Microbial diversity of methanogenic communities in the systems for anaerobic treatment of organic waste

Methane production via anaerobic degradation of organic-contaminated wastewater, semiliquid, or solid municipal waste of complex composition by methanogenic microbial communities is a multistage process involving at least four groups of microorganisms. These are hydrolytic bacteria (polysaccharolytic, proteolytic, and lipolytic), fermentative bacteria, acetogenic bacteria (syntrophic, proton-reducing), and methanogenic archaea; complex trophic interactions exist between these groups. The review provides information concerning the diversity of the major microbial groups identified in the systems for wastewater and concentrated waste treatment, solid-phase anaerobic fermentation, and landfills for disposal of municipal solid waste, and also specifies the sources of isolation of the type strains. The research demonstrates that both new microorganisms and those previously isolated from natural habitats may be found in waste treatment systems. High microbial diversity in the systems for organic waste treatment provides for stable methanogenesis under fluctuating environmental conditions.     

Distribution and activity of microorganisms in the deep repository for liquid radioactive waste at the Siberian Chemical Combine

The physicochemical conditions, composition of microbial communities, and the rates of anaerobic processes in the deep sandy horizons used as a repository for liquid radioactive wastes (LRW) at the Siberian Chemical Combine (Seversk, Tomsk oblast), were studied. Formation waters from the observation wells drilled into the production horizons of the radioactive waste disposal site were found to be inhabited by microorganisms of different physiological groups, including aerobic organotrophs, anaerobic fermentative, denitrifying, sulfate-reducing, and methanogenic bacteria. The density of microbial population, as determined by cultural methods, was low and usually did not exceed 10(4) cells/ml. Enrichment cultures of microorganisms producing gases (hydrogen, methane, carbon dioxide, and hydrogen sulfide) and capable of participation in the precipitation of metal sulfides were obtained from the waters of production horizons. The contemporary processes of sulfate reduction and methanogenesis were assayed; the rates of these terminal processes of organic matter destruction were found to be low. The denitrifying bacteria from the underground repository were capable of reducing the nitrates contained in the wastes, provided sources of energy and biogenic elements were available. Biosorption of radionuclides by the biomass of aerobic bacteria isolated from groundwater was demonstrated. The results obtained give us insight into the functional structure of the microbial community inhabiting the waters of repository production horizons. This study indicates that the numbers and activity of microbial cells are low both inside and outside the zone of radioactive waste dispersion, in spite of the long period of waste discharge.     

Enzyme research and applications in biotechnological intensification of biogas production

Biogas technology provides an alternative source of energy to fossil fuels in many parts of the world. Using local resources such as agricultural crop remains, municipal solid wastes, market wastes and animal waste, energy (biogas), and manure are derived by anaerobic digestion. The hydrolysis process, where the complex insoluble organic materials are hydrolysed by extracellular enzymes, is a rate-limiting step for anaerobic digestion of high-solid organic solid wastes. Biomass pretreatment and hydrolysis are areas in need of drastic improvement for economic production of biogas from complex organic matter such as lignocellulosic material and sewage sludge. Despite development of pretreatment techniques, sugar release from complex biomass still remains an expensive and slow step, perhaps the most critical in the overall process. This paper gives an updated review of the biotechnological advances to improve biogas production by microbial enzymatic hydrolysis of different complex organic matter for converting them into fermentable structures. A number of authors have reported significant improvement in biogas production when crude and commercial enzymes are used in the pretreatment of complex organic matter. There have been studies on the improvement of biogas production from lignocellulolytic materials, one of the largest and renewable sources of energy on earth, after pretreatment with cellulases and cellulase-producing microorganisms. Lipids (characterised as oil, grease, fat, and free long chain fatty acids, LCFA) are a major organic compound in wastewater generated from the food processing industries and have been considered very difficult to convert into biogas. Improved methane yield has been reported in the literature when these lipid-rich wastewaters are pretreated with lipases and lipase-producing microorganisms. The enzymatic treatment of mixed sludge by added enzymes prior to anaerobic digestion has been shown to result in improved degradation of the sludge and an increase in methane production. Strategies for enzyme dosing to enhance anaerobic digestion of the different complex organic rich materials have been investigated. This review also highlights the various challenges and opportunities that exist to improve enzymatic hydrolysis of complex organic matter for biogas production. The arguments in favor of enzymes to pretreat complex biomass are compelling. The high cost of commercial enzyme production, however, still limits application of enzymatic hydrolysis in full-scale biogas production plants, although production of low-cost enzymes and genetic engineering are addressing this issue.     

Enzyme research and applications in biotechnological intensification of biogas production

Biogas technology provides an alternative source of energy to fossil fuels in many parts of the world. Using local resources such as agricultural crop remains, municipal solid wastes, market wastes and animal waste, energy (biogas), and manure are derived by anaerobic digestion. The hydrolysis process, where the complex insoluble organic materials are hydrolysed by extracellular enzymes, is a rate-limiting step for anaerobic digestion of high-solid organic solid wastes. Biomass pretreatment and hydrolysis are areas in need of drastic improvement for economic production of biogas from complex organic matter such as lignocellulosic material and sewage sludge. Despite development of pretreatment techniques, sugar release from complex biomass still remains an expensive and slow step, perhaps the most critical in the overall process. This paper gives an updated review of the biotechnological advances to improve biogas production by microbial enzymatic hydrolysis of different complex organic matter for converting them into fermentable structures. A number of authors have reported significant improvement in biogas production when crude and commercial enzymes are used in the pretreatment of complex organic matter. There have been studies on the improvement of biogas production from lignocellulolytic materials, one of the largest and renewable sources of energy on earth, after pretreatment with cellulases and cellulase-producing microorganisms. Lipids (characterised as oil, grease, fat, and free long chain fatty acids, LCFA) are a major organic compound in wastewater generated from the food processing industries and have been considered very difficult to convert into biogas. Improved methane yield has been reported in the literature when these lipid-rich wastewaters are pretreated with lipases and lipase-producing microorganisms. The enzymatic treatment of mixed sludge by added enzymes prior to anaerobic digestion has been shown to result in improved degradation of the sludge and an increase in methane production. Strategies for enzyme dosing to enhance anaerobic digestion of the different complex organic rich materials have been investigated. This review also highlights the various challenges and opportunities that exist to improve enzymatic hydrolysis of complex organic matter for biogas production. The arguments in favor of enzymes to pretreat complex biomass are compelling. The high cost of commercial enzyme production, however, still limits application of enzymatic hydrolysis in full-scale biogas production plants, although production of low-cost enzymes and genetic engineering are addressing this issue.     

Perspectives and predictions on the microbial ecology of the hyporheic zone

1. Studies of hyporheic microbial ecology have suggested an important role for hyporheic microbial processes in stream ecosystem functioning. Using evidence from microbial communities in other aquatic habitats, some predictions are made concerning the diversity of microbial types and microbial processes likely to occur in the hyporheic zone, and the relative importance of these various types to the hyporheic ecosystem. 2. It is predicted that the biofilm growth form of interstitial micro-organisms will create a variety of microniches, allowing coexistence of a great diversity of microbial types, and promoting the activity of some otherwise poor competitors. It is further predicted that the confluence of reduced groundwaters and aerobic surface waters will favour chemolithotrophic processes in the hyporheic zone, but that these will contribute significantly to hyporheic production only if surface water is very low in dissolved organic carbon, or the groundwater is extremely reduced, such as by the influence of riparian wetlands. A variety of anaerobic respiratory pathways, such as nitrate, ferric ion, sulphate and even methanogenic respiration will be employed in the hyporheic zone, with biofilm dynamics permitting these to occur even in aerobic sediments. Anaerobic pathways may account for a significant proportion of total hyporheic organic matter mineralization. 3. The role of fungi in hyporheic dynamics is, as yet, almost completely unstudied. However, it is expected that they will be important in breaking down buried particulate organic matter (POM), which may account for a large proportion of total stream POM. 4. Physicochemical conditions in hyporheic sediments appear to be highly heterogeneous, and this heterogeneity may be very important in the cycling of certain nutrients, especially nitrogen, which involves a series of steps requiring different conditions. 5. Various new techniques are now available by which biofilm dynamics and in situ microbial processes may be measured. Studies are recommended of intact microbial communities both at the microscale of the biofilm and at the scale of the heterogeneities occurring in hyporheic sediments. Studies are needed that measure actual rates of microbial processes under in situ conditions.     

Distribution and activity of microorganisms in the deep repository for liquid radioactive waste at the Siberian Chemical Combine

The physicochemical conditions, composition of microbial communities, and the rates of anaerobic processes in the deep sand horizons used as a repository for liquid radioactive wastes (LRW) at the Siberian Chemical Combine (Seversk, Tomsk oblast), were studied. Formation waters from the observation wells drilled into the horizons used for the radioactive waste disposal were found to be inhabited by microorganisms of different physiological groups, including aerobic organotrophs, anaerobic fermentative, denitrifying, sulfate-reducing, and methanogenic bacteria. The density of microbial population, as determined by cultural methods, was low and usually did not exceed 10⁴ cells/ml. Enrichment cultures of microorganisms producing gases (hydrogen, methane, carbon dioxide, and hydrogen sulfide) and capable of participation in the precipitation of metal sulfides were obtained from the waters of the disposal site. The contemporary processes of sulfate reduction and methanogenesis were assayed; the rates of these terminal processes of organic matter destruction were found to be low. The denitrifying bacteria from the deep repository were capable of reducing the nitrates contained in the wastes, provided sources of energy and biogenic elements were available. Biosorption of radionuclides by the biomass of aerobic bacteria isolated from groundwater was demonstrated. The results obtained give us insight into the functional structure of the microbial community inhabiting the waters of repository horizons. This study indicates that the numbers and activity of microbial cells are low both inside and outside the zone of radioactive waste dispersion, in spite of the long period of waste discharge.     

Thermophilic anaerobes in Arctic marine sediments induced to mineralize complex organic matter at high temperature

Marine sediments harbour diverse populations of dormant thermophilic bacterial spores that become active in sediment incubation experiments at much higher than in situ temperature. This response was investigated in the presence of natural complex organic matter in sediments of two Arctic fjords, as well as with the addition of freeze‐dried Spirulina or individual high‐molecular‐weight polysaccharides. During 50°C incubation experiments, Arctic thermophiles catalysed extensive mineralization of the organic matter via extracellular enzymatic hydrolysis, fermentation and sulfate reduction. This high temperature‐induced food chain mirrors sediment microbial processes occurring at cold in situ temperatures (near 0°C), yet it is catalysed by a completely different set of microorganisms. Using sulfate reduction rates (SRR) as a proxy for organic matter mineralization showed that differences in organic matter reactivity determined the extent of the thermophilic response. Fjord sediments with higher in situ SRR also supported higher SRR at 50°C. Amendment with Spirulina significantly increased volatile fatty acids production and SRR relative to unamended sediment in 50°C incubations. Spirulina amendment also revealed temporally distinct sulfate reduction phases, consistent with 16S rRNA clone library detection of multiple thermophilic Desulfotomaculum spp. enriched at 50°C. Incubations with four different fluorescently labelled polysaccharides at 4°C and 50°C showed that the thermophilic population in Arctic sediments produce a different suite of polymer‐hydrolysing enzymes than those used in situ by the cold‐adapted microbial community. Over time, dormant marine microorganisms like these are buried in marine sediments and might eventually encounter warmer conditions that favour their activation. Distinct enzymatic capacities for organic polymer degradation could allow specific heterotrophic populations like these to play a role in sustaining microbial metabolism in the deep, warm, marine biosphere.     

Microbial transformation of 2,4,6-trinitrotoluene

The manufacture and decommissioning of explosives has generated, and continues to generate, large quantities of waste material whose primary toxic and mutagenic component is 2,4,6-trinitrotoluene (TNT). The magnitude of this problem has motivated a great deal of research into treatment processes and environmental fate studies, including characterization of microbial transformations of TNT. This work has encompassed studies with mixed cultures and pure cultures of microorganisms derived from either TNT-exposed or unexposed sources, and studies using microorganisms chosen for their known capacities to degrade other pollutants. Several of these studies are discussed with regard to whether they identified a process that may lead to the complete detoxification or mineralization of TNT. Since oxygen can have a significant influence on the types of biochemical reactions that can occur and on the oxidation of intermediates of TNT transformation processes, studies in which oxygen was not excluded are discussed separately from studies conducted under anaerobic conditions. Received 31 October 1995/ Accepted in revised form 29 March 1996     

海洋沉积物微生物介导有机碳转化研究进展

海洋沉积物是地球上最大的有机碳库,其中生存的微生物总量大、分布范围广、类群多样、代谢方式复杂,并共同构成海洋沉积物微生物组。海洋沉积物微生物组介导的有机碳降解与矿化过程不但能为沉积物中的生命活动提供物质和能量,也能参与调控碳循环过程,并在长时间尺度上对地球气候系统产生重大影响。沉积物中的有机碳在复杂多样的微生物代谢活动下被逐步降解,其最终的矿化过程与不同的电子受体消耗相偶合,并形成对应的地球化学分区。研究海洋沉积物微生物及其介导的有机碳转化过程对我们深入认识沉积物中的元素循环过程,并进一步评估其对整个地球系统的影响具有重要科学意义。本文对海洋沉积物微生物组的体量、包含的微生物多样性、代谢活性以及在不同地球化学分区中主要的微生物类群和代谢机制进行综述,最后基于研究现状展望了海洋沉积物微生物组的未来研究方向。     

典型有机固废厌氧消化微生物研究现状与发展方向

经过人工富集和驯化的兼性和严格厌氧微生物是厌氧消化工艺的核心。不同厌氧消化体系中存在的问题大多可以通过改变微生物群落的代谢活性来得到有效改善。得益于微生物组学检测技术的快速发展,对厌氧消化系统中微生物多样性的认识获得了极大的拓展,同时在微生物类群间、微生物与环境的互作关系研究方面也取得了一系列新的进展。然而,有机固废厌氧消化系统中,各种微生物以及微生物和物质的相互作用构成了更为复杂的代谢网络,所以目前对这些互作关系的解析尚不完善。本文重点关注了厌氧消化过程中的典型菌群互作关系,阐述了典型有机固废厌氧消化系统中存在的问题及微生物在其中发挥的作用,最后,立足于现有组学技术推动的微生物组研究进展,对未来有机固废厌氧消化系统微生物组的研究提出展望。     

The mass—Energy balance of anaerobic methane production

A theory was developed for the mass and energy balance of microbial processes, with special reference to the anaerobic production of methane. Interrelations of the bioengineering parameters of the process were delineated substrate quality, biodegrability and biological effciency of anaerobic processing of complex organic waste substrates. Application of the method is demonstrated on practical examples.     

Ratio [13C]/[12C] as an index for express estimation of hydrocarbon-oxidizing potential of microbiota in soil polluted with crude oil

The hydrocarbon-oxidizing potential of soil microbiota and hydrocarbon-oxidizing microorganisms introduced into soil was studied based on the quantitative and isotopic characteristics of carbon in products formed in microbial degradation of oil hydrocarbons. Comparison of CO2 production rates in native soil and that polluted with crude oil showed the intensity of microbial mineralization of soil organic matter (SOM) in the presence of oil hydrocarbons to be higher as compared with non-polluted soil, that is, revealed a priming effect ofoil. The amount of carbon of newly synthesized organic products (cell biomass and exometabolites) due to consumed petroleum was shown to significantly exceed that of SOM consumed for production of CO2. The result of microbial processes in oil-polluted soil was found to be a potent release of carbon dioxide to the atmosphere.     

Structural and dynamic features of microbial complexes of forest-swamp ecosystems in West Siberia

In deep peat soils of forest-swamp ecosystems of West Siberia, the structural, dynamic and functional features of microbial complexes are estimated. Data on biological and chemical activity of peat deposits in the context of processes of organic matter decomposition and humus accumulation are presented. Quantitative indicators of microbiological mineralization and soil oligotrophicity are given, and their coefficients are calculated. The pool of microorganisms of a 8 meter thick layer is calculated for estimating the degree to which the forest-swamp soils are enriched with microflora.     

Microbial communities in anaerobic digestion processes for waste and wastewater treatment: a microbiological update

Anaerobic digestion technology is the biological treatment of organic waste and wastewater without input of external electron acceptors (oxygen), offering the potential to reduce treatment cost and to produce energy as 'biogas' (methane) from organic waste. The technology has become enormously popular in the past two decades, and knowledge of microbiological aspects of the technology has also accumulated significantly. Major advances have been made in elucidating the diversity of yet-to-be cultured microbes in anaerobic digestion processes, and the cultivation of uncultured organisms is of great interest with regard to gaining insights into the function of these organisms. In addition, recent advances have been made in the development of microbial fuel cells as an alternative, direct energy-yielding treatment system.     

奶牛粪酸臭成分降解过程中的微生物群落变化

【目的】研究可降解成年泌乳奶牛粪中主要酸臭物的微生物群落的组成及动态变化。【方法】利用牛粪堆肥环境中的微生物进行了发酵优化、菌种驯化以及酸臭有机物降解规律的研究,结合r DNA高通量测序技术对有益微生物的组成及相对生物量进行了分析。【结果】实验发现,奶牛排泄物中的臭味来源主要为短链有机酸,堆肥自然环境中的微生物可以有效地对有机酸等污染物进行去除,经从低到高浓度的有机酸臭物(W/V,0.1%–0.2%)驯化发酵后,培养物中原核微生物以芽孢杆菌居多,而真核微生物主要由红曲霉及粉状毕赤酵母组成。【结论】进一步推测这几种微生物是耐受并降解有机酸臭物的优势微生物,可以应用于奶牛养殖过程中酸臭排泄物的生物控制。     

Effects of plant species diversity, atmospheric [CO2], and N addition on gross rates of inorganic N release from soil organic matter

A significant challenge in predicting terrestrial ecosystem response to global changes comes from the relatively poor understanding of the processes that control pools and fluxes of plant nutrients in soil. In addition, individual global changes are often studied in isolation, despite the potential for interactive effects among them on ecosystem processes. We studied the response of gross N mineralization and microbial respiration after 6 years of application of three global change factors in a grassland field experiment in central Minnesota (the BioCON experiment). BioCON is a factorial manipulation of plant species diversity (1, 4, 9 and 16 prairie species), atmospheric [CO₂] (ambient and elevated: 560 μmol mol^?1), and N inputs (ambient and ambient +4 g N m^?2 yr^?1). We hypothesized that gross N mineralization would increase with increasing levels of all factors because of stimulated plant productivity and thus greater organic inputs to soils. However, we also hypothesized that N addition would enhance, while elevated [CO₂] and greater diversity would temper, gross N mineralization responses because of increased and reduced plant tissue N concentrations, respectively. In partial support of our hypothesis, gross N mineralization increased with greater diversity and N addition, but not with elevated [CO₂]. The ratio of gross N mineralization to microbial respiration (i.e. the ‘yield’ of inorganic N mineralized per unit C respired) declined with greater diversity and [CO₂] suggesting increasing limitation of microbial processes by N relative to C in these treatments. Based on these results, we conclude that the plant supply of organic matter primarily controls gross N mineralization and microbial respiration, but that the concentration of N in organic matter input secondarily influences these processes. Thus, in systems where N limits plant productivity these global change factors could cause different long‐term ecosystem trajectories because of divergent effects on soil N and C cycling.     

Effect of Crude Oil and Chemical Additives on Metabolic Activity of Mixed Microbial Populations in Fresh Marsh Soils

Abstract Hydrocarbons increase abundance of hydrocarbon-degrading microorganisms, but also decrease microbial diversity. This could disrupt ecosystem dynamics by altering soil organic matter mineralization and resultant nutrient remineralization rates. Crude oil, which is known to contain toxins and reduce microbial diversity, was hypothesized to reduce gross metabolic activity of mixed microbial populations in wetland soils. Soil respiration and Eh were compared, for 6 months, among microcosms containing marsh soils that differed in soil organic matter (Panicum hemitomon Shult. or Sagittaria lancifolia L. dominated marshes), crude oil (Arabian crude, Louisiana crude, or no oil), and additives (a cleaner, a dispersant, fertilizer, or no additive). No treatment slowed activity; instead, Louisiana plus fertilizer and all Arabian treatments temporarily accelerated activity. Additional C respired from oiled microcosms exceeded C added as crude oil by 1.4 to 3.5 times. Thus, much additional C originated from soil organic matter rather than crude oil. Crude oils temporarily lowered soil Eh, which is consistent with accelerated metabolism and demand for electron acceptors. The lack of inhibition observed at the community level does not necessarily indicate an absence of toxicity. Instead, tolerant species with metabolic versatility probably maintained activity. Stimulation probably resulted from removal of micronutrient limitation, rather than removal of grazing pressure or macronutrient limitation. Regardless, accelerated soil organic matter mineralization surely accelerated nutrient remineralization. This might explain some reports of crude oil stimulating plant growth. These results are not inconsistent with theoretical and experimental conclusions regarding effects of biodiversity on ecosystem stability and productivity, nor are they inconsistent with conclusions that crude oils contain components that are toxic to microbes, vegetation, and fauna. However, these data do indicate that crude oils also contain components that temporarily stimulate metabolic activity of surviving microbes. Received: 27 April 1998; Accepted: 15 July 1998     

<Emphasis Type="Italic">Eisenia fetida</Emphasis> (Oligochaeta: Lumbricidae) Modifies the Structure and Physiological Capabilities of Microbial Communities Improving Carbon Mineralization During Vermicomposting of Pig Manure

Although microorganisms are largely responsible for organic matter decomposition, earthworms may also affect the rates of decomposition directly by feeding on and digesting organic matter and microorganisms, or indirectly affect them through their interactions with the microorganisms, basically involving stimulation or depression of the microbial populations. We tested the general hypothesis that microbial populations, and especially fungi, are enhanced by earthworm activity, and also whether earthworms are able to modify the biodiversity of microbial populations, and its relation to the function of the system. In addition, we examined the metabolic quotient and the effect of labile organic C to assess the relationships between earthworm and microbes. We found that decomposition of pig manure has two stages characterized by the presence or absence of earthworms. Thus, the presence of earthworms was related with increases in overall microbial biomass and activity, which decreased when earthworms left the substrate; the same pattern was observed for fungi. Furthermore, earthworms modified the physiological profiles of microbial communities of pig manure, increasing the diversity of substrates utilized. In addition, earthworms promoted a more efficient use of energy of microbial communities, as the metabolic quotient showed. The rate of carbon loss was almost twice where earthworms were present, revealing faster decomposition. Our data match with the recent findings that to maintain essential processes the functional properties of present species are at least as important as the number of species per se. This is in accordance with the “insurance hypothesis,” which states that a large number of species is probably essential for maintaining stable processes in changing environments, as the presence of earthworms would have promoted in pig manure.     

Roles of microorganisms other than Clostridium and Enterobacter in anaerobic fermentative biohydrogen production systems--a review

Anaerobic fermentative biohydrogen production, the conversion of organic substances especially from organic wastes to hydrogen gas, has become a viable and promising means of producing sustainable energy. Successful biological hydrogen production depends on the overall performance (results of interactions) of bacterial communities, i.e., mixed cultures in reactors. Mixed cultures might provide useful combinations of metabolic pathways for the processing of complex waste material ingredients, thereby supporting the more efficient decomposition and hydrogenation of biomass than pure bacteria species would. Therefore, understanding the relationships between variations in microbial composition and hydrogen production efficiency is the first step in constructing more efficient hydrogen-producing consortia, especially when complex and non-sterilized organic wastes are used as feeding substrates. In this review, we describe recent discoveries on bacterial community composition obtained from dark fermentation biohydrogen production systems, with emphasis on the possible roles of microorganisms that co-exist with common hydrogen producers.