Impact of oral administration of compost extract on gene expression in the rat gastrointestinal tract

Oral administration of an extract consisting of compost fermented with thermophiles to pigs reduces the incidence of stillbirth and promotes piglet growth. However, the mechanism by which the compost extract modulates the physiological conditions of the animals remains largely unknown. Here, we investigate the effects of compost extract on the physiological responses in the intestine of a mammalian rat model. The level of fecal immunoglobulin A (IgA), which provides protection against pathogens and is secreted from the small intestine, was significantly higher in rats treated with continuous administration of the compost extract than in untreated rats after 2 months, but not after 1 month. However, the fecal IgA level was not significantly different in rats that received the filtered compost extract compared with the untreated rats or the rats that received the compost extract. Gene expression analyses of the small intestine indicated that several immune-related genes were upregulated following compost exposure. Specifically, the expression levels of lymphocyte chemoattractant chemokine CXCL13 and Granzyme B, which is released within cytotoxic T lymphocytes and natural killer cells, increased in the small intestinal tract following compost exposure. Based on these observations, it was postulated that the increased level of fecal IgA following compost exposure was associated with the expression of CXCL13 and Granzyme B in the intestinal tract. Thus, thermophile-fermented compost could contain microbes or substances that activate the rat's gut mucosal immune response.     

Effect of COD/SO(4)(2-) ratio and sulfide on thermophilic (55 degrees C) sulfate reduction during the acidification of sucrose at pH 6

This study investigated the effect of the COD/SO₄²⁻ ratio (4 and 1) and the sulfide concentration on the performance of thermophilic (55 °C) acidifying (pH 6) upflow anaerobic sludge bed reactors fed with sucrose at an organic loading rate of 4.5 g COD l_reactor⁻¹ day⁻¹. Sulfate reduction efficiencies amounted to 65% and 25-35% for the COD/SO₄²⁻ ratios of 4 and 1, respectively. Acidification was complete at all the tested conditions and the electron flow was similar at the two COD/SO₄²⁻ ratios applied. The stepwise decrease of the sulfide concentrations in the reactors with a COD/SO₄²⁻ ratio of 1 by N₂ stripping caused an immediate stepwise increase in the sulfate reduction efficiencies, indicating a reversible inhibition by sulfide. The degree of reversibility was, however, affected by the growth conditions of the sludge. Acidifying sludge pre-grown at pH 6, at a COD/SO₄²⁻ ratio of 9 and exposed for 150 days to 115 mg l⁻¹ sulfide, showed a slower recovery from the sulfide inhibition than a freshly harvested sludge from a full scale treatment plant (pH 7 and COD/SO₄²⁻=9.5) exposed for a 70 days to 200 mg l⁻¹ sulfide. In the latter case, the decrease of the sulfide concentration from 200 to 45 mg l⁻¹ (35 mg l⁻¹ undissociated sulfide) by N₂ stripping caused an immediate increase of the sulfate reduction efficiency from 35% to 96%.     

Inactivation of Escherichia coli O157:H7 during thermophilic anaerobic digestion of manure from dairy cattle

Inactivation of the pathogenic Escherichia coli serotype O157:H7 and a non-pathogenic E. coli strain isolated from dairy cattle manure was evaluated with batch tests at 50 and 55 degrees C in biosolids from a thermophilic anaerobic digester treating the manure. Using differential-selective plating on sorbitol-MacConkey (SMAC) agar to quantify E. coli, the decline in concentrations of both the sorbitol-negative (putative E. coli O157:H7) and sorbitol-positive (putative non-pathogenic E. coli) organisms followed a model that assumed there was a heat-sensitive fraction and a heat-resistant fraction. Inactivation rates of the heat-sensitive fractions were similar for both colony types at each temperature, suggesting that wild-type E. coli can be used as an indicator of inactivation of serotype O157:H7. The decimal reduction time for the heat-sensitive fractions was in the order of 10min at 55 degrees C and ranged from approximately 1-3h at 50 degrees C. Concentrations of heat-resistant organisms at 55 degrees C were 1.4-1.7log(10)cfu/mL. Confirmatory analyses conducted on 30 randomly selected colonies of heat-resistant sorbitol-negative cells from treatment at 55 degrees C indicated that none were serotype O157:H7, nor were they E. coli. Similar analyses on 10 sorbitol-negative isolates from untreated manure indicated that none were serotype O157:H7, although 16S rRNA gene sequence analysis indicated that eight were E. coli or closely related enteric bacteria. These findings suggest that plating on differential-selective media to quantify E. coli, including serotype O157:H7, in effluent samples from thermophilic anaerobic digestion can lead to false positive results. Therefore, more specific methods should be used to evaluate the extent of thermal inactivation of both pathogenic and non-pathogenic E. coli in manure treatment systems.     

Thermophilic aerobic granular biomass for enhanced settleability

Aerobic biological wastewater treatment at thermophilic (ca. 55 degrees C) temperatures notoriously produces biomass that flocculates poorly or not at all. Contrary to this, thermophilic aerobic biomass that settled well in sequencing batch reactors was cultured with sludge volume index (SVI) values as low as 60mL/g. A mixture of granular and flocculant biomass resulted when closed reactors were sparged with recirculated reactor headspace gas containing some air, whereas a conventionally aerated control reactor sparged with air alone contained dispersed growth that did not flocculate. Maximum granule diameter was from 1.2 to 1.9mm, and granule resistance to disintegration was comparable to aerobic mesophilic granules. Two bacteria were isolated and identified as Anoxybacillus flavothermus and Pseudoxanthomonas taiwanensis as determined by partial 16S rDNA sequencing. Anoxybacilli species are alkaliphilic or alkalitolerant, with the type species having an obligate requirement for carbonate, even when grown on glucose. We postulate that high alkalinity and CO(2) may select for a population of aerobic thermophilies that flocculates and granulates.     

Enrichment of acetogenic bacteria in high rate anaerobic reactors under mesophilic and thermophilic conditions

The objective of the current study was to expand the knowledge of the role of acetogenic Bacteria in high rate anaerobic digesters. To this end, acetogens were enriched by supplying a variety of acetogenic growth supportive substrates to two laboratory scale high rate upflow anaerobic sludge bed (UASB) reactors operated at 37 °C (R1) and 55 °C (R2). The reactors were initially fed a glucose/acetate influent. Having achieved high operational performance and granular sludge development and activity, both reactors were changed to homoacetogenic bacterial substrates on day 373 of the trial. The reactors were initially fed with sodium vanillate as a sole substrate. Although % COD removal indicated that the 55 °C reactor out performed the 37 °C reactor, effluent acetate levels from R2 were generally higher than from R1, reaching values as high as 5023 mg l⁻¹. Homoacetogenic activity in both reactors was confirmed on day 419 by specific acetogenic activity (SAA) measurement, with higher values obtained for R2 than R1.Sodium formate was introduced as sole substrate to both reactors on day 464. It was found that formate supported acetogenic activity at both temperatures. By the end of the trial, no specific methanogenic activity (SMA) was observed against acetate and propionate indicating that the methane produced was solely by hydrogenotrophic Archaea. Higher SMA and SAA values against H₂/CO₂ suggested development of a formate utilising acetogenic population growing in syntrophy with hydrogenotrophic methanogens. Throughout the formate trial, the mesophilic reactor performed better overall than the thermophilic reactor.     

Thermophilic fermentative hydrogen production from various carbon sources by anaerobic mixed cultures

In the present work, various carbon sources, xylose, glucose, galactose, sucrose, cellobiose, and starch were tested for thermophilic (60 °C) fermentative hydrogen production (FHP) by using the anaerobic mixed culture. An inoculum was obtained from a continuously-stirred tank reactor (CSTR) operated at pH 5.5 and HRT 12 h, and fed with tofu processing waste. The dominant species in the CSTR were found to be Thermoanaerobacterium thermosaccharolyticum and Clostridium thermosaccharolyticum, which are well known thermophilic H₂-producers in anaerobic-state, and have the ability to utilize a wide range of carbohydrates. When initial pH was adjusted to 6.8 ± 0.1 but not controlled during fermentation, vigorous pH drop began within 5 h, and finally reached 4.0–4.5 in all carbon sources. Although over 90% of substrate removal was achieved for all carbon sources except cellobiose (71.7%), the fermentation performances were profoundly different with each other. Glucose, galactose, and sucrose exhibited relatively higher H₂ yields whereas lower H₂ yields were observed for xylose, cellobiose, and starch. On the other hand, when pH was controlled (pH ≥ 5.5), the fermentation performance was enhanced in all carbon sources but to a different extent. A substantial increase in H₂ production was observed for cellobiose, a 1.9-fold increase of H₂ yield along with a substrate removal increase to 93.8%, but a negligible increase for xylose. H₂ production capabilities of all carbon sources tested were as follows: sucrose > galactose > glucose > cellobiose > starch > xylose. The maximum H₂ yield of 3.17 mol H₂/mol hexose_added achieved from sucrose is equivalent to a 26.5% conversion of energy content in sucrose to H₂. Acetic and butyric acids were the main liquid-state metabolites of all carbon sources while lactic acid was detected only in cellobiose, starch and xylose exhibiting relatively lower H₂ yields.     

Effects of substrate loading and co-substrates on thermophilic anaerobic conversion of microcrystalline cellulose and microbial communities revealed using high-throughput sequencing

Batch tests were conducted to investigate the effect of co-substrates, including glucose, xylose and starch, on thermophilic anaerobic conversion of microcrystalline cellulose using mixed culture enriched from anaerobic digestion sludge (ADS). Up to 30.9% of cellulose was utilized with xylose as co-substrate. When using glucose as co-substrate, cellulose conversion rate reached the maximum of 0.048 g/l/h at cellulose loading of 5.0 g/l. Illumina high-throughput sequencing of the 16S rRNA gene revealed that the thermophilic consortium exclusively consisted of Clostridium (more than 70% of all sequences). Growth of Thermoanaerobacterium over Clostridium would inhibit cellulose conversion capacity of the consortium. But the growth of Thermoanaerobacterium could be repressed by pH higher than pH 6.0. Co-substrates caused noticeable variation of bacterial community structure. Predominance of Thermoanaerobacterium over Clostridium was observed when monosugars (glucose and xylose) were used as co-substrate without pH control. Starch was ineffective as co-substrate because it competed with cellulose for Clostridium.     

Hydrogen gas production from cheese whey powder (CWP) solution by thermophilic dark fermentation

Hydrogen gas production from cheese whey powder (CWP) solution by thermophilic dark fermentation was investigated at 55 °C. Experiments were performed at different initial total sugar concentrations varying between 5.2 and 28.5 g L⁻¹ with a constant initial bacteria concentration of 1 g L⁻¹. The highest cumulative hydrogen evolution (257 mL) was obtained with 20 g L⁻¹ total sugar (substrate) concentration within 360 h while the highest H₂ formation rate (2.55 mL h⁻¹) and yield (1.03 mol H₂ mol⁻¹ glucose) were obtained at 5.2 and 9.5 g L⁻¹ substrate concentrations, respectively. The specific H₂ production rate (SHPR = 4.5 mL h⁻¹ g⁻¹cells) reached the highest level at 20 g L⁻¹ total sugar concentration. Total volatile fatty acid (TVFA) concentration increased with increasing initial total sugar content and reached the highest level (14.15 g L⁻¹) at 28.5 g L⁻¹ initial substrate concentration. The experimental data was correlated with the Gompertz equation and the constants were determined. The optimum initial total sugar concentration was 20 g L⁻¹ above which substrate and product (VFA) inhibitions were observed.     

Media optimization for biohydrogen production from waste glycerol by anaerobic thermophilic mixed cultures

Media compositions affecting thermophilic biohydrogen production from waste glycerol were optimized using response surface methodology (RSM) with central composite design (CCD). Investigated parameters used were waste glycerol concentration, urea concentration, the amount of Endo-nutrient addition and disodium hydrogen phosphate (Na₂HPO₄) concentration. Waste glycerol concentration and the amount of Endo-nutrient addition had a significant individual effect on the cumulative hydrogen production (HP) (p ≤ 0.05). The interactive effect on HP was found between waste glycerol and urea concentration as well as waste glycerol concentration and the amount of Endo-nutrient addition (p ≤ 0.05). The optimal media compositions were 20.33 g/L of waste glycerol, 0.16 g/L of urea, 3.97 g/L of Na₂HPO₄ and 0.20 mL/L of the amount of Endo-nutrient addition which gave the maximum HP of 1470.19 mL H₂/L. The difference between observed HP (1502.84 mL H₂/L) and predicted HP was 2.22%. The metabolic products from the fermentation process were 1,3-propanediol (1,3-PD), ethanol, acetic, formic, lactic, butyric, and propionic acids. Results from polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) analysis indicated that the hydrogen producers present in the fermentation broth was Thermoanaerobacterium sp.     

Co-culture of Clostridium thermocellum and Clostridium thermosaccharolyticum for enhancing hydrogen production via thermophilic fermentation of cornstalk waste

A strategic method utilizing the co-culture of Clostridium thermocellum and Clostridium thermosaccharolyticum has been developed to improve hydrogen production via the thermophilic fermentation of cornstalk waste. The hydrogen yield in the co-culture fermentation process reached 68.2 mL/g-cornstalk which was 94.1% higher than that in the mono-culture. The hydrogen fermentation process was successfully scaled-up from 125 mL anaerobic bottles to an 8 L continuous stirred tank reactor, and the hydrogen production from cornstalk waste was significantly improved in the bioreactor system due to efficient mixing and mass transfer. The hydrogen yield in the bioreactor reached 74.9 mL/g-cornstalk which was 9.8% higher than that in the 125 mL anaerobic bottle. The present work indicates that the direct microbial conversion of lignocellulosic waste by co-culturing C. thermocellum and C. thermosaccharolyticum is a promising avenue for enhancing hydrogen production.