Progress 09/01/04 to 08/31/09
Outputs
OUTPUTS: The first laboratory experiment we performed to study the link between stability and microbial ecology in animal-waste digesters, was a study with four completely-stirred anaerobic digesters (4.5 liter-digesters) fed with dairy manure. We maintained four different mixing intensities by operating at 1500, 500, 250, and 50 rotations per min (RPM) for 250 days. This was done as part of a new collaboration with Prof. Muthanna Al-Dahhan. His lab had developed an in-situ particle tracking method (i.e., computer automated particle tracking [CARPT]) in combination with computational fluid dynamics (CFD) to quantify the mixing intensities in 3 dimensions of the digester. A molecular biology technique to quantify different methanogens was used to study the effect on the microbial ecology of the four digesters. This project trained one MS student (Rebecca Hoffmann), who wrote a thesis based on this work. Next, for the PhD dissertation-based project of another graduate student (Marcelo Garcia), we operated four anaerobic sequencing batch reactors for approximately 3 years by feeding swine waste. After a stable ecology was obtained, the ammonia concentrations of two reactors was increased to stress the system with the goal to change the metabolic pathway in which methane was formed from acetate. Initially, we had operated the reactors at 25 degrees C, but it became clear that a higher temperature (35 degrees C) was necessary to manipulate the changes in pathways. The project supported four undergraduate students (Elizabeth Campbell, Ryan Mackin, Alex Esche, and James Wexler), who helped with reactor operation and chemical analyses. A new collaboration was initiated with Kevin Yarasheski and Samual Smith, who are affiliated with the mass spectrometry core at Washington University in St. Louis, to quantify the two pathways from acetate to methane by isotopic measurements (13CO2). Several molecular biology techniques were used to monitor the community structure over time: full-length 16S ribosomal RNA (rRNA) gene surveys (~6000 sequences with Sanger sequencig); slot-blot hybridization; FISH-nanoSIMS; metagenomics (with highly-parallel 454 pyrosequencing); and DNA-SIP with 13C-labelled acetate. For the latter technique a new collaboration was initiated with Prof. Eugene Madsen at Cornell University. In addition, to analyze the data for the 16S rRNA gene survey, a new collaboration with Prof. Rob Knight at The University of Colorado was initiated. One of our anaerobic sludge samples that we believed should not have the alternative acetate conversion pathway (and therefore we had planned it to use as a negative control) was found to be positive. This resulted in a new research project; we sampled nine full-scale digesters for one year every month. We then performed isotopic analyses and a 16S rRNA gene survey of 115 samples (with a 454 pyrosequencing platform). The analysis of this project is completed but not the writing of the research paper. Finally, we also developed a novel micro-bioreactor for research purposes. This supported one chapter in the PhD dissertation of Benjamin Steinhaus. I have also written two book chapters. PARTICIPANTS: Undergraduates: James Wexler (objective 1); Alex Esche (objective 1 and 3), Ryan Mackin (objective 1); Liz Campbel (objective 1) Graduate students: Rebecca Hoffmann (MS objective 1); Benjamin Steinhaus (PhD objective 2); Marcelo Garcia (PhD objective 1-3) Post-doctoral research associate: Dr. Sarah Dryden (objective 1) Collaborators and co-PDs: Dr. Amy Shen Lab at Washington University (objective 2) Dr. Christine Floss at Washington University (objective 2) Dr. Eugene Madsen at Cornell University (objective 1) Dr. Rob Knight at the University of Colorado at Boulder (objective 1) Dr. Muthanna Al-Dahhan at Washington University (objective 1) Drs. Kevin Yarasheski and Samual Smith at Washington University (objective 1) TARGET AUDIENCES: Oral/Poster Presentations: Garcia M. G., Dryden S. K. and Angenent L. T. (2008). Treating swine waste with anaerobic sequencing batch reactors: performance and microbial community. 2008 Institute of Biological Engineering Annual Meeting, March 6-9, 2008, Chapel Hill, NC. Garcia M. L., Dryden S. K. and Angenent L. T. (2007). Swine waste treatment with anaerobic sequencing batch reactors: performance and microbial community. In: Abstracts for 2007 IWA, anaerobic digestion in mountain area and isolated rural zones workshop, June 5-7, 2007, Chambery, France. Steinhaus B., Garcia M. L., Shen A. Q. and Angenent L. T. (2006). Using microfluidics to study the optimal growth conditions of pure cultures of Methanosaeta concilii. In: Abstracts for the 106th General Meeting of the American Society for Microbiology, Orlando, FL, May 21-25, 2006. Steinhaus B., Shen A. and Angenent L. (2005). Growth and analysis of anaerobic wastewater methanogens using microfluidics. In: Abstracts of 58th Meeting of the Division of Fluid Dynamics, November 20-22, Chicago, IL, American Physical Society, College Park, MD. Hoffmann R.*, Garcia M. L., Veskivar M., Varma R., Karim K., Al-Dahhan M. H. and Angenent L. T. (2005). Effect of shear on performance and microbial ecology of completely-stirred anaerobic digesters treating animal manure. In: Proceedings of the Animal Agriculture and Processing: Managing Environmental Impacts Conference, Aug. 31 - Sept. 2, 2005, St. Louis, MO, Air and Waste Management Association and Water Environment Federation. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts
We found that the mixing intensity has an influence on the methanogenic populations, which in turn have an effect on the long-term stability of digesters. In short, there are two different groups of methanogens that convert acetate directly into methane through a pathway called acetate cleavage: Methanosaeta concilii and Methanosarcina spp.. When the mixing intensity is high, the conditions for the filamentous M. concilii are not optimal, resulting in a low ratio of M. concilii/Methanosarcina spp.. This ratio is reversed for the reactors with a lower mixing intensity. This result has consequences to the long-term stability of anaerobic digesters on the farm because Methanosarcina spp. growths faster during periods of high acetate concentrations, which are common during unstable periods. Therefore, the latter organism is advantageous for long-term stability (intense mixing). However, the reactors with an intense mixing were less stable during start up. Therefore, during start up the mixing intensity should be low, but this may be increased during the long-term operation. In the other reactor study, we found that the interaction between temperature and ammonia has a large effect of reactor stability and methane yields. Theoretically, a lower temperature of 25 degrees C compared to 35 degrees C should be advantageous for digesters with high total ammonium (i.e., NH3 and NH4+) concentrations because the inhibiting free ammonia concentration is much lower, resulting in a lower toxicity. However, we found the opposite because the anaerobic foodweb performs better at a higher temperature due to advantageous kinetics. This was unexpected and the farmers should, thus, operate the system at the higher temperature. In this study, we found also that at a total ammonium concentration of 5,200 mg N/L and 35 degrees C, 25% of the acetate-to-methane conversion occurs via an alterative pathway than the acetate cleavage (with isotopic studies) - syntrophic acetate oxidation. We found with DNA-SIP that a group of Proteobacteria closely related to Pseudomonas sp. was the bacterial symbiont, but that these levels in the biomass are lower than 1%, which makes them undetectable with FISH-nanoSIMS, 16S rRNA surveys, and even with metagenomics. The latter techniques were still extremely useful to add knowledge on the microbial ecology and community dynamics. The survey showed us that the communities in the independent reactors were similar to each other over the operating period and that they evolved similarly (very different from the community in the swine waste); ammonia hardly had an effect. This is different from the general ecology studies that are typically done over short time periods: communities changed chaotically. Metagenomics did show us that a shift in the ratio of acetate cleavage/acetate oxidation occurred at the same time as a change in the ratio of acetate-utilizing/hydrogen-utilizing methanogens as one would expect. The changing pathways are important for the stability of farm-based digesters because without the presence of the alternative acetate oxidation pathway the bioreactor would have likely crashed.
Publications
- Angenent L. T. and Wrenn B. A. (2008). Optimizing mixed-culture bioprocesses to convert wastes into biofuels (chapter 15, pp. 179-194). In: Bioenergy. Eds.: Wall J. D., Harwood, C. S. and Demain, A. ASM Press, Washington, DC.
- Angenent L. T. and Scott N. R. (2010). Practical aspects of methane production from agricultural wastes (Chapter 14). In: Biofuels from Agricultural Wastes and Byproducts. Eds.: Blaschek H., Ezeji T. and Scheffran, J., Blackwell Publishing, Ames, IA. In press.
- Jeffrey Werner, Marcelo Garcia, Sarah Dryden, Nicholas Scalfone, Kevin Yarasheski, Samual Smith, Ruth Ley, Rob Knight, Jeffrey Gordon, Christine Floss, Eugene Madsen, Largus Angenent. (2010). The alternative pathway of syntrophic acetate oxidation improves the stability of mesophilic anaerobic digesters. In preparation.
- Jeffrey Werner, Marcelo Garcia, Nicholas Scalfone, Kevin Yarasheski, Samual Smith, Rob Knight, Largus Angenent. (2010). Combining metadata with a large-scale 16S rRNA gene survey to understand syntrophic acetate oxidation in full-scale anaerobic digesters. In preparation.
- Steinhaus B., Garcia M. L., Shen A. Q. and Angenent L. T. (2007). A portable anaerobic microbioreactor reveals optimum growth conditions for the methanogen Methanosaeta concilii. Applied and Environmental Microbiology, Vol. 73, No. 5, pp. 1653-1658.
- Hoffmann R. A., Garcia M. L., Veskivar M., Karim K., Al-Dahhan M. H. and Angenent L. T. (2008). Effect of shear on performance and microbial ecology of continuously-stirred anaerobic digesters treating animal manure. Biotechnology and Bioengineering, Vol. 100, No. 1, pp. 38-48.
- Garcia M. L. and Angenent L. T. (2009) Interaction between temperature and ammonia in mesophilic digesters for animal waste treatment. Water Research, Vol. 43, No. 9, pp. 2373-2382.
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Progress 10/01/07 to 09/30/08
Outputs
OUTPUTS: The finding that a direct effect of the anaerobic digestion operating conditions on the types of methanogen populations and its quantities exists is important information to maintain stable farm-based digestion to treat animal manure. In the past a high percentage of farm-based digester failure has hampered full implementation of this technology, and this can be changed with an in-depth knowledge of the link between digester function and microbial ecology. Our goal is to prevent farm-based digestion failure and to promote the technology for simultaneous waste stabilization and bioenergy generation on the farm. Addition of value to animal manure treatment by producing bioenergy (biogas) will make agriculture in the US a more sustainable endeavor and reduce this country's dependency on foreign energy supplies. To disseminate our findings we presented a conference paper and gave presentation at the 2008 Institute of Biological Engineering Annual Meeting on March 6-9, 2008 in Chapel Hill, NC. In addition, a book chapter (focused in part on the stability of anaerobic digestion of animal manures) was published in a book called "Bioenergy" (American Society of Microbiology [ASM] Press) and a research paper was published (in Biotechnology and Bioengineering) on the effect of mixing on digester stability for animal manure treatment. PARTICIPANTS: Largus T. Angenent, PD Marcelo Garcia, was a graduate student during this period and his work was part of his PhD Thesis work. He operated the reactors and performed the molecular biology and isotopic measurement techniques. Alex Esche was an Undergraduate Research Assistant, assisting with the reactor work. Alex learned about anaerobic digester performance and operation. Jim Wexler sas an Undergraduate Research Assistant, assisting with the reactor work. Jim also learned about anaerobic digester performance and operation. Christine Floss, was a co-PD and assisted the graduate student with a technique called FISH-nanoSIMS as part of the study into the microbial ecology of the system. Christine is in the Physics Department at Washington University. All the other individuals were in the Department of Energy, Environmental, and Chemical Engineering during the time of the project period. TARGET AUDIENCES: Farmers and operators of digesters are target audiences of the outcome of digester stability research. One of our conclusions was to increase the temperatures to the maximum to handle increases in ammonium concentrations. This result can be directly implemented in the field. Environmental engineers are also a target audience because these engineers typically design the treatment systems on the farm. Finally, other target audiences are scientist who work with nondefined mixed cultures for energy generation or to study the microbial ecology of nondefined cultures in the environment or engineered systems. We have and will publish research papers with the results to help the field move forward. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts
In the summer of 2005, we had started four identical 5-liter anaerobic sequencing batch reactor (ASBR) systems to treat swine waste (i.e., water, urine, feces, and spilled plant-based feed) at 25 degrees C with a solids concentration of 20 g volatile solids (VS)/L. The design-loading rate of 4 g VS/L/d was reached after approximately 250 days. After a close to 1,000-day operating period, the reactor operation was stopped because the goals of our reactor study had been reached. Several operating changes, including ammonia concentrations and temperature, were made to study the effect on performance and microbial ecology. During period 1 (day 0-378), the methane yield was 0.31 L CH4/g volatile solids (VS) for all digesters (with no statistical differences among them) at a temperature and total ammonium-N levels of 25 degrees C and ~ 1,200 mg ammonium-N/L, respectively. During period 2 (day 379 to 745), the methane yield at 25 degrees C decreased by 45% when total ammonium-N and ammonia-N were increased in two of the four ASBRs to levels > 4,000 mg ammonium-N/L and > 80 mg ammonia-N/L, respectively. During period 3 (day 746-988), this relative inhibition was reduced from 45% to 13% compared to the low-ammonia control reactors when the operating temperature was increased from 25 degrees C to 35 degrees C (while the free ammonia levels increased from ~ 100 to ~ 250 mg ammonia-N/L). The 10 degrees C increase in temperature doubled the kinetic constant for methanogenesis, which overwhelmed the elevated toxicity effects caused by the increasing concentration of free ammonia. Thus, the farmer/operator may alleviate ammonia toxicity by increasing the operating temperature within the mesophilic range. We extrapolated our data to correlate temperature, ammonia, and methane yield and hypothesized that the difference between high- and low-ammonia reactors are negligible at the optimum mesophilic temperature of 38 degrees C. We also monitored the effect of the changing operating conditions on the microbial community and the pathways of methane formation in the four reactors. Nucleic acids- and isotopes-based techniques were used to unravel microbial (archaeal and bacterial) communities with respect to their identity, dynamics, and function in biomass samples collected from the same four reactors over the operating period. The 16S rRNA gene sequencing survey on the bacterial community over the operating period did not show a significant difference between low and high ammonia samples at the phyla level, with Firmicutes (~ 46% of the bacterial community) and Bacteroidetes (~ 33%) as the predominant phyla. We conclude that the community structure of the four ASBRs are similar to each other at each sampling point and were mostly affected by substrate type rather than total ammonium concentrations and/or temperature. This was not anticipated. Members of the family Methanosarcinaceae (from 3 to 8% of the total microbial community) and of the order Methanomicrobiales (also between 3 and 8%) predominated as the acetoclastic and hydrogenotrophic methanogens, respectively.
Publications
- Hoffmann R. A., Garcia M. L., Veskivar M., Karim K., Al-Dahhan M. H. and Angenent L. T. (2008). Effect of shear on performance and microbial ecology of continuously-stirred anaerobic digesters treating animal manure. Biotechnology and Bioengineering, Vol. 100, No. 1, pp. 38-48.
- Angenent L. T. and Wrenn B. A. (2008). Optimizing mixed-culture bioprocesses to convert wastes into biofuels (chapter 15, pp. 179-194). In: Bioenergy. Eds.: Wall J. D., Harwood, C. S. and Demain, A. ASM Press, Washington, DC.
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Progress 10/01/06 to 09/30/07
Outputs
The finding that a direct effect of the anaerobic digestion operating conditions on the methanogen levels exists is important information in order to maintain stable farm-based digestion to treat animal manure. In the past a high percentage of farm-based digester failure has hampered full implementation of this technology, and this can be changed with an in-depth knowledge of the link between digester function and microbial ecology. Our goal is to prevent farm-based digestion failure and to promote the technology for bioenergy generation on the farm. Addition of value to animal manure treatment by producing bioenergy (biogas) will make agriculture in the US a more sustainable endeavor and reduce this country's dependency on foreign energy supplies. To disseminate our findings we presented a conference paper and gave presentation at the 2007 International Water Association workshop on anaerobic digestion in mountain areas and isolated rural zones, which was held on June
5-7, 2007, in Chambery, France. In addition, a book chapter (focused in part on the stability of anaerobic digestion of animal manures) was written during the reporting period for a book called "Bioenergy", to be published by the American Society of Microbiology (ASM) Press. The date of publication is unknown at this time.
Impacts
In the summer of 2005, we had started four identical 5-liter anaerobic sequencing batch reactor (ASBR) systems to treat swine waste (i.e., water, urine, feces, and spilled plant-based feed) at 25 degrees C with a solids concentration of 20 g volatile solids (VS)/L. The design-loading rate of 4 g VS/L/d was reached after approximately 250 days. At the end of the reporting period, these reactors had been operated for two years and two months (or 790 days of operation). During this period, two operating changes were made. First, the ammonia concentrations were increased for two of the four reactors (on day 378 of the operating period). Prior to ammonia addition into the reactors during the second operational year, methanogenic activity tests with the ASBR biomass were conducted to find an optimum total ammonium concentration for which acetoclastic methanogens would be considerably inhibited. We found a 40-50% decrease in methane production from acetate by having a total
ammonium concentration of 4,000 mg NH3-N/L at 25 degrees C, and therefore this ammonia concentration was chosen to select for a syntrophic association of acetate-oxidizing bacteria and hydrogen-utilizing methanogens. The methane yield for all four digesters was 0.32 L CH4/g VS fed in the first year when no ammonia was added. In the second year, the methane yield for the reactors with no ammonia addition remained stable (0.29 L CH4/g VS fed), whereas the methane yield decreased to 0.18 L CH4/g VS fed for the ammonia-augmented ASBRs. The 16S ribosomal RNA (rRNA) gene surveys for the biomass samples from each ASBR at the end of the second year (day 700) revealed no major differences in terms of the dominant bacterial phyla or species between the reactors with and without ammonia augmentation. The fact that ammonia concentration differences at these levels did not have an effect of the bacterial community structure was a surprising result. In accordance, tests with stable-isotopic labeled
acetate to CO2 conversion in serum bottles and gas analysis with GC/MS showed that syntrophic acetate-oxidation was not an important process in the reactors. Therefore, the second change during the operating period was the increase in reactor temperature from 25 to 35 degrees C on day 740 of the operating period to increase the methane yield of the ammonia-augmented ASBRs. Indeed, increasing the temperature did decrease the difference in methane yield to almost zero for the ASBRs with and without ammonia augmentation. We are currently investigating if this change is due to the growth of a syntrophic community that can produce methane though a two-step process. We have developed the protocols for identification of acetate-oxidizing bacteria by using labeled isotope substrate (13C-labeled acetate). By using DNA-SIP we found that many species that are not acetate oxidizers were sequenced and this disappointing result is in agreement with findings in the literature. We will, therefore,
perform rRNA-SIP in the future. Finally, we have obtained promising results with FISH-NanoSIMS with 13C labeled acetate.
Publications
- Hoffmann, R. A., Garcia, M. L., Veskivar, M., Karim, K., Al-Dahhan, M. H. and Angenent, L. T. 2007. Effect of shear on performance and microbial ecology of continuously-stirred anaerobic digesters treating animal manure. Online ahead of print publication: Biotechnol. Bioeng.
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Progress 10/01/05 to 09/30/06
Outputs
In the summer of 2005 we had started four identical 5-liter ASBR systems to treat swine waste (i.e., water, urine, feces, and spilled plant-based feed) with a solids concentration of 20 g volatile solids (VS)/L. The anaerobic bioreators have been operated at a temperature of 25 degrees C. We increased the volumetric loading rate (VLR) from 1 to 4 g VS/L/day by decreasing the hydraulic retention time from 20 to 5 days over a period of 1 year. Performance data was nearly identical for all digesters after reaching a steady-state performance at the design VLR of 4 g VS/L/day (summer 2006). The methane yield corrected for standard temperature and pressure during steady-state periods was not statistically different between reactors ([ANOVA]: n=16, p=0.99). The overall methane yield (y) in liters was y = 0.28x +0.29 (standard error of 0.06, R2=0.98) based on the amount (x in grams) of VS fed. We increased the ammonia levels from 1,000 to 4,000 mg N/L for two of the four
reactors at a pH of 7.6 and will operate them for another year. The capacity of anaerobic systems to produce large quantities of renewable energy from animal wastes has been demonstrated, however, there is a lack of knowledge on the bacterial and archaeal composition in these systems, and for this study we are especially interested in bacteria that can oxidize acetate when living in a syntrophic relationship with hydrogen-utilizing metahnogens. We have performed a 16S ribosomal RNA (rRNA) gene survey on the mixed culture in the four digesters after the one-year operating period. The survey is a culture-independent survey to determine "who is there" when metagenomic DNA is sampled, extracted, and PCR amplified. Based on the similarity of the performance data among digesters, we anticipated a similar bacterial composition. This was indeed the case on the phyla level for the four biomass samples from each of the ASBRs ( 300 bacterial sequences for each digester). The bacterial community
in the inoculum (granular biomass) was predominantly characterized by species from the phyla Chloroflexi (33%) and Proteobacteria (30%), followed by Firmicutes (17%) and Bacteroidetes (11%) ( 300 bacterial sequences). After the one-year operating period, the bacterial community had shifted towards the phyla Firmicutes ( 50%), Bacteroidetes ( 20%), and Spirochaetes ( 10%), with only small differences between the reactors. The work already has shed light on the bacterial community dynamics, because the bacterial composition for the four reactors was found to be similar in all four reactors. It is often thought that the bacterial composition fluctuates chaotically; we have now the results to dispute this idea. After the biomass will acclimate to the high ammonia levels, we will obtain another 1,250 bacterial sequences and 600 archaeal sequences and will perform other molecular techniques, such as stable isotope probing (SIP) in combination with a 16S rRNA gene sequencing survey, to
study syntrophic acetate oxidizing bacteria.
Impacts
The finding that a direct effect of the anaerobic digestion operating conditions on the methanogen levels exists is important information in order to maintain stable farm-based digestion to treat animal manure. In the past a high percentage of farm-based digester failure has hampered full implementation of this technology, and this can be changed with an in-depth knowledge of the link between digester function and microbial ecology. Our goal is to prevent farm-based digestion failure and to promote the technology for bioenergy generation on the farm. Addition of value to animal manure treatment by producing bioenergy (biogas) will make agriculture in the US a more sustainable endeavor and reduce this country's dependency on foreign energy supplies. Development of innovative research tools, such as the anaerobic micro-bioreactor, to study anaerobic bacteria and archaea will be instrumental to understand the link between function and microbial ecology. The techniques
that are being developed for this study can be used for other work involving microbial biotechnology.
Publications
- Steinhaus B., Garcia M. L., Shen A. Q. and Angenent L. T. (2007). A portable anaerobic microbioreactor reveals optimum growth conditions for the methanogen Methanosaeta concilii. Published ahead online on January 12, 2007: Applied and Environmental Microbiology.
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Progress 10/01/04 to 09/30/05
Outputs
In August 2005, we started four identical 5-L anaerobic sequencing batch reactor (ASBR) systems to treat swine waste with a solids concentration of 20 g volatile solids (VS)/L. At the start of the operating period, we operated the systems at a volumetric loading rate (VLR) of 1 g VS/L/day and we will increase the VLR to the design loading rate 4 g VS/L/day over the course of 1 year by decreasing the hydraulic retention time from 20 to 5 days. When a steady-state performance at 4 g VS/L/day is accomplished, we will increase the ammonia levels from 1,000 to 3,000 mg N/L for two of the four reactors and we will study the microbial community dynamics. We planned to operate the digesters for 2 years and we will utilize molecular biology techniques, such as membrane hybridization, to follow changes in the community structure with respect to syntrophic-acetate oxidizing bacteria and methanogens. To develop the published membrane hybridization protocols for use in our lab, we
first grew pure cultures of the following methanogens as a reference for DNA probes that target methanogens belonging to the classification given in brackets: Methanobrevibacter smithii (family of Methanobacteriaceae), Methanococcus voltaei (family of Methanoccaceae), Methanogenium cariaci (order of Methanomicrobiales), Methanosarcina acetivorans (genus of Methanosarcina), and Methanosaeta concilii (species of M. concilii). We then tested our membrane hybridization protocols for a study in which the effect of shear rate on anaerobic digesters performance was investigated. Four continuously-stirred tank reactors operating at different mixing intensities of 1500, 500, 250, and 50 revolutions per minute (RPM) were fed animal manure for half a year. Biomass samples from these four digesters were collected over the operating period and the methanogen levels were analyzed with membrane hybridization. The most important result of our assessment was the direct effect of shear rate on the
methanogenic community structure. At the end of the operating period, the acetate-utilizing methanogens of the species M. concilii showed the lowest levels (<0.5% based on the total 16S rRNA present) in the digesters with the highest shear rate (1500 and 500 rpm), while the levels of these methanogens was 3% in the digesters with the lowest shear rate (250 and 50 rpm). The competing acetate-utilizing methanogens from the genus of Methanosarcina showed the opposite trend, confirming these results. We demonstrated that anaerobic digester operation has a direct effect on the methanogenic ecology even if the performance is not affected and also that membrane hybridization is useful for anaerobic digestion studies. We developed an anaerobic micro-bioreactor with a total volume of 5 micro-L using microfluidics and used it to study pure cultures of M. concilii. An optimum shear level of 6-8 micro-Pa was found in an experiment with varying channel widths. This information is used to design
experiments to assess and evaluate the optimal pH and ammonia levels for M. concilii. The development of the micro-bioreactor will aid our research on the sensitivity of syntrophic acetate-utilizing bacteria.
Impacts
The finding that a direct effect of the anaerobic digestion operating conditions on the methanogen levels exists is important information in order to maintain stable farm-based digestion to treat animal manure. In the past a high percentage of farm-based digester failure has hampered full implementation of this technology, and this can be changed with an in-depth knowledge of the link between digester function and microbial ecology. Our goal is to prevent farm-based digestion failure and to promote the technology for bioenergy generation on the farm. Addition of value to animal manure treatment by producing bioenergy (biogas) will make agriculture in the US a more sustainable endeavor and reduce the US dependency on foreign energy supplies. Development of innovative research tools, such as the anaerobic micro-bioreactor, to study anaerobic bacteria and archaea will be instrumental to understand the link between function and microbial ecology. The techniques that are
being developed for this study can be used for other work involving microbial biotechnology.
Publications
- Hoffmann R., Garcia M. L., Veskivar M., Varma R., Karim K., Al-Dahhan M. H. and Angenent L. T. (2005). Effect of shear on performance and microbial ecology of completely-stirred anaerobic digesters treating animal manure. In: Proceedings of the Animal Agriculture and Processing: Managing Environmental Impacts Conference, Aug. 31-Sept. 2, 2005, St. Louis, MO, Air and Waste Management Association and Water Environment Federation.
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