# Wastewater Treatment with Bacterial Representatives of the _Thiothrix_ Morphotype
## 3 Methods for Identification of _Thiothrix_ in Wastewater Treatment Plants
### Enhanced Biological Removal of Phosphorus
An important factor determining the function of the representatives of the genus _Thiothrix_ as a PAO is the carbon source and the capacity for anaerobic respiration. Due to a rare metabolic feature of _'Ca_. Thiothrix moscovensis' and _'Ca_. Thiothrix singaporeensis', namely the presence of FAD-dependent malate: quinone oxidoreductase (MQO) instead of NAD-dependent malate dehydrogenase (MDH), these two species can effectively grow and function as PAOs in EBPR under organotrophic conditions, using acetate as a substrate (Figure 6) [56]. The same metabolic potential is inherent in all representatives of the genera _Thiothrix_, _Thiolinea_ and _'Ca_. Thiocaldithrix' since their genomes contain the _mqo_ gene instead of the _mdh_ gene [4; 5]. Presumably, these members of _Thiothricaceae_, except for species lacking nitrate reductase genes, are better able to overcome the anaerobic phase in the EBPR cycle in the presence of nitrate, allowing them to activate nitrate-mediated respiration (Figure 6). This may be facilitated by the following factors, which, under certain unfavorable conditions, can presumably create advantages for species with MQO instead of MDH or (MDH+MQO), despite the fact that, formally, these two enzymes catalyze the same reaction. 1. MQO is selectively activated 3.4-4-fold during microbial growth on acetate compared to growth on glucose, whereas the corresponding figure for MDH is 1.8 [100; 101]. 2. In the case of MQO catalysis, but not MDH catalysis, there is no dependence of the reaction progression malate \(\rightarrow\) oxaloacetate on the NAD/NADH balance, because the cofactor of MQO is the FAD/FADH pair, while NAD/NADH is involved in the MDH-catalyzed reaction. 3. Moreover, in the case of MQO catalysis, there is no dependence of the reaction on the malate/oxaloacetate balance because the reaction is exergonic (\(\Delta G^{\circ f}=-18.9\) and \(-55.0\) kJ/mol when the electron acceptor is MQ or UQ, respectively) and proceeds spontaneously. In contrast, the reaction with MDH is endergonic (\(\Delta G^{\circ f}=+28.6\) kJ/mol) and does not proceed in the forward direction under normal conditions according to the laws of chemistry. 4. It follows from (3) that the simultaneously use of MQO and MDH is disadvantageous under conditions of an energy deficit and a lack of oxygen, because they lead to a futile cycle, where oxaloacetate, reduced from malate by MQO, is converted to malate again due to the reverse reaction catalyzed by MDH. The end result of such a reaction is equivalent to the reaction of noncoupled NADH: quinone oxidoreductase type II (common in _Bacillus_ species and many other bacteria), i.e., the production of reduced quinone, but at a higher cost than in the MQO-catalyzed reaction. 5. One possible way to avoid this wastefulness and not waste an \"expensive\" reducing agent on an unprofitable reaction (see (4)) might be to either block the available MDH or replace MDH with MQO. Switching to MQO catalysis preserves oxaloacetate as an essential substrate for citrate synthase in TCA, as well as for gluconeogenesis and the synthesis of aspartate branch amino acids. 6. The outcome of switching individual microorganisms to MQO maintenance may be the more reliable functioning of TCA under some unfavorable conditions, namely anaerbiosis (respiration with less favorable oxidants compared to oxygen, e.g., nitrate), under carbon-containing substrate limitations, and others. The electron acceptor from MQO can be menaquinone, which has a more negative redox potential (\(E^{\prime}{}_{0}=-0.075\) V) compared to ubiquinone (\(E^{\prime}{}_{0}=+0.100\) V) and is almost ideally matched to the FAD/FADH pair (\(E^{\prime}{}_{0}=-0.060\) V) (cofactor in MQO) to form a unit in the electron transport chain. It is worth noting that, in contrast to strictly aerobic and strictly anaerobic bacteria, both ubiquionones and menaquinones are synthesized in membranes in facultatively anaerobic bacteria, which include representatives of _Thiothrix_. As shown in _E.