FG Biotechnologie der Wasseraufbereitung
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- Photolysis (2)
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- FG Biotechnologie der Wasseraufbereitung (149)
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Durch anthropogene Aktivität hat sich der Nährstoffeintrag in die Umwelt stark erhöht. Die Hauptquellen sind die Landwirtschaft, kommunales und industrielles Abwasser. Erhöhte Stickstofffrachten haben einen starken Einfluss auf den Stickstoffkreislauf. Das Überangebot an Stickstoff führt zu Eutrophierung in Gewässern und somit zu einem weniger diversen, instabileren Ökosystem. Die Stickstoffeliminierung aus Abwässern ist essentiell für den Schutz der Ökosysteme. Das etablierte System der Nitrifikation/Denitrifikation ist aufgrund des hohen Sauerstoffbedarfs sehr energie- und somit auch kostenintensiv. Der Anammox-Prozess (anaerobe Ammonium Oxidation) ist eine potentielle Alternative zur konventionellen Stickstoffelimination. Anammox-Bakterien nutzen Ammonium und Nitrit als Elektronenakzeptor. Durch Nutzung der partiellen Nitrifikation in Kombination mit Anammox (PN/A) kann bis zu 60 % der Belüftungsenergie eingespart werden. Trotz Erfolgen in der Teilstrombehandlung ist der Prozess noch nicht erfolgreich in der Hauptstrombehandlung eingesetzt worden. Die größten Herausforderungen sind die niedrigen Temperaturen, geringe Zulaufkonzentrationen und hohe Konzentrationen organischen Kohlenstoffs. Die vorliegende Arbeit konnte die Wirksamkeit von membranbelüfteten Biofilm Reaktoren unter Verwendung von synthetischem Abwasser bei niedrigen Temperaturen (18 - 8 °C) belegen. Die Innovation in diesem System ist die räumliche Trennung von nitrifizierenden Bakterien und Anammox Bakterien. Die regelmäßige mechanische Entfernung des Nitrifikanten-Biofilms ermöglichte einen langzeitstabilen Reaktorbetrieb. Diese Reaktorkonfiguration konnte auch erfolgreich für hochsaline Abwässer (5 -30 g NaCl*L-1) bei einer Reaktortemperatur von 30 °C und auch unter Zugabe von TOC (40-100 mg TOC*L-1) genutzt werden. Weiterhin wurden Untersuchungen zur Aktivität von salzadaptierter Anammox-Biomasse und nicht adaptierter Biomasse in einem Bereich von 0 bis 50 g NaCl*L-1 durchgeführt. Bei Konzentrationen unter 30 g NaCl*L-1 konnten keine signifikanten Unterschiede im Abbauverhalten festgestellt werden. Allerdings war adaptierte Biomasse bei höheren Salzkonzentrationen konkurrenzstärker. Außerdem wurden Langzeitlagerungsversuche durchgeführt. Die Lagerungsbedingungen variierten in der Lagerungstemperatur, im Redoxpuffer und der Nährstoffversorgung. Nach 106 Tagen bei 4 °C konnte die Anammox-Biomasse noch 91 % ihrer ursprünglichen Aktivität erhalten. Somit konnte eine einfache und kostengünstige Lagerungsmethode für Anammo-Bakterien entwickelt werden.
Calculation of hypolimnic denitrification in a dimictic freshwater lake during summer stratification
(2019)
Phosphonates comprise a very large group of organophosphonates including aminophosphonates. Their main chemical feature leads to complexation of earth alkaline metals and transition metals (under stoichiometric). Therefore they are very often applied as complexing agents of detergents, as compounds of industrial cleaning products, or as antiscalants in cooling water systems and desalination processes. The high consumption of phosphonates within the past two decades leads to an increased discharge into the aquatic environment, of which the environmental risk is still uncertain. To date, there are several analytical methods published to determine phosphonates from environmental samples. However, no standard method has been defined until now. Different treatment technologies to break down phosphonates have been studied extensively such as photochemical degradation, chemical degradation and other advanced oxidation processes. The results obtained from those studies gained our chemical understanding with respect to possible implementation of such pre-treatment approaches in wastewater treatment, cooling water treatment and desalination process. The biological degradation of phosphonates has been studied for more than forty years. It was demonstrated that microorganisms are capable to breakdown the C-P bond with at least three different degradation pathways.
Rapid urbanisation in Ghana has resulted in individuals expanding the cities for abodes without considerations of the negative externalities these may have on the environment. One of the major challenges with rapid urbanisation is the formation of urban slums associated with lack of basic sanitation facilities. This has led to recurrent outbreak of cholera and typhoid fever. The use of a single-stage solar-supported hyper-thermophilic anaerobic biogas digester for the treatment of black water has not been investigated, hence this study. The performance of three seeding sludge under three different hyper-thermophilic temperatures (60°C, 65°C and 70°C) were tested in batch tests. The three seeding sludge were sewage sludge, sludge from maize silage and cow manure. The results from the batch tests showed cow manure at 65°C as the preferred seeding sludge and optimal hyper-thermophilic temperature.
A 50 L single-stage laboratory-scale hyper-thermophilic continuous stirred tank reactor (HT-CSTR) was operated to treat only black water for 10 weeks using cow manure at 65°C as the seeding sludge and optimal hyper-thermophilic temperature. Afterwards, co-digestion of blended kitchen food waste and black water was also practised for 12 weeks. With a mean hydraulic retention time (HRT) of 23.3 days, a mean total COD removal of 86.3 % was achieved. The reactor had an average COD volumetric loading rate of 6.22 kgCOD/(m3.d) and remained uninhibited. It also had organic loading rate of 0.3 kgVS/(m3.d) and a degradation performance (R) of 5.43 kgCOD/(m3.d). Treatment of only black water produced biogas with less methane content of 34.9 % even though a stable pH of 6.9 was recorded both in the reactor and in the effluent. Co-digestion with kitchen food waste increased the percentage content of methane in the biogas by 77 % from 34.9 % to 61.8 %.
The effectiveness of the HT-CSTR to hygienise the effluent for agricultural purpose was assessed by spiking the reactor with 200 ml each of 2 x 109 CFU/ml Salmonella senftenbergensis and 8 x 108 CFU/ml Escherichia coli. The HT-CSTR was able to hygienise all bacteria of Salmonella senftenbergensis and E. coli. A simulation test confirmed that between 30 minutes and 1 hour, all the cells of Salmonella senftenbergensis and E. coli in the treatment system were killed at 65 °C. Eubacteria, Methanosarcina spp., Methanomicrobium spp. and Methanococcus spp. were identified in the seeding sludge at the hyper-thermophilic temperature of 65°C. The design, construction and performance of a pilot-scale reactor in Terterkessim slum in Elmina, Ghana was based on results from the laboratory-scale HT-CSTR. It achieved 97 % removal of influent total COD and could produce about 2.52 Nm³CH₄/(kgCOD.d) which could be burned for at least 8 hours. The effluent cannot be used for cultivation of leafy vegetables such as cabbage since it had some concentrations of pathogens like Salmonella spp. and E. coli but can be used for cotton crop.
An enrichment culture of Candidatus Brocadia fulgida was identified by three independent methods: analysis of autofluorescence using different microscope filter blocks and a fluorescence spectrometer, fluorescence in situ hybridization (FISH) with anammox-specific probes and partial sequencing of the 16S rDNA, hydrazine synthase hzsA and hydrazine oxidoreductase hzo. The filter block BV-2A (400–440, 470 LP, Nikon) was suitable for preliminary detection of Ca. B. fulgida. An excitation-emission matrix revealed three pairs of excitation-emission maxima: 288–330 nm, 288–478 nm and 417–478 nm. Several autofluorescent cell clusters could not be stained with DAPI or by FISH, suggesting empty but intact cells (ghost cells) or inhibited permeability. Successful staining of autofluorescent cells with the FISH probes Ban162 and Bfu613, even at higher formamide concentrations, suggested insufficient specificity of Ban162. Under certain conditions, Ca. B. fulgida lost its autofluorescence, which reduced the reliability of autofluorescence for identification and detection. Non-fluorescent Ca. Brocadia cells could not be stained with Ban162, but with Bfu613 at higher formamide concentrations, suggesting a dependency between both parameters. The phylogenetic analysis showed only good taxonomical clustering of the 16S rDNA and hzsA. In conclusion, careful consideration of autofluorescent characteristics is recommended when analysing and presenting FISH observations of Ca. B. fulgida to avoid misinterpretations and misidentifications.
Energieeffizientes Nährstoffsplitting von flüssigen Wirtschaftsdüngern mittels Membranfiltration
(2017)
The Anammox process is an efficient low energy alternative for the elimination of nitrogen from wastewater. The process is already in use for side stream applications. However, some industrial wastewaters, e.g. from textile industry are highly saline. This may be a limit for the application of the Anammox process. The aim of this study was to evaluate the effects of different NaCl concentrations on the efficiency of adapted and non-adapted Anammox biofilms. The tested NaCl concentrations ranged from 0 to 50 g NaCl*L⁻¹. Concentrations below 30 g NaCl*L⁻¹did not significantly result in different nitrogen removal rates between adapted and non-adapted bacteria. However, adapted bacteria were significantly more resilient to salt at higher concentrations (40 and 50g NaCl*L⁻¹). The IC50 for adapted and non-adapted Anammox bacteria were 19.99 and 20.30 g NaCl*L⁻¹, respectively. Whereas adapted biomass depletes the nitrogen in ratios of NO-₂/NH+₄ around 1.20 indicating a mainly Anammox-driven consumption of the nitrogen, the ratio increases to 2.21 at 40 g NaCl*L⁻¹ for non-adapted biomass. This indicates an increase of other processes like denitrification. At lower NaCL concentrations up to 10 g NaCl*L⁻¹, a stimulating effect of NaCl to the Anammox process has been observed.
DTPMP is predominantly utilized as scale inhibitor. We investigated the reaction rates and degradation mechanism of DTPMP with and without addition of Fe²⁺, Mg²⁺ and Ca²⁺ by performing LC/MS and ³¹P-NMR analyses. DTPMP undergoes conversion with and without addition of bivalent metal ions. The initial cleavage of DTPMP is initiated at the C-N bond leading to release of IDMP as its major breakdown product. The release of smaller quantities of EABMP and AMPA confirmed the nucleophilic attack on the DTPMP amines. Oxidation of Fe²⁺ to Fe³⁺ during the initial 30 min indicated an intramolecular electron transfer changing the electron density distribution at the nitrogen centre, which increased the radical attack during UV irradiation. Independent of the fact that Fe acted as catalyst and Mg²⁺ and Ca²⁺ acted as reactants, we found no significant differences in their degradation mechanisms. However, the reaction rates were strongly affected by the addition of the bivalent metal ions as Fe²⁺ accelerated most DTPMP degradation followed by Mg²⁺ and Ca²⁺. The UV treatment without metal ion addition was four times slower compared with Fe²⁺ addition. We conclude that in environments rich in ferrous iron and/or at reduced redox potential, photolysis of DTPMP will be catalysed by iron and will lead to accumulation of IDMP, EABMP and AMPA and several other none-quantifiable breakdown products.
Microbial dynamics and enzymatic activities of activated sludge processes are not completely understood yet. A better understanding about the biology is indispensable for further process
optimization. Since proteins play a key role as catalysts in sludge processes, a protocol for protein extraction and analysis by 2-D PAGE was established. It is based on phenol extraction of alkaline extracts and on a subsequent precipitation with ammonium sulphate. 2-D protein patterns obtained from different sludges collected from membrane bioreactors showed – besides common spots – significant differences. Selected proteins were identified with nano-
HPLC-ESI-MS/MS. All membrane biological reactor (MBR) sludge samples investigated in this study contained elastase 3A, which implies that this human serine protease is a significant constituent of municipal wastewater. Although the identification of proteins from
ammonia-oxidizing bacterium Nitrosomonas europaea was expected, the detection of a protein with homology to the marine bacterium Saprospira grandis in MBR1 was surprising.
The data presented in this article provide supporting information to the related research article “Comparison of ten different DNA extraction procedures with respect to their suitability for environmental samples” (revised manuscript submitted to J. Microbiol. Methods). In that article, we compared the suitability of ten selected DNA extraction methods based on DNA quality, purity, quantity and applicability to universal PCR. Here we provide the data on the specific DNA gel sample load, all unreported gel images of crude DNA and PCR results, and the complete cost analysis for all tested extraction procedures and in addition two commercial DNA extraction kits for soil and water.
The FISH (fluorescence in situ hybridization) method is widely used to identify various types of cells. In comparison to cultivation-dependent methods, identification of microbes by FISH is easier
and generally takes several hours. This paper presents a review of improvements to the FISH method, its advantages and disadvantages, as well as examples of applications. Particular consideration was
given to the efficiency of microbes identification in samples taken from sewerage. The effectiveness of the method was confirmed by the results obtained in samples from the membrane bioreactor (MBR).
DNA extraction for molecular biological applications usually requires target optimized extraction procedures depending on the origin of the samples. For environmental samples, a range of different procedures has been developed. We compared the applicability and efficiency of ten selected DNA extraction methods published in recent literature using four different environmental samples namely: activated sludge from a domestic wastewater treatment plant, river sediment, anaerobic digestion sludge and nitrifying enrichment culture. We assessed the suitability of the extraction procedures based on both DNA yield and quality. DNA quantification was performed by both ultra violet (UV) spectrophotometry and fluorescence spectrophotometry after staining with PicoGreen. In our study, DNA yields based on UV measurement were overestimated in most cases while DNA yields from fluorescence measurements correlated well with the sample load on agarose gels of crude DNA. The quality of the DNA extracts was determined by gel electrophoresis of crude DNA and PCR products from 16S rDNA with the universal primer set 27f/1525r. It was observed that gel electrophoresis of crude DNA was not always suitable to evaluate DNA integrity and purity since interfering background substances (e.g. humic substances) were not visible. Therefore, we strongly recommend examining the DNA quality of both crude DNA and 16S rDNA PCR products by gel electrophoresis when a new extraction method is established.
Summarizing, we found four out of ten extraction procedures being applicable to all tested samples without noticeable restrictions. The procedure G (according to the standard method 432_10401 of the Lower Saxony State Office for Consumer Protection and Food Safety) had the broadest application range over procedure J
(published by Wilson, 2001). These were followed by procedures F (Singka et al., 2012) and A (Bourrain et al., 1999). All four extraction procedures delivered reliable and reproducible crude DNA and PCR products. From an economical point of view, all procedures tested during this study were cheaper compared to commercial DNA extraction kits.
This study aims at investigating the abiotic degradation pathway of
ethylenediaminetetra(methylenephosphonic acid) (EDTMP) simulated applying UV
irradiation. The degradation of EDTMP and formation of degradation products was
determined using LC-MS and ³¹P-NMR. In the laboratory scale experiments, EDTMP was
degraded within 30 min and the degradation products, iminodi(methylenephosphonic
acid) (IDMP), ethylaminobis(methylenephosphonic acid) (EABMP), and amino-
(methylenephosphonic acid) (AMPA), were simultaneously released. IDMP was the
main degradation product of EDTMP. Therefore, we conclude that the initial cleavage of
EDTMP is a heterolytically driven process, which starts the degradation process at the
intramolecular C-N bond. In contrast, the main product of a possible homolytic C-C
cleavage of methylaminobis(methylenephosphonic acid) could not be confirmed with
either LC-MS or ³¹P-NMR. Additionally, there was no evidence for a primary attack on
the C-P bond. All identified degradation products of EDTMP have been mineralized to
carbon dioxide (CO₂). Three additional degradation products (M1, M2, and M3) have
been found using the ³¹P-NMR analysis but have not yet been quantified using LC-MS.
We assume that the unidentified degradation product M1 is related to m/z 312, M2
to m/z 341, and M3 to m/z 409. Thus we concluded that EDTMP undergoes photochemical
conversion to IDMP, the main degradation product. EABMP and AMPA also accumulate,
but in smaller amounts. All intermediates are further mineralized to CO₂.
The anammox process is a potential alternative to the conventional nitrogen removal from wastewater. However, due to large generation times of anammox bacteria, the start-up of treatment reactors may be impeded. An efficient storage technique can handle this drawback and may be also suitable for seasonally operated treatment plants like in touristic areas. In the current study, several storage techniques were investigated with respect to its suitability for the
preservation of the specific anammox activity after long-term storage. Storing conditions differed in terms of temperature, redox buffer and nutrient supplementation. The specific activity of immobilized anammox bacteria (Candidatus Kuenenia stuttgartiensis) was determined three times during a long-term preservation of 78 days and 106 days, respectively. The highest activity was ensured at a storing temperature of 4 WC, providing nitrate as redox buffer and a nutrient
supplement every 23 days. Thus, 91.4% of the initial anammox activity could be preserved after a storage of 106 days. Superiority of the presented treatment condition was confirmed by a calculated nitrate-ammonium consumption rate close to the optimal ratio of 1.32. This technique
provided an economical and simple method suitable for long-term storage of immobilized anammox biomass.
The precision of cell number quantification in environmental samples depends on the complexity of the sample and on the applied technique. We compared fluorescence microscopy after filtration, quantification of gene copies and the cultivation based most probable number technique for their precision. We further analyzed the effect of increasing complexity of the sample material on the precision of the different methods by using pure cultures of Pseudomonas aeruginosa, fresh water samples and sediment slurries with and without ultrasonic treatment for analyses. Microscopy reached the highest precision, which was similar between pure cultures and water samples, but lower for sediment samples due to a higher percentage of cells in clusters and flocks. The PCR based quantification was most precise for pure cultures. Water and sediment samples were similar but less precise, which might be caused by the applied DNA extraction techniques. MPN measurements were equally precise for pure cultures and water samples. For sediment slurries the precision was slightly lower. The applied ultrasonic treatment of the slurries dispersed the cell clusters and flocks, increased the precision of microscopical and MPN measurements and also increased the number of potential colony forming units. However, the culturable cell number decreased by half. For MPN quantification of viable cells in samples with a high proportion of clustered cells we therefore recommend an optimization of ultrasonic treatment and a confirmation by microscopy and cultivation to reach highest possible dispersion of the cells with a minimum of inactivation. As a result of these observations we suggest a correction factor for MPN measurements to consider the effect of sonication on complex samples. The results are most likely applicable to other complex samples such as soil or biofilms.
Unerwünschte Bewohner : Wasserasseln und andere Makroorganismen in Trinkwasserversorgungsnetzen
(2013)
Unerwünschte Bewohner : Wasserasseln und andere Makroorganismen in Trinkwasserversorgungsnetzen
(2012)
Comparative Study of Different Methods for Analyzing Denitrifying Bacteria in Fresh Water Ecosystems
(2014)
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.
Immobilized microorganisms for the degradation of PAH and hydrocarbons from contaminated soil
(2000)