Microbiological parameters as indicators of compost maturity

AIMS: The objectives of this study were to determine the changes of microbial properties of pig manure collected from pens with different management strategies and composted using different turning and moisture regimes; relate their association with humification parameters and compost temperature; and identify the most suitable microbial indicators of compost maturity. METHODS AND RESULTS: Six different microbial parameters, including total bacterial count, oxygen consumption rate, ATP content, dehydrogenase activity, and microbial biomass C and N, along with humification parameters [humic acid (HA), fulvic acid (FA) and HA : FA ratio] and compost temperature were monitored during composting. Significant positive correlations were found between temperature and microbial properties, including O2 consumption rate, ATP content, dehydrogenase activity, and microbial biomass N. The humification parameters also showed significant correlations with microbial properties of the manure compost. For instance, HA contents of pig manures was positively correlated with total aerobic heterotrophs, and microbial biomass N and C; and negatively correlated with O2 consumption rate, ATP content, and dehydrogenase activity. Among the six microbial parameters examined, dehydrogenase activity was the most important factor affecting compost temperature and humification parameters. Composting strategies employed in this study affected the speed of composting and time of maturation. If the moisture content is maintained weekly at 60% with a 4-day turning frequency, the pig manure will reach maturity in 56 days. CONCLUSIONS: The composting process went through predictable changes in temperature, microbial properties and chemical components despite differences in the initial pig manure and composting strategies used. Among the six microbial parameters used, dehydrogenase activity is the most suitable indicator of compost maturity. Compared with respiration rate, ATP content and microbial biomass procedures, dehydrogenase activity is the simplest, quickest, and cheapest method that can be used to monitor the stability and maturity of composts. SIGNIFICANCE AND IMPACT OF THE STUDY: The results presented here show that microbial parameters can be used in revealing differences between composts and compost maturity. The statistical relationship established between humification parameters and microbial parameters, particularly dehydrogenase activity, demonstrates that it is possible to monitor the composting process more easily and rapidly by avoiding longer and more expensive analytical procedures.     

Comparison of signature lipid methods to determine microbial community structure in compost

The microbial community structure changes substantially during the composting process and simple methods to follow these changes can potentially be used to estimate compost maturity. In this study, two such methods, the microbial identification (MIDI) method and the ester-linked (EL) procedure to determine the composition of long-chain fatty acids, were applied to compost samples of different age. The ability of the two methods to describe the microbial succession was evaluated by comparison with phospholipid fatty acid (PLFA) analysis on the same samples.Samples were taken from a 200-l laboratory compost reactor, treating source-separated organic household waste. During the initial stages of the process, the total concentration of fatty acids in compost samples treated with the EL and MIDI methods was many times higher than with the PLFA method. This was probably due to the presence of fatty acids from the organic material in the original waste. However, this substantial difference between PLFA and the other two methods was not found later in composting. Although the PLFA method gave the most detailed information about the growth and overall succession of the microbial community, the much simpler MIDI and EL methods also successfully described the shift from the initially dominating straight chain fatty acids to iso- and anteiso branched, 10 Me branched and cyclopropane fatty acids in the later stages of the process. Thus, the MIDI and EL extraction methods appear to be suitable for analysis of microbial FAME profiles in compost, particularly in the later stages of the process.     

Comparison of characterization and microbial communities in rice straw- and wheat straw-based compost for <Emphasis Type="Italic">Agaricus bisporus</Emphasis> production

Rice straw (RS) is an important raw material for the preparation of Agaricus bisporus compost in China. In this study, the characterization of composting process from RS and wheat straw (WS) was compared for mushroom production. The results showed that the temperature in RS compost increased rapidly compared with WS compost, and the carbon (C)/nitrogen (N) ratio decreased quickly. The microbial changes during the Phase I and Phase II composting process were monitored using denaturing gradient gel electrophoresis (DGGE) and phospholipid fatty acid (PLFA) analysis. Bacteria were the dominant species during the process of composting and the bacterial community structure dramatically changed during heap composting according to the DGGE results. The bacterial community diversity of RS compost was abundant compared with WS compost at stages 4–5, but no distinct difference was observed after the controlled tunnel Phase II process. The total amount of PLFAs of RS compost, as an indicator of microbial biomass, was higher than that of WS. Clustering by DGGE and principal component analysis of the PLFA compositions revealed that there were differences in both the microbial population and community structure between RS- and WS-based composts. Our data indicated that composting of RS resulted in improved degradation and assimilation of breakdown products by A. bisporus, and suggested that the RS compost was effective for sustaining A. bisporus mushroom growth as well as conventional WS compost.     

Temporal and spatial changes in the microbial bioaerosol communities in green-waste composting

In this study, the microbial community within compost, emitted into the airstream, downwind and upwind from a composting facility was characterized and compared through phospholipid fatty acid analysis and 16S rRNA gene analysis using denaturing gradient gel electrophoresis and bar-coded pyrosequencing techniques. All methods used suggested that green-waste composting had a significant impact upon bioaerosol community composition. Daily variations of the on-site airborne community showed how specific site parameters such as compost process activity and meteorological conditions affect bioaerosol communities, although more data are required to qualify and quantify the causes for these variations. A notable feature was the dominance of Pseudomonas in downwind samples, suggesting that this genus can disperse downwind in elevated abundances. Thirty-nine phylotypes were homologous to plant or human phylotypes containing pathogens and were found within compost, on-site and downwind microbial communities. Although the significance of this finding in terms of potential health impact was beyond the scope of this study, it clearly illustrated the potential of molecular techniques to improve our understanding of the impact that green-waste composting emissions may have on the human health.     

Enzyme production-based approach for determining the functions of microorganisms within a community

The functions of specific microorganisms in a microbial community were investigated during the composting process. Cerasibacillus quisquiliarum strain BLx(T) and Bacillus thermoamylovorans strain BTa were isolated and characterized in our previous studies based on their dominance in the composting system. Strain BLx(T) degrades gelatin, while strain BTa degrades starch. We hypothesized that these strains play roles in gelatinase and amylase production, respectively. The relationship between changes in the abundance ratios of each strain and those of each enzyme activity during the composting process was examined to address this hypothesis. The increase in gelatinase activity in the compost followed a dramatic increase in the abundance ratio of strain BLx(T). Zymograph analysis demonstrated that the pattern of active gelatinase bands from strain BLx(T) was similar to that from the compost. Gelatinases from both BLx(T) and compost were partially purified and compared. Homologous N-terminal amino acid sequences were found in one of the gelatinases from strain BLx(T) and that of compost. These results indicate strain BLx(T) produces gelatinases during the composting process. Meanwhile, the increase in the abundance ratio of strain BTa was not concurrent with that of amylase activity in the compost. Moreover, the amylase activity pattern of strain BTa on the zymogram was different from that of the compost sample. These results imply that strain BTa may not produce amylases during the composting process. To our knowledge, this is the first report demonstrating that the function of a specific microorganism is directly linked to a function in the community, as determined by culture-independent and enzyme-level approaches.     

Investigation of the microbial community structure and activity as indicators of compost stability and composting process evolution

In a bid to identify suitable microbial indicators of compost stability, the process evolution during windrow composting of poultry manure (PM), green waste (GW) and biowaste was studied. Treatments were monitored with regard to abiotic factors, respiration activity (determined using the SOUR test) and functional microflora. The composting process went through typical changes in temperature, moisture content and microbial properties, despite the inherent feedstock differences. Nitrobacter and pathogen indicators varied as a monotonous function of processing time. Some microbial groups have shown a potential to serve as fingerprints of the different process stages, but still they should be examined in context with respirometric tests and abiotic parameters. Respiration activity reflected well the process stage, verifying the value of respirometric tests to access compost stability. SOUR values below 1 mg O₂/g VS/h were achieved for the PM and the GW compost.     

Investigating microbial activities in compost using mushroom (Agaricus bisporus) cultivation as an experimental system

Investigations into the dynamic nature of composting environments are necessary to understand and ultimately optimise the complex processes that occur. In this study, various parameters were measured to investigate physical, chemical and biological changes that occur in compost during the production of Agaricus bisporus. In addition to monitoring the compost samples during mushroom cultivation, uninoculated samples were maintained for comparative purposes. Principal components analysis of the variables measured showed a clear distinction between the thermophilic Phase I composts, uninoculated Phase II composts and mushroom inoculated composts. Leucine assimilation, a novel technique to composting environments, is presented as suitable method for assessing microbial activity in such systems. Strict agreement between leucine assimilation and fluorescein diacetate (FDA) hydrolysis, a rarely used technique in composting environments, was not observed, suggesting that neither should be used as a sole measure of microbial activity in compost. Association of FDA hydrolysis with the culturable heterotrophic count suggests that FDA hydrolysis may indicate bacterial as opposed to total microbial activity.     

A statistical method for the analysis of nonlinear temperature time series from compost

Temperature is widely accepted as a critical indicator of aerobic microbial activity during composting but, to date, little effort has been made to devise an appropriate statistical approach for the analysis of temperature time series. Nonlinear, time-correlated effects have not previously been considered in the statistical analysis of temperature data from composting, despite their importance and the ubiquity of such features. A novel mathematical model is proposed here, based on a modified Gompertz function, which includes nonlinear, time-correlated effects. Methods are shown to estimate initial values for the model parameter. Algorithms in SAS are used to fit the model to different sets of temperature data from passively aerated compost. Methods are then shown for testing the goodness-of-fit of the model to data. Next, a method is described to determine, in a statistically rigorous manner, the significance of differences among the time-correlated characteristics of the datasets as described using the proposed model. An extra-sum-of-squares method was selected for this purpose. Finally, the model and methods are used to analyze a sample dataset and are shown to be useful tools for the statistical comparison of temperature data in composting.     

Enzyme Production-Based Approach for Determining the Functions of Microorganisms within a Community

The functions of specific microorganisms in a microbial community were investigated during the composting process. Cerasibacillus quisquiliarum strain BLx^T and Bacillus thermoamylovorans strain BTa were isolated and characterized in our previous studies based on their dominance in the composting system. Strain BLx^T degrades gelatin, while strain BTa degrades starch. We hypothesized that these strains play roles in gelatinase and amylase production, respectively. The relationship between changes in the abundance ratios of each strain and those of each enzyme activity during the composting process was examined to address this hypothesis. The increase in gelatinase activity in the compost followed a dramatic increase in the abundance ratio of strain BLx^T. Zymograph analysis demonstrated that the pattern of active gelatinase bands from strain BLx^T was similar to that from the compost. Gelatinases from both BLx^T and compost were partially purified and compared. Homologous N-terminal amino acid sequences were found in one of the gelatinases from strain BLx^T and that of compost. These results indicate strain BLx^T produces gelatinases during the composting process. Meanwhile, the increase in the abundance ratio of strain BTa was not concurrent with that of amylase activity in the compost. Moreover, the amylase activity pattern of strain BTa on the zymogram was different from that of the compost sample. These results imply that strain BTa may not produce amylases during the composting process. To our knowledge, this is the first report demonstrating that the function of a specific microorganism is directly linked to a function in the community, as determined by culture-independent and enzyme-level approaches.     

DGGE and T-RFLP Analysis of Bacterial Succession during Mushroom Compost Production and Sequence-aided T-RFLP Profile of Mature Compost

The amount of button mushroom (Agaricus bisporus) harvested from compost is largely affected by the microbial processes taking place during composting and the microbes inhabiting the mature compost. In this study, the microbial changes during the stages of this specific composting process were monitored, and the dominant bacteria of the mature compost were identified to reveal the microbiological background of the favorable properties of the heat-treated phase II mushroom compost. 16S ribosomal deoxyribonucleic acid (rDNA)-based denaturing gradient gel electrophoresis (DGGE) and terminal restriction fragment length polymorphism (T-RFLP) molecular fingerprinting methods were used to track the succession of microbial communities in summer and winter composting cycles. DNA from individual DGGE bands were reamplified and subjected to sequence analysis. Principal component analysis of fingerprints of the composting processes showed intensive changes in bacterial community during the 22-day procedure. Peak temperature samples grouped together and were dominated by Thermus thermophilus. Mature compost patterns were almost identical by both methods (DGGE, T-RFLP). To get an in-depth analysis of the mature compost bacterial community, the sequence data from cultivation of the bacteria and cloning of environmental 16S rDNA were uniquely coupled with the output of the environmental T-RFLP fingerprints (sequence-aided T-RFLP). This method revealed the dominance of a supposedly cellulose-degrading consortium composed of phylotypes related to Pseudoxanthomonas, Thermobifida, and Thermomonospora.     

Assessing the efficiency of urban waste biocomposting by analytical pyrolysis (Py-GC/MS)

Analytical pyrolysis (Py-GC/MS) was used to study complex composting processes. The technique was first validated for reproducibility and finally applied to assess the efficiency of a microbial bio-accelerator product (CBB) in composting organic residues with different composition. Fresh lignocellulosic and urban wastes were treated with CBB and the composting kinetics studied to investigate the transformations undergone in the course of biocomposting. Our findings demonstrate that these changes, as well as the efficiency of CBB, can be monitored through the molecular characterization of the released pyrolysis products. The CBB bacterial product effectively seems to favour composting accelerating the process and shorten composting time. Analytical pyrolysis was informative in assessing to which extent compost transformation reached an acceptable stabilization point. The technique could be also developed into a semi-quantitative tool to monitor changes of main organic matter components (polysaccharides, proteins, lignin, lipids, etc.) as composting proceeds.     

Microbial indicators for sawdust and bark compost stability and humification processes

To what extent some microbial index ratios are suitable for use as early criteria for the level of compost stability during aerobic composting of coniferous sawdust and bark at mesophilic conditions was studied. Evolution of the specific respiration activity (CO₂-C/biomass C) and the ratios between some groups of microorganisms were followed as a function of composting time. The specific respiration activity was found to be an early and most reliable indicator of compost stability. The peroxidase and polyphenoloxidase enzyme activity during composting, as well as the composition of newly-formed humus substances were studied. The duration of composting increased the quality of newly-formed humus substances (C_h.a.:C_f.a ratio; Ca-complexed humic acid and resistance of organo-mineral complexes). The quality of humus substances could be used to assess compost stability. However, the results can be applied only under defined conditions.     

Microbiological aspects of biowaste during composting in a monitored compost bin

AIMS: To determine the microbial succession of the dominating taxa and functional groups of microorganisms and the total microbial activity during the composting of biowaste in a monitored process. METHODS AND RESULTS: Biowaste (vegetable, fruit and garden waste) was composted in a monitored composting bin system. During the process, taxonomic and functional subpopulations of microorganisms were enumerated, and dominating colonies were isolated and identified. All counts decreased during the thermophilic phase of the composting, but increased again when the temperature declined. Total microbial activity, measured with an enzyme activity assay, decreased during the thermophilic phase, increased substantially thereafter, and decreased again during maturation. Bacteria dominated during the thermophilic phase while fungi, streptomycetes and yeasts were below the detection limit. Different bacterial populations were found in the thermophilic and mesophilic phases. In fresh wastes and during the peak-heating phase, all bacterial isolates were bacilli. During the cooling and maturation phase the bacterial diversity increased, including also other Gram-positive and Gram-negative bacteria. Among the fungi, Aspergillus spp. and Mucor spp. were predominant after the thermophilic phase. CONCLUSIONS: The microbial abundance, composition and activity changed substantially during composting and compost maturity was correlated with high microbial diversity and low activity. SIGNIFICANCE AND IMPACT OF THE STUDY: A more complete overview of the whole composting process of biowaste, based on microbial counts, species diversity and functional groups and abiotic parameters is presented, and the potential of a simple enzyme assay to measure total microbial activity was demonstrated.     

Effect of bioaugmentation and nitrogen supplementation on composting of paddy straw

A composting experiment was conducted to evaluate the effect of a hyperlignocellulolytic fungal consortium and different nitrogen amendments on paddy straw composting in terms of changes in physicochemical and biological parameters. A fungal consortium comprising four lignocellulolytic mesophilic fungal cultures was used as inoculum for bioaugmentation of paddy straw in perforated pits. The comparative effect of farmyard manure (FYM), soybean trash, poultry litter and urea on the composting process was evaluated at monthly intervals in terms of physicochemical (pH, EC, available P, C:N ratio and humus content) and biological (enzymatic and microbial activity) parameters. The compost prepared from bioaugmented paddy straw composting mixture, with poultry manure as nitrogen supplement attained desirable C:N ratio in 1 month and displayed least phytotoxicity levels along with higher production of β-1,4-Exoglucanase. The combined activity of the autochthonous composting microbiota as well as the externally applied fungal inoculum accelerated the composting process of paddy straw. Supplementation of paddy straw with poultry manure in 8:1 ratio was identified as the best treatment to hasten the composting process. This study highlights the importance of application of fungal inoculum and an appropriate N-amendment such as poultry manure for preparation of compost using a substrate having high C:N ratio, such as paddy straw.     

高通量测序技术分析猪粪堆肥过程中微生物群落结构变化

为了解猪粪堆肥过程中微生物群落结构组成及多样性的变化,采集猪粪堆肥过程的三个代表性样品—新鲜猪粪、高温堆肥、腐熟堆肥,利用Illumina Miseq高通量测序技术对16S rRNA V4～V5可变区序列进行测序,分别获得37 009、42 470、36 713条有效序列及328、280、160个操作分类单元(OTU)。Alpha多样性分析表明,在堆肥过程中微生物群落丰富度呈现降低趋势,而多样性呈现先上升后下降趋势。随着堆肥的进行,在门水平上,厚壁菌门、拟杆菌门和软壁菌门相对丰度降低,而变形菌门和放线菌门相对丰度升高;在属水平上,Turicibacter、Terrisporobacter、Parabacteroides、Clostridium sensu stricto、Corynebacterium等来自动物肠道的微生物相对丰度明显下降,Thermopolyspora、Thermomonospora、Thermobifida、Halocella等耐热耐盐微生物成为最主要优势菌。堆肥过程不同菌群优势度的变化是微生物与堆肥中各理化因子相互作用的结果。     

Changes in Bacterial and Fungal Communities across Compost Recipes,Preparation Methods,and Composting Times

Compost production is a critical component of organic waste handling, and compost applications to soil are increasingly important to crop production. However, we know surprisingly little about the microbial communities involved in the composting process and the factors shaping compost microbial dynamics. Here, we used high-throughput sequencing approaches to assess the diversity and composition of both bacterial and fungal communities in compost produced at a commercial-scale. Bacterial and fungal communities responded to both compost recipe and composting method. Specifically, bacterial communities in manure and hay recipes contained greater relative abundances of Firmicutes than hardwood recipes with hay recipes containing relatively more Actinobacteria and Gemmatimonadetes. In contrast, hardwood recipes contained a large relative abundance of Acidobacteria and Chloroflexi. Fungal communities of compost from a mixture of dairy manure and silage-based bedding were distinguished by a greater relative abundance of Pezizomycetes and Microascales. Hay recipes uniquely contained abundant Epicoccum, Thermomyces, Eurotium, Arthrobotrys, and Myriococcum. Hardwood recipes contained relatively abundant Sordariomycetes. Holding recipe constant, there were significantly different bacterial and fungal communities when the composting process was managed by windrow, aerated static pile, or vermicompost. Temporal dynamics of the composting process followed known patterns of degradative succession in herbivore manure. The initial community was dominated by Phycomycetes, followed by Ascomycota and finally Basidiomycota. Zygomycota were associated more with manure-silage and hay than hardwood composts. Most commercial composters focus on the thermophilic phase as an economic means to insure sanitation of compost from pathogens. However, the community succeeding the thermophilic phase begs further investigation to determine how the microbial dynamics observed here can be best managed to generate compost with the desired properties.     

Characterization of dairy cattle manure/wallboard paper compost mixture

The aim of this research was to evaluate the use of manufacturing wallboard paper scraps as an alternative bulking agent for dairy cattle manure composting. The characteristics of the composting process were studied based on the changes in physico-chemical parameters and final compost quality. Composting of dairy cattle manure with wallboard paper was performed in a 481-L cylindrical reactor with vacuum-type aeration. Rapid degradation of organic matter was observed during the thermophilic stage of composting due to high microbial activity. High temperature and alkaline pH conditions promoted intense ammonia emission during the early stage of composting. The number of mesophilic and thermophilic microorganisms were found to be affected by changes in temperature at different composting stages. The total nitrogen (N), phosphorus (P), potassium (K), and sodium (Na) concentrations of the mixture did not change significantly after 28days of composting. However, the presence of gypsum in the paper scraps increased the calcium content of the final compost. The wallboard paper had no phyto-inhibitory effects as shown by high germination index of final compost (GI=99%).     

Estimation of maturity of compost from food wastes and agro-residues by multiple regression analysis

The composting process of food wastes and tree cuttings was examined on four composting types composed from two kinds of systems and added mixture of microorganisms. The time courses of 32 parameters in each composting type were observed. The efficient composting system was found to be the static aerated reactor system in comparison with the turning pile one. Using the multiple regression analysis of all the data (159 samples) obtained from this study, some parameters were selected to predict the germination index (GI) value, which was adopted as a marker of compost maturity. For example, using the regression model generated from pH, NH(4)(+) concentration, acid phosphatase activity, and esterase activity of water extracts of the compost, GI value was expressed by the multi-linear regression equation (p<0.0001). High correlations between the measured GI value and the predicted one were made in each type of compost. As a result of these observations, the compost maturity might be predicted by only sensing of the water extract at the composting site without any requirements for a large-size equipment and skill, and this prediction system could contribute to the production of a stable compost in wide-spread use for the recycling market.     

Universal ligation-detection-reaction microarray applied for compost microbes

Background  

Composting is one of the methods utilised in recycling organic communal waste. The composting process is dependent on aerobic microbial activity and proceeds through a succession of different phases each dominated by certain microorganisms. In this study, a ligation-detection-reaction (LDR) based microarray method was adapted for species-level detection of compost microbes characteristic of each stage of the composting process. LDR utilises the specifiCity of the ligase enzyme to covalently join two adjacently hybridised probes. A zip-oligo is attached to the 3'-end of one probe and fluorescent label to the 5'-end of the other probe. Upon ligation, the probes are combined in the same molecule and can be detected in a specific location on a universal microarray with complementary zip-oligos enabling equivalent hybridisation conditions for all probes. The method was applied to samples from Nordic composting facilities after testing and optimisation with fungal pure cultures and environmental clones.     

Physical and Chemical Correlates of Microbial Activity and Biomass in Composting Municipal Sewage Sludge

Various physical and chemical parameters were monitored to evaluate their influence on the microbial communities present in composting municipal sewage sludge. Temperature, moisture content, depth, pH, protein content, total nitrogen, total carbon, lipid phosphate biomass, and the rates of microbial incorporation of substrates into lipids were measured at several times throughout the 17- to 19-day composting runs. Temperature was found to have the most consistent and dramatic effect on microbial activity and biomass. When temperatures exceeded 55 to 60°C, microbial activity fell dramatically, usually by more than 1 order of magnitude. Microbial activity was generally greatest in samples taken from the 35 to 50°C areas of the composting piles. Changes in the composition of the compost over time included increased pH, increased protein content, and decreased total organic content. The changes in these parameters appeared to reflect the microbial activity and biomass present. The results of this study indicate that the rate of composting may best be optimized by controlling the composting temperatures, provided that the other parameters fall within reasonable limits in the starting material.