Effects of slatted frame placed in compost pile on enhancing heat generation and organic matter degradation during high-moisture cow manure composting.

Excess moisture content in cow manure inhibits the process of composting. This study examined the effects of introducing a slatted frame on temperature development, organic matter degradation, and moisture content during high-moisture composting. Twelve tons of cow manure was piled individually, and an iron slatted frame with a pipe and a plain frame filled inside with styrofoam was introduced in the treatment and control piles, respectively. Two hours after the start, running leachate via the pipe in the treatment pile was observed, but not in the control pile. The maximum temperatures as well as durations of the temperatures above 55°C of the treatment piles at all seven positions were higher than those of the control piles. In particular, four of the seven positions in the treatment pile exhibited more than twice the duration. Installing the slatted frame improved the organic matter degradation; the values of organic matter loss in the control and treatment were 48.0% and 58.4%, respectively. The final moisture content in the treated compost was 5% lower than that in the control. Overall, the introduction of a slatted frame generated higher temperatures and produced relatively dried and well-degraded compost.

Collection of ear corn residue and its utilization as a bulking agent for cow manure composting.

This study examined the collection, storage, and utilization of ear corn residue as a bulking agent for composting. Ear corn residue left in fields was collected by a sequence of windrowing and round baling operations, which showed a collection efficiency of 53%-56%. More than 70% of the corn stalks had lengths shorter than 15 cm. The moisture content of corn residue collected in late October was approximately 58%; it decreased to 23% during storage. Dried corn residue was mixed with raw manure in two different ways, namely using complete mixing (CM) treatment or creating a bottom layer of residue, and a total of 3.4 t of the manure and ear corn residue mixture were composted using a pilot-scale composting apparatus. The results showed that the CM treatment resulted in higher temperature generation and produced less odorous and well-degraded compost after 2 months of composting, while the odorous compounds in the compost with corn residue as the bottom layer remained similar to those of the initial compost. To sufficiently utilize the advantages of the ear corn residue, thorough mixing of the corn residue with raw manure is preferable; this procedure produced well-degraded and safety compost in a shorter time period.

Dead bacterial biomass-assimilating bacterial populations in compost revealed by high-sensitivity stable isotope probing.

Pathogens are known to survive in compost and to regrow under the influence of certain factors, such as moisture content, temperature and nutrient availability. Dead biomass, by providing available nutrients, is a factor that may affect pathogen regrowth. However, the indigenous microorganisms, including pathogens, that grown on the dead biomass of compost have not yet been identified. Here, the regrowth potential of the pathogenic indicator bacterium Escherichia coli in the presence of dead bacterial biomass was determined, and the biomass metabolizers that grew competitively with E. coli were identified by high-sensitivity stable isotope probing of rRNA. Culture-dependent analysis indicated that the addition of dead bacterial biomass did not stimulate E. coli growth. High-throughput analysis of density-resolved 16S rRNA molecules from compost samples amended with carbon-13-labeled dead bacterial biomass revealed dead bacterial-assimilating bacteria, including Sphingobium sp., myxobacterial lineages and Bacillales. These bacteria are potentially competitive with pathogens due to their preferential assimilation of dead biomass in compost.

Eukaryotic Microbiomes of Membrane-Attached Biofilms in Membrane Bioreactors Analyzed by High-Throughput Sequencing and Microscopic Observations.

Limited information is currently available on the contribution of eukaryotes to the reactor performance of membrane bioreactors (MBRs). Using high-throughput Illumina sequencing of 18S rRNA genes and microscopic observations, we investigated eukaryotic microbiomes in membrane-attached biofilms in MBRs treating piggery wastewater. Protozoa preying on bacteria were frequently detected under stable conditions when membrane clogging was suppressed. However, the eukaryotes preying upon protozoa became predominant in biofilms when membrane fouling rapidly progressed. We herein demonstrated that a comprehensive investigation of eukaryotic microbiomes using high-throughput sequencing contributes to a better understanding of the microbial ecology involved in wastewater treatment.

Characterization of the denitrifying bacterial community in a full-scale rockwool biofilter for compost waste-gas treatment.

The potential denitrification activity and the composition of the denitrifying bacterial community in a full-scale rockwool biofilter used for treating livestock manure composting emissions were analyzed. Packing material sampled from the rockwool biofilter was anoxically batch-incubated with 15N-labeled nitrate in the presence of different electron donors (compost extract, ammonium, hydrogen sulfide, propionate, and acetate), and responses were compared with those of activated sludge from a livestock wastewater treatment facility. Overnight batch-incubation showed that potential denitrification activity for the rockwool samples was higher with added compost extract than with other potential electron donors. The number of 16S rRNA and nosZ genes in the rockwool samples were in the range of 1.64-3.27 × 109 and 0.28-2.27 × 108 copies/g dry, respectively. Denaturing gradient gel electrophoresis analysis targeting nirK, nirS, and nosZ genes indicated that the distribution of nir genes was spread in a vertical direction and the distribution of nosZ genes was spread horizontally within the biofilter. The corresponding denitrifying enzymes were mainly related to those from Phyllobacteriaceae, Bradyrhizobiaceae, and Alcaligenaceae bacteria and to environmental clones retrieved from agricultural soil, activated sludge, freshwater environments, and guts of earthworms or other invertebrates. A nosZ gene fragment having 99% nucleotide sequence identity with that of Oligotropha carboxidovorans was also detected. Some nirK fragments were related to NirK from micro-aerobic environments. Thus, denitrification in this full-scale rockwool biofilter might be achieved by a consortium of denitrifying bacteria adapted to the intensely aerated ecosystem and utilizing mainly organic matter supplied by the livestock manure composting waste-gas stream.

Survival of free-living Acholeplasma in aerated pig manure slurry revealed by (13)C-labeled bacterial biomass probing.

Many studies have been performed on microbial community succession and/or predominant taxa during the composting process; however, the ecophysiological roles of microorganisms are not well understood because microbial community structures are highly diverse and dynamic. Bacteria are the most important contributors to the organic-waste decomposition process, while decayed bacterial cells can serve as readily digested substrates for other microbial populations. In this study, we investigated the active bacterial species responsible for the assimilation of dead bacterial cells and their components in aerated pig manure slurry by using (13)C-labeled bacterial biomass probing. After 3 days of forced aeration, (13)C-labeled and unlabeled dead Escherichia coli cell suspensions were added to the slurry. The suspensions contained (13)C-labeled and unlabeled bacterial cell components, possibly including the cell wall and membrane, as well as intracellular materials. RNA extracted from each slurry sample 2 h after addition of E. coli suspension was density-resolved by isopycnic centrifugation and analyzed by terminal restriction fragment length polymorphism, followed by cloning and sequencing of bacterial 16S rRNA genes. In the heavy isotopically labeled RNA fraction, the predominant (13)C-assimilating population was identified as belonging to the genus Acholeplasma, which was not detected in control heavy RNA. Acholeplasma spp. have limited biosynthetic capabilities and possess a wide variety of transporters, resulting in their metabolic dependence on external carbon and energy sources. The prevalence of Acholeplasma spp. was further confirmed in aerated pig manure slurry from four different pig farms by pyrosequencing of 16S rRNA genes; their relative abundance was ∼4.4%. Free-living Acholeplasma spp. had a competitive advantage for utilizing dead bacterial cells and their components more rapidly relative to other microbial populations, thus allowing the survival and prevalence of Acholeplasma spp. in pig manure slurry.

Reorganization of the bacterial and archaeal populations associated with organic loading conditions in a thermophilic anaerobic digester.

Organic loading conditions are an important factor influencing reactor performances in methanogenic bioreactors. Yet the underlying microbiological basis of the process stability, deterioration, and recovery remains to be understood. Here, structural responses of the bacterial and archaeal populations to the change of organic loading conditions in a thermophilic anaerobic digester were investigated by process analyses and 16S rRNA gene-based molecular approaches. The biogas was produced stably without the accumulation of volatile fatty acids (VFAs) at low organic loading rates (OLRs) in the beginning of reactor operation. Increasing OLR in stages disrupted the stable reactor performance, and high OLR conditions continued the deteriorated performance with slight biogas production and high accumulation of VFAs. Thereafter, the gradual decrease of OLR resulted in the recovery from the deterioration, giving rise to the stable performance again. The stable performances before and after the high OLR conditions conducted were associated with compositionally similar but not identical methanogenic consortia. The bacterial and archaeal populations were synchronously changed at both the transient phases toward the deteriorated performance and in recovery process, during which the dynamic shift of aceticlastic and hydrogenotrophic methanogens including the recently identified Methanomassiliicoccus might contribute to the maintenance of the methanogenic activity. The distinctive bacterial population with a high predominance of Methanobacterium formicicum as archaeal member was found for the deteriorated performance. The results in this study indicate the coordinated reorganization of the bacterial and archaeal populations in response to functional states induced by the change of organic loading conditions in the anaerobic digester.

Comparison of the composting process using ear corn residue and three other conventional bulking agents during cow manure composting under high-moisture conditions.

To elucidate the characteristics of ear corn residue as a bulking agent, the composting process using this residue was compared with processes using three other conventional materials such as sawdust, wheat straw and rice husk, employing a bench-scale composting reactor. As evaluated via biochemical oxygen demand (BOD), ear corn residue contains 3.3 and 2.0 times more easily digestible materials than sawdust and rice husk, respectively. In addition, mixing ear corn residue with manure resulted in reduced bulk density, which was the same as that of wheat straw and was 0.58 and 0.67 times lower than that of sawdust and a rice husk mixture, respectively. To evaluate temperature generation during the composting process, the maximum temperature and area under the temperature curve (AUCTEMP) were compared among the mixed composts of four bulking agents. Maximum temperature (54.3°C) as well as AUCTEMP (7310°C●h) of ear corn residue were significantly higher than those of sawdust and rice husk (P<0.05), and they are similar to that of wheat straw mixed compost. Along with the value of AUCTEMP, the highest organic matter losses of 31.1% were observed in ear corn residue mixed compost, followed by wheat straw, saw dust and rice husk.

Denitrifiers in the surface zone are primarily responsible for the nitrous oxide emission of dairy manure compost.

During the dairy manure composting process, significant nitrous oxide (N2O) emissions occur just after the pile turnings. To understand the characteristics of this N2O emission, samples were taken from the compost surface and core independently, and the N2O production was monitored in laboratory incubation experiments. Equal amounts of surface and core samples were mixed to simulate the turning, and the (15)N isotope ratios within the molecules of produced N2O were analyzed by isotopomer analysis. The results showed that the surface samples emitted significant levels of N2O, and these emissions were correlated with NOx(-)-N accumulation. Moreover, the surface samples and surface-core mixed samples incubated at 30°C produced N2O with a low site preference (SP) value (-0.9 to 7.0‰) that was close to bacteria denitrification (0‰), indicating that denitrifiers in the surface samples are responsible for this N2O production. On the other hand, N2O produced by NO2(-)-amended core samples and surface samples incubated at 60°C showed unrecognized isotopic signatures (SP=11.4-20.3‰). From these results, it was revealed that the N2O production occurring just after the turnings was mainly derived from bacterial denitrification (including nitrifier denitrification) of NOx(-)-N under mesophilic conditions, and surface denitrifying bacteria appeared to be the main contributor to this process.

Microbiology of nitrogen cycle in animal manure compost.

Composting is the major technology in the treatment of animal manure and is a source of nitrous oxide, a greenhouse gas. Although the microbiological processes of both nitrification and denitrification are involved in composting, the key players in these pathways have not been well identified. Recent molecular microbiological methodologies have revealed the presence of dominant Bacillus species in the degradation of organic material or betaproteobacterial ammonia-oxidizing bacteria on nitrification on the surface, and have also revealed the mechanism of nitrous oxide emission in this complicated process to some extent. Some bacteria, archaea or fungi still would be considered potential key players, and the contribution of some pathways, such as nitrifier denitrification or heterotrophic nitrification, might be involved in composting. This review article discusses these potential microbial players in nitrification-denitrification within the composting pile and highlights the relevant unknowns through recent activities that focus on the nitrogen cycle within the animal manure composting process.

Characterization and spatial distribution of bacterial communities within passively aerated cattle manure composting piles.

The bacterial communities in the core, bottom, top, middle-surface, and lower-surface full-scale passively aerated cattle manure compost was investigated using DGGE of PCR-amplified 16S rRNA sequences. Some Bacillus species and strictly anaerobic thermophilic Clostridium species were dominant only in the core and bottom zones. In contrast, bands belonging to mesophilic bacteria such as Bacteroidetes, Clostoridia,alpha and gamma-proteobacteria were detected in surface zones, even in the initial thermophilic stage of the process. Our results clearly show the spatial distribution of the microbial community within full-scale composting piles, which indicates N or C conversion by zone-specific bacterial communities were occurring in each zone of the pile.

Source of nitrous oxide emissions during the cow manure composting process as revealed by isotopomer analysis of and amoA abundance in betaproteobacterial ammonia-oxidizing bacteria.

A molecular analysis of betaproteobacterial ammonia oxidizers and a N(2)O isotopomer analysis were conducted to study the sources of N(2)O emissions during the cow manure composting process. Much NO(2)(-)-N and NO(3)(-)-N and the Nitrosomonas europaea-like amoA gene were detected at the surface, especially at the top of the composting pile, suggesting that these ammonia-oxidizing bacteria (AOB) significantly contribute to the nitrification which occurs at the surface layer of compost piles. However, the (15)N site preference within the asymmetric N(2)O molecule (SP = delta(15)N(alpha) - delta(15)N(beta), where (15)N(alpha) and (15)N(beta) represent the (15)N/(14)N ratios at the center and end sites of the nitrogen atoms, respectively) indicated that the source of N(2)O emissions just after the compost was turned originated mainly from the denitrification process. Based on these results, the reduction of accumulated NO(2)(-)-N or NO(3)(-)-N after turning was identified as the main source of N(2)O emissions. The site preference and bulk delta(15)N results also indicate that the rate of N(2)O reduction was relatively low, and an increased value for the site preference indicates that the nitrification which occurred mainly in the surface layer of the pile partially contributed to N(2)O emissions between the turnings.

Characteristics of the microbial community associated with ammonia oxidation in a full-scale rockwool biofilter treating malodors from livestock manure composting.

The relationship between the activity and community structure of microbes associated with the oxidation of ammonia in a full-scale rockwool biofilter was examined by kinetic, denaturing gradient gel electrophoresis (DGGE), and sequence analyses. The packing materials were sampled from two different depths at 3 sites. Estimated K(m) values were similar among depths at same sampling sites, while V(max) differed in the mid-point sample. The lower depth of this site had the highest V(max). A correspondence analysis showed the DGGE profile of ammonia-oxidizing bacterial amoA of the lower depth of the mid-point sample to be distinguishable from the others. Banding patterns at other sites were similar among depths. Banding patterns of ammonia-oxidizing archaeal amoA of the mid-point sample were also similar among depths. The results suggested an association between the ammonia-oxidizing bacterial community's composition and ammonium oxidation kinetics in samples. Sequence analysis indicated that the ammonia-oxidizing bacterial community mainly belonged to the Nitrosomonas europaea lineage and Nitrosospira cluster 3. The ammonia-oxidizing archaeal amoA-like sequences were related to those belonging to soil and sediment groups, including one with 84% nucleotide similarity with Nitrosopumilus maritimus.

Key odor components responsible for the impact on olfactory sense during swine feces composting.

This study aimed to identify the major odor contributing components produced during swine feces composting which have an impact on the olfactory senses. A total of 64 gas samples collected at different stages of composting were analyzed by both a gas chromatograph and human panel test using the triangle odor bag method. Multiple regression analysis of representative odor substances present in the outlet gas was carried out employing the odor index (OI) as the dependent variable and the odor unit as the independent variable. The recorded changes in OI indicated that turning was an important event during odor evolution, and that the odor emission during the thermophilic phase should be the main target for odor abatement. The model incorporating ammonia, methyl mercaptan and dimethyl sulfide as independent variables confirmed the value of the OI (R(2)=0.70). These compounds were identified to be the key odor components significantly determining the OI.

The impact of using mature compost on nitrous oxide emission and the denitrifier community in the cattle manure composting process.

The diversity and dynamics of the denitrifying genes (nirS, nirK, and nosZ) encoding nitrite reductase and nitrous oxide (N(2)O) reductase in the dairy cattle manure composting process were investigated. A mixture of dried grass with a cattle manure compost pile and a mature compost-added pile were used, and denaturing gradient gel electrophoresis was used for denitrifier community analysis. The diversity of nirK and nosZ genes significantly changed in the initial stage of composting. These variations might have been induced by the high temperature. The diversity of nirK was constant after the initial variation. On the other hand, the diversity of nosZ changed in the latter half of the process, a change which might have been induced by the accumulation of nitrate and nitrite. The nirS gene fragments could not be detected. The use of mature compost that contains nitrate and nitrite promoted the N(2)O emission and significantly affected the variation of nosZ diversity in the initial stage of composting, but did not affect the variation of nirK diversity. Many Pseudomonas-like nirK and nosZ gene fragments were detected in the stage in which N(2)O was actively emitted.

Nitrogen removal by co-occurring methane oxidation, denitrification, aerobic ammonium oxidation, and anammox.

The pathway for removing NO(3)(-) and NH(4)(+) from wastewater in the presence of both CH(4) and O(2) was clarified by studying microbial activity and community. Batch incubation tests were performed to characterize the microbial activity of the sludge, which was acclimatized in a bioreactor in which O(2) and CH(4) were supplied to treat wastewater containing NO(3)(-) and NH(4)(+) . The tests showed that the sludge removed significant amounts of NO(3)(-) and NH(4)(+) in the presence of CH(4) and O(2), and the presence of the activity of methane oxidation, denitrification, nitrification, and anammox in the sludge. It was estimated that the total inorganic nitrogen removal was attributed to denitrification associated with methane oxidation as 53.4%, microbial assimilation as 37.9%, and anammox as 8.7%. Nitrification also contributed to NH(4)(+) decrease as 34.5% and anammox as 6.4%. Anammox activity was unambiguously demonstrated by (29)N(2) production in anaerobic batch incubation with (15)N-labeled inorganic nitrogen compounds. The presence of methane-oxidizing bacteria and candidate denitrifiers in the sludge was shown by denaturing gradient gel electrophoresis of 16S rRNA gene fragments. Clone library analysis of the PCR-amplified 16S rRNA gene fragment using specific primers for aerobic ammonium oxidizer and anammox revealed the presence of these bacteria. The results reveal that complex nitrogen-removal processes occur in the presence of CH(4) and O(2) by methanotroph, denitrifier, aerobic ammonium oxidizer, and anammox.

Evaluation of full-scale biofilter with rockwool mixture treating ammonia gas from livestock manure composting.

NH3 removal by a full-scale biofilter with rockwool packing materials was studied by measuring the gases and potential nitrification and denitrification activities of those materials in order to improve the biofiltration technology used in livestock farms. The rockwool biofilter was a durable and effective system for removing NH3, which was varied with the turning of manure composts. Furthermore, NH3 could be treated in the absence of an extra increase in two greenhouse gases, N2O and CH4. Potential nitrification and denitrification activities of the packing materials were estimated to be 8.2-12.2 mg N, and 1.42-4.69 mg N/100 g dry samples per day, respectively. The results suggested that potential nitrification and denitrification activities would increase within the biofilter where substrates, NH3 or NO3(-), have accumulated as a result of its operation. However, since percolate water contained high concentrations of NH4(+) and NO3(-), further improvement is required by reducing nitrogenous compounds within both the biofilter and percolate water.

Removal and recovery of phosphorous from swine wastewater by demonstration crystallization reactor and struvite accumulation device.

A demonstration crystallization reactor and struvite accumulation device for the removal and recovery of phosphorous was constructed and their performance was evaluated using actual swine wastewater for 3.5 years. The wastewater pH was increased by aeration, and the concentrations of total P and soluble PO(4)-P were reduced by a struvite crystallization reaction induced under a high pH condition. A 30% MgCl(2) addition was effective in enhancing the struvite crystallization reaction. The concentrations of suspended solids, total Zn and total Cu, were also decreased by the settling function of the reactor. On removing the efficiencies of these components, no noticeable seasonal fluctuation in performance was observed during the 3.5-year operation. In terms of maximum yield, 171g struvite was obtained from 1m(3) swine wastewater by the demonstration accumulation device for struvite recovery. The recovered struvite needed only air-drying before use since it was approximately 95% pure even without washing.

Reduction of nitrous oxide emission from pig manure composting by addition of nitrite-oxidizing bacteria.

Nitrous oxide (N2O) is emitted from pig manure composting, and the emission correlates with nitrite (NO2-) accumulation in the composting material. In the present study, we added nitrite-oxidizing bacteria (NOB)to inhibit NO2- accumulation and evaluated its effect on N2O emission in a laboratory-scale composting experiment. Mature pig manure compost (MPMC) containing NOB at 10(6) MPN g(-1) WM or cultured MPMC (cul-MPMC) NOB at 10(11) MPN g(-1) WM was added after the thermophilic phase of composting. The addition of these materials prevented NO2- accumulation, promoting oxidation to nitrate (NO3-), whereas the accumulation of NO2- occurred in the material to which NOB was not added as the result of the delayed growth of indigenous NOB compared with that of ammonia-oxidizing bacteria (AOB). The pattern of NO2- in the material agreed with that of N2O emission; therefore, N2O emission ceased rapidly when NOB was added. Emission rates of N2O were 88.5 (no addition), 17.5 (MPMC addition), and 20.2 (cul-MPMC addition) g N-N2O kg(-1) TNinitial, respectively. Improving composition of nitrifying communities for complete nitrification promotion would be useful to establish a composting method with low N2O emission.

Effect of addition of organic waste on reduction of Escherichia coli during cattle feces composting under high-moisture condition.

To ensure Escherichia coli reduction during cattle feces composting, co-composting with a variety of organic wastes was examined. A mixture of dairy cattle feces and shredded rice straw (control) was blended with organic wastes (tofu residue, rice bran, rapeseed meal, dried chicken feces, raw chicken feces, or garbage), and composted using a bench-scale composter under the high-moisture condition (78%). The addition of organic waste except chicken feces brought about maximum temperatures of more than 55 degrees C and significantly reduced the number of E. coli from 10(6) to below 10(2)CFU/g-wet after seven days composting, while in the control treatment, E. coli survived at the same level as that of raw feces. Enhancements of the thermophilic phase and E. coli reduction were related to the initial amount of easily digestible carbon in mass determined as BOD. BOD value more than 166.2 mg O2/DMg brought about significant E. coli reduction.