Purple Non-sulfur Photosynthetic Bacteria Research Articles

Multidimensional engineering of Rhodobacter sphaeroides for enhanced photo-fermentative hydrogen production

The photo-fermentative hydrogen production by purple non-sulfur photosynthetic bacteria (PNSB) is regarded as a promising approach for sustainable hydrogen production. In PNSB, biohydrogen synthesis is catalyzed by nitrogenase which requires two essential cofactors of adenosine triphosphate (ATP) and electron. To avoid the insufficient ATP supply that limits the nitrogenase activity for hydrogen production, we systematically engineered Rhodobacter sphaeroides to improve the hydrogen production by manipulating the light harvesting (LH) complexes, the electron transfer chain (ETC), the adenosine diphosphate (ADP) synthetic pathway, the F0F1-ATPase expression, and the nitrogenase expression. To improve the photosynthetic efficiency, we engineered the LH complexes by eliminating pucBA and overexpressing pufQ and spbA. Then, the ETC was boosted by deleting qxt-type oxidase and integrating the electron carrier cytochrome cy at the site of cyt cbb3 oxidase. In addition, we increased the supply of precursor ADP by overexpressing adk, and the abundance of F0F1-ATPase was efficiently enhanced through overexpressing atpXF. Further transposon engineering with the integration of nifHDK and atpXF resulted in a mutant YH27 with a substantial improvement of performance over the base strain, and its nitrogenase activity and hydrogen production were enhanced by 404 % and 93 %, respectively. Taken together, the multidimensional strategies enabled the engineered R. sphaeroides with a hydrogen productivity of 61.2 mL/(Lh) and a yield of 3.0 mol/mol acetate under the fed-batch condition, which represents one of the most efficient biohydrogen-producing strains.

Multidimensional optimization for accelerating light-powered biocatalysis in Rhodopseudomonas palustris

BackgroundWhole-cell biocatalysis has been exploited to convert a variety of substrates into high-value bulk or chiral fine chemicals. However, the traditional whole-cell biocatalysis typically utilizes the heterotrophic microbes as the biocatalyst, which requires carbohydrates to power the cofactor (ATP, NAD (P)H) regeneration.ResultsIn this study, we sought to harness purple non-sulfur photosynthetic bacterium (PNSB) as the biocatalyst to achieve light-driven cofactor regeneration for cascade biocatalysis. We substantially improved the performance of Rhodopseudomonas palustris-based biocatalysis using a highly active and conditional expression system, blocking the side-reactions, controlling the feeding strategy, and attenuating the light shading effect. Under light-anaerobic conditions, we found that 50 mM ferulic acid could be completely converted to vanillyl alcohol using the recombinant strain with 100% efficiency, and > 99.9% conversion of 50 mM p-coumaric acid to p-hydroxybenzyl alcohol was similarly achieved. Moreover, we examined the isoprenol utilization pathway for pinene synthesis and 92% conversion of 30 mM isoprenol to pinene was obtained.ConclusionsTaken together, these results suggested that R. palustris could be a promising host for light-powered biotransformation, which offers an efficient approach for synthesizing value-added chemicals in a green and sustainable manner.Graphical

Reconstituted LH2 in multilayer membranes induced by poly-l-lysine: Structure of supramolecular and electronic states

To explore the effect of cell membrane stacking on the function of light-harvesting complex 2 (LH2) in purple nonsulfur photosynthetic bacteria, LH2 from Rhodobacter sphaeroides 2.4.1 (R. sph 2.4.1) was reconstituted into lipid bilayer vesicles (LH2@liposome) and further formed multi-layer structure by electrostatic interaction with poly-l-lysine (LH2@liposome/PLL), which was characterized by cryo-electron microscopy (cryo-EM) and TEM. When embedded in liposomes and additionally in multi-layer liposomes, the absorption band, zero-crossing point of CD signals and fluorescence emission of LH2 B850 excitons were observed to uniformly have 1–2 nm red-shifting. Combining with the corresponding fluorescence quench and the generation of shorter-living fluorescence species, a new excitonic species generated through B850 structural splitting was proposed. By FT-Raman and triplet carotenoid dynamics, the structural mechanism was deduced and discussed. Briefly, all environmental changes, including LH2 aggregating and multi-layer membrane stacking, eventually applied forces on B850 exciton molecules mainly through the hydrogen bonding between the C3-acetyl carbonyl groups of B850 BChls and Tyr44 and 45 residues at C-terminus of α-polypeptides. The effect of multi-layer structure on LH2 could be assigned as a kind of photoprotection.

Biyorafineri konseptiyle Rhodobacter sphaeroides O.U.001 ile biyolojik hidrojen ve bakteriyel karotenoid üretimi

In this study, the goal was to produce biohydrogen and bacterial carotenoids with Rhodobacter sphaeroides O.U.001, a purple non-sulfur photosynthetic bacterium, utilizing sugar beet molasses in the context of biorefinery. First, media with different sugar concentrations (10 g/L, 20 g/L, 30 g/L, 40 g/L, 50 g/L) were prepared for bacterial growth. Then, hydrogen production was carried out using these media in anaerobic conditions in 100 ml bioreactors. After hydrogen gas was collected from the bioreactors, carotenoid extraction was performed from the remaining bacteria. As a result of the analyzes, it was found that the amount of biohydrogen and the amount of bacterial carotenoids obtained were inversely proportional to the increased sugar concentrations. The maximum hydrogen formation was detected in the medium containing 10 g/L of sugar (19.18 mL). According to the results of gas chromatography analysis, the quantity of hydrogen in the total gas was found to be around 23.6%. The highest yield of carotenoids was again obtained from bacteria reproduced in a medium containing 10 g/L of sugar (3.12 mg/g, carotenoid/dry biomass). As a conclusion, this study provides an example for the successful realization of two high value-added products within a biorefinery approach by using molasses obtained at an affordable cost.

Effect and mechanism of the improvement of coastal silt soil by application of organic fertilizer and gravel combined with Sesbania cannabina cultivation.

Jiangsu Province of China has a large area of coastal silt soil (CSS) with poor permeability, high salinity, and poor nutrients, which brings great difficulties to the development and utilization of coastal zones, so that needs to be improved as a matter of urgency. In this study, river-sand, serpentine, and organic fertilizer were used as additives in CSS, and Sesbania cannabina, a salt-tolerant cash crop, was planted in these differently treated soils. Through high-throughput sequencing, analysis of soil physico-chemical properties, and detection of plant growth status, the rhizosphere bacterial diversity of S. cannabina growing in CSS under different treatments and their environmental impact factors were studied, while exploring the effect and mechanism of organic fertilizer combined with gravel as a CSS modifier. The results implied that single application of organic fertilizer could significantly increase the fertility levels of total nitrogen (TN), total organic carbon (TOC) and Avail. P in CSS; then, the application of organic fertilizer with river-sand significantly reduced salt content and alkalinity of soil; meanwhile, in the treatment of single application of organic fertilizer and application of organic fertilizer combined with river-sand, the rhizosphere of S. cannabina enriched the bacterial communities of organic matter degradation and utilization to varying degrees. The soil moisture content and indicators related to saline-alkali soil (including total salt, electrical conductivity (EC), exchangeable sodium percentage (ESP), Avail. Na and Avail. K, etc.) were further reduced significantly by the application of organic fertilizer combined with river-sand and serpentine. The method has greatly improved the growth conditions of S. cannabina and promoted the positive development of its rhizosphere bacterial community. Among them, in the treatment of organic fertilizer combined with river-sand and serpentine, a variety of plant growth-promoting rhizobacteria (PGPR, such as Sphingomonas, Ensifer, and Rhodobacter) and nitrogen-cycle-related bacteria (such as nitrate-reduction-related bacteria, nitrogen-fixing bacteria like Ensifer, and purple non-sulfur photosynthetic bacteria like Rhodobacter) were enriched in the rhizosphere of S. cannabina; moreover, the mutual association and robustness of bacterial co-occurrence networks have been significantly enhanced. The results provide a theoretical basis and reference model for the improvement of coastal saline-alkali silt soil.

Tuning photosynthetic oxygen for hydrogen evolution in synergistically integrated, sulfur deprived consortia of Coccomyxa chodatii and Rhodobium gokarnense at dim and high light

In this work, tuning oxygen tension was targeted to improve hydrogen evolution. To achieve such target, various consortia of the chlorophyte Coccomyxa chodatii with a newly isolated photosynthetic purple non-sulfur bacterium (PNSB) strain Rhodobium gokarnense were set up, sulfur replete/deprived, malate/acetate fed, bicarbonate/sulfur added at dim/high light. C. chodatii and R. gokarnense are newly introduced to biohydrogen studies for the first time. Dim light was applied to avoid the inhibitory drawbacks of photosynthetic oxygen evolution, values of hydrogen are comparable with high light or even more and thus economically feasible to eliminate the costs of artificial illumination. Particularly, the consortium of 2n− (n = 1.9 × 105 cell/ml, sulfur deprived) demonstrated its perfection for the target, i.e., the highest possible cumulative hydrogen. This consortium exhibited negative photosynthesis, i.e., oxygen uptake in the light. Most hydrogen in consortia is from bacterial origin, although algae evolved much more hydrogen than bacteria on per cell basis, but for only one day (the second 24 h), as kinetics revealed. The higher hydrogen in unibacterial culture or consortia results from higher bacterial cell density (20 times). Consortia evolved more hydrogen than their respective separate cultures, further enhanced when bicarbonate and sulfur were supplemented at higher light. The share of algae relatively increased as bicarbonate or sulfur were added at higher light intensity, i.e., PSII activity partially recovered, resulting in a transient autotrophic hydrogen evolution. The addition of acetic acid in mixture with malic acid significantly enhanced the cumulative hydrogen levels, mostly decreased cellular ascorbic acid indicating less oxidative stress and relief of PSII, relative to malic acid alone. Starch, however, decreased, indicating the specificity of acetic acid. Exudates (reducing sugars, amino acids, and soluble proteins) were detected, indicating mutual utilization. Yet, hydrogen evolution is limited; tuning PSII activity remains a target for sustainable hydrogen production.

Plant-Growth-Promoting Effect by Cell Components of Purple Non-Sulfur Photosynthetic Bacteria.

Rhodobacter sphaeroides, a purple non-sulfur photosynthetic bacterium (PNSB), was disrupted by sonication and fractionated by centrifugation into the supernatant and pellet. The effects of the supernatant and pellet on plant growth were examined using Brassica rapa var. perviridis (komatsuna) in the pot experiments. Both fractions showed growth-promoting effects: the supernatant at high concentrations (1 × 107 to 4 × 107 cfu-equivalent mL−1) and the pellet at a low concentration of 2 × 103 cfu-equivalent mL−1). We expected lipopolysaccharide (LPS) to be the active principle of the pellet fraction and examined the effects of LPS on the growth of B. rapa var. perviridis. The growth of the plants was significantly enhanced by the foliar feeding of R. sphaeroides LPS at concentrations ranging from 10 to 100 pg mL−1. The present study is the first report indicating that LPS acts as one of the active principles of the plant-growth-promoting effect of PNSB.

Comparison of the probiotic effects of freshwater and marine photosynthetic bacteria in kuruma shrimp (Marsupenaeus japonicus) culture

ABSTRACT In a field study, the probiotic effects of a freshwater purple nonsulfur photosynthetic bacterium (PNSB), Rhodobacter sphaeroides, and a marine PNSB, Rhodovulum sulfidophilum, on kuruma shrimp (Marsupenaeus japonicus) were compared by using two commercially operating shrimp ponds. Shrimp were fed with feed containing 0.01% fresh weight (106 cfu/g feed) of PNSB cells for 152 d. The marine PNSB showed a better performance than the freshwater PNSB in average body weight at harvest time, survival rate (P < 0.001 by Fisher’s exact test), and feed conversion rate. The effects of freshwater and marine PNSB on shrimp gene expression were also compared by next-generation sequencing and quantitative RT-PCR. Both freshwater and marine PNSB promoted the expression of several genes related to the innate immune system, including NADPH oxidase, superoxide dismutase, lysozyme, antimicrobial peptides, and lectins, and the promotive effect of marine PNSB was stronger than that of freshwater PNSB. The cost of PNSB was calculated to be US$ 0.003–0.005 per kg feed; thus, marine PNSB is a cost-effective and feasible probiotic for shrimp aquaculture.

Engineered Mutants of a Marine Photosynthetic Purple Nonsulfur Bacterium with Increased Volumetric Productivity of Polyhydroxyalkanoate Bioplastics.

Polyhydroxyalkanoates (PHAs) are green and sustainable bioplastics that could replace petrochemical synthetic plastics without posing environmental threats to living organisms. In addition, sustainable PHA production could be achieved using marine photosynthetic purple nonsulfur bacteria (PNSBs) that utilize natural seawater, sunlight, carbon dioxide gas, and nitrogen gas for growth. However, PHA production using marine photosynthetic PNSBs has not been economically feasible yet due to its high cost and low productivity. In this work, strain improvement, using genome-wide mutagenesis coupled with high-throughput screening via fluorescence-activated cell sorting, we were able to create Rhodovulum sulfidophilum mutants with enhanced volumetric PHA productivity, with an up to 1.7-fold increase. The best selected mutants (E6 and E6M4) reached the stationary growth phase 1 day faster and accumulated the maximum PHA content 2 days faster than the wild type. Maximizing volumetric PHA productivity before the stationary growth phase is indeed an additional advantage for R. sulfidophilum as a growth-associated PHA producer.

Extraction of Polyhydroxyalkanoates from Purple Non-Sulfur Bacteria by Non-Chlorinated Solvents.

In this study, non-chlorinated solvents such as cyclohexanone (CYC) and three ionic liquids, (ILs) (1-ethyl-3-methylimidazolium dimethylphosphate, [EMIM][DMP], 1-ethyl-3-methylimidazolium diethylphosphate, [EMIM][DEP] and 1-ethyl-3-methylimidazolium methylphosphite, [EMIM][MP]) were tested to extract polyhydroxyalkanoates (PHAs) from the purple non-sulfur photosynthetic bacterium (PNSB) Rhodovulum sulfidophilum DSM-1374. The photosynthetic bacterium was cultured in a new generation photobioreactor with 4 L of working volume using a lactate-rich medium. The extracted PHAs were characterized using a thermogravimetric analysis, differential scanning calorimetry, infrared spectroscopy, proton nuclear magnetic resonance and gel permeation chromatography. The most promising results were obtained with CYC at 125 °C with an extraction time of above 10 min, obtaining extraction yields higher than 95% and a highly pure poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHB-HV) with around 2.7 mol% of hydroxylvalerate (HV). A similar yield and purity were obtained with chloroform (CHL) at 10 °C for 24 h, which was used as the referent solvent Although the three investigated ILs at 60 °C for 4 and 24 h with biomass/IL up to 1/30 (w/w) obtained PHAs strongly contaminated by cellular membrane residues, they were not completely solubilized by the investigated ILs.

High-Yielding Terpene-Based Biofuel Production in Rhodobacter capsulatus.

Energy crisis and global climate change have driven an increased effort toward biofuel synthesis from renewable feedstocks. Herein, purple nonsulfur photosynthetic bacterium (PNSB) of Rhodobacter capsulatus was explored as a platform for high-titer production of a terpene-based advanced biofuel-bisabolene. A multilevel engineering strategy such as promoter screening, improving the NADPH availability, strengthening the precursor supply, suppressing the side pathways, and introducing a heterologous mevalonate pathway, was used to improve the bisabolene titer in R. capsulatus. The above strategies enabled a 35-fold higher titer of bisabolene than that of the starting strain, reaching 1089.7 mg/L from glucose in a shake flask. The engineered strain produced 9.8 g/L bisabolene with a yield of >0.196 g/g-glucose under the two-phase fed-batch fermentation, which corresponds to >78% of theoretical maximum. Taken together, our work represents one of the pioneering studies to demonstrate PNSB as a promising platform for terpene-based advanced biofuel production.

Biosynthesis of pinene in purple non-sulfur photosynthetic bacteria

BackgroundPinene is a monoterpene, that is used in the manufacture of fragrances, insecticide, fine chemicals, and renewable fuels. Production of pinene by metabolic-engineered microorganisms is a sustainable method. Purple non-sulfur photosynthetic bacteria belong to photosynthetic chassis that are widely used to synthesize natural chemicals. To date, researches on the synthesis of pinene by purple non-sulfur photosynthetic bacteria has not been reported, leaving the potential of purple non-sulfur photosynthetic bacteria synthesizing pinene unexplored.ResultsRhodobacter sphaeroides strain was applied as a model and engineered to express the fusion protein of heterologous geranyl diphosphate synthase (GPPS) and pinene synthase (PS), hence achieving pinene production. The reaction condition of pinene production was optimized and 97.51 μg/L of pinene was yielded. Then, genes of 1-deoxy-d-xylulose 5-phosphate synthase, 1-deoxy-d-xylulose 5-phosphate reductoisomerase and isopentenyl diphosphate isomerase were overexpressed, and the ribosome binding site of GPPS-PS mRNA was optimized, improving pinene titer to 539.84 μg/L.Conclusions In this paper, through heterologous expression of GPPS-PS, pinene was successfully produced in R. sphaeroides, and pinene production was greatly improved by optimizing the expression of key enzymes. This is the first report on pinene produce by purple non-sulfur photosynthetic bacteria, which expands the availability of photosynthetic chassis for pinene production.

The effects of a marine photosynthetic bacteria Rhodovulum sulfidophilum on the growth and survival rate of Marsupenaeus japonicus (kuruma shrimp)

The sustainability of the shrimp aquaculture depends largely on disease control and the health status of shrimp. Probiotics, which make shrimps healthier and more resistant to pathogens, are promising countermeasure for shrimp diseases. In this study, the effects of the marine purple non-sulfur photosynthetic bacterium (PNSB) Rhodovulum sulfidophilum on Marsupenaeus japonicus (kuruma shrimp) growth and survival were examined in 177 m2 aquaria (140 tons of water) for 70 days. The shrimp received feed containing 0.01 % fresh weight (106 colony forming unit/g) of R. sulfidophilum cells. The survival rate significantly improved (P &lt; 0.001) (R. sulfidophilum-fed = 81.9 %; control = 71.5 %), the feed conversion rate improved (R. sulfidophilum-fed = 1.83; control = 2.11), and there was no difference in the shrimp average body weight. The approximate bacterial cell cost was $0.003 to $0.005 per 1 kg feed, indicating that the R. sulfidophilum approach is economically feasible and a promising candidate for probiotic bacteria in shrimp aquaculture. Keywords: photosynthetic bacteria, Rhodovulum sulfidophilum, Marsupenaeus japonicus, shrimp, probiotics

Production of Biohydrogen and/or Poly-β-hydroxybutyrate by Rhodopseudomonas sp. Using Various Carbon Sources as Substrate.

The polyhydroxyalkanoates (PHA) are family of biopolyesters synthesized by numerous bacteria which are attracting a great attention due to their thermoplastic properties. Polyhydroxybutyrate (PHB) is the most common type of PHA which presents thermoplastic and biodegradable properties. It is synthesized under stressful conditions by heterotrophic bacteria and many photosynthetic microorganisms such as purple non-sulfur bacteria and cyanobacteria. Biological hydrogen (H2) production is being evaluated for use as a fuel since it is a promising substitute for carbonaceous fuels owing to its high conversion efficiency and high specific content. In the present work, the purple non-sulfur photosynthetic bacterium Rhodopseudomonas sp. for the simultaneous H2 photo-evolution and poly-β-hydroxybutyrate (PHB) production has been investigated. Three different types of carbon sources were tested in the presence of glutamate as a nitrogen source in a batch cultivation system, under continuous irradiance. The results indicated the fact that the type of carbon source in the culture broth affects in various ways the metabolic activity of the bacterial biomass, as evidenced by the production of PHB and/or H2 and biomass. The best carbon source for PHB accumulation and H2 production by Rhodopseudomonas sp. turned out to be the acetate, having the highest H2 production (2286 mL/L) and PHB accumulation (68.99 mg/L, 18.28% of cell dry weight).

Use of Purple Non-Sulfur Photosynthetic Bacteria (Rhodobacter sphaeroides) in Promoting Ciliated Protozoa Growth.

Photosynthetic bacterium (PSB) was isolated from sediment samples of Yamagawa Bay, Kagoshima, Japan. Phylogenetic analysis results of PSB isolate were closely related to Rhodobacter sphaeroides, purple non-sulfur photosynthetic bacteria (PNSB). Pink-colored smooth edges of single bacterial colonies were observed after 3-5 days of incubation period on Basic I medium agar plates. Rhodobacter sphaeroides microscopic examination showed a short rod cell (1-2 µm length) with round ends. Sediment and water samples used for ciliates cultivation were collected from Kuwano-ura Bay, Koshiki Island, Japan. Ciliates were cultivated using fish meal with radish leaves medium (MI), with sediment into MI (MII) and algae media (MIII). The use of the algae media (MIII) in cultivation mixture produced the highest total number of ciliates. Big size ciliates were identified as Euplotes minuta and Cyclidium varibonneti, while small size was identified as Micrometopion nutans, based on PCR-DGGE. When ciliates were cultured with the PSB isolate, Rhodobacter sphaeroides as a feed, ciliates grow to 2,081 individual ml-1 72 hrs later. These findings indicate that PNSB can be used to promote ciliates growth.

Engineering the transcriptional activator NifA for the construction of Rhodobacter sphaeroides strains that produce hydrogen gas constitutively.

Purple non-sulfur photosynthetic bacteria such as Rhodobacter sphaeroides and Rhodopseudomonas palustris produce hydrogen gas (H2) via proton reduction, which is catalyzed by nitrogenase. Although the expression of nitrogenase is usually repressed under nitrogen-sufficient conditions, a partial deletion of nifA, which encodes a transcriptional activator of nitrogen-fixation genes, has been reported to enable the constitutive expression of nitrogenase in R. palustris. In this study, we evaluated the effects of a similar mutation (nifA* mutation) on H2 production during the photoheterotrophic growth of R. sphaeroides, based on the notion that H2 production by nitrogenase compensates for the loss of CO2 fixation via the Calvin cycle, thereby restoring the redox balance. The chromosomal nifA* mutation resulted in the slight restoration of the photoheterotrophic growth of a mutant strain lacking ribulose-1,5-bisphosphate carboxylase/oxygenase (RubisCO), the key enzyme of the Calvin cycle, when the strain was cultured in van Niel's yeast medium. In addition, the strain with the nifA* mutation produced detectable levels of H2 during photoheterotrophic growth with acetate and ammonium; however, the H2 production was considerably lower than that observed during the photoheterotrophic growth of the strain with acetate and L-glutamate, where L-glutamate serves as a poor nitrogen source, thereby causing nitrogenase derepression. On the other hand, introduction of a multicopy plasmid harboring nifA* markedly restored the photoheterotrophic growth of the RubisCO-deletion mutant in van Niel's yeast medium and resulted in efficient H2 production during the photoheterotrophic growth with acetate and ammonium.

Inactivation of the uptake hydrogenase in the purple non-sulfur photosynthetic bacterium Rubrivivax gelatinosus CBS enables a biological water-gas shift platform for H2 production.

Biological H2 production has potential to address energy security and environmental concerns if produced from renewable or waste sources. The purple non-sulfur photosynthetic bacterium Rubrivivax gelatinosus CBS produces H2 while oxidizing CO, a component of synthesis gas (Syngas). CO-linked H2 production is facilitated by an energy-converting hydrogenase (Ech), while a subsequent H2 oxidation reaction is catalyzed by a membrane-bound hydrogenase (MBH). Both hydrogenases contain [NiFe] active sites requiring 6 maturation factors (HypA-F) for assembly, but it is unclear which of the two annotated sets of hyp genes are required for each in R. gelatinosus CBS. Herein, we report correlated expression of hyp1 genes with Ech genes and hyp2 expression with MBH genes. Moreover, we find that while Ech H2 evolving activity is only delayed when hyp1 is deleted, hyp2 deletion completely disrupts MBH H2 uptake, providing a platform for a biologically driven water-gas shift reaction to produce H2 from CO.

Novel adjuvants derived from attenuated lipopolysaccharides and lipid As of purple non-sulfur photosynthetic bacteria

Adjuvants are substances that enhance adaptive immune response to antigen. Development of a safe and effective immunostimulant adjuvant is essential for the efficacy of a vaccine to protect against infectious pathogens. Purple non-sulfur photosynthetic bacteria exhibited nontoxic natural lipid A variants that are distinct in their chemical structures from that of the Escherichia coli-type lipid A. In this study, the adjuvant efficacy of attenuated lipid A variants and their corresponding lipopolysaccharides (LPSs), derived from purple photosynthetic bacteria (Rhodocyclus tenuis and Rhodobacter sphaeroides) were evaluated. LPS was extracted using modified phenol-chloroform-petroleum ether method and lipid A was separated by mild acid hydrolysis. Trinitrophenol (TNP) was conjugated to hen egg albumin (TNP-HEA) and used as haptenic antigen. The LPS and lipid A adjuvant candidates were formulated in oil-in-water emulsion (OIWE) and evaluated to elicit anti-TNP IgG against TNP-HEA conjugate in BALB/c female mice. The anti-TNP IgG titers were measured using ELISA. The intact LPS-based adjuvants present in OIWE formulation showed significantly higher efficacy to elicit anti-TNP IgG titers against TNP-HEA conjugate compared to their corresponding lipid A-based adjuvants. As expected, the OIWE formulations of all LPS- and lipid A-based adjuvant candidates showed higher activities compared to the aqueous formulations. Slow reduction in the levels of anti-TNP IgG antibodies in the serum was observed over 4 months after immunization using the LPS- and lipid A-based adjuvant candidates which may provide a long protection against pathogens. The attenuated LPSs and lipid A’s from the photosynthetic bacteria showed promising results to develop novel safe and effective adjuvants that can evoke the immune response. The most promising adjuvant candidate was the LPS-based adjuvant from R. tenuis.

Optimal iron concentrations for growth-associated polyhydroxyalkanoate biosynthesis in the marine photosynthetic purple bacterium Rhodovulum sulfidophilum under photoheterotrophic condition.

Polyhydroxyalkanoates (PHAs) are a group of natural biopolyesters that resemble petroleum-derived plastics in terms of physical properties but are less harmful biologically to the environment and humans. Most of the current PHA producers are heterotrophs, which require expensive feeding materials and thus contribute to the high price of PHAs. Marine photosynthetic bacteria are promising alternative microbial cell factories for cost-effective, carbon neutral and sustainable production of PHAs. In this study, Rhodovulum sulfidophilum, a marine photosynthetic purple nonsulfur bacterium with a high metabolic versatility, was evaluated for cell growth and PHA production under the influence of various media components found in previous studies. We evaluated iron, using ferric citrate, as another essential factor for cell growth and efficient PHA production and confirmed that PHA production in R. sulfidophilum was growth-associated under microaerobic and photoheterotrophic conditions. In fact, a subtle amount of iron (1 to 2 μM) was sufficient to promote rapid cell growth and biomass accumulation, as well as a high PHA volumetric productivity during the logarithmic phase. However, an excess amount of iron did not enhance the growth rate or PHA productivity. Thus, we successfully confirmed that an optimum concentration of iron, an essential nutrient, promotes cell growth in R. sulfidophilum and also enhances PHA utilization.

Fermentative utilization of glycerol residue for the production of acetic acid

Glycerol residue, frequently known as pitch, is a waste produced from the downstream product of crude glycerine distillation. With the increasing need of pure glycerine in the world, the glycerol residue produced is also increasing. Glycerol residue is a solid waste at room temperature, highly alkaline (pH > 13), corrosive, and categorized as hazardous and poisonous waste. In this research, acetic acid was produced from glycerol residue through the anaerobic fermentation process by using purple non-sulphur photosynthetic bacteria. The purpose of this study was to find out the influence of concentration change of glycerol residue on time and to find out the possibility of glycerol residue to be utilized as acetic acid. In this research, at first 400 g of glycerol residue was diluted with 200 ml of distilled water to change the glycerine phase, from solid to liquid at room temperature, acidified by using hydrochloric acid until pH 2. The top layer formed was fatty acid and triglycerides that should be removed. Meanwhile, the bottom layer was diluted glycerol residue which was then neutralized with caustic soda. To produce acetic acid, glycerol residue with various concentrations, salt, and purple non-sulphur photosynthetic bacteria were put together into a 100 ml bottle which had been previously sterilized, then incubated for four weeks under the light of 40-watt bulb. The result showed that on the 28th day of fermentation, the produced acetic acid were 0.28, 1.85, and 0.2% (w/w) by using glycerine with the concentration of 0.5, 1.0, and 1.5% (w/w), respectively.