Community analysis of ammonia and nitrite oxidizers during start-up of nitritation reactors

Partial nitrification of ammonium to nitrite under oxic conditions (nitritation) is a critical process for the effective use of alternative nitrogen removal technologies from wastewater. Here we investigated the conditions which promote establishment of a suitable microbial community for performing nitritation when starting from regular sewage sludge. Reactors were operated in duplicate under different conditions (pH, temperature, and dilution rate) and were fed with 50 mM ammonium either as synthetic medium or as sludge digester supernatant. In all cases, stable nitritation could be achieved within 10 to 20 days after inoculation. Quantitative in situ hybridization analysis with group-specific fluorescent rRNA-targeted oligonucleotides (FISH) in the different reactors showed that nitrite-oxidizing bacteria of the genus Nitrospira were only active directly after inoculation with sewage sludge (up to 4 days and detectable up to 10 days). As demonstrated by quantitative FISH and restriction fragment length polymorphism (RFLP) analyses of the amoA gene (encoding the active-site subunit of the ammonium monooxygenase), the community of ammonia-oxidizing bacteria changed within the first 15 to 20 days from a more diverse set of populations consisting of members of the Nitrosomonas communis and Nitrosomonas oligotropha sublineages and the Nitrosomonas europaea-Nitrosomonas eutropha subgroup in the inoculated sludge to a smaller subset in the reactors. Reactors operated at 30 degrees C and pH 7.5 contained reproducibly homogeneous communities dominated by one amoA RFLP type from the N. europaea-N. eutropha group. Duplicate reactors at pH 7.0 developed into diverse communities and showed transient population changes even within the ammonia oxidizer community. Reactors at pH 7.5 and 25 degrees C formed communities that were indistinguishable by the applied FISH probes but differing in amoA RFLP types. Communities in reactors fed with sludge digester supernatant exhibited a higher diversity and were constantly reinoculated with ammonium oxidizers from the supernatant. Therefore, such systems could be maintained at a higher dilution rate (0.75 day(-1) compared to 0.2 day(-1) for the synthetic wastewater reactors). Despite similar reactor performance with respect to chemical parameters, the underlying community structures were different, which may have an influence on stability during perturbations.     

Interpreting 16S rDNA T-RFLP Data: Application of Self-Organizing Maps and Principal Component Analysis to Describe Community Dynamics and Convergence

Interpreting the large amount of data generated by rapid profiling techniques, such as T-RFLP, DGGE, and DNA arrays, is a difficult problem facing microbial ecologists. This study compares the ability of two very different ordination methods, principal component analysis (PCA) and self-organizing map neural networks (SOMs), to analyze 16S-DNA terminal restriction-fragment length polymorphism (T-RFLP) profiles from microbial communities in glucose-fed methanogenic bioreactors during startup and changes in operational parameters. Our goal was not only to identify which samples were similar, but also to decipher community dynamics and describe specific phylotypes, i.e., phylogenetically similar organisms, that behaved similarly in different reactors. Fifteen samples were taken over 56 volume changes from each of two bioreactors inoculated from river sediment (S2) and anaerobic digester sludge (M3) and from a well-established control reactor (R1). PCA of bacterial T-RFLP profiles indicated that both the S2 and M3 communities changed rapidly during the first nine volume changes, and then became relatively stable. PCA also showed that an HRT of 8 or 6 days had no effect on either reactor communtity, while an HRT of 2 days changed community structure significantly in both reactors. The SOM clustered the terminal restriction fragments according to when each fragment was most abundant in a reactor community, resulting in four clearly discernible groups. Thirteen fragments behaved similarly in both reactors, eight of which composed a significant proportion of the microbial community as judged by the relative abundance of the fragment in the T-RFLP profiles. Six Bacteria terminal restriction fragments shared between the two communities matched cloned 16S rDNA sequences from the reactors related to Spirochaeta, Aminobacterium, Thermotoga, and Clostridium species. Convergence also occurred within the acetoclastic methanogen community, resulting in a predominance of Methanosarcina siciliae-related organisms. The results demonstrate that both PCA and SOM analysis are useful in the analysis of T-RFLP data; however, the SOM was better at resolving patterns in more complex and variable data than PCA ordination.     

Community Analysis of Ammonia and Nitrite Oxidizers during Start-Up of Nitritation Reactors

Partial nitrification of ammonium to nitrite under oxic conditions (nitritation) is a critical process for the effective use of alternative nitrogen removal technologies from wastewater. Here we investigated the conditions which promote establishment of a suitable microbial community for performing nitritation when starting from regular sewage sludge. Reactors were operated in duplicate under different conditions (pH, temperature, and dilution rate) and were fed with 50 mM ammonium either as synthetic medium or as sludge digester supernatant. In all cases, stable nitritation could be achieved within 10 to 20 days after inoculation. Quantitative in situ hybridization analysis with group-specific fluorescent rRNA-targeted oligonucleotides (FISH) in the different reactors showed that nitrite-oxidizing bacteria of the genus Nitrospira were only active directly after inoculation with sewage sludge (up to 4 days and detectable up to 10 days). As demonstrated by quantitative FISH and restriction fragment length polymorphism (RFLP) analyses of the amoA gene (encoding the active-site subunit of the ammonium monooxygenase), the community of ammonia-oxidizing bacteria changed within the first 15 to 20 days from a more diverse set of populations consisting of members of the Nitrosomonas communis and Nitrosomonas oligotropha sublineages and the Nitrosomonas europaea-Nitrosomonas eutropha subgroup in the inoculated sludge to a smaller subset in the reactors. Reactors operated at 30°C and pH 7.5 contained reproducibly homogeneous communities dominated by one amoA RFLP type from the N. europaea-N. eutropha group. Duplicate reactors at pH 7.0 developed into diverse communities and showed transient population changes even within the ammonia oxidizer community. Reactors at pH 7.5 and 25°C formed communities that were indistinguishable by the applied FISH probes but differing in amoA RFLP types. Communities in reactors fed with sludge digester supernatant exhibited a higher diversity and were constantly reinoculated with ammonium oxidizers from the supernatant. Therefore, such systems could be maintained at a higher dilution rate (0.75 day⁻¹ compared to 0.2 day⁻¹ for the synthetic wastewater reactors). Despite similar reactor performance with respect to chemical parameters, the underlying community structures were different, which may have an influence on stability during perturbations.     

Phylogenetic Comparison of the Methanogenic Communities from an Acidic, Oligotrophic Fen and an Anaerobic Digester Treating Municipal Wastewater Sludge

Methanogens play a critical role in the decomposition of organics under anaerobic conditions. The methanogenic consortia in saturated wetland soils are often subjected to large temperature fluctuations and acidic conditions, imposing a selective pressure for psychro- and acidotolerant community members; however, methanogenic communities in engineered digesters are frequently maintained within a narrow range of mesophilic and circumneutral conditions to retain system stability. To investigate the hypothesis that these two disparate environments have distinct methanogenic communities, the methanogens in an oligotrophic acidic fen and a mesophilic anaerobic digester treating municipal wastewater sludge were characterized by creating clone libraries for the 16S rRNA and methyl coenzyme M reductase alpha subunit (mcrA) genes. A quantitative framework was developed to assess the differences between these two communities by calculating the average sequence similarity for 16S rRNA genes and mcrA within a genus and family using sequences of isolated and characterized methanogens within the approved methanogen taxonomy. The average sequence similarities for 16S rRNA genes within a genus and family were 96.0 and 93.5%, respectively, and the average sequence similarities for mcrA within a genus and family were 88.9 and 79%, respectively. The clone libraries of the bog and digester environments showed no overlap at the species level and almost no overlap at the family level. Both libraries were dominated by clones related to uncultured methanogen groups within the Methanomicrobiales, although members of the Methanosarcinales and Methanobacteriales were also found in both libraries. Diversity indices for the 16S rRNA gene library of the bog and both mcrA libraries were similar, but these indices indicated much lower diversity in the 16S digester library than in the other three libraries.     

Acclimation of denitrifying activated sludge to a single vs. complex external carbon source during a start-up of sequencing batch reactors treating ammonium-rich anaerobic sludge digester liquors

In this study, denitrification of ammonium-reach anaerobic sludge digester liquor was investigated during start-up periods of two laboratory-scale “fill-and-draw” reactors. One reactor was fed with a single carbon source (ethanol), whereas the other reactor was fed with a complex carbon source (fusel oil). During two acclimation experiments, the structure of microbial community involved in denitrification was analyzed using 16S rDNA polymerase chain reaction-denaturing gradient gel electrophoresis fingerprints and fluorescent in situ hybridization. The characteristics of the mixed liquor were additionally supported by regular measurements of nitrate uptake rates. The addition of fusel oil and ethanol resulted in a significant enhancement of the denitrification rate and efficiency combined with the increasing volumetric addition of sludge digester liquor up to 15 % of the reactor volume. The microbiological analyses revealed that the addition of sludge digester liquor as well as both external carbon sources (fusel oil and ethanol) did not affect the structure of microbial communities in a severe way. In both reactors, Curvibacter sp. and Azoarcus sp. were found as the most abundant representatives of denitrifiers.     

Methanogenic population dynamics during start-up of anaerobic digesters treating municipal solid waste and biosolids

An aggressive start-up strategy was used to initiate codigestion in two anaerobic, continuously mixed bench-top reactors at mesophilic (37 degrees C) and thermophilic (55 degrees C) conditions. The digesters were inoculated with mesophilic anaerobic sewage sludge and cattle manure and were fed a mixture of simulated municipal solid waste and biosolids in proportions that reflect U.S. production rates. The design organic loading rate was 3.1 kg volatile solids/m3/day and the retention time was 20 days. Ribosomal RNA-targeted oligonucleotide probes were used to determine the methanogenic community structure in the inocula and the digesters. Chemical analyses were performed to evaluate digester performance. The aggressive start-up strategy was successful for the thermophilic reactor, despite the use of a mesophilic inoculum. After a short start-up period (20 days), stable performance was observed with high gas production rates (1.52 m3/m3/day), high levels of methane in the biogas (59%), and substantial volatile solids (54%) and cellulose (58%) removals. In contrast, the mesophilic digester did not respond favorably to the start-up method. The concentrations of volatile fatty acids increased dramatically and pH control was difficult. After several weeks of operation, the mesophilic digester became more stable, but propionate levels remained very high. Methanogenic population dynamics correlated well with performance measures. Large fluctuations were observed in methanogenic population levels during the start-up period as volatile fatty acids accumulated and were subsequently consumed. Methanosaeta species were the most abundant methanogens in the inoculum, but their levels decreased rapidly as acetate built up. The increase in acetate levels was paralleled by an increase in Methanosarcina species abundance (up to 11.6 and 4.8% of total ribosomal RNA consisted of Methanosarcina species ribosomal RNA in mesophilic and thermophilic digesters, respectively). Methanobacteriaceae were the most abundant hydrogenotrophic methanogens in both digesters, but their levels were higher in the thermophilic digester.     

Characterisation of community structure of bacteria in parallel mesophilic and thermophilic pilot scale anaerobe sludge digesters

The aim of the present work was to compare the microbial communities of a mesophilic and a thermophilic pilot scale anaerobe sludge digester. For studying the communities cultivation independent chemotaxonomical methods (RQ and PLFA analyses) and T-RFLP were applied. Microbial communities of the mesophilic and thermophilic pilot digesters showed considerable differences, both concerning the species present, and their abundance. A Methanosarcina sp. dominated the thermophilic, while a Methanosaeta sp. the mesophilic digester among Archaea. Species diversity of Bacteria was reduced in the thermophilic digester. Based on the quinone patterns in both digesters the dominance of sulphate reducing respiratory bacteria could be detected. The PLFA profiles of the digester communities were similar though in minor components characteristic differences were shown. Level of branched chain fatty acids is slightly lower in the thermophilic digester that reports less Gram positive bacteria. The relative ratio of fatty acids characteristic to Enterobacteriaceae, Bacteroidetes and Clostridia shows differences between the two digesters: their importance generally decreased under thermophilic conditions. The sulphate reducer marker (15:1 and 17:1) fatty acids are present in low quantity in both digesters.     

Carbon source — A strong determinant of microbial community structure and performance of an anaerobic reactor

Industrial effluents differ in their organic composition thereby providing different carbon sources to the microbial communities involved in its treatment. This study aimed to investigate the correlation of microbial community structure with wastewater composition and reactor's performance. Self-immobilized granules were developed in simulated wastewater based on different carbon sources (glucose, sugarcane molasses, and milk) in three hybrid anaerobic reactors operated at 37 °C. To study archaeal community structure, a polyphasic approach was used with both qualitative and quantitative analysis. While PCR-denaturing gradient gel electrophoresis of 16S rRNA gene did not reveal major shifts in diversity of archaea with change in substrate, quantification of different groups of methanogens and total bacteria by real-time PCR showed variations in relative abundances with the dominance of Methanosaetaceae and Methanobacteriales. These data were supported by differences in the ratio of total counts of archaea and bacteria analyzed by catalyzed reporter deposition – fluorescence in situ hybridization. During hydraulic and organic shocks, the molasses-based reactor showed the best performance followed by the milk- and the glucose-based reactor. The study indicates that carbon source shapes the microbial community structure more in terms of relative abundance with distinct metabolic capacities rather than its diversity itself.     

Formation of filamentous aerobic granules: role of pH and mechanism

Filamentous overgrowth in aerobic granular sludge processes can cause reactor failure. In this work, aerobic granules were cultivated in five identical sequencing batch reactors with acetate or glucose as the carbon source with various values of influent pH (4.5–8). Microscopic observations revealed that acidic pH, rather than the species of carbon source, epistatically controls the aerobic granules with filamentous structure. An acidic pH shifted the structure of the microbial community in the granules, such that the fungus Geotrichum fragrans was the predominant filamentous microorganism therein. The acidic pH reduced the intracellular cyclic diguanylate (c-di-GMP) content for increasing the motility of the bacteria to washout and increase the growth rate of G. fragrans on glucose or acetate, together causing overgrowth of the fungus. Maintaining the suspension under alkaline condition is proposed as an effective way to suppress filamentous overgrowth and maintain granule stability.     

Microbial community structure and activity linked to contrasting biogeochemical gradients in bog and fen environments of the glacial lake agassiz peatland

The abundances, compositions, and activities of microbial communities were investigated at bog and fen sites in the Glacial Lake Agassiz Peatland of northwestern Minnesota. These sites contrast in the reactivity of dissolved organic matter (DOM) and the presence or absence of groundwater inputs. Microbial community composition was characterized using pyrosequencing and clone library construction of phylogenetic marker genes. Microbial distribution patterns were linked to pH, concentrations of dissolved organic carbon and nitrogen, C/N ratios, optical properties of DOM, and activities of laccase and peroxidase enzymes. Both bacterial and archaeal richness and rRNA gene abundance were >2 times higher on average in the fen than in the bog, in agreement with a higher pH, labile DOM content, and enhanced enzyme activities in the fen. Fungi were equivalent to an average of 1.4% of total prokaryotes in gene abundance assayed by quantitative PCR. Results revealed statistically distinct spatial patterns between bacterial and fungal communities. Fungal distribution did not covary with pH and DOM optical properties and was vertically stratified, with a prevalence of Ascomycota and Basidiomycota near the surface and much higher representation of Zygomycota in the subsurface. In contrast, bacterial community composition largely varied between environments, with the bog dominated by Acidobacteria (61% of total sequences), while the Firmicutes (52%) dominated in the fen. Acetoclastic Methanosarcinales showed a much higher relative abundance in the bog, in contrast to the dominance of diverse hydrogenotrophic methanogens in the fen. This is the first quantitative and compositional analysis of three microbial domains in peatlands and demonstrates that the microbial abundance, diversity, and activity parallel with the pronounced differences in environmental variables between bog and fen sites.     

Phylogenetic description of immobilized methanogenic community using real-time PCR in a fixed-bed anaerobic digester

After immobilization of anaerobes on polyurethane foam in a thermophilic, fixed-bed, anaerobic digester supplied with acetate, the results of real-time PCR analysis indicated that the major immobilized methanogenic archaea were Methanosarcina spp., and that the major free-living methanogenic archaea were Methanosarcina and Methanobacterium spp. 16S rRNA gene densities of Methanosarcina spp. and Methanobacterium spp. immobilized on the polyurethane foam were 7.6x10(9) and 2.6x10(8) copies/cm3, respectively. Immobilized methanogenic archaea could be concentrated 1000 times relative to those in the original anaerobically digested sludge from a completely mixed thermophilic digester supplied with cattle waste. On the other hand, immobilized bacteria could be concentrated only 10 times. The cell densities of the immobilized methanogenic archaea and bacteria were higher than those of the free-living methanogenic archaea and bacteria in the reactor. The results of clone analysis indicate that the major methanogenic archaea of the original thermophilic sludge are members of the order Methanomicrobiales, and that the major methanogenic archaea immobilized on the polyurethane foam are Methanosarcina spp., and those of the liquid phase are Methanobacterium spp. The results of the real time PCR analysis approximately agree with those of the clone analysis. These results indicate that real-time PCR analysis is useful for quantitatively describing methanogenic communities.     

Effect of pH on anoxic sulfide oxidizing reactor performance

The effects of pH on the performance of anoxic sulfide oxidizing (ASO) reactor were evaluated. Performance was investigated under various operational conditions at influent pH range of 4-11. At the influent pH of 7-7.5 during loading tests and HRT tests, the sulfide oxidation was partial. In general, the amount of sulfate formed decreased with the increasing sulfide and nitrite loadings. The bacterial communities in ASO reactors were more sensitive to acidic pH compared with alkaline pH, as nitrite and sulfide removal rates dropped significantly when exposed to acidic pH 3. High dissolved bisulfide ions, nitrite and excess of sulfate (>300 mg/L) might have inhibited the sulfide oxidation under highly acidic and alkaline conditions in the ASO reactor. Based on sulfide and nitrite removal efficiencies, the ASO reactor can be operated in a wide range of pH, i.e. 5-11.     

16S rDNA characterisation of bacterial and archaeal communities during start-up of anaerobic thermophilic digestion of cattle manure

A laboratory-scale continuously stirred anaerobic thermophilic batch digester was inoculated with cattle manure. Bacterial and archaeal communities, as well as digester performances, were analysed during reactor start-up for about 20 days. Polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) was used for overall detection and for study of the dynamics of microbial populations. Dominant bacteria and archaea 16S rDNAs were sequenced from the sample on day 12. Ten bacteria and 3 archaea OTUs (operational taxonomic units) were identified from the 52 clones sequenced. Sequences corresponding to the dominant bacterial SSCP peak were phylogenetically close to the 16S rDNA sequence of Bacillus thermoterrestris, whereas sequences corresponding to the two dominant archaeal SSCP peaks were phylogenetically close to the 16S rDNA sequence of Methanoculleus thermophilicus and Methanosarcina thermophila.     

Microwave radiation and reactor design influence microbial communities during methane fermentation

The effect of reactor design and method of heating on the efficiency of methane fermentation and composition of microbial communities, especially methanogenic Archaea, were determined. The research was carried out using submerge- and trickling-bed reactors fed with wastewater and the heat supply into the reactors included a convection heating method and microwave radiation. The polymerase chain reaction-denaturing gradient gel electrophoresis and relative real-time PCR were used in order to assess the biofilm communities. The best fermentation results and the highest abundance of methanogenic Archaea in biomass were observed in microwave heated trickling-bed reactors. The research proved that in reactors of identical design, the application of microwaves enabled a higher fermentation efficiency to be obtained and simultaneously increased the diversity of methanogenic Archaea communities that favors process stability. All the identified sequences of Archaea belonged to Methanosarcina sp., suggesting that species from this genera are susceptible to non-thermal effects of microwaves. There were no effects from microwaves on the bacterial communities in both types of reactors, however, the bacterial species composition varied in the reactors of different design.     

DIATOM COMMUNITY DYNAMICS IN STREAMS OF CHRONIC AND EPISODIC ACIDIFICATION: THE ROLES OF ENVIRONMENT AND TIME1

Community dynamics of epiphytic diatoms were studied for 3 years in a chronically and an episodically acidified tributary of Buck Creek, Adirondacks. Both streams experienced pulses of acidity during hydrologic events but these pulses were more pronounced in the episodically acidified stream, where pH decreased over two units (between 4.53 and 6.62) and the acid‐neutralizing capacity (ANC) became negative. In the chronically acidified stream, pH was below 4.9 and the ANC was negative 94% of the time. In this stream, high inorganic acidity following SO₄^2? enrichment from snowmelt or rainstorms alternated with high organic acidity derived from a headwaters wetland during base flow. The fluctuating water chemistry generated shifts in diatom community composition: from exclusive dominance of Eunotia bilunaris (Ehrenberg) Mills during periods of high inorganic acidity to proliferation of several subdominant species during periods of high organic acidity. In the episodically acidified stream, the pulses of acidity were associated with high NO₃^? concentrations and the corresponding high ratios of inorganic monomeric Al (Al_im) to organic monomeric Al (Al_om). Diatom communities there were dominated exclusively by E. exigua (Brébisson) Rabenhorst year round; however, this species peaked during periods of low acidity. Periods of high acidity and Al_im:Al_om ratios were marked by a decline in E. exigua and a concomitant increase in the subdominant species. Variance partitioning into terms of environmental and temporal variance, and their covariance, suggested that diatom communities in the chronically acidic stream were governed primarily by environmental factors while in the episodically acidic stream environmental and temporal factors had equal contributions.     

Impact of crop species on bacterial community structure during anaerobic co-digestion of crops and cow manure

The bacterial communities in three continuously stirred tank reactors co-digesting cow manure with grass silage, oat straw, and sugar beet tops, respectively, were investigated by 16S rRNA gene-based fingerprints and clone libraries. The analyses revealed both clearly distinct and similar phylotypes in the bacterial communities between the reactors. The major groups represented in the three reactors were Clostridia, unclassified Bacteria, and Bacteroidetes. Phylotypes affiliated with Bacilli or Deltaproteobacteria were unique to the sugar beet and straw reactor, respectively. Unclassified Bacteria dominated in sugar beet reactor while in the straw and grass reactor Clostridia was the dominant group. An increase in organic loading rate from 2 to 3 kg volatile solids m^?3 d^?1 resulted in larger changes in the bacterial community in the straw compared to grass reactor. The study shed more light on the evolution of bacterial community during anaerobic co-digestion of different crops and manure to methane.     

Benthic community response (primarily Chironomidae) to nutrient enrichment and alkalinization in shallow,soft water humic lakes

A comparison of the benthic fauna found in two shallow lakes in the New Jersey Pinelands (USA) illustrated the impact of elevated pH and nutrients caused by residential and agricultural disturbance on a naturally acidic, poorly buffered aquatic environment. Detailed community analysis suggested that change in community composition was a better indicator of response to disturbance than biological diversity indices. Chironomidae (insecta) were the predominant components of the benthic macroinvertebrate assemblage of both undisturbed Oswego Lake (low pH, low nutrients) and Nescochague Lake (fluctuating pH, elevated nutrients). The genera Procladius, Tribelos, and Pagastiella dominated Oswego Lake, where as Zalutschia zalutschicola, Procladius, Dicrotendipes, and Tanytarsus dominated Nescochague Lake. Glyptotendipes was a common and unique member of the Nescochague Lake assemblage. Cluster analysis indicated that the chemical differences between lakes were the most important community determinants, although, within each lake, depth and substrate affected the local communities. Oswego Lake exhibited a depauperate nonchironomid benthic fauna typical of low nutrient, acid lakes. In turn, Nescochague Lake exhibited an enriched nonchironomid fauna including mollusks and planaria which were not found in Oswego Lake.     

Comparison of two-stage thermophilic (68 degrees C/55 degrees C) anaerobic digestion with one-stage thermophilic (55 degrees C) digestion of cattle manure

A two-stage 68 degrees C/55 degrees C anaerobic degradation process for treatment of cattle manure was studied. In batch experiments, an increase of the specific methane yield, ranging from 24% to 56%, was obtained when cattle manure and its fractions (fibers and liquid) were pretreated at 68 degrees C for periods of 36, 108, and 168 h, and subsequently digested at 55 degrees C. In a lab-scale experiment, the performance of a two-stage reactor system, consisting of a digester operating at 68 degrees C with a hydraulic retention time (HRT) of 3 days, connected to a 55 degrees C reactor with 12-day HRT, was compared with a conventional single-stage reactor running at 55 degrees C with 15-days HRT. When an organic loading of 3 g volatile solids (VS) per liter per day was applied, the two-stage setup had a 6% to 8% higher specific methane yield and a 9% more effective VS-removal than the conventional single-stage reactor. The 68 degrees C reactor generated 7% to 9% of the total amount of methane of the two-stage system and maintained a volatile fatty acids (VFA) concentration of 4.0 to 4.4 g acetate per liter. Population size and activity of aceticlastic methanogens, syntrophic bacteria, and hydrolytic/fermentative bacteria were significantly lower in the 68 degrees C reactor than in the 55 degrees C reactors. The density levels of methanogens utilizing H2/CO2 or formate were, however, in the same range for all reactors, although the degradation of these substrates was significantly lower in the 68 degrees C reactor than in the 55 degrees C reactors. Temporal temperature gradient electrophoresis profiles (TTGE) of the 68 degrees C reactor demonstrated a stable bacterial community along with a less divergent community of archaeal species.     

HRT and nutrients affect bacterial communities grown on recirculation aquaculture system effluents

In a recirculation aquaculture system the drumfilter effluent can be used as substrate for heterotrophic bacterial production, which can be recycled as feed. Because the bacteria might contain pathogens, which could reduce its suitability as feed, it is important to characterize these communities. Bacteria were produced in growth reactors under different conditions: 7 h hydraulic retention time (HRT) vs. 2 h, sodium acetate vs. molasses, and ammonia vs. nitrate. The community of the drumfilter effluent was different from those found in the reactors. However, all major community components were present in the effluent and reactor broths. HRT influenced the bacteria community, resulting in a DGGE profile dominated by a band corresponding to an Acinetobacter sp.-related population at 2 h HRT compared to 7 h HRT, where bands indicative of alpha-proteobacterial populations most closely related to Rhizobium and Shinella spp. were most abundant. Molasses influenced the bacterial community. It was dominated by an Aquaspirillum serpens-related population. Providing total ammonia nitrogen (TAN) in addition to nitrate led to the occurrence of bacteria close to Sphaerotilus spp., Flavobacterium mizutaii and Jonesia spp. It was concluded from these results that a 6-7 h HRT is recommended, and that the type of substrate is less important, and results in communities with a comparably low pathogenic risk.     

The effect of temperature and retention time on methane production and microbial community composition in staged anaerobic digesters fed with food waste

Background

Food waste is a large bio-resource that may be converted to biogas that can be used for heat and power production, or as transport fuel. We studied the anaerobic digestion of food waste in a staged digestion system consisting of separate acidogenic and methanogenic reactor vessels. Two anaerobic digestion parameters were investigated. First, we tested the effect of 55 vs. 65 °C acidogenic reactor temperature, and second, we examined the effect of reducing the hydraulic retention time (HRT) from 17 to 10 days in the methanogenic reactor. Process parameters including biogas production were monitored, and the microbial community composition was characterized by 16S amplicon sequencing.

Results

Neither organic matter removal nor methane production were significantly different for the 55 and 65 °C systems, despite the higher acetate and butyrate concentrations observed in the 65 °C acidogenic reactor. Ammonium levels in the methanogenic reactors were about 950 mg/L NH₄ ⁺ when HRT was 17 days but were reduced to 550 mg/L NH₄ ⁺ at 10 days HRT. Methane production increased from ~ 3600 mL/day to ~ 7800 when the HRT was decreased. Each reactor had unique environmental parameters and a correspondingly unique microbial community. In fact, the distinct values in each reactor for just two parameters, pH and ammonium concentration, recapitulate the separation seen in microbial community composition. The thermophilic and mesophilic digesters were particularly distinct from one another. The 55 °C acidogenic reactor was mainly dominated by Thermoanaerobacterium and Ruminococcus, whereas the 65 °C acidogenic reactor was initially dominated by Thermoanaerobacterium but later was overtaken by Coprothermobacter. The acidogenic reactors were lower in diversity (34–101 observed OTU_0.97, 1.3–2.5 Shannon) compared to the methanogenic reactors (472–513 observed OTU_0.97, 5.1–5.6 Shannon). The microbial communities in the acidogenic reactors were > 90% Firmicutes, and the Euryarchaeota were higher in relative abundance in the methanogenic reactors.

Conclusions

The digestion systems had similar biogas production and COD removal rates, and hence differences in temperature, NH₄ ⁺ concentration, and pH in the reactors resulted in distinct but similarly functioning microbial communities over this range of operating parameters. Consequently, one could reduce operational costs by lowering both the hydrolysis temperature from 65 to 55 °C and the HRT from 17 to 10 days.