-
Applied and Environmental Microbiology Aug 2020The enzymatic production of 2,5-furandicarboxylic acid (FDCA) from 5-hydroxymethylfurfural (HMF) has gained interest in recent years, as FDCA is a renewable precursor of...
The enzymatic production of 2,5-furandicarboxylic acid (FDCA) from 5-hydroxymethylfurfural (HMF) has gained interest in recent years, as FDCA is a renewable precursor of poly(ethylene-2,5-furandicarboxylate) (PEF). 5-Hydroxymethylfurfural oxidases (HMFOs) form a flavoenzyme family with genes annotated in a dozen bacterial species but only one enzyme purified and characterized to date (after heterologous expression of a sp. HMFO gene). This oxidase acts on both furfuryl alcohols and aldehydes and, therefore, is able to catalyze the conversion of HMF into FDCA through 2,5-diformylfuran (DFF) and 2,5-formylfurancarboxylic acid (FFCA), with only the need of oxygen as a cosubstrate. To enlarge the repertoire of HMFO enzymes available, genetic databases were screened for putative HMFO genes, followed by heterologous expression in After unsuccessful trials with other bacterial HMFO genes, HMFOs from two species were produced as active soluble enzymes, purified, and characterized. The sp. enzyme was also produced and purified in parallel for comparison. Enzyme stability against temperature, pH, and hydrogen peroxide, three key aspects for application, were evaluated (together with optimal conditions for activity), revealing differences between the three HMFOs. Also, the kinetic parameters for HMF, DFF, and FFCA oxidation were determined, the new HMFOs having higher efficiencies for the oxidation of FFCA, which constitutes the bottleneck in the enzymatic route for FDCA production. These results were used to set up the best conditions for FDCA production by each enzyme, attaining a compromise between optimal activity and half-life under different conditions of operation. HMFO is the only enzyme described to date that can catalyze by itself the three consecutive oxidation steps to produce FDCA from HMF. Unfortunately, only one HMFO enzyme is currently available for biotechnological application. This availability is enlarged here by the identification, heterologous production, purification, and characterization of two new HMFOs, one from and one from an unidentified species. Compared to the previously known HMFO, the new enzyme from exhibits better performance for FDCA production in wider pH and temperature ranges, with higher tolerance for the hydrogen peroxide formed, longer half-life during oxidation, and higher yield and total turnover numbers in long-term conversions under optimized conditions. All these features are relevant properties for the industrial production of FDCA. In summary, gene screening and heterologous expression can facilitate the selection and improvement of HMFO enzymes as biocatalysts for the enzymatic synthesis of renewable building blocks in the production of bioplastics.
Topics: Bacterial Proteins; Dicarboxylic Acids; Escherichia coli; Furaldehyde; Furans; Methylophilaceae; Microorganisms, Genetically-Modified; Oxidoreductases; Pseudomonas
PubMed: 32503910
DOI: 10.1128/AEM.00842-20 -
Applied Microbiology and Biotechnology Jun 2017The surfactant sodium lauryl ether sulfate (SLES) is widely used in the composition of detergents and frequently ends up in wastewater treatment plants (WWTPs). While...
The surfactant sodium lauryl ether sulfate (SLES) is widely used in the composition of detergents and frequently ends up in wastewater treatment plants (WWTPs). While aerobic SLES degradation is well studied, little is known about the fate of this compound in anoxic environments, such as denitrification tanks of WWTPs, nor about the bacteria involved in the anoxic biodegradation. Here, we used SLES as sole carbon and energy source, at concentrations ranging from 50 to 1000 mg L, to enrich and isolate nitrate-reducing bacteria from activated sludge of a WWTP with the anaerobic-anoxic-oxic (A/O) concept. In the 50 mg L enrichment, Comamonas (50%), Pseudomonas (24%), and Alicycliphilus (12%) were present at higher relative abundance, while Pseudomonas (53%) became dominant in the 1000 mg L enrichment. Aeromonas hydrophila strain S7, Pseudomonas stutzeri strain S8, and Pseudomonas nitroreducens strain S11 were isolated from the enriched cultures. Under denitrifying conditions, strains S8 and S11 degraded 500 mg L SLES in less than 1 day, while strain S7 required more than 6 days. Strains S8 and S11 also showed a remarkable resistance to SLES, being able to grow and reduce nitrate with SLES concentrations up to 40 g L. Strain S11 turned out to be the best anoxic SLES degrader, degrading up to 41% of 500 mg L. The comparison between SLES anoxic and oxic degradation by strain S11 revealed differences in SLES cleavage, degradation, and sulfate accumulation; both ester and ether cleavage were probably employed in SLES anoxic degradation by strain S11.
Topics: Aeromonas; Biodegradation, Environmental; Carbon; Comamonadaceae; Comamonas; Denitrification; Gram-Negative Bacteria; Oxidation-Reduction; Pseudomonas; Sewage; Sodium Dodecyl Sulfate; Surface-Active Agents
PubMed: 28299401
DOI: 10.1007/s00253-017-8212-x -
ISME Communications Sep 2023Food safety of leafy greens is an emerging public health issue as they can harbor opportunistic human pathogens (OHPs) and expose OHPs to consumers. Protists are an...
Food safety of leafy greens is an emerging public health issue as they can harbor opportunistic human pathogens (OHPs) and expose OHPs to consumers. Protists are an integral part of phyllosphere microbial ecosystems. However, our understanding of protist-pathogen associations in the phyllosphere and their consequences on public health remains poor. Here, we examined phyllosphere protists, human pathogen marker genes (HPMGs), and protist endosymbionts from four species of leafy greens from major supermarkets in Xiamen, China. Our results showed that Staphylococcus aureus and Klebsiella pneumoniae were the dominant human pathogens in the vegetable phyllosphere. The distribution of HPMGs and protistan communities differed between vegetable species, of which Chinese chive possessed the most diverse protists and highest abundance of HPMGs. HPMGs abundance positively correlated with the diversity and relative abundance of phagotrophic protists. Whole genome sequencing further uncovered that most isolated phyllosphere protists harbored multiple OHPs which carried antibiotic resistance genes, virulence factors, and metal resistance genes and had the potential to HGT. Colpoda were identified as key phagotrophic protists which positively linked to OHPs and carried diverse resistance and virulence potential endosymbiont OHPs including Pseudomonas nitroreducens, Achromobacter xylosoxidans, and Stenotrophomonas maltophilia. We highlight that phyllosphere protists contribute to the transmission of resistant OHPs through internalization and thus pose risks to the food safety of leafy greens and human health. Our study provides insights into the protist-OHP interactions in the phyllosphere, which will help in food safety surveillance and human health.
PubMed: 37660098
DOI: 10.1038/s43705-023-00302-z -
Molecular Plant-microbe Interactions :... Apr 2022
Topics: Acyl-Butyrolactones; Pseudomonas; Quorum Sensing
PubMed: 35285671
DOI: 10.1094/MPMI-12-21-0310-A -
Applied and Environmental Microbiology Nov 2016Pseudomonas nitroreducens TX1 is of special interest because of its ability to utilize 0.05% to 20% octylphenol polyethoxylates (OPEO) as a sole source of carbon. In...
Pseudomonas nitroreducens TX1 is of special interest because of its ability to utilize 0.05% to 20% octylphenol polyethoxylates (OPEO) as a sole source of carbon. In this study, a library containing 30,000 Tn5-insertion mutants of the wild-type strain TX1 was constructed and screened for OPEO utilization, and 93 mutants were found to be unable to grow on OPEO In total, 42 separate disrupted genes were identified, and the proteins encoded by the genes were then classified into various categories, namely, information storage and processing (14.3%), cellular processes and signaling (28.6%), metabolism (35.7%), and unknown proteins (21.4%). The individual deletion of genes encoding isocitrate lyase (aceA), malate synthase (aceB), and glycolate dehydrogenase (glcE) was carried out, and the requirement for aceA and aceB but not glcE confirmed the role of the glyoxylate cycle in OPEO degradation. Furthermore, acetaldehyde dehydrogenase and acetyl-coenzyme A (acetyl-CoA) synthetase activity levels were 13.2- and 2.1-fold higher in TX1 cells grown on OPEO than in TX1 cells grown on succinate, respectively. Growth of the various mutants on different carbon sources was tested, and based on these findings, a mechanism involving exoscission to liberate acetaldehyde from the end of the OPEO chain during degradation is proposed for the breakdown of OPEO IMPORTANCE: Octylphenol polyethoxylates belong to the alkylphenol polyethoxylate (APEO) nonionic surfactant family. Evidence based on the analysis of intermediate metabolites suggested that the primary biodegradation of APEO can be achieved by two possible pathways for the stepwise removal of the C ethoxylate units from the end of the chain. However, direct evidence for these hypotheses is still lacking. In this study, we described the use of transposon mutagenesis to identify genes critical to the catabolism of OPEO by P. nitroreducens TX1. The exoscission of the ethoxylate chain leading to the liberation of acetaldehyde is proposed. Isocitrate lyase and malate synthase in glyoxylate cycle are required in the catabolism of ethoxylated surfactants. Our findings also provide many gene candidates that may help elucidate the mechanisms in stress responses to ethoxylated surfactants by bacteria.
Topics: Acetaldehyde; Alcohol Oxidoreductases; Aldehyde Oxidoreductases; Bacterial Proteins; DNA Transposable Elements; Gene Deletion; Isocitrate Lyase; Malate Synthase; Mutagenesis, Insertional; Phenols; Pseudomonas; Succinic Acid; Surface-Active Agents
PubMed: 27590807
DOI: 10.1128/AEM.01907-16 -
BMC Infectious Diseases Feb 2024Pseudomonas nitroreducens is a non-fermenting, gram-negative, rod-shaped bacterium commonly inhabiting soil, particularly soil contaminated with oil brine. To our...
BACKGROUND
Pseudomonas nitroreducens is a non-fermenting, gram-negative, rod-shaped bacterium commonly inhabiting soil, particularly soil contaminated with oil brine. To our knowledge, no cases of human infection with P. nitroreducens have been previously reported. Here, we present the first documented case of cholangitis caused by P. nitroreducens in a patient with bacteremia.
CASE PRESENTATION
A 46-year-old Japanese man with an advanced pancreatic neuroendocrine tumor was hospitalized with fever and chills. Four days before admission, the patient developed right upper abdominal pain. Two days later, he also experienced fever and chills. Endoscopic retrograde cholangiopancreatography was performed on the day of admission, and the patient was diagnosed as having cholangitis associated with stent dysfunction. Gram-negative rods were isolated from blood cultures, but attempts to identify the bacteria using VITEK2 and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) with VITEK MS ver. 4.7.1 (bioMérieux Japan Co. Ltd., Tokyo, Japan) were unsuccessful. Finally, the organism was identified as P. nitroreducens using MALDI-TOF MS with a MALDI Biotyper (Bruker Daltonics Co., Ltd., Billerica, MA, USA) and 16 S ribosomal RNA sequencing. Despite thorough interviews with the patient, he denied any exposure to contaminated soil. The patient was treated with intravenous cefepime and oral ciprofloxacin for 16 days based on susceptibility results, achieving a good therapeutic outcome. At the outpatient follow-up on day 28, the patient was in good general condition.
CONCLUSIONS
This is the first reported human case of cholangitis with bloodstream infection caused by P. nitroreducens. This report provides clinicians with novel insights into the clinical manifestations and diagnostic methods necessary for the accurate diagnosis of P. nitroreducens, along with guidance on treatment.
Topics: Male; Humans; Middle Aged; Neuroendocrine Tumors; Bacteremia; Bacteria; Pseudomonas; Bacteria, Aerobic; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Cholangitis; Soil; Pancreatic Neoplasms
PubMed: 38336644
DOI: 10.1186/s12879-024-09092-8 -
Biotechnology Reports (Amsterdam,... Dec 2021The mechanisms of tolerance to heavy metals used by some microorganisms identified by bioprospection processes are useful for the development and implementation of...
The mechanisms of tolerance to heavy metals used by some microorganisms identified by bioprospection processes are useful for the development and implementation of bioremediation strategies for contaminated environments with high toxic load caused by heavy metals. A total of seven native microbial isolates were obtained from wastewater bodies from an industrial zone in the municipality of Girardota, Antioquia, Colombia. Subsequently, they were selected to evaluate their lead tolerance capacity at different concentrations. In addition, some parameters were determined, such as the capacity to produce exopolysaccharides and their biosorption to understand potential mechanisms associated to lead tolerance. According to the biocehemical test (Vitek) and the molecular analysis of sequences of 16S rDNA, bacterial were identified as , and . We determined that the seven isolates had the capacity to tolerate concentrations higher than 50 mg/ml of lead, and that the concentration and exposure time (40 h) to this metal significantly affect the spp. isolates. Statistically significant differences were detected ( < 0.05) in the production of the exopolysaccharide (EPS) among the isolates. (P16) was the strain with the maximum absorbance exopolysaccharide measured. We evidenced that (P14) and (P20) have 80% capacity to biosorber lead using live mass (minimum range from 80.9% to 87%). It is suggested that the tolerance to lead exhibited by the environmental isolates of spp. can be attributed to the production of exopolysaccharides and biosorption, which are protection factors for its survival in contaminated places. Finally, it was determined that the adsorption measured from dead biomass was significant ( < 0.05) from 40 h of exposure to metal (Average 182.2 ± 7). We generated new knowledge about the potential use of the spp. genus to bioremediate affluent contaminated with heavy metals.
PubMed: 34765463
DOI: 10.1016/j.btre.2021.e00685 -
International Journal of Molecular... Nov 2022The B12-producing strains DSM 1650 and sp. CCUG 2519 (both formerly ), with the most distributed pathway among bacteria for exogenous choline/betaine utilization, are...
The B12-producing strains DSM 1650 and sp. CCUG 2519 (both formerly ), with the most distributed pathway among bacteria for exogenous choline/betaine utilization, are promising recombinant hosts for the endogenous production of B12 precursor betaine by direct methylation of bioavailable glycine or non-proteinogenic -alanine. Two plasmid-based de novo betaine pathways, distinguished by their enzymes, have provided an expression of the genes encoding for -methyltransferases of the halotolerant cyanobacterium or plant to synthesize the internal glycine betaine or -alanine betaine, respectively. These betaines equally allowed the recombinant pseudomonads to grow effectively and to synthesize a high level of cobalamin, as well as to increase their protective properties against abiotic stresses to a degree comparable with the supplementation of an exogenous betaine. Both de novo betaine pathways significantly enforced the protection of bacterial cells against lowering temperature to 15 °C and increasing salinity to 400 mM of NaCl. However, the expression of the single plant-derived gene for the -alanine-specific -methyltransferase additionally increased the effectiveness of exogenous glycine betaine almost twofold on cobalamin biosynthesis, probably due to the ' ability to use two independent pathways, their own choline/betaine pathway and the plant -alanine betaine biosynthetic pathway.
Topics: Betaine; Choline; Pseudomonas; Stress, Physiological; Methyltransferases; beta-Alanine; Vitamin B 12
PubMed: 36430408
DOI: 10.3390/ijms232213934 -
Heliyon Jun 2023Polystyrene is a plastic that leads to environmental pollution. In particular, expanded polystyrene is very light and takes up much space, causing additional...
OBJECTIVES
Polystyrene is a plastic that leads to environmental pollution. In particular, expanded polystyrene is very light and takes up much space, causing additional environmental problems. The aim of this study was to isolate new symbiotic bacteria which degraded polystyrene from mealworms.
METHODS
The population of polystyrene degrading bacteria was increased by enrichment culture of intestinal bacteria from mealworms with polystyrene as a sole carbon source. The degradation activity of isolated bacteria was evaluated by morphological change of micro-polystyrene particles and the surface change of polystyrene films.
RESULTS
Eight isolated species (, , , , , , , and ) were identified that degrade polystyrene.
CONCLUSION
Bacterial identification shows that a broad spectrum of bacteria decomposing polystyrene coexists in the intestinal tract of mealworms.
PubMed: 37426801
DOI: 10.1016/j.heliyon.2023.e17352 -
AoB PLANTS Apr 2019Gametophytes of the fern develop into either hermaphrodites or males. As hermaphrodites develop, they secrete antheridiogen, or A, into the environment, inducing male...
Gametophytes of the fern develop into either hermaphrodites or males. As hermaphrodites develop, they secrete antheridiogen, or A, into the environment, inducing male development in undifferentiated gametophytes. Hermaphrodites are composed of archegonia, antheridia, rhizoids and a notch meristem, while males consist of antheridia and rhizoids. Much of the research on sexual and morphological development concerns gametophytes grown in sterile environments. Using biochemical and molecular techniques we identify a soil bacterium and explore its effects on sexual and rhizoid development. Hermaphrodite and male gametophytes were exposed to this bacterium and the effects on sexual development, rhizoid length and rhizoid number were explored. The bacterium was identified as a pseudomonad, . Gametophytes grown in the presence of the pseudomonad were more likely to develop into hermaphrodites across all gametophyte densities. Across all gametophyte sizes, hermaphrodites had rhizoids that were 2.95× longer in the presence of the pseudomonad while males had rhizoids that were 2.72× longer in the presence of the pseudomonad. Both hermaphrodite and male gametophytes developed fewer rhizoids in the presence of the pseudomonad. Control hermaphrodites produced 1.23× more rhizoids across all gametophyte sizes. For male gametophytes grown in the absence of the pseudomonad, the rate of increase in the number of rhizoids was greater with increasing size in the control than the rate of increase in males grown in the presence of the pseudomonad. The pseudomonad may be acting on gametophyte sexual development via several potential mechanisms: degradation of A, changes in nutrient availability or phytohormone production. The pseudomonad may also increase rhizoid number through production of phytohormones or changes in nutrient availability.
PubMed: 31019671
DOI: 10.1093/aobpla/plz012