Transcriptomic and metabolomic analyses revealed regulation mechanism of mixotrophic Cylindrotheca sp. glycerol utilization and biomass promotion.

BACKGROUND: Diatoms have been viewed as ideal cell factories for production of some high-value bioactive metabolites, such as fucoxanthin, but their applications are restrained by limited biomass yield. Mixotrophy, by using both CO2 and organic carbon source, is believed effective to crack the bottleneck of biomass accumulation and achieve a sustainable bioproduct supply. RESULTS: Glycerol, among tested carbon sources, was proved as the sole that could significantly promote growth of Cylindrotheca sp. with illumination, a so-called growth pattern, mixotrophy. Biomass and fucoxanthin yields of Cylindrotheca sp., grown in medium with glycerol (2 g L-1), was increased by 52% and 29%, respectively, as compared to the autotrophic culture (control) without compromise in photosynthetic performance. As Cylindrotheca sp. was unable to use glycerol without light, a time-series transcriptomic analysis was carried out to elucidate the light regulation on glycerol utilization. Among the genes participating in glycerol utilization, GPDH1, TIM1 and GAPDH1, showed the highest dependence on light. Their expressions decreased dramatically when the alga was transferred from light into darkness. Despite the reduced glycerol uptake in the dark, expressions of genes associating with pyrimidine metabolism and DNA replication were upregulated when Cylindrotheca sp. was cultured mixotrophically. Comparative transcriptomic and metabolomic analyses revealed amino acids and aminoacyl-tRNA metabolisms were enhanced at different timepoints of diurnal cycles in mixotrophic Cylindrotheca sp., as compared to the control. CONCLUSIONS: Conclusively, this study not only provides an alternative for large-scale cultivation of Cylindrotheca, but also pinpoints the limiting enzymes subject to further metabolic manipulation. Most importantly, the novel insights in this study should aid to understand the mechanism of biomass promotion in mixotrophic Cylindrotheca sp.

Metabolic Responses of a Model Green Microalga Euglena gracilis to Different Environmental Stresses.

Euglena gracilis, a green microalga known as a potential candidate for jet fuel producers and new functional food resources, is highly tolerant to antibiotics, heavy metals, and other environmental stresses. Its cells contain many high-value products, including vitamins, amino acids, pigments, unsaturated fatty acids, and carbohydrate paramylon as metabolites, which change contents in response to various extracellular environments. However, mechanism insights into the cellular metabolic response of Euglena to different toxic chemicals and adverse environmental stresses were very limited. We extensively investigated the changes of cell biomass, pigments, lipids, and paramylon of E. gracilis under several environmental stresses, such as heavy metal CdCl2, antibiotics paromomycin, and nutrient deprivation. In addition, global metabolomics by Ultra-high-performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) was applied to study other metabolites and potential regulatory mechanisms behind the differential accumulation of major high-valued metabolites. This study collects a comprehensive update on the biology of E. gracilis for various metabolic responses to stress conditions, and it will be of great value for Euglena cultivation and high-value [154mm][10mm]Q7metabolite production.

Rhizobium flavescens sp. nov., Isolated from a Chlorothalonil-Contaminated Soil.

A Gram-negative, facultative anaerobic, non-lagellated and rod-shaped bacterium FML-4T was isolated from a chlorothalonil-contaminated soil in Nanjing, China. Phylogenetic analyses of 16S rRNA genes revealed that the strain FML-4T shared the highest sequence similarity of 97.1% with Ciceribacter thiooxidans KCTC 52231T, followed by Rhizobium rosettiformans CCM 7583T (97.0%) and R. daejeonense KCTC 12121T (96.8%). Although the sequence similarities of the housekeeping genes thrC, rceA, glnII, and atpD between strain FML-4T and C. thiooxidans KCTC 52231T were 83.8%, 88.7%, 86.2%, and 92.0%, respectively, strain FML-4T formed a monophyletic clade in the cluster of Rhizobium species. Importantly, the feature gene of the genus Rhizobium, nifH gene (encoding the dinitrogenase reductase), was detected in strain FML-4T but not in C. thiooxidans KCTC 52231T. In addition, strain FML-4T contained the summed feature 8 (C18:1ω7c and/or C18:1ω6c), C19:0 cyclo ω8c and C16:0 as the major fatty acids. Genome sequencing of strain FML-4T revealed a genome size of 7.3 Mbp and a G+C content of 63.0 mol%. Based on the results obtained by phylogenetic and chemotaxonomic analyses, phenotypic characterization, average nucleotide identity (ANI, similarity 77.3-75.4%), and digital DNA-DNA hybridization (dDDH, similarity 24.5-22.3%), it was concluded that strain FML-4T represented a novel species of the genus Rhizobium, for which the name Rhizobium flavescens sp. nov. was proposed (type strain FML-4T = CCTCC AB 2019354T = KCTC 62839T).

DNA functionalized metal and metal oxide nanoparticles: principles and recent advances in food safety detection.

The frequent occurrence of food safety incidents has given rise to unprecedented concern about food contamination issues for both consumers and the industry. Various contaminations in food pose serious threats to food safety and human health. Many detection methods were studied to address the challenge. Recently, biosensors relying on deoxyribonucleic acid (DNA)-functionalized nanoparticles have been developed as an efficient and effective detection method. In the current review, the strategies for DNA assembly metal and metal oxide nanoparticles are elaborated, recent applications of the sensors based on DNA-functionalized nanoparticles in food contaminant detection are discussed. Pathogenic bacteria, heavy metal ions, mycotoxins, antibiotics, and pesticides are covered as food contaminants. Additionally, limitations and future trends of functionalized nanoparticles-based technology are also presented. The current review indicates that DNA-functionalized metal and metal oxide nanoparticles are a novel nanomaterial with unique biological and physical properties for developing electrochemical, fluorescent, colourimetric and surface-enhanced Raman spectroscopy (SERS) sensors, etc. Compared with conventional detection techniques, DNA-functionalized metal and metal oxide nanoparticles have considerable advantages with high accuracy, high specificity, micro-intelligence, and low cost. Nevertheless, the stability of these sensors and the limitations of real-time detection are still under discussion. Therefore, more tolerant, portable, and rapid DNA sensors should be developed to better the real-time monitoring of harmful contaminants.

Enantioselective Catabolism of the Two Enantiomers of the Phenoxyalkanoic Acid Herbicide Dichlorprop by Sphingopyxis sp. DBS4.

Dichlorprop [(RS)-2-(2,4-dichlorophenoxy)propanoic acid; DCPP], an important phenoxyalkanoic acid herbicide (PAAH), is extensively used in the form of racemic mixtures (Rac-DCPP), and the environmental fates of both DCPP enantiomers [(R)-DCPP and (S)-DCPP] mediated by microorganisms are of great concern. In this study, a bacterial strain Sphingopyxis sp. DBS4 was isolated from contaminated soil and was capable of utilizing both (R)-DCPP and (S)-DCPP as the sole carbon source for growth. Strain DBS4 preferentially catabolized (S)-DCPP as compared to (R)-DCPP. The optimal conditions for Rac-DCPP degradation by strain DBS4 were 30 °C and pH 7.0. In addition to Rac-DCPP, other PAAHs such as (RS)-2-(4-chloro-2-methylphenoxy)propanoic acid, 2,4-dichlorophenoxyacetic acid, 4-chloro-2-methylphenoxyacetic acid, and 2,4-dichlorophenoxyacetic acid butyl ester could also be catabolized by strain DBS4. Bioremediation of Rac-DCPP-contaminated soil by inoculation of strain DBS4 exhibited an effective removal of both (R)-DCPP and (S)-DCPP from the soil. Due to its broad substrate spectrum, strain DBS4 showed great potential in the bioremediation of PAAH-contaminated sites.

Mineralization of the herbicide swep by a two-strain consortium and characterization of a new amidase for hydrolyzing swep.

BACKGROUND: Swep is an excellent carbamate herbicide that kills weeds by interfering with metabolic processes and inhibiting cell division at the growth point. Due to the large amount of use, swep residues in soil and water not only cause environmental pollution but also accumulate through the food chain, ultimately pose a threat to human health. This herbicide is degraded in soil mainly by microbial activity, but no studies on the biotransformation of swep have been reported. RESULTS: In this study, a consortium consisting of two bacterial strains, Comamonas sp. SWP-3 and Alicycliphilus sp. PH-34, was enriched from a contaminated soil sample and shown to be capable of mineralizing swep. Swep was first transformed by Comamonas sp. SWP-3 to the intermediate 3,4-dichloroaniline (3,4-DCA), after which 3,4-DCA was mineralized by Alicycliphilus sp. PH-34. An amidase gene, designated as ppa, responsible for the transformation of swep into 3,4-DCA was cloned from strain SWP-3. The expressed Ppa protein efficiently hydrolyzed swep and a number of other structural analogues, such as propanil, chlorpropham and propham. Ppa shared less than 50% identity with previously reported arylamidases and displayed maximal activity at 30 °C and pH 8.6. Gly449 and Val266 were confirmed by sequential error prone PCR to be the key catalytic sites for Ppa in the conversion of swep. CONCLUSIONS: These results provide additional microbial resources for the potential remediation of swep-contaminated sites and add new insights into the catalytic mechanism of amidase in the hydrolysis of swep.

Sphingomonas flavalba sp. nov., isolated from a procymidone-contaminated soil.

A novel Gram-stain-negative, strictly aerobic, non-spore-forming, motile with one polar flagellum and short rod-shaped bacterium, designated strain ZLT-5T, was isolated from procymidone-contaminated soil sampled in Nanjing, Jiangsu, PR China. Growth occurred at 26-37 °C (optimum, 37 °C), at pH 6.0-9.0 (pH 7.0) and in the presence of 0-1.5 % NaCl (0.5 %). Phylogenetic analysis based on the 16S rRNA gene sequences showed that strain ZLT-5T belonged to the genus Sphingomonas, with the highest sequence similarity to Sphingomonas kyeonggiensis THG-DT81T (96.6 %), followed by Sphingomonas dokdonensis DSM 21029T (96.5 %) and Sphingomonas silvisoli RP18T (96.3 %). The G+C content of strain ZLT-5T was 68.0 mol% (draft genome sequence). The average nucleotide identity value of the draft genomes between strain ZLT-5T and S. kyeonggiensis THG-DT81T was 75.4 %. Strain ZLT-5T contained ubiquinone-10 as the predominant isoprenoid quinone and sym-homospermidine as the major polyamine. The major polar lipids consisted of diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol, phosphatidylcholine, phosphoaminolipid and sphingoglycolipid. The main cellular fatty acids (>10 % of the total fatty acids) of strain ZLT-5T were C17 : 1ω6c, summed feature 3 (C16 : 1ω7c and/or C15 : 0 ISO 2-OH) and summed feature 8 (C18 : 1ω7c and/or C18 : 1ω6c). Based on phylogenetic analysis and physiological and biochemical characterization, strain ZLT-5T is described as a novel species of the genus Sphingomonas, for which the name Sphingomonas flavalba sp. nov. is proposed. The type strain is ZLT-5T (=CCTCC AB 2018188T=KCTC 62840T).

Spirosoma sordidisoli sp. nov., a propanil-degrading bacterium isolated from a herbicide-contaminated soil.

A novel Gram-stain negative, aerobic, rod-shaped, asporogenous, propanil-degrading bacterial strain, TY50T, was isolated from a herbicide-contaminated soil in Nanjing, China. Strain TY50T was found to grow optimally at pH 9.0, 30 °C and in the absence of NaCl. The G + C content of the total DNA was determined to be 55.5 mol%. The 16S rRNA gene sequence of strain TY50T shows high identity to that of Spirosoma lacussanchae CPCC 100624T (99.3%), Spirosoma metallicum PR1014kT (94.8%) and Spirosoma soli MIMBbqt12T (94.6%). DNA-DNA hybridization indicated that the isolate had relatively low levels of DNA-DNA relatedness with S. lacussanchae CPCC 100624T (48.3%). Average nucleotide identity and the digital DNA-DNA hybridizations for draft genomes between strain TY50T and S. lacussanchae CPCC 100624T were 93.2% and 51.0%, respectively. The major cellular fatty acids of strain TY50T were identified as C16:1ω5c (24.5%) and summed feature 3 (C16:1ω6c/C16:1ω7c, 40.7%). MK-7 was found to be the predominant respiratory quinone. The major polar lipid profile includes phosphatidylethanolamine, an unidentified lipid and an unidentified aminolipid. These chemotaxonomic data support the affiliation of strain TY50T with the members of the genus Spirosoma. Strain TY50T can be distinguished from its close phylogenetic neighbours based on its phenotypic, genotypic, and chemotaxonomic features. Therefore, strain TY50T represents a novel member of the genus Spirosoma, for which the name Spirosoma sordidisoli sp. nov. is proposed. The type strain is TY50T (= KCTC 62494T = CCTCC AB 2018041T).

Rhizobium album sp. nov., isolated from a propanil-contaminated soil.

A novel Gram-stain negative, facultatively anaerobic, non-spore-forming, motile and rod-shaped bacterium (NS-104T) was isolated from a propanil-contaminated soil in Nanjing, China. Growth occurred at pH 5.0-9.0 (optimum 6.0), 16-37 °C (optimum 30 °C) and in the presence of 0-2.0% (w/v) NaCl (optimum, without NaCl). Strain NS-104T showed high 16S rRNA gene sequence identity to Rhizobium azooxidifex DSM 100211T (96.7%). The phylogenetic analysis of the 16S rRNA gene as well as the housekeeping genes recA, atpD and glnA demonstrated that strain NS-104T belongs to the genus Rhizobium. Strain NS-104T did not form nodules on six different legumes, and the nodD, nodC and nifH genes were neither amplified by PCR nor found in the draft genome of strain NS-104T. The sole respiratory quinone was ubiquinone Q-10. The polar lipid profile included the major amounts phosphatidylmonomethylethanolamine, phosphatidylglycerol and moderate amounts of phosphatidylethanolamine, phosphatidylcholine, diphosphatidylglycerol and unidentified aminolipids. The major cellular fatty acids were C18:1ω7c (39.6%), C19:0 cyclo ω8c (29.8%) and C16:0 (11.5%). The G + C content of strain NS-104T was 61.9 mol%. Strain NS-104T therefore represents a new species, for which the name Rhizobium album sp. nov. is proposed, with the type strain NS-104T (= KCTC 62327T = CCTCC AB 2017250T).

Characterization of an arylamidase from a newly isolated propanil-transforming strain of Ochrobactrum sp. PP-2.

Propanil, one of the most extensively used post-emergent contact herbicides, has also been reported to have adverse effect on environmental safety. A bacterial strain of Ochrobactrum sp. PP-2, which was capable of transforming propanil, was isolated from a propanil-contaminated soil collected from a chemical factory. An arylamidase gene mah responsible for transforming propanil to 3,4-dichloroaniline (3,4-DCA) was cloned from strain PP-2 by shotgun method and subsequently confirmed by function expression. The arylamidase Mah shares low amino acid sequence identity (27-50%) with other biochemically characterized amidases and shows less than 30% identities to other reported propanil hydrolytic enzymes. Mah was most active at pH 8 and 35 °C. Mah had a remarkable activity toward propanil (Km = 6.3 ±â€¯1.2 µM), showing the highest affinity efficiency for propanil as compared with other reported propanil hydrolytic enzymes. Our study also provides a new arylamidase for the hydrolysis of propanil.

Degradation of Phenylurea Herbicides by a Novel Bacterial Consortium Containing Synergistically Catabolic Species and Functionally Complementary Hydrolases.

Phenylurea herbicides (PHs) are frequently detected as major water contaminants in areas where there is extensive use. In this study, Diaphorobacter sp. strain LR2014-1, which initially hydrolyzes linuron to 3,4-dichloroanaline, and Achromobacter sp. strain ANB-1, which further mineralizes the produced aniline derivatives, were isolated from a linuron-mineralizing consortium despite being present at low abundance in the community. The synergistic catabolism of linuron by the consortium containing these two strains resulted in more efficient catabolism of linuron and growth of both strains. Strain LR2014-1 harbors two evolutionary divergent hydrolases from the amidohydrolase superfamily Phh and the amidase superfamily TccA2, which functioned complementarily in the hydrolysis of various types of PHs, including linuron ( N-methoxy- N-methyl-substituted), diuron, chlorotoluron, fluomethuron ( N, N-dimethyl-substituted), and siduron. These findings show that a bacterial consortium can contain catabolically synergistic species for PH mineralization, and one strain could harbor functionally complementary hydrolases for a broadened substrate range.

Xinfangfangia soli gen. nov., sp. nov., isolated from a diuron-polluted soil.

A novel Gram-staining-negative, non-motile and rod-shaped bacterial strain ZQBWT, which was isolated from a diuron-polluted soil collected near Nanjing, PR China, was investigated for its taxonomic position by a polyphasic approach. ZQBWT grew well at pH 6.0-12.0 (optimum, pH 7.0), 26-35 °C (optimum, 30 °C) and up to 0.5 % NaCl (optimally the absence of NaCl) in R2A broth. The major fatty acids of ZQBWT were C18 : 1ω7c (82.7 %) and C18 : 0 (5.3 %). The polar lipid profile included the major compounds phophatidylcholine, phosphatidylglycerol and phosphatidylmethylethanolamine. The only respiratory quinone was ubiquinone Q-10. The G+C content of genomic DNA was 67.0 mol%. Comparisons with 16S rRNA gene sequences revealed that ZQBWT has the highest sequence similarities with members of the genus Tabrizicola (≤95.97 %), followed by Rhodobacter(≤95.96 %) and Falsirhodobacter(95.95 %) which all belong to the family Rhodobacteraceaein the phylum Proteobacteria. Photosynthesis genes pufLM were not found and photosynthesis pigments were not formed in ZQBWT. On the basis of the results from chemotaxonomic, phenotypic and phylogenetic analysis, ZQBWT represents a novel species of a novel genus, for which the name Xinfangfangia soli gen. nov., sp. nov. is proposed. The type strain is ZQBWT (=KCTC 62102T=CCTCC AB 2017177T).