Assimilation Genes Research Articles

Transcriptional Downregulation of Methanol Metabolism Key Genes During Yeast Death in Engineered Pichia pastoris.

Pichia pastoris possesses the unique ability to utilize methanol as its sole carbon source, which makes it a proper host for producing various high-value-added products via metabolic engineering. Nevertheless, cell death has been observed during the fermentation of modified P. pastoris, with limited literature elucidating the underlying causes and mechanisms. Understanding the death mechanisms during methanol-based fermentation is crucial for optimizing fermentation strategies, enhancing the accumulation of target products, and reducing production costs. Here, we first sought to eliminate the potential causes of cell death during fermentation, such as inadequate inorganic salts and toxic by-product accumulation. The elimination of these potential causes was achieved efficiently utilizing the high-throughput fermentation equipment. Subsequently, we established a correlation between yeast cell death and the duration of the methanol metabolism period by monitoring the growth of the yeast at different fermentation stages. A critical revelation from this work came from analyzing the yeast's transcriptomic data at various stages of methanol metabolism. It was observed that a significant characteristic of yeast cell death during fermentation was the marked down-regulation of transcript levels of key enzymes involved in the methanol assimilation pathway and genes related to their biosynthesis process. The findings of this work are crucial for better understanding the causes and mechanisms of cell death for engineered P. pastoris during methanol-utilized fermentation.

Synthesis and properties of the natural ultraviolet absorber gadusol in Komagataella phaffii

Gadusol, an efficient natural ultraviolet (UV) absorbing substance with antioxidant capacity, is ubiquitous in aquatic organisms such as microorganisms, algae, and fish eggs. In order to address issues such as its low natural extraction yield and environmental unfriendliness, we introduced the gadusol synthesis pathway from zebrafish into Komagataella phaffii and successfully constructed the recombinant strain capable of synthesizing gadusol. The xylose assimilation genes derived from Scheffersomyces stipitis were further introduced into the recombinant strain to increase the content of the key substrate sedoheptulose 7-phosphate (S7P). The results showed that the utilization of xylose was an effective strategy to increase the yield of gadusol. In the medium with only xylose as the substrate, the yield of gadusol reached 141.8 mg/L (32.3 mg/g dry cell weight, DCW), which was about 46 times of that in the medium with only glucose as the substrate. The product showed obvious absorption in the range of 275-305 nm, with the maximum absorption at 290 nm. Moreover, the product demonstrated antioxidant capacity. After reaction for 5 h, the ferric ion reducing antioxidant power (FRAP), 2, 2'-azino- bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS), and 1, 1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging rates changed by 90.83%, 25.50%, and 131.80%, respectively, which suggested that the product may be used as a long-acting antioxidant. This study demonstrated the great potential of K. phaffii as a chassis for the biosynthesis of natural UV absorbers. The biosynthesized gadusol has good UV absorption and antioxidant properties, which provides a theoretical basis for the industrial production and application of gadusol.

Conversion of monocropping to intercropping promotes rhizosphere microbiome functionality and soil nitrogen cycling

Intercropping can increase soil nutrient availability and provide greater crop yields for intensive agroecosystems. Despite its multiple benefits, how intercropping influences rhizosphere microbiome assemblages, functionality, and complex soil nitrogen cycling is not fully understood. Here, a three-year field experiment was carried out on different cropping system with five fertilization treatments at the main soybean production regions. We found that soybean yields in intercropped systems were on average 17 % greater than in monocropping system, regardless of fertilization treatments. We also found that intercropping systems significant increased network modularity (by 46 %) and functional diversity (by 11 %) than monocropping systems. Metagenomics analyses further indicated intercropping promotes microbiome functional adaptation, particularly enriching core functions related to nitrogen metabolism. Cropping patterns had a stronger influence on the functional genes associated with soil nitrogen cycling (R2 = 0.499). Monocropping systems increased the abundance of functional genes related to organic nitrogen ammonification, nitrogen fixation, and denitrification, while functional guilds of nitrate assimilation (by 28 %), nitrification (by 31 %), and dissimilatory nitrate reduction (by 10.1 %) genes were enriched in intercropping systems. Furthermore, we found that abiotic factors (i.e. AP, pH, and Moisture) are important drivers in shaping soil microbial community assemblage and nitrogen cycling. The functional genes include hzsB, and nrfA, and nxrA that affected by these biotic and abiotic variables were strongly related to crop yield (R2 = 0.076 ~ R2 = 0.249), suggesting a key role for maintaining crop production. We demonstrated that land use conversion from maize monocropping to maize-soybean intercropping diversify rhizosphere microbiome and functionality signatures, and intercropping increased key gene abundance related to soil nitrogen cycling to maintain the advantage of crop yield. The results of this study significantly facilitate our understanding of the complex soil nitrogen cycling processes and lay the foundation for manipulating desired specific functional taxa for improved crop productivity under sustainable intensification.

Arabidopsis WRKY1 promotes monocarpic senescence by integrative regulation of flowering, leaf senescence, and nitrogen remobilization

Monocarpic senescence, characterized by whole-plant senescence following a single flowering phase, is widespread in seed plants, particularly in crops, determining seed harvest time and quality. However, how external and internal signals are systemically integrated into monocarpic senescence remains largely unknown. Here, we report that the Arabidopsis thaliana transcription factor WRKY1 plays essential roles in multiple key steps of monocarpic senescence. WRKY1 expression is induced by age, salicylic acid (SA), and nitrogen (N) deficiency. Flowering and leaf senescence are accelerated in the WRKY1 overexpression lines but are delayed in the wrky1 mutants. The combined DNA affinity purification sequencing and RNA sequencing analyses uncover the direct target genes of WRKY1. Further studies show that WRKY1 coordinately regulates three processes in monocarpic senescence: (1) suppressing FLOWERING LOCUS C gene expression to initiate flowering, (2) inducing SA biosynthesis genes to promote leaf senescence, and (3) activating the N assimilation and transport genes to trigger N remobilization. In summary, our study reveals how one stress-responsive transcription factor, WRKY1, integrates flowering, leaf senescence, and N remobilization processes into monocarpic senescence, providing important insights into plant lifetime regulation.

Genomic organization and expression profiles of nitrogen assimilation genes in Glycine max

Background Glutamine synthetase (GS), glutamate synthase (GOGAT), and nitrate reductase (NR) are key enzymes involved in nitrogen assimilation and metabolism in plants. However, the systematic analysis of these gene families lacked reports in soybean (Glycine max (L.) Merr.), one of the most important crops worldwide. Methods In this study, we performed genome-wide identification and characterization of GS, GOGAT, and NR genes in soybean under abiotic and nitrogen stress conditions. Results We identified a total of 10 GS genes, six GOGAT genes, and four NR genes in the soybean genome. Phylogenetic analysis revealed the presence of multiple isoforms for each gene family, indicating their functional diversification. The distribution of these genes on soybean chromosomes was uneven, with segmental duplication events contributing to their expansion. Within the nitrogen assimilation genes (NAGs) group, there was uniformity in the exon-intron structure and the presence of conserved motifs in NAGs. Furthermore, analysis of cis-elements in NAG promoters indicated complex regulation of their expression. RT-qPCR analysis of seven soybean NAGs under various abiotic stresses, including nitrogen deficiency, drought-nitrogen, and salinity, revealed distinct regulatory patterns. Most NAGs exhibited up-regulation under nitrogen stress, while diverse expression patterns were observed under salt and drought-nitrogen stress, indicating their crucial role in nitrogen assimilation and abiotic stress tolerance. These findings offer valuable insights into the genomic organization and expression profiles of GS, GOGAT, and NR genes in soybean under nitrogen and abiotic stress conditions. The results have potential applications in the development of stress-resistant soybean varieties through genetic engineering and breeding.

Nitrate reduction to ammonium: a phylogenetic, physiological, and genetic aspects in Prokaryotes and eukaryotes.

The microbe-mediated conversion of nitrate (NO3-) to ammonium (NH4+) in the nitrogen cycle has strong implications for soil health and crop productivity. The role of prokaryotes, eukaryotes and their phylogeny, physiology, and genetic regulations are essential for understanding the ecological significance of this empirical process. Several prokaryotes (bacteria and archaea), and a few eukaryotes (fungi and algae) are reported as NO3- reducers under certain conditions. This process involves enzymatic reactions which has been catalysed by nitrate reductases, nitrite reductases, and NH4+-assimilating enzymes. Earlier reports emphasised that single-cell prokaryotic or eukaryotic organisms are responsible for this process, which portrayed a prominent gap. Therefore, this study revisits the similarities and uniqueness of mechanism behind NO3- -reduction to NH4+ in both prokaryotes and eukaryotes. Moreover, phylogenetic, physiological, and genetic regulation also shed light on the evolutionary connections between two systems which could help us to better explain the NO3--reduction mechanisms over time. Reports also revealed that certain transcription factors like NtrC/NtrB and Nit2 have shown a major role in coordinating the expression of NO3- assimilation genes in response to NO3- availability. Overall, this review provides a comprehensive information about the complex fermentative and respiratory dissimilatory nitrate reduction to ammonium (DNRA) processes. Uncovering the complexity of this process across various organisms may further give insight into sustainable nitrogen management practices and might contribute to addressing global environmental challenges.

Phosphorus limitation intensifies heat-stress effects on the potential mutualistic capacity in the coral-derived Symbiodinium

Coral reef ecosystems have been severely ravaged by global warming and eutrophication. Eutrophication often originates from nitrogen (N) overloading that creates stoichiometric phosphorus (P) limitation, which can be aggravated by sea surface temperature rises that enhances stratification. However, how P-limitation interacts with thermal stress to impact coral-Symbiodiniaceae mutualism is poorly understood and underexplored. Here, we investigated the effect of P-limitation (P-depleted vs. P-replete) superimposed on heat stress (31 °C vs. 25 °C) on a Symbiodinium strain newly isolated from the coral host by a 14-day incubation experiment. The heat and P-limitation co-stress induced an increase in alkaline phosphatase activity and reppressed cell division, photosynthetic efficiency, and expression of N uptake and assimilation genes. Moreover, P limitation intensified downregulation of carbon fixation (light and dark reaction) and metabolism (glycolysis) pathways in heat stressed Symbiodinium. Notably, co-stress elicited a marked transcriptional downregulation of genes encoding photosynthates transporters and microbe-associated molecular patterns, potentially undermining the mutualism potential. This work sheds light on the interactive effects of P-limitation and heat stress on coral symbionts, indicating that nutrient imbalance in the coral reef ecosystem can intensify heat-stress effects on the mutualistic capacity of Symbiodiniaceae.

Morphological, Metabolomic and Genomic Evidences on Drought Stress Protective Functioning of the Endophyte Bacillus safensis Ni7.

The metabolomic and genomic characterization of an endophytic Bacillus safensis Ni7 was carried out in this study. This strain has previously been isolated from the xerophytic plant Nerium indicum L. and reported to enhance the drought tolerance in Capsicum annuum L. seedlings. The effects of drought stress on the morphology, biofilm production, and metabolite production of B. safensis Ni7 are analyzed in the current study. From the results obtained, the organism was found to have multiple strategies such as aggregation and clumping, robust biofilm production, and increased production of surfactin homologues under the drought induced condition when compared to non-stressed condition. Further the whole genome sequencing (WGS) based analysis has demonstrated B. safensis Ni7 to have a genome size of 3,671,999bp, N50 value of 3,527,239, and a mean G+C content of 41.58%. Interestingly the organism was observed to have the presence of various stress-responsive genes (13, 20U, 16U,160, 39, 17M, 18, 26, and ctc) and genes responsible for surfactin production (srfAA, srfAB, srfAC, and srfAD), biofilm production (epsD, epsE, epsF, epsG, epsH, epsI, epsK, epsL, epsM, epsN, and pel), chemotaxis (cheB_1, cheB_2, cheB_3, cheW_1, cheW_2 cheR, cheD, cheC, cheA, cheY, cheV, and cheB_4), flagella synthesis (flgG_1, flgG_2, flgG_3, flgC, and flgB) as supportive to the drought tolerance. Besides these, the genes responsible for plant growth promotion (PGP), including the genes for nitrogen (nasA, nasB, nasC, nasD, and nasE) and sulfur assimilation (cysL_1&L_2, cysI) and genes for phosphate solubilization (phoA, phoP_1& phoP_2, and phoR) could also be predicted. Along with the same, the genes for catalase, superoxide dismutase, protein homeostasis, cellular fitness, osmoprotectants production, and protein folding could also be predicted from its WGS data. Further pan-genome analysis with plant associated B. safensis strains available in the public databases revealed B. safensis Ni7 to have the presence of a total of 5391 gene clusters. Among these, 3207 genes were identified as core genes, 954 as shell genes and 1230 as cloud genes. This variation in gene content could be taken as an indication of evolution of strains of Bacillus safensis as per specific conditions and hence in the case of B. safensis Ni7 its role in habitat adaptation of plant is well expected. This diversity in endophytic bacterial genes may attribute its role to support the plant system to cope up with stress conditions. Overall, the study provides genomic evidence on Bacillus safensis Ni7 as a stress alleviating microbial partner in plants.

Orphan response regulator NnaR is critical for nitrate and nitrite assimilation in Mycobacterium abscessus.

Mycobacterium abscessus (Mab) is an opportunistic pathogen afflicting individuals with underlying lung disease such as Cystic Fibrosis (CF) or immunodeficiencies. Current treatment strategies for Mab infections are limited by its inherent antibiotic resistance and limited drug access to Mab in its in vivo niches resulting in poor cure rates of 30-50%. Mab's ability to survive within macrophages, granulomas and the mucus laden airways of the CF lung requires adaptation via transcriptional remodeling to counteract stresses like hypoxia, increased levels of nitrate, nitrite, and reactive nitrogen intermediates. Mycobacterium tuberculosis (Mtb) is known to coordinate hypoxic adaptation via induction of respiratory nitrate assimilation through the nitrate reductase narGHJI. Mab, on the other hand, does not encode a respiratory nitrate reductase. In addition, our recent study of the transcriptional responses of Mab to hypoxia revealed marked down-regulation of a locus containing putative nitrate assimilation genes, including the orphan response regulator nnaR (nitrate/nitrite assimilation regulator). These putative nitrate assimilation genes, narK3 (nitrate/nitrite transporter), nirBD (nitrite reductase), nnaR, and sirB (ferrochelatase) are arranged contiguously while nasN (assimilatory nitrate reductase identified in this work) is encoded in a different locus. Absence of a respiratory nitrate reductase in Mab and down-regulation of nitrogen metabolism genes in hypoxia suggest interplay between hypoxia adaptation and nitrate assimilation are distinct from what was previously documented in Mtb. The mechanisms used by Mab to fine-tune the transcriptional regulation of nitrogen metabolism in the context of stresses e.g. hypoxia, particularly the role of NnaR, remain poorly understood. To evaluate the role of NnaR in nitrate metabolism we constructed a Mab nnaR knockout strain (MabΔnnaR ) and complement (MabΔnnaR+C ) to investigate transcriptional regulation and phenotypes. qRT-PCR revealed NnaR is necessary for regulating nitrate and nitrite reductases along with a putative nitrate transporter. Loss of NnaR compromised the ability of Mab to assimilate nitrate or nitrite as sole nitrogen sources highlighting its necessity. This work provides the first insights into the role of Mab NnaR setting a foundation for future work investigating NnaR's contribution to pathogenesis.

Energy deprivation affects nitrogen assimilation and fatty acid biosynthesis leading to leaf chlorosis under waterlogging stress in the endangered Abies koreana.

Energy deprivation triggers various physiological, biochemical and molecular changes in plants under abiotic stress. We investigated the oxidative damages in the high altitude grown conifer Korean fir (Abies koreana) exposed to waterlogging stress. Our experimental results showed that waterlogging stress led to leaf chlorosis, 35days after treatment. A significant decrease in leaf fresh weight, chlorophyll and sugar content supported this phenotypic change. Biochemical analysis showed a significant increase in leaf proline, lipid peroxidase and 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical content of waterlogged plants. To elucidate the molecular mechanisms, we conducted RNA-sequencing (RNA-seq) and de novo assembly. Using RNA-seq analysis approach and filtering (P<0.05 and false discovery rate<0.001), we obtained 134 unigenes upregulated and 574 unigenes downregulated. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis placed the obtained differentially expressed unigenes in α-linoleic pathway, fatty acid degradation, glycosis, glycolipid metabolism and oligosaccharide biosynthesis process. Mapping of unigenes with Arabidopsis using basic local alignment search tool for nucleotides showed several critical genes in photosynthesis and carbon metabolism downregulated. Following this, we found the repression of multiple nitrogen (N) assimilation and nucleotide biosynthesis genes including purine metabolism. In addition, waterlogging stress reduced the levels of polyunsaturated fatty acids with a concomitant increase only in myristic acid. Together, our results indicate that the prolonged snowmelt may cause inability of A. koreana seedlings to lead the photosynthesis normally due to the lack of root intercellular oxygen and emphasizes a detrimental effect on the N metabolic pathway, compromising this endangered tree's ability to be fully functional under waterlogging stress.

Efficient nitrogen removal by microalgal-bacterial granular sludge-marimo coupling process

Current biological wastewater treatment processes usually have a drawback of insufficient nitrogen (N) removal, contributing to the ubiquitous eutrophication of aquatic ecosystems globally. To address such a challenging situation, this study explored an innovative microalgal-bacterial granular sludge-marimo (MBGS-MA) coupling process. The process removed 83.4 % of N with the effluent N concentration of 4.0 mg/L. With the growth of MBGS, there was a shift towards genes associated with nitrification and denitrification, and away from ammonia assimilation genes, revealing internal mechanism of the shift of N removal pathway. Contrarily, MA could use gaseous N2 with the N fixing genes in MA enriched, and the genes abundance related to assimilatory nitrate reduction were also raised under the mutualistic interactions between Proteobacteria and Cyanobacteria, which was beneficial to achieve efficient N removal. These findings may open a new horizon for developing innovative hybrid microalgal-bacterial processes aimed at high-efficiency N removal from wastewater.

Enhancing nitrogen removal through macrophyte harvest and installation of woodchips-based floating beds in surface-flow constructed wetlands

Wetland management maintains nitrogen (N) removal capacity in mature and overgrown constructed wetlands (CWs). We evaluated whether CW management by macrophyte harvesting, and subsequent installation of woodchips-based floating beds (WFBs) planted with Glyceria maxima and Filipendula ulmaria improved N removal. In sixteen heavily overgrown experimental CWs, we applied four treatments: i) only macrophyte harvesting, ii) 5% of the harvested-CW surface covered with WFBs, iii) 20% WFBs cover, and iv) a control treatment (heavily overgrown). N removal was determined in all wetlands at nine occasions. Plant biomass accrual, N assimilation, and denitrification genes nirS, nirK, nosZI and nosZII on plant roots and woodchips from WFBs were estimated. Macrophyte harvesting improved N removal of heavily overgrown CWs, whereas subsequent WFB installation only sometimes improved N removal. Mean N removal efficiencies (± standard deviation) overall were 41 ± 15 %, 45 ± 20 %, 46 ± 16 % and 27 ± 8.3 % for treatments i to iv, respectively. Relative biomass production, root length and root surface area for G.maxima (mean ± standard deviation: 234 ± 114 %, 40 ± 6.5 cm, 6308 ± 1059 cm2g-1, respectively) were higher than those for F. ulmaria (63 ± 86 %, 28 ± 12 cm, 3131 ± 535 cm2g-1, respectively) whereas biomass N assimilation was higher for F. ulmaria (1.8 ± 0.9 gNm−2 of WFB) than for G. maxima (1.3 ± 0.5 gNm−2 of WFB). Denitrification gene abundance was higher on plant roots than on woodchips while G. maxima hosted higher root denitrification gene abundance than F. ulmaria. We conclude that macrophyte harvesting improves N removal in heavily overgrown CWs. WFBs installation has the potential to support plant growth and denitrification in surface-flow constructed wetlands. Further studies need to evaluate the long-term effects of macrophyte harvesting and WFB installation on N removal in CWs.

Impacts of AlaAT3 transgenic poplar on rhizosphere soil chemical properties, enzyme activity, bacterial community, and metabolites under two nitrogen conditions

ABSTRACT Poplar stands as one of the primary afforestation trees globally. We successfully generated transgenic poplar trees characterized by enhanced biomass under identical nutrient conditions, through the overexpression of the pivotal nitrogen assimilation gene, pxAlaAT3. An environmental risk assessment was conducted for investigate the potential changes in rhizosphere soil associated with these overexpressing lines (OL). The results show that acid phosphatase activity was significantly altered under ammonium in OL compared to the wild-type control (WT), and a similar difference was observed for protease under nitrate. 16SrDNA sequencing indicated no significant divergence in rhizosphere soil microbial community diversity between WT and OL. Metabolomics analysis revealed that the OL caused minimal alterations in the metabolites of the rhizosphere soil, posing no potential harm to the environment. With these findings in mind, we anticipate that overexpressed plants will not adversely impact the surrounding soil environment.

Genetic and Functional Diversity Help Explain Pathogenic, Weakly Pathogenic, and Commensal Lifestyles in the Genus Xanthomonas.

The genus Xanthomonas has been primarily studied for pathogenic interactions with plants. However, besides host and tissue-specific pathogenic strains, this genus also comprises nonpathogenic strains isolated from a broad range of hosts, sometimes in association with pathogenic strains, and other environments, including rainwater. Based on their incapacity or limited capacity to cause symptoms on the host of isolation, nonpathogenic xanthomonads can be further characterized as commensal and weakly pathogenic. This study aimed to understand the diversity and evolution of nonpathogenic xanthomonads compared to their pathogenic counterparts based on their cooccurrence and phylogenetic relationship and to identify genomic traits that form the basis of a life history framework that groups xanthomonads by ecological strategies. We sequenced genomes of 83 strains spanning the genus phylogeny and identified eight novel species, indicating unexplored diversity. While some nonpathogenic species have experienced a recent loss of a type III secretion system, specifically the hrp2 cluster, we observed an apparent lack of association of the hrp2 cluster with lifestyles of diverse species. We performed association analysis on a large data set of 337 Xanthomonas strains to explain how xanthomonads may have established association with the plants across the continuum of lifestyles from commensals to weak pathogens to pathogens. Presence of distinct transcriptional regulators, distinct nutrient utilization and assimilation genes, transcriptional regulators, and chemotaxis genes may explain lifestyle-specific adaptations of xanthomonads.

Variability in the phytoplankton response to upwelling across an iron limitation mosaic within the California current system

AbstractCoastal upwelling currents such as the California Current System (CCS) comprise some of the most productive biological systems on the planet. Diatoms dominate these upwelling events in part due to their rapid response to nutrient entrainment. In this region, they may also be limited by the micronutrient iron (Fe), an important trace element primarily involved in photosynthesis and nitrogen assimilation. The mechanisms behind how diatoms physiologically acclimate to the different stages of the upwelling conveyor belt cycle remain largely uncharacterized. Here, we explore their physiological and metatranscriptomic response to the upwelling cycle with respect to the Fe limitation mosaic that exists in the CCS. Subsurface, natural plankton assemblages that would potentially seed surface blooms were examined over wide and narrow shelf regions. The initial biomass and physiological state of the phytoplankton community had a large impact on the overall response to simulated upwelling. Following on‐deck incubations under varying Fe physiological states, our results suggest that diatoms quickly dominated the blooms by “frontloading” nitrogen assimilation genes prior to upwelling. However, diatoms subjected to induced Fe limitation exhibited reductions in carbon and nitrogen uptake and decreasing biomass accumulation. Simultaneously, they exhibited a distinct gene expression response which included increased expression of Fe‐starvation induced proteins and decreased expression of nitrogen assimilation and photosynthesis genes. These findings may have significant implications for upwelling events in future oceans, where changes in ocean conditions are projected to amplify the gradient of Fe limitation in coastal upwelling regions.

Investigation on comparative transcriptome profiling of resistant and susceptible non-CMS maize genotypes during Bipolaris maydis race O infection

Maydis leaf blight is a significant disease of maize caused by Bipolaris maydis race T, O and C. Molecular mechanisms regulating defense responses in non-CMS maize towards race O fungus are not fully known. In the present investigation, comparative transcriptome profiling was conducted on a highly resistant maize genotype SC-7-2-1-2-6-1 against a standard susceptible variety CM 119 at 48 h post inoculation (h PI) along with non-infected control. mRNA sequencing generated 38.4 Gb data, where 9349602 reads were mapped uniquely in SC-7, whereas 2714725 reads were mapped uniquely in CM-119. In inoculated SC-7, the total number of differentially expressed genes (DEGs) against control was 1413, where 1011 were up-regulated, and 402 were down-regulated. In susceptible inoculated genotype CM 119, the number of DEGs against control was 2902, where 1703 were up-, and 1199 were down-regulated. DEGs between inoculated resistant and susceptible genotypes were 10745, where 5343 were up-, and 5402 were down-regulated. The RNA-seq data were validated using RT-qPCR. The key findings are that SC-7 poses a robust plant signaling system mainly induced by oxidation–reduction process and calcium-mediated signaling. It regulates its fitness-related genes efficiently, viz., aldolase 2 gene, isopropanoid, phyto hormones, P450 cytochrome, amino acid synthesis, nitrogen assimilation genes etc. These findings showed more transcriptional changes in the SC-7 genotype, which contains many defence-related genes. They can be explored in future crop development programmes to combat multiple maize diseases. The current finding provides information to elucidate molecular and cellular processes occurring in maize during B. maydis race O infection.

Genome analysis and hyphal movement characterization of the hitchhiker endohyphal Enterobacter sp. from Rhizoctonia solani.

Bacterial-fungal interactions are pervasive in the rhizosphere. While an increasing number of endohyphal bacteria have been identified, little is known about their ecology and impact on the associated fungal hosts and the surrounding environment. In this study, we characterized the genome of an Enterobacter sp. Crenshaw (En-Cren), which was isolated from the generalist fungal pathogen Rhizoctonia solani, and examined the genetic potential of the bacterium with regard to the phenotypic traits associated with the fungus. Overall, the En-Cren genome size was typical for members of the genus and was capable of free-living growth. The genome was 4.6 MB in size, and no plasmids were detected. Several prophage regions and genomic islands were identified that harbor unique genes in comparison with phylogenetically closely related Enterobacter spp. Type VI secretion system and cyanate assimilation genes were identified from the bacterium, while some common heavy metal resistance genes were absent. En-Cren contains the key genes for indole-3-acetic acid (IAA) and phenylacetic acid (PAA) biosynthesis, and produces IAA and PAA in vitro, which may impact the ecology or pathogenicity of the fungal pathogen in vivo. En-Cren was observed to move along hyphae of R. solani and on other basidiomycetes and ascomycetes in culture. The bacterial flagellum is essential for hyphal movement, while other pathways and genes may also be involved.IMPORTANCEThe genome characterization and comparative genomics analysis of Enterobacter sp. Crenshaw provided the foundation and resources for a better understanding of the ecology and evolution of this endohyphal bacteria in the rhizosphere. The ability to produce indole-3-acetic acid and phenylacetic acid may provide new angles to study the impact of phytohormones during the plant-pathogen interactions. The hitchhiking behavior of the bacterium on a diverse group of fungi, while inhibiting the growth of some others, revealed new areas of bacterial-fungal signaling and interaction, which have yet to be explored.

Incorporating microbial inoculants to reduce nitrogen loss during sludge composting by suppressing denitrification and promoting ammonia assimilation

To address the challenge of increasing nitrogen retention in compost, this study investigated the effects of microbial communities on denitrification and ammonia assimilation during sludge composting by inoculating microbial inoculants. The results showed that the retention rates of total Kjeldahl nitrogen (TKN) and humic acid (HA) in MIs group (with microbial inoculants) were 4.94 % and 18.52 % higher than those in the control group (CK), respectively. Metagenomic analysis showed that Actinobacteria and Proteobacteria were identified as main microorganisms contributing to denitrification and ammonia assimilation. The addition of microbial agents altered the structure of the microbial community, which in turn stimulated the expression of functional genes. During cooling period, the ammonia assimilation genes glnA, gltB and gltD in MIs were 15.98 %, 24.84 % and 32.88 % higher than those in CK, respectively. Canonical correspondence analysis revealed a positive correlation between the dominant bacterial genera from the cooling stage to the maturity stage and the levels of NO3−-N, NH4+-N, HA, and TKN contents. NH4+-N was positively correlated with HA, indicating NH4+-N might be incorporated into HA. Heat map and network analyses revealed NH4+-N as a key factor affecting functional genes of denitrification and ammonia assimilation, with Nitrospira identified as the core bacteria in the microbial network. Therefore, the addition of microbial agents could increase nitrogen retention and improve compost product quality.

The synthesis of ectoine enhance the assimilation of ammonia nitrogen in hypersaline wastewater by the salt-tolerant assimilation bacteria sludge

In contrast to nitrification-denitrification microorganisms that convert ammonia nitrogen in hypersaline wastewater into nitrogen for discharge, this research utilizes sludge enriched with salt-tolerant assimilation bacteria (STAB) to assimilate organic matter and ammonia nitrogen in hypersaline wastewater into ectoine - a biomass with high economic value and resistance to external osmotic pressure. The study investigates the relationship between the synthesis of ectoine and nitrogen removal efficiency of STAB sludge in three sequencing batch reactors (SBR) operated at different salinities (50, 75, and 100 g/L) and organic matter concentrations. The research reveals that, under low concentration carbon sources (TOC/N = 4, NH4+-N = 60 mg/L), the ammonia nitrogen removal efficiency of SBR reactors increased by 14.51 % and 17.25 % within 5 d and 2 d, respectively, when salinity increased from 50 g/L to 75 g/L and 100 g/L. Under high concentration carbon sources (TOC/N = 8, NH4+-N = 60 mg/L), the ammonia nitrogen removal efficiency of STAB sludge in the three reactors stabilized at 80.20 %, 76.71 %, and 72.87 %, and the total nitrogen removal efficiency was finally stabilized at 80.47 %, 73.15 %, and 65.53 %, respectively. The nitrogen removal performance by ammonium-assimilating of STAB sludge is more sustainable under low salinity, while it is more short-term explosive under high salinity. Moreover, the intracellular ectoine concentration of STAB sludge was found to be related to this behavior. Empirical formulas confirm that STAB sludge synthesizes ectoine from nutrients in wastewater through assimilation, and intracellular ectoine has a threshold defect (150 mg/gVss). The ectoine metabolism pathways of STAB sludge was constructed using the Kyoto Encyclopedia of Genes and Genomes (KEGG). The ammonia nitrogen in sewage is converted into glutamic acid under the action of assimilation genes. It then undergoes a tricarboxylic acid cycle to synthesize the crucial precursor of ectoine - aspartic acid. Subsequently, ectoine is produced through ectoine synthase. The findings suggest that when the synthesis of intracellular ectoine reaches saturation, it inhibits the continuous nitrogen removal performance of STAB sludge under high salinity. STAB sludge does not actively release ectoine through channels under stable external osmotic pressure.

Elucidating microbial mechanisms of microcystin-LR degradation in Lake Erie beach sand through metabolomics and metatranscriptomics

As one of five Laurentian Great Lakes, Lake Erie ranks among the top freshwater drinking sources and ecosystems globally. Historical and current agriculture mismanagement and climate change sustains the environmental landscape for late summer cyanobacterial harmful algal blooms, and consequently, cyanotoxins such as microcystin (MC). Microcystin microbial degradation is a promising mitigation strategy, however the mechanisms controlling the breakdown of MCs in Lake Erie are not well understood. Pelee Island, Ontario, Canada is located in the western basin of Lake Erie and the bacterial community in the sand has demonstrated the capacity of metabolizing the toxin. Through a multi-omic approach, the metabolic, functional and taxonomical signatures of the Pelee Island microbial community during MC-LR degradation was investigated over a 48-hour period to comprehensively study the degradation mechanism. Cleavage of bonds surrounding nitrogen atoms and the upregulation of nitrogen deamination (dadA, alanine dehydrogenase, leucine dehydrogenase) and assimilation genes (glnA, gltB) suggests a targeted isolation of nitrogen by the microbial community for energy production. Methylotrophic pathways RuMP and H4MPT control assimilation and dissimilation of carbon, respectively and differential abundance of Methylophilales indicates an interconnected role through electron exchange of denitrification and methylotrophic pathways. The detected metabolites did not resolve a clear breakdown pathway, but rather the diversity of products in combination with taxonomic and functional results supports that a variety of strategies are applied, such as epoxidation, hydroxylation, and aromatic degradation. Annual repeated exposure to the toxin may have allowed the community to adaptatively establish a novel pathway through functional plasticity and horizontal gene transfer. The culmination of these results reveals the complexity of the Pelee Island sand community and supports a dynamic and cooperative metabolism between microbial species to achieve MC degradation.