Identity and physiology of glycogen accumulating organisms in activated sludge
Date
2018
Authors
Onetto Carvallo, Cristobal Andres
Editors
Advisors
Grbin, Paul Ramon
Eales, Kathryn
Eales, Kathryn
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Abstract
Glycogen accumulating organisms (GAOs) form part of the core microbial
component of the activated sludge community of both industrial and domestic
wastewater treatment plants. A plethora of research has focused on the activity of
GAOs in enhanced biological phosphorus removal (EBPR) domestic wastewater
treatment plants due to their competition for carbon with the polyphosphate
accumulating organisms (PAOs) at the expense of EBPR efficiency. However, in
industrial systems such as winery wastewater (WWW), where GAOs are known to
proliferate and cause settling problems, limited research has focused on their
identification and ecophysiology.
Initial metagenomics and fluorescence in situ hybridization (FISH) surveys of WWW
treatment plants revealed that Defluviicoccus-related GAOs belonging to cluster II
(DF2) and III (DF3) appeared as highly dominant with abundances reaching up to
52% of the bacterial population. GAOs belonging to the Competibacteraceae
(CPB_S18 and CPB_S60) were also observed in notable numbers along with putative
GAOs belonging to the Actinobacteria (Micropruina spp. and Nakamurella spp.) and
Gammaproteobacteria CCM19a.
Previous investigations have suggested that high carbon loads along with low nitrogen
levels are a combination of factors that enhance the proliferation of GAOs. In order to
elucidate if the low nitrogen levels of WWW are the underlying cause of DF2
proliferation, incubations under different COD:N ratios using ¹³C - acetate and ¹⁵N -
urea were performed. Cell substrate assimilation was quantified using an innovative
technique, FISH-NanoSIMS, revealing that low (100:1) or null nitrogen
concentrations enhanced DF2 carbon uptake while ratios of 60:1 and 20:1 reduced
carbon uptake. Nitrogen dosing at COD:N ratios of 60:1 or higher was demonstrated
as a feasible strategy for controlling the excessive DF2 growth in WWW treatment
plants.
The unique filamentous morphology of DF3 is important as its proliferation leads to
severe bulking issues. Although previously reported as abundant in industrial and
domestic activated sludge, limited research has focused on understanding the
physiology of these organisms. The first genome of a filamentous DF3 was extracted
from a WWW activated sludge metagenome. Annotation revealed interesting
metabolic features that help to understand the competitiveness and abundance of this
microorganism in WWW activated sludge. The genetic potential to cycle trehalose
through glycogen, nitrogen fixation, hydrogenase activity and urea uptake appear as
adaptive strategies of DF3 to the WWW nitrogen limited environment.
In a recent 16S rRNA survey of EBPR plants, Micropruina spp. were identified as the
most abundant GAO, yet little is known about their ecophysiology. To further
elucidate the ecophysiology of this putative GAO observed in WWW and EBPR
treatment plants, genomic and metabolomic studies were made in pure culture
Micropruina glycogenica str. Lg2T [T superscript] and compared to the in situ physiology of the genus using state-of-the-art single cell techniques. Micropruina spp. were observed to
take up carbon substrates under anaerobic conditions, which were partly fermented to
lactate, acetate, propionate and ethanol, and partly stored as glycogen for aerobic use.
This physiology is markedly different from the classical GAO model, suggesting a
need to reconsider current understanding of the GAO phenotype.
Metagenomics analyses revealed a codominance of filamentous Thiothrix spp. and
GAOs in a particular WWW treatment plant with bulking issues. In an attempt to
understand this problem, a further experiment based on raw wastewater feeding was
performed. Raw feeding reduced the Thiothrix spp. population and improved settling,
therefore, direct feeding is proposed as a control method for industrial plants with
surge/anaerobic lagoons in order to manage the bulking problems caused
by Thiothrix spp..
School/Discipline
School of Agriculture, Food and Wine
Dissertation Note
Thesis (Ph.D.) (Research by Publication) -- University of Adelaide, School of Agriculture, Food and Wine, 2018
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