chosen according to the aim of the study: nested-PCR-DGGE for
community complexity analysis, while PCR-DGGE for identification of the
dominant bacteria.
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Authors and Affiliations
Aleksandra Ziembińska-Buczyńska