Organic acid production from potato starch waste fermentation by rumen microbial communities from Dutch and Thai dairy cows

Background: Exploring different microbial sources for biotechnological production of organic acids is important. Dutch and Thai cow rumen samples were used as inocula to produce organic acid from starch waste in anaerobic reactors. Organic acid production profiles were determined and microbial communities were compared using 16S ribosomal ribonucleic acid gene amplicon pyrosequencing. Results: In both reactors, lactate was the main initial product and was associated with growth of Streptococcus spp. (86% average relative abundance). Subsequently, lactate served as a substrate for secondary fermentations. In the reactor inoculated with rumen fluid from the Dutch cow, the relative abundance of Bacillus and Streptococcus increased from the start, and lactate, acetate, formate and ethanol were produced. From day 1.33 to 2, lactate and acetate were degraded, resulting in butyrate production. Butyrate production coincided with a decrease in relative abundance of Streptococcus spp. and increased relative abundances of bacteria of other groups, including Parabacteroides, Sporanaerobacter, Helicobacteraceae, Peptostreptococcaceae and Porphyromonadaceae. In the reactor with the Thai cow inoculum, Streptococcus spp. also increased from the start. When lactate was consumed, acetate, propionate and butyrate were produced (day 3-4). After day 3, bacteria belonging to five dominant groups, Bacteroides, Pse udoramibacter_Eubacterium, Dysgonomonas, Enterobacteriaceae and Porphyromonadaceae, were detected and these showed significant positive correlations with acetate, propionate and butyrate levels. Conclusions: The complexity of rumen microorganisms with high adaptation capacity makes rumen fluid a suitable source to convert organic waste into valuable products without the addition of hydrolytic enzymes. Starch waste is a source for organic acid production, especially lactate.Peer reviewe

The thermoacidophilic methanotroph Methylacidiphilum fumariolicum SolV oxidizes subatmospheric H₂ with a high-affinity, membrane-associated [NiFe] hydrogenase

The trace amounts (0.53 ppmv) of atmospheric hydrogen gas (H2) can be utilized by microorganisms to persist during dormancy. This process is catalyzed by certain Actinobacteria, Acidobacteria, and Chloroflexi, and is estimated to convert 75 × 1012 g H2 annually, which is half of the total atmospheric H2. This rapid atmospheric H2 turnover is hypothesized to be catalyzed by high-affinity [NiFe] hydrogenases. However, apparent high-affinity H2 oxidation has only been shown in whole cells, rather than for the purified enzyme. Here, we show that the membrane-associated hydrogenase from the thermoacidophilic methanotroph Methylacidiphilum fumariolicum SolV possesses a high apparent affinity (Km(app) = 140 nM) for H2 and that methanotrophs can oxidize subatmospheric H2. Our findings add to the evidence that the group 1h [NiFe] hydrogenase is accountable for atmospheric H2 oxidation and that it therefore could be a strong controlling factor in the global H2 cycle. We show that the isolated enzyme possesses a lower affinity (Km = 300 nM) for H2 than the membrane-associated enzyme. Hence, the membrane association seems essential for a high affinity for H2. The enzyme is extremely thermostable and remains folded up to 95 °C. Strain SolV is the only known organism in which the group 1h [NiFe] hydrogenase is responsible for rapid growth on H2 as sole energy source as well as oxidation of subatmospheric H2. The ability to conserve energy from H2 could increase fitness of verrucomicrobial methanotrophs in geothermal ecosystems with varying CH4 fluxes. We propose that H2 oxidation can enhance growth of methanotrophs in aerated methane-driven ecosystems. Group 1h [NiFe] hydrogenases could therefore contribute to mitigation of global warming, since CH4 is an important and extremely potent greenhouse gas.</p

(Per)chlorate reduction by an acetogenic bacterium, Sporomusa sp., isolated from an underground gas storage

A mesophilic bacterium, strain An4, was isolated from an underground gas storage reservoir with methanol as substrate and perchlorate as electron acceptor. Cells were Gram-negative, spore-forming, straight to curved rods, 0.5–0.8 μm in diameter, and 2–8 μm in length, growing as single cells or in pairs. The cells grew optimally at 37°C, and the pH optimum was around 7. Strain An4 converted various alcohols, organic acids, fructose, acetoin, and H2/CO2 to acetate, usually as the only product. Succinate was decarboxylated to propionate. The isolate was able to respire with (per)chlorate, nitrate, and CO2. The G+C content of the DNA was 42.6 mol%. Based on the 16S rRNA gene sequence analysis, strain An4 was most closely related to Sporomusa ovata (98% similarity). The bacterium reduced perchlorate and chlorate completely to chloride. Key enzymes, perchlorate reductase and chlorite dismutase, were detected in cell-free extracts

Influence of inoculum source on gas production profiles

Gas production equipment is used in many locations to study rate and extent of fermentation of feeds in the rumen. Since data obtained with rumen fluid from sheep are often used for dairy cattle, rates and extents of fermentation were compared from cow and sheep rumen fluid. However, since it is often impossible for many industries and schools to retain ruminally fistulated animals, use of fresh bovine faeces as an alternative inoculum was also compared with rumen fluid. Rumen fluid was collected from rumen fistulated sheep and cows kept under the same conditions and, at the same time, fresh faeces were collected from the same cows. Incubations with rumen fluid from cows and sheep resulted in gas productions with comparable profiles. Incubations with sheep rumen fluid and bovine faeces as inoculum resulted in decreased total gas productions, compared with incubations with cows rumen fluid. The correlation between rumen fluid from cows and sheep was high for gas production at 24 and 48 h incubation, calculated maximal gas production (Am) and sharpness of the profiles (Cm) (R2 = 0.90 to 0.96). For rate of gas production (Bm) the correlation between rumen fluid from cows and sheep was lower (R2 = 0.62). Comparison of gas production profiles from bovine rumen fluid and faeces also resulted in a high correlation for gas production at 48 h incubation and Am (R2 = 0.82 to 0.88), but a low correlation for Bm, Cm and gas production at 24h incubation (R2 = 0.24 to 0.61). Rumen fluid from sheep and faeces from cows can be used as an alternative to rumen fluid from cows to accurately determine differences in total gas production. These alternative inocula, however, did not accurately estimate the rate of gas production (Bm) of different feed samples obtained when incubated with rumen fluid from cows. © 2002 Elsevier Science B.V. All rights reserved

Comparison of organic matter degradation in several feedstuffs in the rumen as determined with the nylon bag and gas production techniques

Organic matter (OM) degradation of 21 feedstuffs was investigated with rumen fluid using a rumen in situ technique and a gas production technique. Fitting the nylon bag data to an exponential model showed that there was a high variation in the rate of OM degradation ranging from 1.7% h-1 for protected solvent extracted soybean meal to 10.5% h-1 for potato pulp. The percentage fermentable OM (FOM), calculated from the nylon bag data, ranged from 26.9% for maize gluten meal to 76.4% for pea meal. Gas production was recorded with fully automated equipment using twice-diluted rumen fluid. The gas production profiles were fitted to a mono-phasic and a tri-phasic model. The aim of the study described is to investigate the possibilities to estimate in situ degradation characteristics using gas production characteristics and chemical composition. The in situ washout fraction (W), degradable fraction (D) and undegradable fraction (U) could be predicted from chemical composition and gas production parameters with R2 ranging from 0.50 to 0.72. There was a closer relationship between in situ degradation rate of OM (kd) and the incubation period halfway to maximum gas production (B), using the tri-phasic model (R2 = 0.58) than the mono-phasic model (R2 = 0.43). Accounting for an in situ lag-period slightly improved prediction of kd by gas production parameters (R2 = 0.47-0.62). Percentage FOM, calculated from in situ results, could be predicted from chemical composition and gas production parameters with R2 ranging from 0.50 to 0.75. Transformation of kd (determined in situ) to its half-life value of degradation ((ln 2/kd) × 100) provided a slight improvement of kd prediction by chemical composition and gas production parameters, with R2 ranging from 0.56 to 0.81. There was only a moderate relationship for OM degradation in these feedstuffs determined using an in situ and a gas production technique. © 2002 Elsevier Science B.V. All rights reserved

Comparative proteomics of Geobacter sulfurreducens PCAT in response to acetate, formate and/or hydrogen as electron donor

Geobacter sulfurreducens is a model bacterium to study the degradation of organic compounds coupled to the reduction of Fe(III). The response of G. sulfurreducens to the electron donors acetate, formate, hydrogen and a mixture of all three with Fe(III) citrate as electron acceptor was studied using comparative physiological and proteomic approaches. Variations in the supplied electron donors resulted in differential abundance of proteins involved in the citric acid cycle (CAC), gluconeogenesis, electron transport, and hydrogenases and formate dehydrogenase. Our results provided new insights into the electron donor metabolism of G. sulfurreducens. Remarkably, formate was the preferred electron donor compared to acetate, hydrogen, or acetate plus hydrogen. When hydrogen was the electron donor, formate was formed, which was associated with a high abundance of formate dehydrogenase. Notably, abundant proteins of two CO2 fixation pathways (acetyl-CoA pathway and the reversed oxidative CAC) corroborated chemolithoautotrophic growth of G. sulfurreducens with formate or hydrogen and CO2, and provided novel insight into chemolithoautotrophic growth of G. sulfurreducens. This article is protected by copyright. All rights reserved.This work was performed in the TTIW-cooperation framework of Wetsus, European Centre of Excellence for Sustain able Water Technology (www.wetsus.nl). Wetsus is funded by the Dutch Ministry of Economic Affairs, the European Union Regional Development Fund, the Province of Fryslân, the City of Leeuwarden and the EZ/Kompas program of the “Samenwerkingsverband Noord-Nederland”. The authors like to thank the participants of the research theme ‘Resource Recovery’ for fruitful discussions and their financial support. Research of AJMS is financed by an advanced grant of the European Research Council under the European Union’s Seventh Framework Programme (FP/2007e2013)/ERC Grant Agreement (project 323009). Research of AJMS and PHAT is supported by a Gravitation grant (project 024.002.002) of the Netherlands Ministry of Education, Culture and Science.info:eu-repo/semantics/publishedVersio

The thermoacidophilic methanotroph Methylacidiphilum fumariolicum SolV oxidizes subatmospheric H-2 with a high-affinity, membrane-associated [NiFe] hydrogenase

The trace amounts (0.53 ppmv) of atmospheric hydrogen gas (H2) can be utilized by microorganisms to persist during dormancy. This process is catalyzed by certain Actinobacteria, Acidobacteria, and Chloroflexi, and is estimated to convert 75 × 1012 g H2 annually, which is half of the total atmospheric H2. This rapid atmospheric H2 turnover is hypothesized to be catalyzed by high-affinity [NiFe] hydrogenases. However, apparent high-affinity H2 oxidation has only been shown in whole cells, rather than for the purified enzyme. Here, we show that the membrane-associated hydrogenase from the thermoacidophilic methanotroph Methylacidiphilum fumariolicum SolV possesses a high apparent affinity (Km(app) = 140 nM) for H2 and that methanotrophs can oxidize subatmospheric H2. Our findings add to the evidence that the group 1h [NiFe] hydrogenase is accountable for atmospheric H2 oxidation and that it therefore could be a strong controlling factor in the global H2 cycle. We show that the isolated enzyme possesses a lower affinity (Km = 300 nM) for H2 than the membrane-associated enzyme. Hence, the membrane association seems essential for a high affinity for H2. The enzyme is extremely thermostable and remains folded up to 95 °C. Strain SolV is the only known organism in which the group 1h [NiFe] hydrogenase is responsible for rapid growth on H2 as sole energy source as well as oxidation of subatmospheric H2. The ability to conserve energy from H2 could increase fitness of verrucomicrobial methanotrophs in geothermal ecosystems with varying CH4 fluxes. We propose that H2 oxidation can enhance growth of methanotrophs in aerated methane-driven ecosystems. Group 1h [NiFe] hydrogenases could therefore contribute to mitigation of global warming, since CH4 is an important and extremely potent greenhouse gas

Ammonia oxidation at pH 2.5 by a new gammaproteobacterial ammonia-oxidizing bacterium

Ammonia oxidation was considered impossible under highly acidic conditions, as the protonation of ammonia leads to decreased substrate availability and formation of toxic nitrogenous compounds. Recently, some studies described archaeal and bacterial ammonia oxidizers growing at pH as low as 4, while environmental studies observed nitrification at even lower pH values. In this work, we report on the discovery, cultivation, and physiological, genomic, and transcriptomic characterization of a novel gammaproteobacterial ammonia-oxidizing bacterium enriched via continuous bioreactor cultivation from an acidic air biofilter that was able to grow and oxidize ammonia at pH 2.5. This microorganism has a chemolithoautotrophic lifestyle, using ammonia as energy source. The observed growth rate on ammonia was 0.196 day−1, with a doubling time of 3.5 days. The strain also displayed ureolytic activity and cultivation with urea as ammonia source resulted in a growth rate of 0.104 day−1 and a doubling time of 6.7 days. A high ammonia affinity (Km(app) = 147 ± 14 nM) and high tolerance to toxic nitric oxide could represent an adaptation to acidic environments. Electron microscopic analysis showed coccoid cell morphology with a large amount of intracytoplasmic membrane stacks, typical of gammaproteobacterial ammonia oxidizers. Furthermore, genome and transcriptome analysis showed the presence and expression of diagnostic genes for nitrifiers (amoCAB, hao, nor, ure, cbbLS), but no nirK was identified. Phylogenetic analysis revealed that this strain belonged to a novel bacterial genus, for which we propose the name “Candidatus Nitrosacidococcus tergens” sp. RJ19.</p