Advance on the production of polyhydroxyalkanoates by mixed cultures

Polyhydroxyalkanoates (PHAs) are the polymers of hydroxyalkanoates that accumulate as carbon/energy or reducing-power storage material in various microorganisms. PHAs have attracted considerable attention as biodegradable substitutes for conventional polymers. Until now, however, industrial production of PHAs has encountered only limited success. The main barrier to the replacement of synthetic plastics by PHAs has been the higher cost. The use of mixed cultures and renewable sources obtained from waste organic carbon can substantially decrease the cost of PHA and increase their market potential. This work reviews two main methods of PHA production by mixed cultures, anaerobic-aerobic processing and aerobic transient feeding processing, and analyzed the metabolic and effective factors.     

Advance on the production of polyhydroxyalkanoates by mixed cultures

Polyhydroxyalkanoates (PHAs)are the polymers of hydroxyalkanoates that accumulate as carbon/energy or reducing-power storage material in various microorganisms.PHAs have attracted considerable attention as biodegradable substitutes for conventional polymers.Until now,however,industrial production of PHAs has encountered only limited success.The main barrier to the replacement of synthetic plastics by PHAs has been the higher cost.The use of mixed cultures and renewable sources obtained from waste organic carbon can substantially decrease the cost of PHA and increase their market potential.This work reviews two main methods of PHA production by mixed cultures,anaerobicaerobic processing and aerobic transient feeding processing,and analyzed the metabolic and effective factors.     

The impact of pH control on the volumetric productivity of mixed culture PHA production from fermented molasses

Polyhydroxyalkanoate (PHA) production via mixed microbial cultures (MMCs) can potentially decrease process operational costs as compared to conventional pure culture techniques. However, the volumetric productivity of PHA by MMCs must be augmented to increase its cost competitiveness. For this purpose, a three‐stage bioreactor system was operated in this study, with (i) anaerobic fermentation of molasses, (ii) culture selection, and (iii) PHA accumulation and harvesting stages. In stage 2, bioreactor operation with pH control at 8 led to twice the biomass concentration (up to 8 g VSS L^?1, where VSS is the volatile suspended solids) as compared to operation without pH control (maximum pH 9). No loss in the specific PHA storage efficiency was observed (PHA content up to 57.5% and PHA storage rate up to 0.27 Cmol PHA Cmol X^?1 h^?1, where X is the active biomass), thereby resulting in twice the volumetric PHA production rate. The limited biomass growth at the higher pH level was not due to nutrient limitation, but likely to a shift in the microbial community. It is hypothesized that the increased enrichment of Azoarcus at pH 8 led to higher PHA productivity. pH control in the culture selection stage can lead to enhanced PHA production from MMCs, improving the viability of the process.     

Segregated flux balance analysis constrained by population structure/function data: The case of PHA production by mixed microbial cultures

In this study we developed a segregated flux balance analysis (FBA) method to calculate metabolic flux distributions of the individual populations present in a mixed microbial culture (MMC). Population specific flux data constraints were derived from the raw data typically obtained by the fluorescence in situ hybridization (FISH) and microautoradiography (MAR)‐FISH techniques. This method was applied to study the metabolic heterogeneity of a MMC that produces polyhydroxyalkanoates (PHA) from fermented sugar cane molasses. Three populations were identified by FISH, namely Paracoccus sp., Thauera sp., and Azoarcus sp. The segregated FBA method predicts a flux distribution for each of the identified populations. The method is shown to predict with high accuracy the average PHA storage flux and the respective monomeric composition for 16 independent experiments. Moreover, flux predictions by segregated FBA were slightly better than those obtained by nonsegregated FBA, and also highly concordant with metabolic flux analysis (MFA) estimated fluxes. The segregated FBA method can be of high value to assess metabolic heterogeneity in MMC systems and to derive more efficient eco‐engineering strategies. For the case of PHA‐producing MMC considered in this work, it becomes apparent that the PHA average monomeric composition might be controlled not only by the volatile fatty acids (VFA) feeding profile but also by the population composition present in the MMC. Biotechnol. Bioeng. 2013; 110: 2267–2276. © 2013 Wiley Periodicals, Inc.     

Batch tests for assessing decolourisation of azo dyes by methanogenic and mixed cultures

Most of the published studies on azo dye colour removal involve anaerobic mixed cultures and there is some interest in the knowledge of how dye reduction occurs, if by facultative, strictly anaerobic or both bacterial trophic groups present in classic anaerobic digestors. This paper describes the behaviour of methanogenic and mixed bacteria cultures on the colour removal in batch systems, of a commercial azo dye, C.I. Acid Orange 7, used in paper and textile industries. The aim of this study is to demonstrate, by analysing dye decolourisation, that it occurs with mixed cultures as well as with strictly anaerobic (methanogenic) cultures. Tests were performed with a range of dye concentrations between 60 and 300 mg l⁻¹. The influence of dye concentration on the carbon source removal and decolourisation processes was studied. The effect of carbon source concentration on colour removal was also analysed for both cultures. The degradation rates in mixed and methanogenic cultures were compared. The consumption of carbon source was monitored by COD analysis and dye degradation by ultraviolet-visible spectrophotometry and thin layer chromatography.     

Optimization of polyhydroxybutyrate production by mixed cultures submitted to aerobic dynamic feeding conditions

Activated sludge submitted to aerobic dynamic feeding conditions showed a good and stable capacity to store polyhydroxybutyrate (PHB). The system, working for 2 years, selected a microbial population with a high PHB storage capacity. The influence of carbon and nitrogen concentrations on the PHB accumulation yield was studied in a range of 15-180 Cmmol/l for acetate and 0-2.8 Nmmol/l for ammonia. Low ammonia concentrations favored PHB accumulation. The maximum PHB content, 67.5%, was obtained for 180 Cmmol/l of acetate supplied in one pulse. However, such high substrate concentration proved to be inhibitory for the storage mechanism, causing a slowdown of the specific PHB storage rate. In order to avoid substrate inhibition, 180 Cmmol/l of acetate was supplied in different ways: continuously fed and in three pulses of 60 Cmmol/l each. In both cases the specific PHB storage rate increased and the PHB content obtained were 56.2% and 78.5%, respectively. The latter value of PHB content is similar to that obtained by pure cultures and was never reported for mixed cultures. Addition of acetate by pulses controlled by the oxygen concentration was kept for 16 days, the PHB content being always above 70% of cell dry weight.     

The utilization of glycogen accumulating organisms for mixed culture production of polyhydroxyalkanoates

Production of polyhydroxyalkanoates (PHAs) by an open mixed culture enriched in glycogen accumulating organisms (GAOs) under alternating anaerobic–aerobic conditions with acetate as carbon source was investigated. The culture exhibited a stable enrichment performance over the 450‐day operating period with regards to phenotypic behavior and microbial community structure. Candidatus Competibacter phosphatis dominated the culture at between 54% and 70% of the bacterial biomass throughout the study, as determined by fluorescence in situ hybridization. In batch experiments under anaerobic conditions, PHA containing 3‐hydroxybutyrate (3HB) and 27 mol‐% 3‐hydroxyvalerate (3HV) was accumulated up to 49% of cell dry weight utilizing the glycogen pool stored in the SBR cycle. Under aerobic and ammonia limited conditions, PHA comprising only 3HB was accumulated to 60% of cell dry weight. Glycogen was consumed during aerobic PHA accumulation as well as under anaerobic conditions, but with different stoichiometry. Under aerobic conditions 0.31 C‐mol glycogen was consumed per consumed C‐mol acetate compared to 0.99 under anaerobic conditions. Both the PHA biomass content and the specific PHA production rate obtained were similar to what is typically obtained using the more commonly applied aerobic dynamic feeding strategy. Biotechnol. Bioeng. 2009; 104: 698–708 © 2009 Wiley Periodicals, Inc.     

Modeling and experimental study on the anaerobic/aerobic/anoxic process for simultaneous nitrogen and phosphorus removal: The effect of acetate addition

A mathematical model based on the simulation software AQUASIM was developed to validate an anaerobic/aerobic/anoxic (AOA) process that enables simultaneous nitrogen and phosphorus removal in a single reactor by adding external organic carbon to preclude excess aerobic phosphate uptake by polyphosphate-accumulating organisms (PAOs) and provide phosphate for denitrifying PAOs (DNPAOs). Aerobic batch tests after anaerobic phosphate release with different chemical oxygen demand (COD) concentrations indicated that the effect of COD concentration on the phosphate uptake preclusion could be expressed by a simple formula. The reduction factor reflecting the formula, which retards the aerobic phosphate uptake in the presence of COD, was added to the process rates of aerobic polyphosphate storage and PAOs growth in the model. The improved model, which included the reduction factor, reasonably matched the experimental result regarding aerobic phosphate uptake behavior whereas the model without it did not; thus, the former precisely predicts the AOA process behavior.     

不同生境微生物转化H2/CO2产乙酸及其在合成气发酵中应用

【目的】合成气发酵对大力开发可再生资源和促进国家可持续发展具有重要意义,研究旨在探究不同生境微生物转化H2/CO2产乙酸及其合成气发酵的潜力。【方法】采集剩余污泥、牛粪、产甲烷污泥和河道底物样品在中温(37 °C)条件下生物转化H2/CO2气体,将来源于牛粪样品的H2/CO2转化富集物用于合成气发酵,通过454高通量技术和定量PCR技术分析复杂微生物群落的组成,GC气相色谱法检测气体转化产生的挥发性脂肪酸(VFAs)浓度。【结果】牛粪和剩余污泥微生物利用H2/CO2气体生成乙酸、乙醇和丁酸等,最高乙酸浓度分别为63 mmol/L和40 mmol/L,明显高于河道底物和产甲烷污泥样品的最高乙酸浓度3 mmol/L和16 mmol/L。牛粪和剩余污泥微生物中含有种类多样化的同型产乙酸菌,剩余污泥中同型产乙酸菌主要为Clostridium spp.、Sporomusa malonica和Acetoanaerobium noterae,牛粪中则为Clostridium spp.、Treponema azotonutricium和Oxobacter pfennigii。【结论】同型产乙酸菌的丰富度和数量两个因素都对复杂微生物群落转化H2/CO2产乙酸效率至关重要;转化H2/CO2得到的富集物可用于合成气发酵产乙酸和乙醇,这为基于混合培养技术的合成气发酵提供了依据。     

Assessment of biotransformation of organic matter during co-composting of sewage sludge-lignocelullosic waste by chemical,FTIR analyses,and phytotoxicity tests

The co-composting of activated sludge and lignocellulosic waste (palm tree waste) was monitored to study the evolution of two mixtures, referred to as A (2/3 palm waste + 1/3 sludge) and B (1/2 palm waste + 1/2 sludge) for 6 mo. The biotransformation during the co-composting process was evaluated by physicochemical and spectroscopy analyses. The final composts exhibited a higher degree of decomposition than the controls as shown by a decrease of C/N and ${\text{NH}}_4^{+}/{\text{NO}}_3^{-}$ ratios, and a 43% decrease in total lipid content. The decrease of aliphatic absorbance bands at 2964, 2922, and 2850 cm-1 and the increase of structure aromatic absorbance bands at 1514, 1426, and 1386 cm-1 reflect the progress of the humification process, which judging by the increase in the humification index, is about 60%. This efficiency of co-composting in reducing phytotoxicity was confirmed by the germination index, which reached over 90%, and by metallic trace element concentration.     

Geographical and technological differences in life cycle inventories shown by the use of process models for waste incinerators part I. technological and geographical differences

Goal and Background  Geographical and technological differences in Life Cycle Inventory data are an important source for uncertainty in the result of Life Cycle Assessments. Knowledge on their impact on the result of an LCA is scarce, and also knowledge on how to manage them in an LCA case study. Objective  Goal of this paper is to explore these differences for municipal solid waste incinerator plants, and to develop recommendations for managing technological and geographical differences. Methodology  The paper provides a definition of technological and geographical differences, and analyses their possible impacts. In a case study, the differences are caused intentionally in ‘games’, by virtually transplanting incineration plants to a different location and by changing parameters such as the composition of the waste input incinerated. The games are performed by using a modular model for municipal solid waste incinerator plants. In each case, an LCA including an Impact Assessment is calculated to trace the impact of these changes, and the results are compared. Conclusions  The conclusions of the paper are two-fold: (1) reduce the differences in inventory data where their impact on the result is high; where it is possible reducing them to a great extent, and the effort for performing the change acceptable; in the case of incineration plants: Adapt the flue gas treatment, especially a possible DeNOx step, to the real conditions; (2) make use of modular process models that allow adapting plant parameters to better meet real conditions, but be aware of possible modelling errors. The paper invites the scientific community to validate the model used for a waste incinerator plant, and suggest putting up similar models for other processes, preferably those of similar relevance for Life Cycle Inventories.     

Production of fuels and chemicals from renewable resources using engineered Escherichia coli

Biotechnological production of fuels and chemicals from renewable resources is an appealing way to move from the current petroleum-based economy to a biomass-based green economy. Recently, the feedstocks that can be used for bioconversion or fermentation have been expanded to plant biomass, microbial biomass, and industrial waste. Several microbes have been engineered to produce chemicals from renewable resources, among which Escherichia coli is one of the best studied. Much effort has been made to engineer E. coli to produce fuels and chemicals from different renewable resources. In this paper, we focused on E. coli and systematically reviewed a range of fuels and chemicals that can be produced from renewable resources by engineered E. coli. Moreover, we proposed how can we further improve the efficiency for utilizing renewable resources by engineered E. coli, and how can we engineer E. coli for utilizing alternative renewable feedstocks. e.g. C1 gases and methanol. This review will help the readers better understand the current progress in this field and provide insights for further metabolic engineering efforts in E. coli.     

Mechanism for verification of mismatched and homoduplex DNAs by nucleotides‐bound MutS analyzed by molecular dynamics simulations

In order to understand how MutS recognizes mismatched DNA and induces the reaction of DNA repair using ATP, the dynamics of the complexes of MutS (bound to the ADP and ATP nucleotides, or not) and DNA (with mismatched and matched base‐pairs) were investigated using molecular dynamics simulations. As for DNA, the structure of the base‐pairs of the homoduplex DNA which interacted with the DNA recognition site of MutS was intermittently disturbed, indicating that the homoduplex DNA was unstable. As for MutS, the disordered loops in the ATPase domains, which are considered to be necessary for the induction of DNA repair, were close to (away from) the nucleotide‐binding sites in the ATPase domains when the nucleotides were (not) bound to MutS. This indicates that the ATPase domains changed their structural stability upon ATP binding using the disordered loop. Conformational analysis by principal component analysis showed that the nucleotide binding changed modes which have structurally solid ATPase domains and the large bending motion of the DNA from higher to lower frequencies. In the MutS–mismatched DNA complex bound to two nucleotides, the bending motion of the DNA at low frequency modes may play a role in triggering the formation of the sliding clamp for the following DNA‐repair reaction step. Moreover, MM‐PBSA/GBSA showed that the MutS–homoduplex DNA complex bound to two nucleotides was unstable because of the unfavorable interactions between MutS and DNA. This would trigger the ATP hydrolysis or separation of MutS and DNA to continue searching for mismatch base‐pairs. Proteins 2016; 84:1287–1303. © 2016 Wiley Periodicals, Inc.