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In recent decades, antibiotics have been widely used in sea urchin aquaculture to prevent diseases and improve water quality. However, their use leads to the emergence of resistant strains and environmental problems. Probiotics can serve as a green and pollution-free alternative. In this study, we mixed probiotics (Rhodopseudomonas palustris) (3 × 10 8  CFU/mL) and composite probiotics (5 × 10 8  CFU/mL) in a 1:1 ratio, establishing four supplementation levels: 0 % (FC, control group), 0.5 % (FL group), 1 % (FM group), and 2 % (FH group). At the end of the 50-day experiment, compared to the control group (FC), supplementation with R. palustris and composite probiotics (containing 60.13 % Lactobacillaceae and 20.79 % Acetobacteraceae) in the FL, FM, and FH groups significantly improved the growth performance of sea urchins (Strongylocentrotus intermedius), including final body weight (FBW), weight gain rate (WGR), specific growth rate (SGR), and gonadosomatic index (GSI). Additionally, antioxidant indicators (e.g., total antioxidant capacity (T-AOC), superoxide dismutase (SOD), and catalase (CAT)), immune indicators (e.g., acid phosphatase (ACP), alkaline phosphatase (AKP), and lysozyme (LZM)), and digestive enzyme activity were significantly increased in FM and FH groups. Probiotic supplementation also increased colonization of beneficial bacteria (Rhodobacteraceae) and enhanced microbiota diversity. The FM group, in particular, showed significantly upregulated expression of heat shock protein 70 (HSP70), glutathione peroxidase (GPX), glutathione (GST), toll-like receptor (TLR), and lysozyme (LYZ) genes. Overall, 1 % supplementation significantly enhanced growth, antioxidant capacity, digestive ability, microbiota stability, and immune-related gene expression in S. intermedius., Competing Interests: Declaration of competing interest The authors have no relevant financial or non-financial interests to disclose., (Copyright © 2024 Elsevier Inc. All rights reserved.)

The biomethanation process is widely recognized as a significant approach to mitigating carbon dioxide emissions while simultaneously generating methane. However, only a few microorganisms that required intricate culturing conditions were identified for biomethanation. Here, Escherichia coli that featured easy cultivation and versatile chassis was genetically modified for biomethanation for the first time. The nif-related gene cluster with methanogenic capability from Rhodopseudomonas palustris was systematically analyzed, cloned, and integrated into a synthetic biomethanation module. As a result, E. coli BL21 (DE3) and Rosetta (DE3) carrying this synthetic biomethanation module exhibited significant methane production activity, with methane yields reaching 50 nmol/mL and 159 nmol/mL, respectively. This finding provided a simple route to construct synthetic strain for biomethanation, which would advance the fundamental research and be beneficial to further harness the power of biomethanation for practical application., Competing Interests: Declaration of competing interest The authors declare no competing financial interests., (Copyright © 2025 Elsevier Inc. All rights reserved.)

A biohydrogen and polyhydroxyalkanoates (PHA)-producing natural photoheterotrophic mixed culture composed mainly by Rhodopseudomonas palustris and Clostridium sp was studied by a proteomic analysis under non-growth conditions (nitrogen-absence and organic acids). Proteins in C. pasteurianum were upregulated, particularly those related to stress response. In contrast, C. pasteurianum in the consortium did not present such proteins, showing the advantage of being part of it. Both cultures showed proteins involved in organic acid metabolism and biohydrogen production, such as lactate dehydrogenase, ferredoxins, and hydrogenases. Proteomes of R. palustris as single culture and in consortium showed that organic acids were redirected into central carbon pathways to generate reduced equivalents for biohydrogen production. Light-harvesting proteins and fatty acid metabolism linked to PHA accumulation were also upregulated. This study provides insights into how the proteomes of individual organisms and their consortium counterparts adapt to non-growth conditions, shedding light on how microbial interactions influence protein expression., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Utilizing Rhodopseudomonas palustris ( R. pal ), this study constructed a dual-chamber microbial electrosynthesis system, based on microbial electrolysis cells, that was capable of producing lycopene. Cultivation within the electrosynthesis chamber yielded a lycopene concentration of 282.3722 mg/L when the optical density (OD) reached 0.6, which was four times greater than that produced by original strains. The mutant strain showed significantly higher levels of extracted riboflavin compared to the wild-type strain, and the riboflavin content of the mutant strain was 61.081 mg/L, which was more than 10 times that of the original strain. Furthermore, sequencing and analyses were performed on the mutant strains observed during the experiment. The results indicated differences in antibiotic resistance genes, carbohydrate metabolism-related genes, and the frequencies of functional genes between the mutant and original strains. The mutant strain displayed potential advantages in specific antibiotic resistance and carbohydrate degradation capabilities, likely attributable to its adaptation to electrogenic growth conditions. Moreover, the mutant strain demonstrated an enrichment of gene frequencies associated with transcriptional regulation, signal transduction, and amino acid metabolism, suggesting a complex genetic adaptation to electrogenic environments. This study presents a novel approach for the efficient and energy-conserving production of lycopene while also providing deeper insights into the genetic basis of electro-resistance genes.

Rhodopseudomonas palustris R1 is a phototrophic, purple, non-sulfur Gram-negative bacterium first isolated from the soils of rice fields, known for its polylactic acid degradability. Here, we present the complete circular genome sequence of this bacterium, spanning 5.32 Mbp and 4,949 protein-coding sequences., Competing Interests: The authors declare no conflict of interest.

Rice blast is one of the most hazardous diseases affecting rice production. Previously, we discovered that the Atp2 protein of Rhodopseudomonas palustris could significantly inhibit the appressorium formation and pathogenicity of Magnaporthe oryzae. However, the molecular mechanism of this fungus has remained unknown. This study revealed that Atp2 can enter the cell and interact with the ribosomal protein MoRpl12 of M. oryzae , directly affecting the expression of the MoRpl12 protein. Silencing the MoRPL12 gene can affect cell wall integrity, growth, conidiogenesis, and fungal pathogenicity. The quantitative reverse transcription PCR results showed significant changes in the expression of conidiation-related genes in the MoRPL12 gene-silenced mutants or in the Atp2 protein-treated plants. We further found that Atp2 treatment can influence the expression of ribosomal-related genes, such as RPL , in M. oryzae . Our study revealed a novel antifungal mechanism by which the Atp2 protein binds to the ribosomal protein MoRpl12 and inhibits the pathogenicity of rice blast fungus, providing a new potential target for rice blast prevention and control., Competing Interests: The author(s) declare no conflict of interest.

Sea urchin contains physiologically active substances, such as amino acids and unsaturated fatty acids, and an important aquatic organism. Purple sea urchin, one of the common edible sea urchins, is an important aquatic product. In order to supply the vast seafood market, large-scale aquaculture of sea urchins is very important. The aim of this study was to optimize the rearing of the Anthocidaris crassipina larvae enhancing the nutrition by mixing feed to improve their growth and survival. The survival rate of Chaetoceros muelleri feeding alone is only 40%. If the survival rate is improved through nutrient enrichment, the large-scale aquaculture of larvae can be promoted. The experiment was divided into two parts. Experiment 1: Two types of commonly used microalgae, Isochrysis galbana tml (I), C. muelleri (C) and two types of probiotics, Rhodopseudomonas palustris (R), and Saccharomyces cerevisiae (S) were used in the. Feeding amounts are 5000, 10,000, and 20,000 cell mL -1 , and the control group (N) did not eat. Experiment 2: C. muelleri 20,000 cell mL -1 was mixed with I. galbana tml, R. palustris (R) and S. cerevisiae (S) at 5000 and 10,000 cell mL -1 . After the experiment, body length, body width, stomach length, rudiment length, rudiment length, body composition, digestive enzymes and survival rate were measured to evaluate the best feed. The results showed that the mixed feeding of C. muelleri 20,000 cell mL -1 and R. palustris 5000 cell mL -1 can achieve the best development and survival of larval embryos and can promote metamorphosis into juveniles in the shortest time. The research results will be applied to the large-scale aquaculture of A. crassipina larvae to promote the diversity of aquaculture., (© 2024 Wiley‐VCH GmbH. Published by John Wiley & Sons Ltd.)

Photosynthetic bacteria (PSB) are suitable to live and remediate cadmium (Cd) in the slightly oxygenated or anaerobic flooding paddy field. However, there is currently limited study on the inhibition of Cd accumulation in rice by PSB, and the relevant mechanisms has yet to be elucidated. In the current study, we firstly used Rhodopseudomonas palustris SC06 (a typical PSB) as research target and combined physiology, biochemistry, microbiome and metabolome to evaluate the mechanisms of remeding Cd pollution in paddy field and inhibiting Cd accumulation in rice. Microbiome analysis results revealed that intensive inoculation with R. palustris SC06 successfully survived and multiplied in flooding paddy soil, and significantly increased the relatively abundance of anaerobic bacteria including Desulfobacterota, Anaerolineaceae, Geobacteraceae, and Gemmatimonadaceae by 46.40 %, 45.00 %, 50.12 %, and 21.30 %, respectively. Simultaneously, the structure of microbial community was regulated to maintain relative stability in the rhizosphere soil of rice under Cd stress. In turn, these bacteria communities reduced bioavailable Cd and enhanced residual Cd in soil, and induced the upregulation of sugar and organic acids in the rice roots, which further inhibited Cd uptake in rice seedlings, and dramatically improved the photosynthetic efficiency in the leaves and the activities of antioxidative enzymes in the roots. Finally, Cd content of the roots, stems, leaves, and grains significantly decreased by 38.14 %, 69.10 %, 83.40 %, and 37.24 % comparing with the control, respectively. This study provides a new strategy for the remediation of Cd-contaminated flooding paddy fields and the safe production of rice., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier B.V.)

Halogenated aromatic compounds are used in a variety of industrial applications but can be harmful to humans and animals when released into the environment. Microorganisms that degrade halogenated aromatic compounds anaerobically have been isolated but the evolutionary path that they may have taken to acquire this ability is not well understood. A strain of the purple nonsulfur bacterium, Rhodopseudomonas palustris , RCB100, can use 3-chlorobenzoate (3-CBA) as a carbon source whereas a closely related strain, CGA009, cannot. To reconstruct the evolutionary events that enabled RCB100 to degrade 3-CBA, we isolated an evolved strain derived from CGA009 capable of growing on 3-CBA. Comparative whole-genome sequencing of the evolved strain and RCB100 revealed both strains contained large deletions encompassing badM , a transcriptional repressor of genes for anaerobic benzoate degradation. It was previously shown that in strain RCB100, a single nucleotide change in an alicyclic acid coenzyme A ligase gene, named aliA , gives rise to a variant AliA enzyme that has high activity with 3-CBA. When the RCB100 aliA allele and a deletion in badM were introduced into R. palustris CGA009, the resulting strain grew on 3-CBA at a similar rate as RCB100. This work provides an example of pathway evolution in which regulatory constraints were overcome to enable the selection of a variant of a promiscuous enzyme with enhanced substrate specificity.IMPORTANCEBiodegradation of man-made compounds often involves the activity of promiscuous enzymes whose native substrate is structurally similar to the man-made compound. Based on the enzymes involved, it is possible to predict what microorganisms are likely involved in biodegradation of anthropogenic compounds. However, there are examples of organisms that contain the required enzyme(s) and yet cannot metabolize these compounds. We found that even when the purple nonsulfur bacterium, Rhodopseudomonas palustris , encodes all the enzymes required for degradation of a halogenated aromatic compound, it is unable to metabolize that compound. Using adaptive evolution, we found that a regulatory mutation and a variant of promiscuous enzyme with increased substrate specificity were required. This work provides insight into how an environmental isolate evolved to use a halogenated aromatic compound., Competing Interests: The authors declare no conflict of interest.

Background: Pickled mustard, the largest cultivated vegetable in China, generates substantial waste annually, leading to significant environmental pollution due to challenges in timely disposal, leading to decomposition and sewage issues. Consequently, the imperative to address this concern centers on the reduction and comprehensive resource utilization of raw mustard waste (RMW). To achieve complete and quantitative resource utilization of RMW, this study employs novel technology integration for optimizing its higher-value applications., Results: Initially, subcritical hydrothermal technology was applied for rapid decomposition, with subsequent ammonia nitrogen removal via zeolite. Thereafter, photosynthetic bacteria, Rhodopseudomonas palustris, were employed to maximize hydrogen and methane gas production using various fermentation enhancement agents. Subsequent solid-liquid separation yielded liquid fertilizer from the fermented liquid and soil amendment from solid fermentation remnants. Results indicate that the highest glucose yield (29.6 ± 0.14) was achieved at 165-173℃, with a total sugar content of 50.2 g/L and 64% glucose proportion. Optimal ammonia nitrogen removal occurred with 8 g/L zeolite and strain stable growth at 32℃, with the highest OD 600 reaching 2.7. Several fermentation promoters, including FeSO 4 , Neutral red, Na 2 S, flavin mononucleotide, Nickel titanate, Nickel oxide, and Mixture C, were evaluated for hydrogen production. Notably, Mixture C resulted in the maximum hydrogen production (756 mL), a production rate of 14 mL/h, and a 5-day stable hydrogen production period. Composting experiments enhanced humic acid content and organic matter (OM) by 17% and 15%, respectively., Conclusions: This innovative technology not only expedites RMW treatment and hydrogen yield but also substantially enriches soil fertility. Consequently, it offers a novel approach for low-carbon, zero-pollution RMW management. The study's double outcomes extend to large-scale RMW treatment based on the aim of full quantitative resource utilization of RMW. Our method provides a valuable reference for waste management in similar perishable vegetable plantations., (© 2024. The Author(s).)

Rhodopseudomonas palustris CGA009 is a Gram-negative, purple non-sulfur, metabolically diverse bacterium with wide-ranging habitats. The extraordinary ability of R. palustris to decompose a variety of raw materials and convert them into high-value products makes it an attractive host for biotechnology and industrial applications. However, being a freshwater bacterium R. palustris has limited application in highly-saline environments. Therefore, it is of great significance to obtain the salt-tolerant strain of R. palustris and understand its tolerance mechanism. In this study, R. palustris CGA009 was successfully evolved into eight salt-tolerant strains using an adaptive laboratory evolution technique. RPAS-11 ( R. palustris anti-salt strain 11) was selected as the best salt-tolerant strain and was used in further studies to explore the salt-tolerance mechanism. The expression of most genes associated with the carotenoid synthesis in RPAS-11 increased significantly under high concentration of salt stress, suggesting that carotenoid synthesis is one of the reasons for the salt tolerance of RPAS-11. Gene overexpression and knockout experiments were performed to get clear about the role of carotenoids in salt stress tolerance. RPAS-11-IDI, the mutant with overexpression of IDI (Isopentenyl diphosphate isomerase) exhibited enhanced salt tolerance, whereas the knockout mutant CGA009-∆ crtI showed a decline in salt tolerance. In addition, the results indicated that rhodopin, a carotenoid compound, was the key pigment responsible for the salt tolerance in R. palustris . Furthermore, the production of lycopene, a widely-used carotenoid, was also increased. Taken together, our research helps to deepen the understanding of the salt tolerance mechanism of R. palustris and also widens the application of R. palustris in highly-saline environments., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Li, Zhu, Yang, Wang, Liu and Yang.)

Background: Whole-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., Results: In 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., Conclusions: Taken 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., (© 2023. The Author(s).)

The use of waste as a carbon source can significantly reduce the cost of production of Polyhydroxyalkanoates (PHAs). In this study, an acidified hydrolysate solution derived from food waste (FW) was used as a carbon source for the synthesis of PHAs by Rhodopseudomonas palustris (R. palustris) and optimized the process parameters. The results showed that the PHAs yield reached 48.62% under optimal conditions (an incubation time of 30 days, volatile fatty acids (VFAs) in substrate concentration of 2202.21 mg⋅L -1 , an initial pH of 8.0, and inoculum concentration of 15%). The fraction of VFAs affects the composition of PHAs, R. palustris first uses VFAs with an even number of carbons to synthesize poly(3-hydroxybutyrate)(3HB), and later uses VFAs with an odd number of carbons to synthesize poly-3-hydroxyvalerate (3HV). Pathways for the synthesis of PHAs by R. palustris were inferred. R. palustris is a strain with the potential to synthesize PHAs., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier Ltd.)

Rhodopseudomonas palustris is a purple non-sulfide bacterium (PNSB), and some strains have been proven to promote plant growth. However, the mechanism underlying the effect of these PNSBs remains limited. Based on genetic information, R. palustris possesses the ability to produce pyrroloquinoline quinone (PQQ). PQQ is known to play a crucial role in stimulating plant growth, facilitating phosphorous solubilization, and acting as a reactive oxygen species scavenger. However, it is still uncertain whether growth conditions influence R. palustris 's production of PQQ and other characteristics. In the present study, it was found that R. palustris exhibited a higher expression of genes related to PQQ synthesis under autotrophic culture conditions as compared to acetate culture conditions. Moreover, similar patterns were observed for phosphorous solubilization and siderophore activity, both of which are recognized to contribute to plant-growth benefits. However, these PNSB culture conditions did not show differences in Arabidopsis growth experiments, indicating that there may be other factors influencing plant growth in addition to PQQ content. Furthermore, the endophytic bacterial strains isolated from Arabidopsis exhibited differences according to the PNSB culture conditions. These findings imply that, depending on the PNSB's growing conditions, it may interact with various soil bacteria and facilitate their infiltration into plants.

Photofermentative hydrogen production has gained increasing attention as a source of green energy. To make such photofermentation processes economically competitive, operating costs need to be reduced, possibly through outdoor operation. Because photofermentation processes are light dependent, the emission spectrum and intensity of light both have a significant influence on the hydrogen production and merit investigation. This study investigates the effect of light sources on the hydrogen production and growth of Rhodopseudomonas palustris, comparing the organism's productivity under longer-wavelength light and light mimicking sunlight. Hydrogen production is enhanced under longer-wavelength light, producing 26.8% (± 7.3%) more hydrogen as compared to under light mimicking that of sunlight; however, R. palustris is still able to produce a considerable volume of hydrogen under light with a spectrum mimicking that of sunlight, providing a promising avenue for future research., (© 2023. The Author(s).)

Glutamine synthetase (GS) is responsible for the ammonium assimilation into glutamine, which serves as an important nitrogen donor for the synthesis of biomolecules and also plays a key role in regulating the nitrogen fixation catalyzed by nitrogenase. Rhodopseudomonas palustris, whose genome encodes 4 putative GSs and 3 nitrogenases, is an attractive photosynthetic diazotroph for studies of nitrogenase regulation, as it can produce the powerful greenhouse gas (methane) by iron-only (Fe-only) nitrogenase using light energy. However, the primary GS enzyme for ammonium assimilation and its role in nitrogenase regulation remain elusive in R. palustris. Here, we show that GlnA1, whose activity is finely regulated by reversible adenylylation/deadenylylation of Tyr 398 residue, is primarily responsible for ammonium assimilation as the preferred GS in R. palustris. The inactivation of GlnA1 makes R. palustris shift to use the alternative GlnA2 for ammonium assimilation, resulting in the expression of Fe-only nitrogenase even in the presence of ammonium. We present a model, showing how R. palustris responds to ammonium availability and further regulates the expression of Fe-only nitrogenase. These data may contribute to the design of promising strategies for a better control of greenhouse gas emissions. IMPORTANCE The photosynthetic diazotrophs, such as Rhodopseudomonas palustris, can utilize light energy to drive the conversion of carbon dioxide (CO 2 ) to a much more powerful greenhouse gas methane (CH 4 ) by Fe-only nitrogenase, which is strictly regulated in response to the ammonium, a substrate of glutamine synthetase for the biosynthesis of glutamine. However, the primary glutamine synthetase for ammonium assimilation and its role in nitrogenase regulation remain unclear in R. palustris. This study shows that GlnA1 is the primary glutamine synthetase for ammonium assimilation, and also plays a key role in Fe-only nitrogenase regulation in R. palustris. For the first time, a R. palustris mutant capable of expressing Fe-only nitrogenase even in the presence of ammonium is obtained by inactivation of GlnA1. A better understanding of the Fe-only nitrogenase regulation achieved in this study provide us with new insights into the efficient control of CH 4 emissions.

Hydrogen production via microbial photofermentation shows great promise as a method for sustainable hydrogen production; however, operating costs associated with photofermentative hydrogen production need to be reduced. Costs can be reduced using a passive circulation system like the thermosiphon photobioreactor, and by operating it under natural sunlight. In this study, an automated system was implemented to investigate the effect of diurnal light cycles on the hydrogen productivity and growth of Rhodopseudomonas palustris and on the operation of a thermosiphon photobioreactor, under controlled conditions. Diurnal light cycles, simulating daylight times, were found to reduce hydrogen production in the thermosiphon photobioreactor demonstrating a low maximum production rate of 0.015 mol m -3  h -1 (± 0.002 mol m -3  h -1 ) as compared to 0.180 mol m -3  h -1 (± 0.0003 mol m -3  h -1 ) under continuous illumination. Glycerol consumption as well as hydrogen yield also decreased under diurnal light cycles. Nonetheless, hydrogen production in a thermosiphon photobioreactor under outdoor conditions was demonstrated as possible avenue for further investigation., (© 2023. The Author(s).)

Nitrogenase is the key enzyme involved in nitrogen fixation and uses low potential electrons delivered by ferredoxin (Fd) or flavodoxin (Fld) to reduce dinitrogen gas (N 2 ) to produce ammonia, generating hydrogen gas (H 2 ) as an obligate product of this activity. Although the phototrophic alphaproteobacterium Rhodopseudomonas palustris encodes multiple proteins that can reduce Fd, the FixABCX complex is the only one shown to support nitrogen fixation, and R. palustris Fix - mutants grow poorly under nitrogen-fixing conditions. To investigate how native electron transfer chains (ETCs) can be redirected toward nitrogen fixation, we leveraged the strong selective pressure of nitrogen limitation to isolate a suppressor of an R. palustris Δ fixC strain that grows under nitrogen-fixing conditions. We found two mutations were required to restore growth under nitrogen-fixing conditions in the absence of functional FixABCX. One mutation was in the gene encoding the primary Fd involved in nitrogen fixation, fer1 , and the other mutation was in aadN , which encodes a homolog of NAD + -dependent Fd:NADPH oxidoreductase (Nfn). We present evidence that AadN plays a role in electron transfer to benzoyl coenzyme A reductase, the key enzyme involved in anaerobic aromatic compound degradation. Our data support a model where the ETC for anaerobic aromatic compound degradation was repurposed to support nitrogen fixation in the Δ fixC suppressor strain. IMPORTANCE There is increasing evidence that protein electron carriers like Fd evolved to form specific partnerships with select electron donors and acceptors to keep native electron transfer pathways insulated from one another. This makes it challenging to integrate a Fd-dependent pathway such as biological nitrogen fixation into non-nitrogen-fixing organisms and provide the high-energy reducing power needed to fix nitrogen. Here, we show that amino acid substitutions in an electron donor for anaerobic aromatic compound degradation and an Fd involved in nitrogen fixation enabled electron transfer to nitrogenase. This study provides a model system to understand electron transfer chain specificity and how new electron transfer pathways can be evolved for biotechnologically valuable pathways like nitrogen fixation.

Novel peripheral light-harvesting (LH) complex designated as LL LH2 was isolated along with LH4 complex from Rhodopseudomonas palustris cells grown under low light intensity (LL). FPLC-MS/MS allowed to reveal PucABd and PucBabc apoproteins in LL LH2 complex, which is different from previously described LH4 complex containing PucABd, PucABa and PucBb. The main carotenoids in LL LH2 complex were rhodopin and 3,4-didehydrorhodopin. Three-dimensional modeling demonstrated which amino acid residues of all the β -subunits could interact with carotenoids (Car) and bacteriochlorophyll a (BChl a). Analysis of amino acid sequences of α -subunits of both LL complexes showed presence of different C-terminal motifs, IESSVNVG in α a subunit and IESSIKAV in α d subunit, in the same positions of C-termini, which could reflect different retention force of LL LH2 and LH4 on hydroxyl apatite, facilitating successful isolation of these complexes. Differences of these LL complexes in protein and carotenoid composition, in efficiency of energy transfer from Car to BChl a, which is two times lower in LL LH2 than in LH4, allow to assign it to a novel type of light-harvesting complex in Rhodopseudomonas palustris ., Competing Interests: The authors declare there are no competing interests., (©2023 Serdyuk et al.)

One of the main factors affecting hydrogen production and growth of photofermentative microorganisms is light; low light penetration and utilization are significant bottlenecks in photofermentative hydrogen production systems. In this study, light distribution in a thermosiphon photobioreactor operated with Rhodopseudomonas palustris was investigated. Radiation fields were modelled and simulated using computational fluid dynamics (ANSYS® Fluent, 2019 R2) and a reflector system was evaluated for the enhancement of light distribution in a thermosiphon photobioreactor. The effect of the reflector system was investigated experimentally in terms of hydrogen production, carbon substrate consumption and biomass circulation in the reactor. With the addition of the reflector system, hydrogen production was increased by 48% while glycerol consumption was increased from approximately 24% to 32%. After 336 h, the concentration of R. palustris cells still in suspension ranged from 0.13 to 0.18 g∙L -1 , with no discernible difference in concentration between the systems with and without reflectors. Collectively, the reflector system was shown to be a viable option in enhancing light distribution in photobioreactors, with an associated increase in both hydrogen production as well as glycerol consumption., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Purple nonsulfur bacteria (PNSB) were investigated for their carotenoid production and anti-vibrio activity against acute hepatopancreatic necrosis disease (AHPND)-causing Vibrio parahaemolyticus. To test carotenoid production, selected strains were cultivated in basic isolation medium (BIM), glutamate acetate medium, G5 medium and artificial acetic acid wastewater (AAW) medium. From 144 PNSB, Rhodopseudomonas palustris KTSSG46 was selected to produce carotenoids under microaerobic light conditions in BIM. When the culture medium was optimized, strain KTSSG46 grown in BIM modified with l-glutamate at 1 g/L more effectively inhibited AHPND-causing V. parahaemolyticus strains than standard BIM with 1 g/L (NH 4 ) 2 SO 4 . BIM was further modified with 1.23 g/L MgSO 4 ·7H 2 O and carotenoid production increased 40.22%. Carotenoid production at day 2 by strain KTSSG46 grown in BIM modified with l-glutamate at 1 and 1.23 g/L MgSO 4 ·7H 2 O was the same as production in BIM modified with monosodium glutamate (MSG). Culture supernatants from all BIM formulations showed similar activity against the resistant AHPND strain SR2. Based on high-performance liquid chromatography, carotenoids of strain KTSSG46 might be canthaxanthin. Grown in BIM modified with MSG, strain KTSSG46 could produce inexpensive carotenoids and release anti-vibrio compounds that, applied as shrimp feed additive, would prevent AHPND strains., (© 2021 International Union of Biochemistry and Molecular Biology, Inc.)

Rhodopseudomonas palustris is an attractive option for biotechnical applications and industrial engineering due to its metabolic versatility and its ability to catabolize a wide variety of feedstocks and convert them to several high-value products. Given its adaptable metabolism, R. palustris has been studied and applied in an extensive variety of applications such as examining metabolic tradeoffs for environmental perturbations, biodegradation of aromatic compounds, environmental remediation, biofuel production, agricultural biostimulation, and bioelectricity production. This review provides a holistic summary of the commercial applications for R. palustris as a biotechnology chassis and suggests future perspectives for research and engineering., Competing Interests: Declaration of Competing Interest The authors report no conflict of interest., (Copyright © 2022 The Author(s). Published by Elsevier Inc. All rights reserved.)

5-Aminolevulinic acid (5-ALA), a vital precursor for the biosynthesis of tetrapyrrole compounds, has been widely applied in agriculture and medicine, while extremely potential for the treatment of cancers, corona virus disease 2019 (COVID-19) and metabolic diseases in recent years. With the development of metabolic engineering and synthetic biology, the biosynthesis of 5-ALA has attracted increasing attention. 5-Aminolevulinic acid synthase (ALAS), the key enzyme for 5-ALA synthesis in the C4 pathway, is subject to stringent feedback inhibition by heme. In this work, cysteine-targeted mutation of ALAS was proposed to overcome this drawback. ALAS from Rhodopseudomonas palustris (RP-ALAS) and Rhodobacter capsulatus (RC-ALAS) were selected for mutation and eight variants were generated. Variants RP-C132A and RC-C201A increased enzyme activities and released hemin inhibition, respectively, maintaining 82.5% and 81.9% residual activities in the presence of 15 μM hemin. Moreover, the two variants exhibited higher stability than that of their corresponding wild-type enzymes. Corynebacterium glutamicum overexpressing RP-C132A and RC-C201A produced 14.0% and 21.6% higher titers of 5-ALA than the control, respectively. These results strongly suggested that variants RP-C132A and RC-C201A obtained by utilizing cysteine-targeted mutation strategy released hemin inhibition, broadening their applications in 5-ALA biosynthesis., (Copyright © 2022 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.)

The genomes of some purple photosynthetic bacteria contain a multigene puc family encoding a series of α- and β-polypeptides that together form a heterogeneous antenna of light-harvesting 2 (LH2) complexes. To unravel this complexity, we generated four sets of puc deletion mutants in Rhodopseudomonas palustris , each encoding a single type of pucBA gene pair and enabling the purification of complexes designated as PucA-LH2, PucB-LH2, PucD-LH2, and PucE-LH2. The structures of all four purified LH2 complexes were determined by cryogenic electron microscopy (cryo-EM) at resolutions ranging from 2.7 to 3.6 Å. Uniquely, each of these complexes contains a hitherto unknown polypeptide, γ, that forms an extended undulating ribbon that lies in the plane of the membrane and that encloses six of the nine LH2 αβ-subunits. The γ-subunit, which is located near to the cytoplasmic side of the complex, breaks the C9 symmetry of the LH2 complex and binds six extra bacteriochlorophylls (BChls) that enhance the 800-nm absorption of each complex. The structures show that all four complexes have two complete rings of BChls, conferring absorption bands centered at 800 and 850 nm on the PucA-LH2, PucB-LH2, and PucE-LH2 complexes, but, unusually, the PucD-LH2 antenna has only a single strong near-infared (NIR) absorption peak at 803 nm. Comparison of the cryo-EM structures of these LH2 complexes reveals altered patterns of hydrogen bonds between LH2 αβ-side chains and the bacteriochlorin rings, further emphasizing the major role that H bonds play in spectral tuning of bacterial antenna complexes.

The current work provides insights for improving the hydrogen output while degrading emerging contaminants using Rhodopseudomonas palustris. The changes in the growth rate of a microorganism due to different substrate inputs affects the hydrogen production due to metabolic route changes. The different ratios of glutamate and glycerol as nitrogen and carbon sources along with the presence of ethinylestradiol (EE2) in the photofermenter affected the flux of electrons being directed towards biosynthesis and biohydrogen generation. The combination of glutamate and glycerol in different ratios (Glu:Gly; 0, 0.20 and 0.54) along with estrogen showed no significant difference in the bacteriochlorophyll concentrations. The highest biomass concentration (0.013 h -1 ) was in ratio of 0.54 while maximum specific hydrogen production (1.9 ± 0.05 ml g -1 biomass h -1 ) was observed under complete suppression of nitrogen (0; without Glu; non-growing condition) with resultant improved estrogen degradation of about 78% in 168 h by R. palustris strain MDOC01., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Microorganisms that carry out Fe(II) oxidation play a major role in biogeochemical cycling of iron in environments with low oxygen. Fe(II) oxidation has been largely studied in the context of autotrophy. Here, we show that the anoxygenic phototroph, Rhodopseudomonas palustris CGA010, carries out Fe(II) oxidation during photoheterotrophic growth with an oxidized carbon source, malate, leading to an increase in cell yield and allowing more carbon to be directed to cell biomass. We probed the regulatory basis for this by transcriptome sequencing (RNA-seq) and found that the expression levels of the known pioABC Fe(II) oxidation genes in R. palustris depended on the redox-sensing two-component system, RegSR, and the oxidation state of the carbon source provided to cells. This provides the first mechanistic demonstration of mixotrophic growth involving reducing power generated from both Fe(II) oxidation and carbon assimilation. IMPORTANCE The simultaneous use of carbon and reduced metals such as Fe(II) by bacteria is thought to be widespread in aquatic environments, and a mechanistic description of this process could improve our understanding of biogeochemical cycles. Anoxygenic phototrophic bacteria like Rhodopseudomonas palustris typically use light for energy and organic compounds as both a carbon and an electron source. They can also use CO 2 for carbon by carbon dioxide fixation when electron-rich compounds like H 2 , thiosulfate, and Fe(II) are provided as electron donors. Here, we show that Fe(II) oxidation can be used in another context to promote higher growth yields of R. palustris when the oxidized carbon compound malate is provided. We further established the regulatory mechanism underpinning this observation.

A thermosiphon photobioreactor (TPBR) can potentially be used for biohydrogen production, circumventing the requirement for external mixing energy inputs. In this study, a TPBR is evaluated for photofermentative hydrogen production by Rhodopseudomonas palustris ( R. palustris ). Experiments were conducted in a TPBR, and response surface methodology (RSM), varying biomass concentration, and light intensity and temperature were employed to determine the operating conditions for the enhancement of both hydrogen production as well as biomass suspension. Biomass concentration was found to have had the most pronounced effect on both hydrogen production as well as biomass suspension. RSM models predicted maximum specific hydrogen production rates of 0.17 mol m -3 h -1 and 0.21 mmol g CDW -1 h -1 at R. palustris concentrations of 1.21 and 0.4 g L -1 , respectively. The experimentally measured hydrogen yield was in the range of 45 to 77% (±3.8%), and the glycerol consumption was 8 to 19% (±0.48). At a biomass concentration of 0.40 g L -1 , the highest percentage of biomass (72.3%), was predicted to remain in suspension in the TPBR. Collectively, the proposed novel photobioreactor was shown to produce hydrogen as well as passively circulate biomass.

Rhodopseudomonas palustris ferredoxin:NADP + oxidoreductase ( Rp FNR) belongs to a novel group of thioredoxin reductase-type FNRs with partly characterized redox properties. Based on the reactions of Rp FNR with the 3-acetylpyridine adenine dinucleotide phosphate redox couple, we estimated the two-electron reduction midpoint potential of the FAD cofactor to be -0.285 V. 5-Deaza-FMN-sensitized photoreduction revealed -0.017 V separation of the redox potentials between the first and second electron transfer events. We examined the mechanism of oxidation of Rp FNR by several different groups of nonphysiological electron acceptors. The k cat / K m values of quinones and aromatic N -oxides toward Rp FNR increase with their single-electron reduction midpoint potential. The lower reactivity, mirroring their lower electron self-exchange rate, is also seen to have a similar trend for nitroaromatic compounds. A mixed single- and two-electron reduction was characteristic of quinones, with single-electron reduction accounting for 54% of the electron flux, whereas nitroaromatics were reduced exclusively via single-electron reduction. It is highly possible that the FADH· to FAD oxidation reaction is the rate-limiting step during the reoxidation of reduced FAD. The calculated electron transfer distances in the reaction with quinones and nitroaromatics were close to those of Anabaena and Plasmodium falciparum FNRs, thus demonstrating their similar "intrinsic" reactivity.

Rhodopseudomonas palustris , a purple nonsulfur bacterium, is a bacterium with the properties of extraordinary metabolic versatility, carbon source diversity and metabolite diversity. Due to its biodetoxification and biodegradation properties, R. palustris has been traditionally applied in wastewater treatment and bioremediation. R. palustris is rich in various metabolites, contributing to its application in agriculture, aquaculture and livestock breeding as additives. In recent years, R. palustris has been engineered as a microbial cell factory to produce valuable chemicals, especially photofermentation of hydrogen. The outstanding property of R. palustris as a microbial cell factory is its ability to use a diversity of carbon sources. R. palustris is capable of CO 2 fixation, contributing to photoautotrophic conversion of CO 2 into valuable chemicals. R. palustris can assimilate short-chain organic acids and crude glycerol from industrial and agricultural wastewater. Lignocellulosic biomass hydrolysates can also be degraded by R. palustris . Utilization of these feedstocks can reduce the industry cost and is beneficial for environment. Applications of R. palustris for biopolymers and their building blocks production, and biofuels production are discussed. Afterward, some novel applications in microbial fuel cells, microbial electrosynthesis and photocatalytic synthesis are summarized. The challenges of the application of R. palustris are analyzed, and possible solutions are suggested., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Li, Ning, Sun, Luo and Yang.)

Synthetic estrogenic compounds such as 17α-ethinylestradiol (EE2) are significant environmental contaminants. This research studied the biodegradation of EE2 utilizing the EE2 adapted cells isolated from a dairy farm waste site in suspension flask vis-a-vis Bioelectrochemical System (BES) and compared the power output in the BES with and without EE2 as a co-substrate. 78% removal of EE2 was observed in the BES as against 60% removal in suspension flasks. The maximum power density in the BES increased about 53% when EE2 is used as a co-substrate. The EE2 biodegradation studied using HPLC and Q-TOF methods, also proposes a hypothetical pathway for EE2 degradation by the newly isolated strain Rhodopseudomonas palustris MDOC01 and reports the significant metabolites like nicotinic acid and oxoproline being detected during bioelectrochemical treatment process of EE2. Study also suggests that Plasma peroxide treatment of anode material enhanced the overall performance in terms of biodegradation efficiency and power output., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Photosynthetic bacteria can be useful biotechnological tools-they produce a variety of valuable products, including high purity hydrogen, and can simultaneously treat recalcitrant wastewaters. However, while photobioreactors have been designed and modeled for photosynthetic algae and cyanobacteria, there has been less work on understanding the effect of light in photosynthetic bacterial fermentations. To design photobioreactors, and processes using these organisms, robust models of light penetration, utilization, and conversion are needed. This stydy uses experimental data from a tubular photobioreactor designed to focus in on light intensity effects, to model the effect of light intensity on the growth of Rhodopseudomonas palustris, a model photosynthetic bacterium. The work demonstrates that growth is controlled by light intensity, and that this organism does experience photolimitation below 200 W/m 2 and photoinhibition above 600 W/m 2 . This has implications for outdoor applications, as there will be low growth during the periods of limited light, and growth may be inhibited during the light intensive hours of mid-day. Further, the work presents a model for light penetration in cylindrical photobioreactors, which tends to be the most common geometry. The model developed showed good fit to the experimental data for each light intensity investigated, with high R 2 values and NRMSE values all below 20%. The work extends the modeling tools for these organisms, and will allow for better photobioreactor design, and the integration of modeling tools in designing processes which use photosynthetic bacteria., (© 2021 Wiley Periodicals LLC.)

The photosynthetic bacterium Rhodopseudomonas palustris converts nitrogen gas (N 2 ) to fertilizer ammonia (NH 3 ) and also produces clean energy hydrogen gas (H 2 ) from protons (H + ) when it is grown anaerobically in nitrogen fixing medium with illumination, a condition that promotes the expression of active nitrogenase. Compared with quantitative real-time PCR (qRT-PCR) and the lacZ reporter system, two methods commonly used for in vivo study of nitrogenase regulation in photosynthetic bacteria, the fluorescent protein reporter system has advantages in terms of its simplicity and sensitivity. However, little is known concerning if the fluorescent protein reporter system can be used in bacterial cells that need to grow anaerobically. Here, we developed an RFP-based method to measure the nitrogenase gene expression in photosynthetic bacteria grown anaerobically. This method was able to determine the levels of both the genome-based and the plasmid-based nitrogenase expression under anaerobic conditions, providing a better method for in vivo study of gene expression affected by oxygen. The RFP reporter system developed here will promote a better understanding of the molecular mechanism of nitrogenase regulation and will be used on other genes of interest in a wider range of anaerobic bacteria.

The organic polymer lignin is a component of plant cell walls, which like (hemi)-cellulose is highly abundant in nature and relatively resistant to degradation. However, extracellular enzymes released by natural microbial consortia can cleave the β-aryl ether linkages in lignin, releasing monoaromatic phenylpropanoids that can be further catabolised by diverse species of bacteria. Biodegradation of lignin is therefore important in global carbon cycling, and its natural abundance also makes it an attractive biotechnological feedstock for the industrial production of commodity chemicals. Whilst the pathways for degradation of lignin-derived aromatics have been extensively characterised, much less is understood about how they are recognised and taken up from the environment. The purple phototrophic bacterium Rhodopseudomonas palustris can grow on a range of phenylpropanoid monomers and is a model organism for studying their uptake and breakdown. R. palustris encodes a tripartite ATP-independent periplasmic (TRAP) transporter (TarPQM) linked to genes encoding phenylpropanoid-degrading enzymes. The periplasmic solute-binding protein component of this transporter, TarP, has previously been shown to bind aromatic substrates. Here, we determine the high-resolution crystal structure of TarP from R. palustris as well as the structures of homologous proteins from the salt marsh bacterium Sagittula stellata and the halophile Chromohalobacter salexigens, which also grow on lignin-derived aromatics. In combination with tryptophan fluorescence ligand-binding assays, our ligand-bound co-crystal structures reveal the molecular basis for high-affinity recognition of phenylpropanoids by these TRAP transporters, which have potential for improving uptake of these compounds for biotechnological transformations of lignin., (© 2021 The Authors. The FEBS Journal published by John Wiley & Sons Ltd on behalf of Federation of European Biochemical Societies.)

Rhodopseudomonas palustris CGA009 is a metabolically robust microbe that can utilize lignin breakdown products to produce polyhydroxyalkanoates (PHAs), biopolymers with the potential to replace conventional plastics. Our recent efforts suggest PHA granule formation is a limiting factor for maximum production of the bioplastic poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) by R. palustris. The Phap1 phasin ( phaP1 ) from the PHB-producing model bacterium Cupriavidus necator H16 was expressed in R. palustris with the aim of overproducing PHBV from the lignin breakdown product p- coumarate by fostering smaller and more abundant granules. Expression of phaP1 yielded PHBV production from R. palustris aerobically (0.7 g/L), which does not occur in the wild-type strain, and led to a significantly higher PHBV titer than wild-type anaerobic production (0.41 g/L). The 3HV fractions were also significantly increased under both anaerobic and aerobic conditions, which boosts thermomechanical properties and potential for application. Thus, heterologous phasin expression in R. palustris provides flexibility for industrial processing and could foster compositional changes in copolymers with better thermomechanical properties compared to PHB alone., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2021 The Authors.)

Background: GroEL, which is a chaperone, plays a key role in maintaining protein homeostasis and, among other functions, serves to prevent protein misfolding and aggregation. In addition, the GroEL protein also has a significant effect on enhancing plant resistance and inhibiting plant diseases. However, the function of the GroEL protein in the inhibition of rice blast remains unknown., Results: Field experiment results show that photosynthetic bacteria PSB-06 have a good control effect on Magnaporthe oryzae. PSB-06 also can promote rice growth and enhance stress resistance. A GroEL protein which was separated and purified from photosynthetic bacteria had a significant antagonistic effect on appressorial formation and pathogenicity of Magnaporthe oryzae, meanwhile transcriptional analysis demonstrated that the GroEL protein could improve the expression of defense gene of rice., Conclusion: Our results show that the photosynthetic bacteria Rhodopseudomonas palustris significantly controls rice blast disease. Its action involves an extracellular GroEL protein, which inhibits appressoria formation, antagonizes the pathogenicity of Magnaporthe oryzae and promotes a host defense response. The research results provide evidence of the potential of this photosynthetic bacterium as a biocontrol agent at least for rice blast control. © 2021 Society of Chemical Industry., (© 2021 Society of Chemical Industry.)

The phototrophic bacterium Rhodopseudomonas palustris is emerging as a promising biotechnological chassis organism, due to its resilience to a range of harsh conditions, a wide metabolic repertoire, and the ability to quickly regenerate ATP using light. However, realization of this promise is impeded by a lack of efficient, rapid methods for genetic modification. Here, we present optimized tools for generating chromosomal insertions and deletions employing electroporation as a means of transformation. Generation of markerless strains can be completed in 12 days, approximately half the time for previous conjugation-based methods. This system was used for overexpression of alternative nitrogenase isozymes with the aim of improving biohydrogen productivity. Insertion of the pucBa promoter upstream of vnf and anf nitrogenase operons drove robust overexpression up to 4000-fold higher than wild-type. Transcript quantification was facilitated by an optimized high-quality RNA extraction protocol employing lysis using detergent and heat. Overexpression resulted in increased nitrogenase protein levels, extending to superior hydrogen productivity in bioreactor studies under nongrowing conditions, where promoter-modified strains better utilized the favorable energy state created by reduced competition from cell division. Robust heterologous expression driven by the pucBa promoter is thus attractive for energy-intensive biosyntheses suited to the capabilities of R. palustris . Development of this genetic modification toolset will accelerate the advancement of R. palustris as a biotechnological chassis organism, and insights into the effects of nitrogenase overexpression will guide future efforts in engineering strains for improved hydrogen production.

This study presents the biodegradation potential of ethinylestradiol (EE2) in anaerobic environments using exoelectrogenic activity of Rhodopseudomonas palustris. EE2, a basic ingredient in oral contraceptives, is a significant estrogenic micropollutant in various wastewaters and is considered highly recalcitrant. This recalcitrance of EE2 has caused anoxic areas to become repositories for these pollutants. Thus, it is essential to find the microorganisms and suitable methods to degrade this compound. An initial EE2 concentration of 1 mg/L, used in an anaerobic photobioreactor, resulted in 70% EE2 degradation over a period of 16 days with an increase of 63% in hydrogen production when EE2 was used with glycerol as the main carbon source in the culture medium. Furthermore, in the novel setup of hybrid photo-assisted microbial fuel cell (h-PMFC) employed here, EE2 degradation enhanced to 89.82% with a maximum power density of 0.633 ± 0.04 mW/m 2 . The hybrid MFC employed here could metabolize EE2 and sustained the bio-hydrogen production for 14 days to run the hydrogen fuel cell which otherwise could not be sustained with glycerol only and thus increased the overall power output. The current work highlights the use of R. palustris and the significance of co-metabolism in bioremediation of pollutants and bioenergy generation., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Rhodopseudomonas palustris is a model microorganism for studying the anaerobic metabolism of aromatic compounds. While it is well documented which aromatics can serve as sole organic carbon sources, co-metabolism of other aromatics is poorly understood. This study used kinetic modeling to analyze the simultaneous degradation of aromatic compounds present in corn stover hydrolysates and model the co-metabolism of aromatics not known to support growth of R. palustris as sole organic substrates. The simulation predicted that p-coumaroyl amide and feruloyl amide were hydrolyzed to p-coumaric acid and ferulic acid, respectively, and further transformed via p-coumaroyl-CoA and feruloyl-CoA. The modeling also suggested that metabolism of p-hydroxyphenyl aromatics was slowed by substrate inhibition, whereas the transformation of guaiacyl aromatics was inhibited by their p-hydroxyphenyl counterparts. It also predicted that substrate channeling may occur during degradation of p-coumaroyl-CoA and feruloyl-CoA, resulting in no detectable accumulation of p-hydroxybenzaldehyde and vanillin, during the transformation of these CoA ligated compounds to p-hydroxybenzoic acid and vanillic acid, respectively. While the simulation correctly represented the known transformation of p-hydroxybenzoic acid via the benzoyl-CoA pathway, it also suggested co-metabolism of vanillic acid and syringic acid, which are known not to serve as photoheterotrophic growth substrate for R. palustris.

G-negative bacteria produce myriad N -acyl-homoserine lactones (AHLs) that can function as quorum sensing (QS) signaling molecules. AHLs are also known to regulate various plant biological activities. p -Coumaroyl-homoserine lactone ( p C-HSL) is the only QS molecule produced by a photosynthetic bacterium, Rhodopseudomonas palustris . The role of p C-HSL in the interaction between R. palustris and plant has not been investigated. In this study, we investigated the effect of p C-HSL on plant immunity and found that this QS molecule can induce a systemic resistance to Tobacco mosaic virus (TMV) infection in Nicotiana benthamiana . The results show that p C-HSL treatment can prolong the activation of two mitogen-associated protein kinase genes (i.e., NbSIPK and NbWIPK ) and increase the expression of transcription factor WRKY8 as well as immune response marker genes NbPR1 and NbPR10 , leading to an increased accumulation of reactive oxygen species (ROS) in the TMV-infected plants. Our results also show that p C-HSL treatment can increase activities of two ROS-scavenging enzymes, peroxidase and superoxide dismutase. Knockdown of NbSIPK or NbWIPK expression in N. benthamiana plants through virus-induced gene silencing nullified or attenuated p C-HSL-induced systemic resistance, indicating that the functioning of p C-HSL relies on the activity of those two kinases. Meanwhile, p C-HSL-pretreated plants also showed a strong induction of kinase activities of NbSIPK and NbWIPK after TMV inoculation. Taken together, our results demonstrate that p C-HSL treatment increases plant resistance to TMV infection, which is helpful to uncover the outcome of interaction between R. palustris and its host plants.

One water exopolysaccharide, designated G-EPS, was secreted by Rhodopseudomonas palustris GJ-22 culture media. The structure of G-EPS was characterized with HPGPC, GC-MS, methylation, 1D and 2D NMR, along with UV and FT-IR spectrum. The G-EPS molecular weight was 10.026 kilodalton, and is composed of D-mannose (92.8%) and d-glucose (7.2%). The purified G-EPS promoted plant growth and induced systemic resistance against TMV in Nicotiana benthamiana. These results suggested that G-EPS is an important active component of the bio-control capacity of GJ-22., Competing Interests: Declaration of competing interest We declare that we have no conflict of interest., (Copyright © 2020. Published by Elsevier B.V.)

Transfer RNA (tRNA) is an essential link between the genetic code and proteins. During the process of translation, tRNA is charged with its cognate amino acid and delivers it to the ribosome, thus serving as a substrate of protein synthesis. To analyze the charging state of a particular tRNA, total RNA is purified and analyzed on an acid-urea gel. Separated RNA is then transferred to a membrane and detected with a probe for the tRNA of interest. Here, we present an improved protocol to analyze the tRNA charging state in the α-proteobacterium Rhodopseudomonas palustris . Compared to the classical method, the RNA isolation step is optimized to suit this organism. Additionally, a non-radioactive platform is used for electrophoresis and Northern blots. This significantly reduces the time and the effort required for this protocol., Competing Interests: Competing interestsThe authors claim no financial or non-financial competing interest., (Copyright © 2020 The Authors; exclusive licensee Bio-protocol LLC.)

Ribosomes are an integral part of cellular life. They are complex molecular machines consisting of multiple ribosomal proteins and RNAs. To study different aspects of ribosome composition, many methods have been developed over the decades. Here, we describe how to purify ribosomes from the α-proteobacterium Rhodopseudomonas palustris . Following this protocol, RNA can be extracted from either purified ribosomes or directly from cell cultures, and ribosomal RNAs quantified using Northern blot. This protocol gives an example of studying ribosomes in a bacterium other than the commonly used E. coli . The challenge of performing Northern blots with rRNA is also addressed in detail., Competing Interests: Competing interestsThe authors claim no financial or non-financial competing interests., (Copyright © 2020 The Authors; exclusive licensee Bio-protocol LLC.)

Biological nitrogen fixation is catalyzed by the enzyme nitrogenase. Two forms of this metalloenzyme, the vanadium (V)- and iron (Fe)-only nitrogenases, were recently found to reduce small amounts of carbon dioxide (CO 2 ) into the potent greenhouse gas methane (CH 4 ). Here, we report carbon ( 13 C/ 12 C) and hydrogen ( 2 H/ 1 H) stable isotopic compositions and fractionations of methane generated by V- and Fe-only nitrogenases in the metabolically versatile nitrogen fixer Rhodopseudomonas palustris The stable carbon isotope fractionation imparted by both forms of alternative nitrogenase are within the range observed for hydrogenotrophic methanogenesis ( 13 α CO2/CH4 = 1.051 ± 0.002 for V-nitrogenase and 1.055 ± 0.001 for Fe-only nitrogenase; values are means ± standard errors). In contrast, the hydrogen isotope fractionations ( 2 α H2O/CH4 = 2.071 ± 0.014 for V-nitrogenase and 2.078 ± 0.018 for Fe-only nitrogenase) are the largest of any known biogenic or geogenic pathway. The large 2 α H2O/CH4 shows that the reaction pathway nitrogenases use to form methane strongly discriminates against 2 H, and that 2 α H2O/CH4 distinguishes nitrogenase-derived methane from all other known biotic and abiotic sources. These findings on nitrogenase-derived methane will help constrain carbon and nitrogen flows in microbial communities and the role of the alternative nitrogenases in global biogeochemical cycles. IMPORTANCE All forms of life require nitrogen for growth. Many different kinds of microbes living in diverse environments make inert nitrogen gas from the atmosphere bioavailable using a special enzyme, nitrogenase. Nitrogenase has a wide substrate range, and, in addition to producing bioavailable nitrogen, some forms of nitrogenase also produce small amounts of the greenhouse gas methane. This is different from other microbes that produce methane to generate energy. Until now, there was no good way to determine when microbes with nitrogenases are making methane in nature. Here, we present an isotopic fingerprint that allows scientists to distinguish methane from microbes making it for energy versus those making it as a by-product of nitrogen acquisition. With this new fingerprint, it will be possible to improve our understanding of the relationship between methane production and nitrogen acquisition in nature., (Copyright © 2020 Luxem et al.)

Background: Phototrophic purple non-sulfur bacteria (PNSB) have gained attention for their ability to produce a valuable clean energy source in the form biohydrogen via photofermentation of a wide variety of organic wastes. For maturation of these phototrophic bioprocesses towards commercial feasibility, development of suitable immobilisation materials is required to allow continuous production from a stable pool of catalytic biomass in which energy is not diverted towards biomass accumulation, and optimal hydrogen production rates are realised. Here, the application of transparent polyvinyl-alcohol (PVA) cryogel beads to immobilisation of Rhodopseudomonas palustris for long-term hydrogen production is described. PVA cryogel properties are characterised and demonstrated to be well suited to the purpose of continuous photofermentation. Finally, analysis of the long-term biocompatibility of the material is illustrated., Results: The addition of glycerol co-solvent induces favourable light transmission properties in normally opaque PVA cryogels, especially well-suited to the near-infrared light requirements of PNSB. Material characterisation showed high mechanical resilience, low resistance to diffusion of substrates and high biocompatibility of the material and immobilisation process. The glycerol co-solvent in transparent cryogels offered additional benefit by reinforcing physical interactions to the extent that only a single freeze-thaw cycle was required to form durable cryogels, extending utility beyond only phototrophic bioprocesses. In contrast, conventional PVA cryogels require multiple cycles which compromise viability of entrapped organisms. Hydrogen production studies of immobilised Rhodopseudomonas palustris in batch photobioreactors showed higher specific hydrogen production rates which continued longer than planktonic cultures. Continuous cultivation yielded hydrogen production for at least 67 days from immobilised bacteria, demonstrating the suitability of PVA cryogel immobilisation for long-term phototrophic bioprocesses. Imaged organisms immobilised in cryogels showed a monolithic structure to PVA cryogels, and demonstrated a living, stable, photofermentative population after long-term immobilisation., Conclusion: Transparent PVA cryogels offer ideal properties as an immobilisation matrix for phototrophic bacteria and present a low-cost photobioreactor technology for the further advancement of biohydrogen from waste as a sustainable energy source, as well as development of alternative photo-bioprocesses exploiting the unique capabilities of purple non-sulfur bacteria., Competing Interests: Competing interestsThe authors declare that they have no competing interests., (© The Author(s) 2020.)

The genus Rhodopseudomonas , containing purple nonsulfur photosynthetic Proteobacteria, has a number of strains that belong to different species, although many of them are collectively called Rhodopseudomonas palustris . The type species R. palustris and closely related species are the focus of this paper. The comparison of available genome sequences indicate that the following Rhodopseudomonas species are well recognized: R. palustris (strains ATH 2.1.6 T =DSM 123 T =NBRC 100419 T and BisB5), Rhodopseudomonas rutila (strains R1 T , DSM 126, CGA009, ATH 2.1.37, Eli 1980, ATCC 17001 and TIE1), Rhodopseudomonas pentothenatexigens JA575 T and Rhodopseudomonas faecalis JCM 11668 T . Other strains for which genome sequences are available are distinct from these four species. Evidence is presented that R. palustris strain ATH 2.1.6 T -KCM as obtained directly from the van Niel collection by one of us (T.E.M.) is identical to the DSMZ deposit DSM 123 T of ATH 2.1.6 T , but not to the deposit at ATCC 17001. The amino acid sequences of the cytochromes C2 and C556 from R. palustris strain ATH 2.1.6 T -KCM are in complete agreement with the translated genome sequences of R. palustris DSM 123 T . In addition, the 16S rRNA gene sequence of R. palustris NBRC 100419 T completely matches that of strain DSM 123 T . In conclusion, the type strain of R. palustris ATH 2.1.6 T is correctly represented by DSM 123 T and NBRC 100419 T . However, the deposit at ATCC 17001 has properties that do not conform with properties of authentic R. palustris , but rather indicate that this is a strain of R. rutila . The previously suggested assignment of the type strain of R. palustris DSM 123 T to the new species R. pseudopalustris was incorrect because strain DSM 123 T is the authentic type strain of R. palustris .

The purple nonsulfur phototrophic bacterium Rhodopseudomonas palustris strain CGA009 uses the three-carbon dicarboxylic acid malonate as the sole carbon source under phototrophic conditions. However, this bacterium grows extremely slowly on this compound and does not have operons for the two pathways for malonate degradation that have been detected in other bacteria. Many bacteria grow on a spectrum of carbon sources, some of which are classified as poor growth substrates because they support low growth rates. This trait is rarely addressed in the literature, but slow growth is potentially useful in biotechnological applications where it is imperative for bacteria to divert cellular resources to value-added products rather than to growth. This prompted us to explore the genetic and physiological basis for the slow growth of R. palustris with malonate as a carbon source. There are two unlinked genes annotated as encoding a malonyl coenzyme A (malonyl-CoA) synthetase (MatB) and a malonyl-CoA decarboxylase (MatA) in the genome of R. palustris , which we verified as having the predicted functions. Additionally, two tripartite ATP-independent periplasmic transporters (TRAP systems) encoded by rpa2047 to rpa2049 and rpa2541 to rpa2543 were needed for optimal growth on malonate. Most of these genes were expressed constitutively during growth on several carbon sources, including malonate. Our data indicate that R. palustris uses a piecemeal approach to growing on malonate. The data also raise the possibility that this bacterium will evolve to use malonate efficiently if confronted with an appropriate selection pressure. IMPORTANCE There is interest in understanding how bacteria metabolize malonate because this three-carbon dicarboxylic acid can serve as a building block in bioengineering applications to generate useful compounds that have an odd number of carbons. We found that the phototrophic bacterium Rhodopseudomonas palustris grows extremely slowly on malonate. We identified two enzymes and two TRAP transporters involved in the uptake and metabolism of malonate, but some of these elements are apparently not very efficient. R. palustris cells growing with malonate have the potential to be excellent biocatalysts, because cells would be able to divert cellular resources to the production of value-added compounds instead of using them to support rapid growth. In addition, our results suggest that R. palustris is a candidate for directed evolution studies to improve growth on malonate and to observe the kinds of genetic adaptations that occur to make a metabolic pathway operate more efficiently., (Copyright © 2020 American Society for Microbiology.)

We conducted this research in order to investigate the potential of a new material called glass pumice for use as a microorganism immobilization carrier to improve aquaculture pond water quality. The pH adjustment capacity and the Rhodopseudomonas palustris P1 cell adsorption capacity of glass pumice were measured. The immobilized Rps. palustris P1 and the free sample were compared to determine which had an enhanced NH 4 + -N and NO 2 - -N removal efficiency. The results showed that glass pumice significantly affected the pH of the acid solution (P < 0.05); the pH increased from 3.0 ± 0.08 to 7.21 ± 0.13 in 12 H. Rps. palustris P1 adsorption to glass pumice was rapid and reached equilibrium within 60 Min. The Langmuir adsorption parameter data showed that glass pumice had a higher affinity for Rps. palustris P1 than SiO 2 powder, with an adsorption capacity of 4.02 × 10 8  cells g -1 . The maximum NH 4 + -N and NO 2 - -N removal rates by immobilized Rps. palustris P1 were 134.82 ± 0.67% and 93.68 ± 0.14% higher than those of nonimmobilized P1, respectively. Based on the above results, we propose that glass pumice is potential as a microorganism carrier material in aquaculture water treatment., (© 2019 International Union of Biochemistry and Molecular Biology, Inc.)

This study investigated the regulatory role of Rhodopseudomonas palustris RP11 in alleviating TBBPA-induced harmful effects in soybean seedlings. In this study, the characteristics of growth promotion by strain RP11 were studied by analysing 5-aminolevulinic acid (ALA) and indole-3-acetic acid (IAA) production, as well as phosphorus-solubilizing and potassium-solubilizing ability. In the pot culture conditions, we tested whether strain RP11 improved soybean seedlings tolerance against TBBPA by measuring the root length and physiological parameters of the seedlings treated with strain RP11 and different concentration of TBBPA (0, 5, 50, 100, and 1000 mg/kg) together. The results showed that strain RP11 secreted IAA and ALA, and solubilized phosphate and potassium. In pot trials, strain RP11 increased the root length, chlorophyll content, carotenoid content, soluble sugar and protein content of soybean seedlings treated with TBBPA, in comparison with the seedlings treated only with TBBPA. Furthermore, strain RP11 induced the activities of superoxide dismutase (SOD), catalase (CAT) and peroxidase (POD), decreased the malondialdehyde (MDA) content in soybean seedlings under TBBPA stress. It was concluded that strain RP11 alleviated TBBPA-induced harmful effects in soybean seedlings by the secretion of IAA and ALA, the accumulation of carotenoid, soluble sugar and soluble protein, and the induction of SOD, CAT and POD as well as nutrient adjustment of phosphorus and potassium levels.

The predominance of the maximum at 800 nm for the light-harvesting complex LH4 (B800) and at 850 nm for LH2 (B800-850) from Rps. palustris is determined by the composition of αβ-polypeptides and pigments. In low light (LL) for Rps. palustris, strain KM 286 (1e5), along with LH4, the LL LH2 complex was synthesized with the same absorption at 800 and 850 nm. It differed from the LH4 and LH2 complex, which is synthesized under high illumination, in the composition and content of carotenoids (Car) and bacteriochlorophyll a (BChl a). LH4 differed from LL LH2 and LH2 by an additional emission maximum at 766 nm in the BChl a fluorescence spectra. All three complexes had approximately the same level (about 45%) of the energy transfer efficiency from Car to BChl a. Isolation of LL LH2 complex from Rps. palustris confirms the hypothesis of the synthesis in these bacteria under low light conditions of other types of complexes, except LH4, which is due to the multiple biosynthesis genes of αβ-polypeptides and the possibility of their various combinations.

The operation of bioelectrochemical systems (BESs) relies on the ability of microbes to export electrons outside of their cells. However, microorganisms are not evolutionary conceived to power BESs as most of the redox processes occur within. In this study, a low cost strategy equivalent to the one used to improve hydrogen production is employed to divert electrons from the metabolism to an electrode. Varying the ratio of nitrogen to carbon concentration (0, 0.20 and 0.54) determines what fraction of the electron flux is directed towards biosynthesis, biohydrogen generation and extracellular electron transfer. The ratio of 0.54 produced a higher specific growth rate while the ratio of 0.20 resulted in combined higher maximum specific hydrogen production and exoelectrogenic activity, translating into a maximum power density of 2.39 ± 0.13 mW m -2 in a novel hybrid hydrogen-photosynthetic microbial fuel cell. The current work sets a framework for the optimisation of R. palustris for bioenergy recovery., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2019. Published by Elsevier Ltd.)