Engineered reduction of S-adenosylmethionine alters lignin in sorghum.

BACKGROUND: Lignin is an aromatic polymer deposited in secondary cell walls of higher plants to provide strength, rigidity, and hydrophobicity to vascular tissues. Due to its interconnections with cell wall polysaccharides, lignin plays important roles during plant growth and defense, but also has a negative impact on industrial processes aimed at obtaining monosaccharides from plant biomass. Engineering lignin offers a solution to this issue. For example, previous work showed that heterologous expression of a coliphage S-adenosylmethionine hydrolase (AdoMetase) was an effective approach to reduce lignin in the model plant Arabidopsis. The efficacy of this engineering strategy remains to be evaluated in bioenergy crops. RESULTS: We studied the impact of expressing AdoMetase on lignin synthesis in sorghum (Sorghum bicolor L. Moench). Lignin content, monomer composition, and size, as well as biomass saccharification efficiency were determined in transgenic sorghum lines. The transcriptome and metabolome were analyzed in stems at three developmental stages. Plant growth and biomass composition was further evaluated under field conditions. Results evidenced that lignin was reduced by 18% in the best transgenic line, presumably due to reduced activity of the S-adenosylmethionine-dependent O-methyltransferases involved in lignin synthesis. The modified sorghum features altered lignin monomer composition and increased lignin molecular weights. The degree of methylation of glucuronic acid on xylan was reduced. These changes enabled a ~20% increase in glucose yield after biomass pretreatment and saccharification compared to wild type. RNA-seq and untargeted metabolomic analyses evidenced some pleiotropic effects associated with AdoMetase expression. The transgenic sorghum showed developmental delay and reduced biomass yields at harvest, especially under field growing conditions. CONCLUSIONS: The expression of AdoMetase represents an effective lignin engineering approach in sorghum. However, considering that this strategy potentially impacts multiple S-adenosylmethionine-dependent methyltransferases, adequate promoters for fine-tuning AdoMetase expression will be needed to mitigate yield penalty.

Fine-tuning the element dose in nanoparticle synthesis is the critical factor determining nanoparticle's impact on plant growth.

This study elucidates the impact of element dose during nanoparticle (NPs) synthesis on plant growth indices. Novel NPs containing two essential micro-nutrients, zinc (Zn) and manganese (Mn), were co-doped on cerium oxide (CeO2) (ZnMnCe) with different ratios (1, 2, and 3%). The synthesized NPs were characterized by advanced analytical techniques (EDX, TEM, SEM, XPS, and XRD) and hydroponically applied to barley (Hordeum vulgare L.). The impact of ZnMnCe NPs on growth indices and plant nutrients was examined. SEM, HRTEM, and confocal microscopy were used to show the morphological and structural influences of ZnMnCe NPs. Results showed that the plant growth indices (root/leaf length, chlorophyll fluorescence, pigmentation, and biomass) were remarkably improved with a 1% Mn/Zn addition. Conversely, growth retardation, cell membrane damage, root morphology deformation, and genotoxicity were apparent by 3% of Mn/Zn addition. Overall, a significant improvement in growth was revealed when Mn and Zn were included at 1%. However, increasing concentrations (2% and 3%) impaired the growth. These results show that the element ratio used in NPs synthesis is essential in the plant's physiological response. Precise adjustment of element dosage during NPs synthesis determines whether the NPs are beneficial or harmful. This must be well-balanced for nanofertilizer production and plant applications.

Defined synthetic microbial communities colonize and benefit field-grown sorghum.

The rhizosphere constitutes a dynamic interface between plant hosts and their associated microbial communities. Despite the acknowledged potential for enhancing plant fitness by manipulating the rhizosphere, the engineering of the rhizosphere microbiome through inoculation has posed significant challenges. These challenges are thought to arise from the competitive microbial ecosystem where introduced microbes must survive, and the absence of adaptation to the specific metabolic and environmental demands of the rhizosphere. Here, we engineered a synthetic rhizosphere community (SRC1) with the anticipation that it would exhibit a selective advantage in colonizing the host Sorghum bicolor, thereby potentially fostering its growth. SRC1 was assembled from bacterial isolates identified either for their potential role in community cohesion through network analysis or for their ability to benefit from host-specific exudate compounds. The growth performance of SRC1 was assessed in vitro on solid media, in planta under gnotobiotic laboratory conditions, and in the field. Our findings reveal that SRC1 cohesion is most robust when cultivated in the presence of the plant host under laboratory conditions, with lineages being lost from the community when grown either in vitro or in a native field setting. We establish that SRC1 effectively promotes the growth of both above- and below-ground plant phenotypes in both laboratory and native field contexts. Furthermore, in laboratory conditions, these growth enhancements correlate with the transcriptional dampening of lignin biosynthesis in the host. Collectively, these results underscore the potential utility of synthetic microbial communities for modulating crop performance in controlled and native environments alike.

Expression of dehydroshikimate dehydratase in poplar induces transcriptional and metabolic changes in the phenylpropanoid pathway.

Modification of lignin in feedstocks via genetic engineering aims to reduce biomass recalcitrance to facilitate efficient conversion processes. These improvements can be achieved by expressing exogenous enzymes that interfere with native biosynthetic pathways responsible for the production of the lignin precursors. In planta expression of a bacterial 3-dehydroshikimate dehydratase in poplar trees reduced lignin content and altered the monomer composition, which enabled higher yields of sugars after cell wall polysaccharide hydrolysis. Understanding how plants respond to such genetic modifications at the transcriptional and metabolic levels is needed to facilitate further improvement and field deployment. In this work, we acquired fundamental knowledge on lignin-modified poplar expressing 3-dehydroshikimate dehydratase using RNA-seq and metabolomics. The data clearly demonstrate that changes in gene expression and metabolite abundance can occur in a strict spatiotemporal fashion, revealing tissue-specific responses in the xylem, phloem, or periderm. In the poplar line that exhibited the strongest reduction in lignin, we found that 3% of the transcripts had altered expression levels and ~19% of the detected metabolites had differential abundance in the xylem from older stems. The changes affected predominantly the shikimate and phenylpropanoid pathways as well as secondary cell wall metabolism, and resulted in significant accumulation of hydroxybenzoates derived from protocatechuate and salicylate.

Impact of magnetic field on the translocation of iron oxide nanoparticles (Fe3O4) in barley seedlings (Hordeum vulgare L.).

The effect and contribution of an external magnetic field (MF) on the uptake and translocation of nanoparticles (NPs) in plants have been investigated in this study. Barley was treated with iron oxide NPs (Fe3O4, 500 mg/L, 50-100 nm) and grown under various MF strengths (20, 42, 125, and 250 mT). The root-to-shoot translocation of NPs was assessed using a vibrating sample magnetometer (VSM) and inductively coupled plasma optical emission spectrometry (ICP-OES). Additionally, plant phenological parameters, such as germination, protein and chlorophyll content, and photosynthetic and nutritional status, were examined. The results demonstrated that the external MF significantly enhances the uptake of NPs through the roots. The uptake was higher at lower MF strengths (20 and 42 mT) than at higher MF strengths (125 and 250 mT). The root and shoot iron (Fe) contents were approximately 2.5-3-fold higher in the 250 mT application compared to the control. Furthermore, the MF treatments significantly increased micro-elements such as Mn, Zn, Cu, Mo, and B (P < 0.005). This effect could be attributed to the disruption of cell membranes at the root tip cells caused by both the MF and NPs. Moreover, the MF treatments improved germination rates by 28%, total protein content, and photosynthetic parameters. These findings show that magnetic field application helps the effective transport of magnetic NPs, which could be essential for NPs-mediated drug delivery, plant nutrition, and genetic transformation applications. Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-023-03727-4.

Phylogenetic analysis based on the ITS, matK and rbcL DNA barcodes and comparison of chemical contents of twelve Paeonia taxa in Türkiye.

BACKGROUD: Twelve taxa of herbaceous Paeonia species were recorded in Türkiye. All definitions were performed morphologically and/or anatomically and there is no study based on DNA barcode sequences. Three barcode regions were sequenced to determine the phylogenetic relationships of Turkish Paeonia taxa. The chemical comparison of roots was also investigated. METHODS AND RESULTS: The taxons were collected between May and June 2021 from nine cities. Leaf materials were used for DNA isolation and ITS, matK and rbcL regions were amplified and sequenced. There was no difference among taxa in terms of rbcL sequences. But the ITS and matK regions distinguished 12 taxa and structured them in two groups. ITS region distinguished P. peregrina, P. arietina, and P. tenuifolia from other taxa, while matK region distinguished P. arietina and P. witmanniana from other taxa. Both barcode sequences actually showed that the registration of P. mascula subsp. arasicola was actually 100% similar to P. arietina. ITS was the most polymorphic region (n = 54) followed by matK (n = 9). These sequences could successfully discriminate Paoenia species from each other and diploid P. tenuifolia. The methanolic root (100 gr) extracts were examined for total phenolic and flavonoid content, and antioxidant activities. Significant variation was found for polyphenolic content, and antioxidant properties (TPC from 204.23 to 2343.89 mg, TFC from 7.73 to 66.16 mg, and FRAP from 523.81 to 4338.62 mg). SC50 values of ABTS and DPPH were ranged from 115.08 to 1115.52 µg/ml and 73.83 to 963.59 µg/ml, respectively. CONCLUSION: It was concluded that 11 of 12 taxa had differences in terms of ITS and matK sequences and these region must be used for the correct identification of Turkish Paeonia.

Whole-genome sequencing-based characteristics of Escherichia coli Rize-53 isolate from Turkey.

BACKGROUND: Urinary tract infections (UTIs) are one of the most common infectious diseases. Inappropriate and excessive administration of antibiotics has led to the increased antibiotic resistance in the pathogens that cause UTIs. This work focused on identifying genetic determinants of antibiotic resistance in a clinical isolate of UTI-causing Escherichia coli. OBJECTIVES: A clinical isolate of E. coli resistant to ß-lactam, tetracycline and aminoglycoside antibiotics was analyzed using whole-genome sequencing (WGS) to identify genes that contribute to its resistance. MATERIAL AND METHODS: The clinical isolate was obtained from a urine sample of a UTI patient in Turkey and identified via 16S rDNA sequencing. Antimicrobial susceptibility test was performed for 17 antibiotics using VITEK® 2 and the results were confirmed using minimum inhibitory concentration assay. Whole-genome sequencing of the isolate was performed using Illumina sequencing and analyzed with bioinformatic tools for multilocus sequence typing, replicon types, virulence factors, and antimicrobial resistance genes. RESULTS: Whole-genome datum was submitted to the National Center for Biotechnology Information (NCBI; accession No. JAKSGM000000000). The isolate was only found to be resistant to piperacillin in the ß-lactam class of antibiotics. While the isolate was also resistant to aminoglycoside and tetracycline antibiotics, it was sensitive to other antibiotics tested. Ten antibiotic resistance genes were identified in the genome of the isolate: blaOXA-1, blaOXA-2, aac(6')-II, aac(6')-Ib-cr, tetB, catB3, qacE, sitABCD, mdfA, and sul-2. Clonal subtype (ST) and serotype of the isolate were identified as ST2141 and O107/H39, respectively. Plasmid replicon typing was used to identify 5 plasmid types in the genome of E. coli Rize-53 (Col(BS512), IncC, IncIA, IncFIB(AP1918), and IncFII(pRSB107)); however, none of the resistance genes were encoded on the plasmid. CONCLUSIONS: Genetic determinants of resistance to tetracycline, ß-lactam and aminoglycoside antibiotics were identified using WGS in a uropathogenic E. coli from ST2141 lineage and O107:H39 serotype, isolated in Turkey.

Characteristics in the whole-genome sequence of Klebsiella pneumoniae ST147 from Turkey.

The study aimed to analyze antibiotic resistance determinants in a carbapenem-resistant Klebsiella pneumoniae by whole-genome sequencing (WGS). K. pneumoniae was isolated from a urine sample and it was characterized by 16S rDNA sequencing in Turkey. This strain was named as Kpn Rize-53-TR. Antimicrobial susceptibility testing was performed for seventeen antibiotics by VITEK-2 and the result was confirmed by MIC. The whole genome of isolate was sequenced by Illumina and was analysed by bioinformatic tools for MLST, replicon types, and antimicrobial resistance genes. The whole genome data was submitted to NCBI. The isolate was found to be resistant to all tested ß-lactam antibiotics and the highest MIC values were found for piperacillin, piperacillin/tazobactam (≥128). No resistance to colistin and moderate susceptibility to amikacin and tetracycline was observed. The isolate carried 12 resistance genes belonging to 10 resistance classes; ere(A), fosA, oqxB, cmlA1, aac(a)-IIa, bla KPC-2, bla TEM-1A, bla SHV-67, bla CTX-M-15, bla OXA-1-2-9. Mutations were detected in gyrA (83Y) and parC (80I) genes. Clonal subtype of the isolate was ST147, and it had wzi420 and wzc38 alleles. Its serotype was O3/O3a. The bla KPC-2 was firstly found in both ST147 clonal group in Turkey and in serotype O3/O3a in the world. By plasmid replicon typing, five plasmids IncFII(K), Col(BS512), IncR, IncFIA(HI1) and IncFIB(pQil) were determined in Kpn Rize-53-TR and bla KPC-2 was located on IncFII(K) plasmid. The presence of bla KPC-2 on the plasmid with other resistance genes accelerates its own spread together with other resistance genes.

PIRIN2 suppresses S-type lignin accumulation in a noncell-autonomous manner in Arabidopsis xylem elements.

PIRIN (PRN) genes encode cupin domain-containing proteins that function as transcriptional co-regulators in humans but that are poorly described in plants. A previous study in xylogenic cell cultures of Zinnia elegans suggested a role for a PRN protein in lignification. This study aimed to identify the function of Arabidopsis (Arabidopsis thaliana) PRN proteins in lignification of xylem tissues. Chemical composition of the secondary cell walls was analysed in Arabidopsis stems and/or hypocotyls by pyrolysis-gas chromatography/mass spectrometry, 2D-nuclear magnetic resonance and phenolic profiling. Secondary cell walls of individual xylem elements were chemotyped by Fourier transform infrared and Raman microspectroscopy. Arabidopsis PRN2 suppressed accumulation of S-type lignin in Arabidopsis stems and hypocotyls. PRN2 promoter activity and PRN2:GFP fusion protein were localised specifically in cells next to the vessel elements, suggesting a role for PRN2 in noncell-autonomous lignification of xylem vessels. Accordingly, PRN2 modulated lignin chemistry in the secondary cell walls of the neighbouring vessel elements. These results indicate that PRN2 suppresses S-type lignin accumulation in the neighbourhood of xylem vessels to bestow G-type enriched lignin composition on the secondary cell walls of the vessel elements. Gene expression analyses suggested that PRN2 function is mediated by regulation of the expression of the lignin-biosynthetic genes.

ß-lactamase genes in carbapenem resistance Acinetobacter baumannii isolates from a Turkish university hospital.

INTRODUCTION: The spread of Acinetobacter baumannii, resistant to most of the available antimicrobial agents, is a serious health problem. The high rate of carbapenem resistance among Acinetobacter baumannii isolates is considered as a threat to public health. In this study, we aimed to determine the antibiotic resistance and related genes in carbapenem-resistant Acinetobacter baumannii isolates. METHODOLOGY: Ninety six isolates of A. baumannii were included. Antimicrobial susceptibility was performed by Phoenix Automated System and disk diffusion method. Carbapenem resistane was characterized by scrneeing of resistance genes such as blaTEM, blaSHV, blaCTX-M1-2, blaPER, blaVEB, blaKPC, blaGES, blaNDM, blaVIM, blaIMP and blaOXA23-24-51-58 using multiplex polymerase chain reaction. RESULTS: Resistance for the levofloxacin, gentamicin, amikacin, and tigecycline were determined as 96.9%, 93.7%, 72.9% and 45.8% respectively. Colistin was the only susceptible antibiotic against all clinical isolates. All isolates were defined as multidrug resistance and of these, 31.2% were extensively drug-resistant (sensitive only to colistin). BlaOXA-51-  and blaOXA-23 genes were detected in 100% strains while blaTEM was found in only 2% strains. There was no amplification for the blaSHV, blaCTX-M1-2, blaPER, blaVEB, blaKPC, blaGES blaNDM, blaVIM, blaIMP and blaOXA24-58 genes. CONCLUSIONS: The high frequency of blaOXA-23 and low frequency of blaTEM gene was observed that indicate prevalence of a variety of A. baumannii strains. The rates of resistance genes vary from region to region. Studies are required for the prevention and control of A. baumannii infection and to formulate the strategies of antibiotic usage.

Gram (-) microorganisms DNA polymerase inhibition, antibacterial and chemical properties of fruit and leaf extracts of Sorbus acuparia and Sorbus caucasica var. yaltirikii.

Investigation of novel plant-based agents might provide alternative antibiotics and thus fight antibiotic resistance. Here, we measured the ability of fruit and leaf extracts of Sorbus aucuparia (Sauc) and endemic Sorbus caucasica var. yaltirikii (Scau) to inhibit nonreplicative (Klenow Fragment-KF and Bacillus Large Fragment-BLF) and replicative (DnaE and PolC) bacterial DNA polymerases along with their antimicrobial, DPPH free radical scavenging activity (RSA), and chemical contents by total phenolic content and HPLC-DAD analysis. We found that leaf extracts had nearly 10-fold higher RSA and 5-fold greater TPC than the corresponding fruit extracts. All extracts had large amounts of chlorogenic acid (CGA) and rutin, while fruit extracts had large amounts of quercetin. Hydrolysis of fruit extracts revealed mainly caffeic acid from CGA (caffeoylquinic acid) and quercetin from rutin (quercetin-3-O-rutinoside), as well as CGA and derivatives of CGA and p-coumaric acid. Plant extracts of Sorbus species showed antimicrobial activity against Gram-negative microorganisms. Scau leaf extracts exhibited strong inhibition of KF activity. Sauc and Scau leaf extracts also strongly inhibited two replicative DNA polymerases. Thus, these species can be considered a potential source of novel antimicrobial agents specific for Gram-negative bacteria.

Accumulation of N-acetylglucosamine oligomers in the plant cell wall affects plant architecture in a dose-dependent and conditional manner.

To study the effect of short N-acetylglucosamine (GlcNAc) oligosaccharides on the physiology of plants, N-ACETYLGLUCOSAMINYLTRANSFERASE (NodC) of Azorhizobium caulinodans was expressed in Arabidopsis (Arabidopsis thaliana). The corresponding enzyme catalyzes the polymerization of GlcNAc and, accordingly, ß-1,4-GlcNAc oligomers accumulated in the plant. A phenotype characterized by difficulties in developing an inflorescence stem was visible when plants were grown for several weeks under short-day conditions before transfer to long-day conditions. In addition, a positive correlation between the oligomer concentration and the penetrance of the phenotype was demonstrated. Although NodC overexpression lines produced less cell wall compared with wild-type plants under nonpermissive conditions, no indications were found for changes in the amount of the major cell wall polymers. The effect on the cell wall was reflected at the transcriptome level. In addition to genes encoding cell wall-modifying enzymes, a whole set of genes encoding membrane-coupled receptor-like kinases were differentially expressed upon GlcNAc accumulation, many of which encoded proteins with an extracellular Domain of Unknown Function26. Although stress-related genes were also differentially expressed, the observed response differed from that of a classical chitin response. This is in line with the fact that the produced chitin oligomers were too small to activate the chitin receptor-mediated signal cascade. Based on our observations, we propose a model in which the oligosaccharides modify the architecture of the cell wall by acting as competitors in carbohydrate-carbohydrate or carbohydrate-protein interactions, thereby affecting noncovalent interactions in the cell wall or at the interface between the cell wall and the plasma membrane.