TITLE:
Comparative Study of Different Methods for Analyzing Denitrifying Bacteria in Fresh Water Ecosystems
AUTHORS:
Kristina Rathsack, Jörg Böllmann, Marion Martienssen
KEYWORDS:
PCR, MPN, Quantification, Denitrification
JOURNAL NAME:
Journal of Water Resource and Protection,
Vol.6 No.6,
April
29,
2014
ABSTRACT: Bacteria
capable of denitrification play a significant role in the nitrogen cycle of
freshwater ecosystems. By metabolizing nitrogen compounds they e.g. counteract
the eutrophication of natural waters. To get detailed insights into the in situ turnover rates of nitrogen a
reliable tool of quantification for active microorganisms is essential. In the
present investigation, quantification capabilities of a molecular tool
(Polymerase Chain Reaction—PCR) and a cultivation based
tool (Most probable number—MPN) were investigated and compared. The total
bacterial concentration yielded by the molecular PCR approach was up to 6-fold
higher compared to the results of the MPN approach. However, the portion of
culturable denitrifying bacteria compared to the number of specific gene copies
(nirS) was much lower. Depending on the environmental conditions, the difference
between the PCR and the MPN approach was up to three orders of magnitude. From
lab scale experiments with a pure P. aeroginosa strain it can be concludes,
that these differences are not the result of inappropriate culture conditions
but rather reflect the portion of so called viable but not culturable bacteria
(VBNC). Low nitrate concentrations as found in many fresh water ecosystems
induced a significant increase in the portion of non culturable denitrifying
bacteria. Referred to the investigation of dynamic populations, the number of
metabolic active bacteria is represented by the MPN rather than by the PCR
approach.