Comparative Genomic Analysis of Sulfurospirillum cavolei MES Reconstructed from the Metagenome of an Electrosynthetic Microbiome

Sulfurospirillum spp. play an important role in sulfur and nitrogen cycling, and contain metabolic versatility that enables reduction of a wide range of electron acceptors, including thiosulfate, tetrathionate, polysulfide, nitrate, and nitrite. Here we describe the assembly of a Sulfurospirillum genome obtained from the metagenome of an electrosynthetic microbiome. The ubiquity and persistence of this organism in microbial electrosynthesis systems suggest it plays an important role in reactor stability and performance. Understanding why this organism is present and elucidating its genetic repertoire provide a genomic and ecological foundation for future studies where Sulfurospirillum are found, especially in electrode-associated communities. Metabolic comparisons and in-depth analysis of unique genes revealed potential ecological niche-specific capabilities within the Sulfurospirillum genus. The functional similarities common to all genomes, i.e., core genome, and unique gene clusters found only in a single genome were identified. Based upon 16S rRNA gene phylogenetic analysis and average nucleotide identity, the Sulfurospirillum draft genome was found to be most closely related to Sulfurospirillum cavolei. Characterization of the draft genome described herein provides pathway-specific details of the metabolic significance of the newly described Sulfurospirillum cavolei MES and, importantly, yields insight to the ecology of the genus as a whole. Comparison of eleven sequenced Sulfurospirillum genomes revealed a total of 6246 gene clusters in the pan-genome. Of the total gene clusters, 18.5% were shared among all eleven genomes and 50% were unique to a single genome. While most Sulfurospirillum spp. reduce nitrate to ammonium, five of the eleven Sulfurospirillum strains encode for a nitrous oxide reductase (nos) cluster with an atypical nitrous-oxide reductase, suggesting a utility for this genus in reduction of the nitrous oxide, and as a potential sink for this potent greenhouse gas.     

初始pH值对产氢产乙酸/耗氢产乙酸两段耦合工艺定向生产乙酸的影响

以葡萄糖为底物,以经加热预处理并活化过的厌氧污泥为种泥,研究了初始pH值对产氢产乙酸/耗氢产乙酸两段耦合工艺厌氧发酵定向生产乙酸的影响。实验考察了7个初始pH值(5、6、7、8、9、10、11)条件下的底物降解、产物产生和发酵过程pH值的变化。结果表明:产氢产乙酸段初始pH值的变化不仅影响本阶段产酸,而且影响耗氢产乙酸段产酸。初始pH=5时主要进行乙醇型发酵;pH=6和7时主要进行丁酸型发酵;pH=8时混合酸型发酵类型逐渐占优势,pH=8~11时均以乙酸为主要产物,耦合系统生产乙酸最优初始pH值为10。在初始pH=8~11范围内,产氢产乙酸段初期的乙醇浓度一般较高,但到后期因乙醇被微生物进一步代谢转化成乙酸而使其含量下降。     

Effects of pH and Lactate on Hydrogen Sulfide Production by Oral Veillonella spp.

Indigenous oral bacteria in the tongue coating such as Veillonella have been identified as the main producers of hydrogen sulfide (H₂S), one of the major components of oral malodor. However, there is little information on the physiological properties of H₂S production by oral Veillonella such as metabolic activity and oral environmental factors which may affect H₂S production. Thus, in the present study, the H₂S-producing activity of growing cells, resting cells, and cell extracts of oral Veillonella species and the effects of oral environmental factors, including pH and lactate, were investigated. Type strains of Veillonella atypica, Veillonella dispar, and Veillonella parvula were used. These Veillonella species produced H₂S during growth in the presence of l-cysteine. Resting cells of these bacteria produced H₂S from l-cysteine, and the cell extracts showed enzymatic activity to convert l-cysteine to H₂S. H₂S production by resting cells was higher at pH 6 to 7 and lower at pH 5. The presence of lactate markedly increased H₂S production by resting cells (4.5- to 23.7-fold), while lactate had no effect on enzymatic activity in cell extracts. In addition to H₂S, ammonia was produced in cell extracts of all the strains, indicating that H₂S was produced by the catalysis of cystathionine γ-lyase (EC 4.4.1.1). Serine was also produced in cell extracts of V. atypica and V. parvula, suggesting the involvement of cystathionine β-synthase lyase (EC 4.2.1.22) in these strains. This study indicates that Veillonella produce H₂S from l-cysteine and that their H₂S production can be regulated by oral environmental factors, namely, pH and lactate.     

Acetic Acid Production by Clostridium thermoaceticum in pH-Controlled Batch Fermentations at Acidic pH

Four strains of the homofermentative, obligately anaerobic thermophile Clostridium thermoaceticum were compared in pH-controlled batch fermentation for their tolerance to acetic acid, efficiency of converting glucose to acetic acid and cell mass, and growth rate. At pH 6 (and pH 7) and initial acetic acid concentrations of less than 10 g/liter, the four strains had mass doubling times of 5 to 7 h and conversion efficiencies to acetic acid and cell mass of about 90% (70 to 110%) and 10%, respectively. At pH 6 and initial acetic acid concentrations of greater than 10 g/liter, only two of the strains grew, the mass doubling time increased to 18 h, and the conversion efficiencies to acetic acid and cell mass remained unchanged. Both of these strains had been selected for their ability to grow in the presence of acetate at neutral pH. The highest acetic acid concentrations reached were about 15 and 20 g/liter at pH 6 and 7, respectively. C. thermoaceticum is apparently more sensitive to free acetic acid than to either acetate ion or pH. It was also shown that, at pH 6 and 7, the redox potential must be at least as low as −300 and −360 mV, respectively, for growth to occur.     

Hydrogen Gas Production by an Ectothiorhodospira vacuolata Strain

A hydrogen gas (H₂)-producing strain of Ectothiorhodospira vacuolata isolated from Soap Lake, Washington, possessed nitrogenase activity. Increasing evolution of H₂ with decreasing ammonium chloride concentrations provided evidence that nitrogenase was the catalyst in gas production. Cells were grown in a mineral medium plus 0.2% acetate with sodium sulfide as an electron donor. Factors increasing H₂ production included addition of reduced carbon compounds such as propionate and succinate, increased reducing power by increasing sodium sulfide concentrations, and increased energy charge (ATP) by increasing light intensity.     

沼泽红假单胞菌乙酸光合放氢研究

依据光合细菌生长代谢特性和有机废水降解主要产物类型,11种有机物被用于沼泽红假单胞菌（Rhodopseudomonas palustris）Z菌株的光合产氢研究,其中,乙酸反应体系产氢活性最高。在此基础上,研究了该菌株的生长与产氢动力学行为,探求了影响该菌株光合放氢的主要限制性影响因素。结果表明,该菌株产氢与生长部分相关。种子培养基和菌龄对产氢活性有明显影响。细胞最适产氢和生长所需要的光照强度和温度基本一致。当种子来源于硫酸铵高菌龄预培养物或谷氨酸钠对数期预培养物时,该菌株产氢活性显著增加,产氢延滞期明显缩短。氧浓度和接种量对产氢活性也有显著影响。供氢体和氮源浓度直接决定细胞的生长与光放氢活性。在低于70 mmol/L乙酸钠和15 mmol/L谷氨酸钠时,产氢活性随底物浓度的增加而增强。谷氨酸钠浓度高于15mmol/L时,由于游离NH₄⁺的出现,产氢活性受到抑制,但却明显刺激细胞的生长。在标准状况下,该菌株的最大产氢速率可达19.4 mL·L^-1·h^-1。     

Acetate Production by Methanogenic Bacteria

Methanosarcina barkeri MS and 227 and Methanosarcina mazei S-6 produced acetate when grown on H₂-CO₂, methanol, or trimethylamine. Marked differences in acetate production by the two bacterial species were found, even though methane and cell yields were nearly the same. M. barkeri produced 30 to 75 μmol of acetate per mmol of CH₄ formed, but M. mazei produced only 8 to 9 μmol of acetate per mmol of CH₄.     

Influence of Sulfate-Reducing Bacteria on Outdoor Hydrogen Production by Photosynthetic Bacterium with Seawater

The application of seawater for bacterial fermentative production is a cost-effective technology. Hydrogen production by marine photosynthetic bacterium with seawater failed to continue after more than 10 days, and was accompanied by the formation of hydrogen sulfide and a change in culture color from red to black. However, substrate consumption in the blackish culture was comparable to that in a hydrogen-producing culture. A decrease in hydrogen production occurred upon the addition of sodium sulfide at concentrations of 1.5 mM or higher. PCR analysis targeted at the 16S rDNA sequence selective for sulfate-reducing bacteria revealed the existence of sulfate-reducing bacteria in inoculation cultures of the phototrophic bacterium and medium for hydrogen production. Hence, the high sulfate concentration of seawater, the low oxidation-reduction potential under hydrogen-producing conditions, and the presence of electron donors such as acetate might promote the metabolic activities of sulfate-reducing bacteria, resulting in the deterioration of hydrogen production with seawater. Received: 15 September 1999 / Accepted: 14 October 1999     

Influence of Culture Parameters on Biological Hydrogen Production by Clostridium saccharoperbutylacetonicum ATCC 27021

Summary Various medium components (carbon and nitrogen sources, iron, inoculum size) and environmental factors (initial pH and the agitation speed) were evaluated for their effects on the rate and the yield of hydrogen production by Clostridium saccharoperbutylacetonicum. Among the carbon sources assessed, cells grown on disaccharides (lactose, sucrose and maltose) produced on the average more than twice (2.81 mol-H2/mol sugar) as much hydrogen as monosaccharides (1.29 mol-H2/mol sugar), but there was no correlation between the carbon source and the production rate. The highest yield (2.83 mol/mol) was obtained in lactose and sucrose but the highest production rate (1.75 mmol/h) in sucrose. Using glucose as carbon source, yeast extract was the best nitrogen source. A parallel increase between the production rate and the yield was obtained by increasing glucose concentration up to 40 g/l (1.76 mol-H2/mol, 3.39 mmol/h), total nitrogen as yeast extract up to 0.1% (1.41 mol/mol, 1.91 mmol/h) and agitation up to 100 rev/min (1.66 mol-H2/mol, 1.86 mmol/h). On the other hand, higher production rates were favoured in preference to the yield at a neutral initial pH 7 (2.27 mmol/h), 1000 mg iron/l or more (1.99 mmol/h), and a larger inoculum size, 10%, (2.36 mmol/h) whereas an initial alkaline pH of 8.5 (1.72 mol/mol), a lower iron concentration of 25 mg/l (1.74 mol/mol) and smaller inoculum size, 1%, (1.85 mol/mol) promoted higher yield over production rate.     

pH控制对热凝胶发酵的影响

热凝胶 (Ｃｕｒｄｌａｎ)是一种直链结构的 β 1,3 葡聚糖 ,由Ａｌｃａｌｉｇｅｎｅｓｆａｅｃａｌｉｓｖａｒ.ｍｙｘｏｇｅｎｅｓ发酵生产而来 ,是一种新型的微生物胞外多糖[1 ] ,其分子量在 5 0万左右。热凝胶在中性条件下不溶于水 ,但能溶于碱溶液中。加热含有热凝胶的水浊液可形成两种类型的凝胶 ,一种是弹性较低的类似琼脂的可逆胶 ;另外一种是凝胶强度大、弹性好的热不可逆胶。由于热凝胶具有独特的热成胶性能 ,在食品工业 ,特别是高温制作的食品领域具有广阔的应用前景。热凝胶的胶体可以包容和控制药物的扩散 ,所以可以用来作为药物…     

Influence of Hyperthyroidism on Superoxide Radical and Hydrogen Peroxide Production by Rat Liver Submitochondrial Particles

Administration of daily doses of 0.1 mg of 3, 5, 3'-triiodothyronine (T₃)/kg body weight for 3 consecutive days to fed rats elicited a calorigenic response in the animals, in concomitance with a 36% increase in the rate of O₂ consumption by the liver. In these conditions, liver submitochondrial particles (SMP) from T₃-treated rats exhibited marked increases in the rate of superoxide radical generation, both in the presence of NADH (142%) or succinate (152%). Furthermore, liver SMP from hyperthyroid animals released hydrogen peroxide at higher rates than those of euthyroid rats, either under basal conditions or in the succinate-supported process, both in the absence and presence of antimycin-A. It is concluded that the hyperthyroid state in the rat leads to a drastic enhancement in the capacity of liver mitochondria to produce active oxygen species, which correlates with the elevated respiratory rate observed in the intact organ.     

State-dependent Inhibition of TRPM2 Channel by Acidic pH

Transient receptor potential melastatin 2 (TRPM2) channel fulfills an important role in oxidative stress signaling in immune and other cells, to which local extracellular acidosis is known to occur under physiological or pathological conditions and impose significant effects on their functions. Here, we investigated whether the ADP-ribose-activated TRPM2 channel is a target for modulation by extracellular acidic pH by patch clamp recording of HEK293 cells expressing hTRPM2 channel. Induced whole cell or single channel currents were rapidly inhibited upon subsequent exposure to acidic pH. The inhibition in the steady state was complete, voltage-independent, and pH-independent in the range of pH 4.0–6.0. The inhibition was irreversible upon returning to pH 7.3, suggesting channel inactivation. In contrast, exposure of closed channels to acidic pH reduced the subsequent channel activation in a pH-dependent manner with an IC₅₀ for H⁺ of 20 μm (pH 4.7) and rendered subsequent current inhibition largely reversible, indicating differential or state-dependent inhibition and inactivation. Alanine substitution of residues in the outer vestibule of the pore including Lys⁹⁵² and Asp¹⁰⁰² significantly slowed down or reduced acidic pH-induced inhibition and prevented inactivation. The results suggest that acidic pH acts as a negative feedback mechanism where protons bind to the outer vestibule of the TRPM2 channel pore and inhibit the TRPM2 channels in a state-dependent manner.     

Influence of Veratryl Alcohol and Hydrogen Peroxide on Ligninase Activity and Ligninase Production by Phanerochaete chrysosporium

Veratryl alcohol, added as a supplement to cultures of Phanerochaete chrysosporium, enhanced ligninase activity through protection of the ligninase against inactivation by hydrogen peroxide produced by this fungus in cultures. In the presence of veratryl alcohol, the loss of ligninase activity observed in non-protein-synthesizing cultures (cycloheximide-treated) equaled the extracellular protein turnover. When cultures were not supplemented with veratryl alcohol, inactivation of ligninase by hydrogen peroxide added to protein turnover, resulting in a more rapid loss of ligninase activity. Although all ligninase isoenzymes are sensitive to inactivation by hydrogen peroxide, only the isoenzyme of the highest specific activity (80.6 nkat · mg of protein⁻¹; M_r, 41,800; pI, 3.96) was found to be protected by veratryl alcohol. The concentration of veratryl alcohol necessary for full protection of ligninase activity varied according to the concentration of hydrogen peroxide present in the medium, which depended on the nature of the carbon source (glucose or glycerol). It is proposed that the nature of the carbon source influences the overall ligninase activity not only directly, by affecting the rate and the type of synthesized ligninase, but also by affecting the rate of hydrogen peroxide production, bringing about different rates of inactivation.     

Influence of Oxygen and pH on Methanethiol Production from l-Methionine by Brevibacterium linens CNRZ 918

The effects of dissolved oxygen concentration and pH on the growth of Brevibacterium linens CNRZ 918 and its production of methanethiol from l-methionine were investigated. Optimal specific methanethiol production was obtained at 25% saturation of dissolved oxygen and at a pH between 8 and 9, whereas optimal cell growth occurred at 50% oxygen saturation and when the pH was maintained constantly at 7. Methanethiol production by nonproliferating bacteria required the presence of l-methionine (7 mM) in the culture medium. This was probably due to the induction of enzyme systems involved in the process. The intracellular concentration of l-methionine seemed to play a key role in this process. B. linens CNRZ 918 tolerated alkaline pHs with a maximal growth pH of approximately 9. Its orange pigmentation seemed to depend on the presence of l-methionine in the culture medium and on the concentration of dissolved oxygen.     

Effects of Hydrogen Pressure during Growth and Effects of Pregrowth with Hydrogen on Acetate Degradation by Methanosarcina Species

Methanosarcina barkeri 227 and Methanosarcina mazei S-6 grew with acetate as the substrate; we found little effect of H₂ on the rate of aceticlastic growth in the presence of various H₂ pressures between 2 and 810 Pa. We used physical (H₂ addition or flushing the headspace to remove H₂) and biological (H₂-producing or -utilizing bacteria in cocultures) methods for controlling H₂ pressure in Methanosarcina cultures growing on acetate. Added H₂ (ca. 100 Pa) was removed rapidly (a few hours) by M. barkeri and slowly (within a day) by M. mazei. When the H₂ produced by the aceticlastic methanogens was removed by coculturing with an H₂-using Desulfovibrio sp., the H₂ pressure was about 2.2 Pa. Under these conditions the stoichiometry of aceticlastic methanogenesis did not change. H₂-grown inocula of M. barkeri grew with acetate as the sole catabolic substrate if the inoculum culture was transferred during logarithmic growth to acetate-containing medium or if the transfer was accomplished within 1 or 2 days after exhaustion of H₂. H₂-grown cultures incubated for 4 or more days after exhaustion of H₂ were able to grow with H₂ but not with acetate as the sole catabolic substrate. Addition of small quantities of H₂ to acetate-containing medium permitted these cultures to initiate growth on acetate.     

Influence of pH on Terminal Carbon Metabolism in Anoxic Sediments from a Mildly Acidic Lake

The carbon and electron flow pathways and the bacterial populations responsible for transformation of H₂-CO₂, formate, methanol, methylamine, acetate, glycine, ethanol, and lactate were examined in sediments collected from Knaack Lake, Wis. The sediments were 60% organic matter (pH 6.2) and did not display detectable sulfate-reducing activity, but they contained the following average concentration (in micromoles per liter of sediment) of metabolites and end products: sulfide, 10; methane, 1,540; CO₂, 3,950; formate, 25; acetate, 157; ethanol, 174; and lactate, 138. Methane was produced predominately from acetate, and only 4% of the total CH₄ was derived from CO₂. Methanogenesis was limited by low environmental temperature and sulfide levels and more importantly by low pH. Increasing in vitro pH to neutral values enhanced total methane production rates and the percentage of CO₂ transformed to methane but did not alter the amount of ¹⁴CO₂ produced from [2-¹⁴C]acetate (~24%). Analysis of both carbon transformation parameters with ¹⁴C-labeled tracers and bacterial trophic group enumerations indicated that methanogenesis from acetate and both heterolactic- and acetic acid-producing fermentations were important to the anaerobic digestion process.     

The Effects of pH on a Periphyton Community in an Acidic Wetland, USA

Despite the importance of peatlands, the algal ecology of peatlands and the periphyton communities which are abundant in these habitats are relatively understudied. We performed an in situ manipulation of pH in an intermediate fen in northern lower Michigan in order to examine how hydrogen ion concentrations structure an epiphytic algal community. Levels of pH were manipulated in enclosures from the control level (pH = 5) to an acid treatment (pH = 4) by adding H₂SO₄ and a neutral treatment (pH = 7) by adding NaOH. Algal communities growing on sections of Chamaedaphne calyculata (L.) Moench stems were examined after 22 days of colonization. Chlorophyll a concentration was significantly greater only in the acid treatment (~5.5 mg m⁻²) relative to the control (~3.5 mg m⁻²). Taxa richness was lower in the acid treatment. The algal assemblages were dominated by filamentous green algae and a filamentous taxon, Mougeotia spp., was significantly greater in the acid treatment relative to the control. Increases in Zygnemataceae and Oedogonium spp. most likely account for the higher chlorophyll a in the acid treatment. Most treatment differences were detected in the neutral treatment, including increased abundances of Closterium polystichum Nygaard, Cosmarium sp., Peridinium inconspicuum Lemmermann, and Synedra acus Kütz. Unexpectedly, there was no strong response of the desmid community. These data can be informative in the development of algal monitoring programs in peatlands when assessment of acidification is desired.     

Temperature Compensation in Methanosarcina barkeri by Modulation of Hydrogen and Acetate Affinity

The affinity of Methanosarcina barkeri 227 for acetate and hydrogen at different incubation temperatures was investigated. Increasing the temperature from 20 to 37°C resulted in a 4.5-fold increase in K_m for acetate and a 4.8-fold increase for hydrogen. The corresponding increase in V_max for acetate was 8.3-fold (5.4-fold for hydrogen). This response implied a decrease in the temperature coefficient (Q₁₀) and hence a decrease in the temperature dependency as a function of decreasing substrate concentration.     

Acidic Nanoparticles Are Trafficked to Lysosomes and Restore an Acidic Lysosomal pH and Degradative Function to Compromised ARPE-19 Cells

Lysosomal enzymes function optimally in acidic environments, and elevation of lysosomal pH can impede their ability to degrade material delivered to lysosomes through autophagy or phagocytosis. We hypothesize that abnormal lysosomal pH is a key aspect in diseases of accumulation and that restoring lysosomal pH will improve cell function. The propensity of nanoparticles to end up in the lysosome makes them an ideal method of delivering drugs to lysosomes. This study asked whether acidic nanoparticles could traffic to lysosomes, lower lysosomal pH and enhance lysosomal degradation by the cultured human retinal pigmented epithelial cell line ARPE-19. Acidic nanoparticles composed of poly (DL-lactide-co-glycolide) (PLGA) 502 H, PLGA 503 H and poly (DL-lactide) (PLA) colocalized to lysosomes of ARPE-19 cells within 60 min. PLGA 503 H and PLA lowered lysosomal pH in cells compromised by the alkalinizing agent chloroquine when measured 1 hr. after treatment, with acidification still observed 12 days later. PLA enhanced binding of Bodipy-pepstatin-A to the active site of cathepsin D in compromised cells. PLA also reduced the cellular levels of opsin and the lipofuscin-like autofluorescence associated with photoreceptor outer segments. These observations suggest the acidification produced by the nanoparticles was functionally effective. In summary, acid nanoparticles lead to a rapid and sustained lowering of lysosomal pH and improved degradative activity.     

Mercury Methylation by Interspecies Hydrogen and Acetate Transfer between Sulfidogens and Methanogens

Cocultures of Desulfovibrio desulfuricans and Methanococcus maripaludis grew on sulfate-free lactate medium while vigorously methylating Hg²⁺. Individually, neither bacterium could grow or methylate mercury in this medium. Similar synergistic growth of sulfidogens and methanogens may create favorable conditions for Hg²⁺ methylation in low-sulfate anoxic freshwater sediments.