Potential of Variovorax paradoxus isolate BFB1_13 for bioremediation of BTEX contaminated sites

Here, we report and discuss the applicability of Variovorax paradoxus strain BFB1_13 in the bioremediation of BTEX contaminated sites. Strain BFB1_13 was capable of degrading all the six BTEX-compounds under both aerobic (O2 conc. 8 mg l−1) and micro-aerobic/oxygen-limited (O2 conc. 0.5 mg l−1) conditions using either individual (8 mg‧l−1) or a mixture of compounds (~ 1.3 mg‧l−1 of each BTEX compound). The BTEX biodegradation capability of SBP-encapsulated cultures (SBP—Small Bioreactor Platform) was also assessed. The fastest degradation rate was observed in the case of aerobic benzene biodegradation (8 mg l−1 per 90 h). Complete biodegradation of other BTEX occurred after at least 168 h of incubation, irrespective of the oxygenation and encapsulation. No statistically significant difference was observed between aerobic and microaerobic BTEX biodegradation. Genes involved in BTEX biodegradation were annotated and degradation pathways were predicted based on whole-genome shotgun sequencing and metabolic analysis. We conclude that V. paradoxus strain BFB1_13 could be used for the development of reactive biobarriers for the containment and in situ decontamination of BTEX contaminated groundwater plumes. Our results suggest that V. paradoxus strain BFB1_13—alone or in co-culture with other BTEX degrading bacterial isolates—can be a new and efficient commercial bioremediation agent for BTEX contaminated sites.

Hybrid Assembly of the Quorum-Quenching Isolate Variovorax paradoxus VAI-C Genome Sequence

ABSTRACT Variovorax paradoxus VAI-C was isolated due to its ability to utilize acyl-homoserine lactones (AHLs) as the sole source of carbon, energy, and nitrogen. Here, we present a hybrid assembly of the V. paradoxus VAI-C genome sequence, consisting of a primary chromosome, a secondary chromid, and a plasmid.

Conservation of isolate substrate preferences in mixed communities revealed through ribosomal marker protein profiling

Assessment of structure-function relationships is a central theme in microbial ecology. However, the degree that isolate metabolic activities are conserved in communities remains unclear. This is because tracking population dynamics and substrate partitioning in microbial communities remains technically challenging. Here, we describe the application of a mass spectrometry-based ribosomal marker protein profiling with stable isotope probing approach that allows for concurrent monitoring of community structure dynamics and resource assimilation within a five-member synthetic soil bacterial community. Using this approach, we find that isolate substrate preferences for glutamine and phenylalanine are largely conserved in the community and can be predicted using a weighted-sum model. However, time-series monitoring revealed a significant delay in phenylalanine incorporation by two of the strains, as well as enhanced growth for Variovorax paradoxus presumably due to interspecies interactions. The unique utility of this approach to temporally probe resource incorporation and community structure enables deciphering the dynamic interactions occurring within the community. Extension of this approach to other communities under various environmental perturbations is needed to reveal the generality of microbial conservation of substrate preferences.

Structure and function of rhizosphere and root endophyte microbial communities associated with healthy and root rot diseased Panax notoginseng

Background: Panax notoginseng (Burkill) F. H. Chen is a Chinese medicinal plant of the Araliaceae family commonly used in the treatment of cardiovascular and cerebrovascular diseases in Asia and elsewhere. To meet an increase in Chinese herbal medicine market demand, most P. notoginseng is planted artificially, and is vulnerable to various plant diseases. Root rot disease, in particular, causes substantial P. notoginseng yield reduction and economic losses. High-depth next-generation sequencing technology was used to analyze the rhizosphere and root endophyte microbial communities of P. notoginseng to compare the characteristics of these two communities between healthy and root rot diseased P. notoginseng plants, and to clarify the relationship between these microbial communities and root rot disease.Results: The P. notoginseng rhizosphere microbial community was more diverse than the root endophyte community, and the difference in functional pathways between healthy and diseased P. notoginseng plants was greater in the root endophyte than in the rhizosphere communities. Multi-database annotation results showed that the highest number of endophytic bacteria occurred in the roots of diseased plants. The number of carbohydrate-active enzymes database families was also higher in diseased roots. The RND antibiotic efflux function was higher in the healthy samples. A high abundance of Variovorax paradoxus and Pseudomonas fluorescens occurred in the healthy and diseased root endophyte communities, respectively. Ilyonectria mors-panacis and Pseudopyrenochaeta lycopersici were most abundant in the diseased samples. In addition, the complete genome of two unknown Flavobacteriaceae species and one unknown Bacteroides species were obtained based on binning analysis.Conclusions: The rhizosphere and root endophyte microbial communities of healthy and root rot diseased P. notoginseng showed marked differences in diversity and functional pathways. The higher mapping values obtained for the diseased samples reflected the occurrence of root rot disease at the molecular level. Variovorax paradoxus and Pseudomonas fluorescens may be antagonistic bacteria of root rot in P. notoginseng, whereas Ilyonectria mors-panacis and Pseudopyrenochaeta lycopersici appear to be P. notoginseng root rot pathogens. Our study provides a theoretical basis for understanding the occurrence of root rot in P. notoginseng and for further research on potential biological control agents.

3,3′-Thiodipropionic acid (TDP), a possible precursor for the synthesis of polythioesters: identification of TDP transport proteins in Variovorax paradoxus TBEA6

Abstract3,3′-Thiodipropionic acid (TDP) is an antioxidant, which can be used as precursor carbon source to synthesize polythioesters. The bacterium Variovorax paradoxus TBEA6 strain can use TDP as a single source of carbon and energy. In the present study, experiments were carried out to identify proteins involved in the transport of TDP into the cells of strain TBEA6. Hence, eight putative tctC genes, which encode for the TctC proteins, were amplified from genomic DNA of TBEA6 strain using polymerase chain reaction and expressed in E. coli BL21 cells. Cells were grown in auto-induction medium, and protein purification was done using His Spin Trap affinity columns. Purity and molecular weight of each protein were confirmed by SDS-PAGE analysis. Protein-ligand interactions were monitored in thermoshift assays using the real-time PCR system. Two TctC proteins (locus tags VPARA-44430 and VPARA-01760) out of eight proteins showed a significant shift in their melting temperatures when they interact with the ligand (TDP or gluconate). The responsible genes were deleted in the genome of TBEA6 using suicide plasmid pJQ200mp18Tc, and single deletion mutants of the two candidate genes were subsequently generated. Finally, growth of the wild-type strain (TBEA6) and the two mutant strains (ΔVPARA-44430 and ΔVPARA-01760) were monitored and compared using TDP or gluconate as carbon sources. Wild type strains were successfully grown with TDP or gluconate. From the two mutant strains, one (ΔVPARA-44430) was unable to grow with TDP indicating that the tctC gene with locus tag VPARA-44430 is involved in the uptake of TDP.Key Points• Putative tctC genes from V. paradoxus TBEA6 were heterologously expressed in E. coli.• Protein-ligand interactions monitored in thermoshift assays using the real-time PCR.• tctC gene with locus tag VPARA-44430 is involved in the uptake of TDP.

Microalgal–bacterial consortia unveil distinct physiological changes to facilitate growth of microalgae

Physiological changes that drive the microalgal–bacterial consortia are poorly understood so far. In the present novel study, we initially assessed five morphologically distinct microalgae for their ability in establishing consortia in Bold's basal medium with a bacterial strain, Variovorax paradoxus IS1, all isolated from wastewaters. Tetradesmus obliquus IS2 and Coelastrella sp. IS3 were further selected for gaining insights into physiological changes including those of metabolomes in consortia involving V. paradoxus IS1. The distinct parameters investigated were pigments (chlorophyll a, b, and carotenoids), reactive oxygen species (ROS), lipids, and metabolites that are implicated in major metabolic pathways. There was a significant increase (>1.2-fold) in pigments, viz., chlorophyll a, b and carotenoids, decrease in ROS, and enhanced lipid yield (>2-fold) in consortia than in individual cultures. In addition, the differential regulation of cellular metabolites such as sugars, amino acids, organic acids, and phytohormones was distinct among the two microalgal–bacterial consortia. Our results thus indicate that the selected microalgal strains, T. obliquus IS2 and Coelastrella sp. IS3, developed efficient consortia with V. paradoxus IS1 by effecting the required physiological changes including metabolomics. Such microalgal–bacterial consortia could largely be used in wastewater treatment and for production of value-added metabolites.

Transcriptome profiling of Variovorax paradoxus EPS under different growth conditions reveals regulatory and structural novelty in biofilm formation

We used transcriptome analysis by paired-end strand-specific RNA-seq to evaluate the specific changes in gene expression associated with the transition to static biofilm growth in the rhizosphere plant growth-promoting bacterium Variovorax paradoxus EPS. Triplicate biological samples of exponential growth, stationary phase and static biofilm samples were examined. DESeq2 and Rockhopper were used to identify robust and widespread shifts in gene expression specific to each growth phase. We identified 1711 protein-coding genes (28%) using DESeq2 that had altered expression greater than twofold specifically in biofilms compared to exponential growth. Fewer genes were specifically differentially expressed in stationary-phase culture (757, 12%). A small set of genes (103/6020) were differentially expressed in opposing fashions in biofilm and stationary phase, indicating potentially substantial shifts in phenotype. Gene-ontology analysis showed that the only class of genes specifically upregulated in biofilms was associated with nutrient transport, highlighting the importance of nutrient uptake in the biofilm. The biofilm-specific genes did not overlap substantially with the loci identified by mutagenesis studies, although some were present in both sets. The most highly upregulated biofilm-specific gene is predicted to be a part of the RNA degradosome, which indicates that RNA stability is used to regulate the biofilm phenotype. Two small putative proteins, Varpa_0407 and Varpa_3832, are highly expressed specifically in biofilms and are predicted to be secreted DNA-binding proteins, which may stabilize extracellular DNA as a component of the biofilm matrix. An flp/tad type-IV pilus locus (Varpa_5148–60) is strongly downregulated specifically in biofilms, in contrast with results from other systems for these pili. Mutagenesis confirms that this locus is important in surface motility rather than biofilm formation. These experimental results suggest that V. paradoxus EPS biofilms have substantial regulatory and structural novelty.