Enzyme Gene Research Articles

Extracytoplasmic polysaccharides control cellulosomal and non-cellulosomal systems in Herbivorax saccincola A7

Herbivorax saccincola A7 is an anaerobic alkali-thermophilic lignocellulolytic bacterium that possesses a cellulosome and high xylan degradation ability. To understand the expression profile of extracellular enzymes by carbon sources, quantitative real-time PCR was performed on all cellulosomal and non-cellulosomal enzyme genes of H. saccincola A7 using cellulose and xylan as carbon sources. The results confirmed that the scaffolding proteins of H. saccincola A7 were expressed. In general, the cellulosomal genes belonging to the glycoside hydrolase families 9, 10, 11, and 48 were repressed when xylan was the sole carbon source, but these genes were significantly induced in the presence of cellulose. These results indicate that cellulose, not xylan, is a key inducer of cellulosomal genes in H. saccincola A7. The RsgI-like proteins, which regulate a carbohydrate-sensing mechanism in Clostridium thermocellum, were also found to be encoded in the H. saccincola A7 genome. To confirm the regulation by RsgI-like proteins, the relative expression of σI1–σI4 factors was analyzed on both carbon sources. The expression of alternative σI1 and σI2 factors was enhanced by the presence of cellulose. By contrast, the expression of σI3 and σI4 factors was activated by both cellulose and xylan. Taken together, the results reveal that the cellulosomal and non-cellulosomal genes of H. saccincola A7 are regulated through a carbohydrate-sensing mechanism involving anti-σ regulator RsgI-like proteins.Key points• qRT-PCR performed on cellulosomal and non-cellulosomal genes of H. saccincola A7• Cellulose is a key inducer of the cellulosome of H. saccincola A7• H. saccincola A7 possesses a similar system of anti-σ regulator RsgI-like proteins

Low temperature and high humidity affect dynamics of chlorophyll biosynthesis and secondary metabolites in Cucumber.

During the cold season, low temperature (LT) and high relative humidity (HRH) are significant environmental factors in greenhouses and plastic tunnels, often hindering plant growth and development. The chlorophyll (Chl) biosynthesis inhibitory mechanisms under LT and HRH stress are still widely unclear. To understand how cucumbers seedlings respond to LT and HRH stress, we investigated the impact of these stressors on Chl biosynthesis. Our results revealed that individual LT, HRH and combined LT + HRH stress conditions affected chlorophyll a, b, total chlorophyll and carotenoid content, reducing the levels of these pigments. The levels of Chlorophyll precursors were also markedly reduced under LT and HRH stresses, with the greatest reduction observed in cucumber seedlings exposed to LT + HRH conditions (9/5℃, 95%HRH). The activities of glutamate-1-semialdehyde transaminase (GSA-AT), ALA dehydratase (ALAD), Mg-chelatase, and protochlorophyllide oxidoreductase (POR) were increased under individual LT, HRH, conditions but decreased by combination of LT + HRH stress condition. In addition, Chl biosynthesis related genes (except PBG) were upregulated by the HRH stress but were significantly downregulated under the LT + HRH stress condition in cucumber seedlings. Furthermore, the content of phenols, flavonoids and phenolic acids (cinnamic acid and caffeic acid) were significantly surged under LT + HRH treatment over the control. Histochemical observation showed higher O2- and H2O2 content in cucumber leaves during the LT and HRH stress. The results indicate that LT + HRH stress significantly impairs chlorophyll biosynthesis in cucumber seedlings by drastically reducing pigment accumulation, altering enzyme activity and gene expression. Additionally, LT + HRH stress induces oxidative damage, which further exacerbates the decline in chlorophyll content and affects overall cucumber metabolism.

Genetic polymorphisms associated with preeclampsia risk in Nigerian women

BackgroundPreeclampsia, a complex hypertensive disorder unique to pregnancy, significantly impacts maternal and fetal health worldwide, with a prevalence of 2–8%. This condition results from a complex interplay of genetic, environmental, and immunological factors.Aim and objectivesThis study aims to investigate the genetic predispositions to preeclampsia, focusing on specific gene polymorphisms among pregnant women at Central Hospital Auchi, Nigeria.Materials and methodsWe examined the endothelial nitric oxide synthase (eNOS), vascular endothelial growth factor (VEGF), angiotensin-converting enzyme (ACE), and tumor necrosis factor-alpha (TNF-α) genes in 200 pregnant women, equally divided between preeclamptic patients and normotensive controls.ResultsThe eNOS G894T polymorphism was significantly associated with preeclampsia, with the T allele nearly doubling the risk. The VEGF C936T polymorphism's T allele also indicated a higher risk. The D allele in the ACE gene's insertion/deletion (I/D) polymorphism significantly increased the risk, as did the A allele in the TNF-α G308A polymorphism.ConclusionsThese findings highlight the importance of genetic factors in preeclampsia and suggest that genetic screening could improve risk stratification and early detection. Future research should integrate genetic, epigenetic, and environmental data to understand preeclampsia's multifaceted nature and develop targeted therapies. This study underscores the potential of personalized medicine in managing and reducing the risks associated with preeclampsia.

Microbial inoculation accelerates rice straw decomposition by reshaping structure and function of lignocellulose-degrading microbial consortia in paddy fields

Inoculating lignocellulose-degrading microorganisms can accelerate straw decomposition in paddy field; however, the relationship between indigenous and inoculated microorganisms remains unclear. This study explored the effects of microbial inoculation on straw decomposition, microbial community, lignocellulose-degrading consortia, and associated functional genes. After inoculation, straw degradation rate increased by up to 4.9 %, and the rice yield increased by 790 kg/ha. Microbial inoculation restructured soil microbial community, influencing key taxa and interactions within the microbial network. A lignocellulose-degrading consortia consisting 37 genera was established, with a notable increase in the relative abundance of lignocellulose-degrading bacteria following inoculation. Among them, Pseudarthrobacter, with high lignin-degrading enzyme activity, emerged as a key genus after inoculation. Additionally, the abundance of lignin-degrading enzyme genes also increased significantly after inoculation. These findings offer new insights into how microbial inoculation accelerates the in situ decomposition of rice straw by reshaping the structure and function of lignocellulose-degrading consortia within the soil ecosystem.

RNAseq-Based Carboxylesterase Nl-EST1 Gene Expression Plasticity Identification and Its Potential Involvement in Fenobucarb Resistance in the Brown Planthopper Nilaparvata lugens.

Carbamate insecticides have been used for over four decades to control brown planthopper, Nilaparvata lugens, but resistance has been reported in many countries, including the Republic of Korea. The bioassay results on resistance to fenobucarb showed that the LC50 values were 3.08 for the susceptible strain, 10.06 for the 2015 strain, and 73.98 mg/L for the 2019 strain. Compared to the susceptible strain, the 2015 and 2019 strains exhibited resistance levels 3.27 and 24.02 times higher, respectively. To elucidate the reason for the varying levels of resistance to fenobucarb in these strains, mutations in the acetylcholinesterase 1 (ACE1) gene, the target gene of carbamate, were investigated, but no previously reported mutations were confirmed. Through RNA-seq analysis focusing on the expression of detoxification enzyme genes as an alternative resistance mechanism, it was found that the carboxylesterase gene Nl-EST1 was overexpressed 2.4 times in the 2015 strain and 4.7 times in the 2019 strain compared to the susceptible strain. This indicates a strong correlation between the level of resistance development in each strain and the expression level of Nl-EST1. Previously, Nl-EST1 was reported in an organophosphorus insecticide-resistant strain of Sri Lanka 2000. Thus, Nl-EST1 is crucial for developing resistance to organophosphorus and carbamate insecticides. Resistance-related genes such as Nl-EST1 could serve as expression markers for resistance diagnosis, and can apply to integrated resistance management of N. lugens.

Role of anthocyanin metabolic diversity in bract coloration of Curcuma alismatifolia varieties

Anthocyanins are one of the key metabolites influencing the coloration of ornamental bracts in plants. Curcuma alismatifolia is an emerging ornamental plant, known for the rich diversity in the coloration of its bracts and the variety of anthocyanins present. However, the specific anthocyanin metabolites contributing to this diversity are not entirely clear. This study examines the bract color variation across 19 C. alismatifolia varieties using colorimetric analysis and spectrophotometric determination of total anthocyanin content. The 19 accessions were categorized into four color groups: white, light pink, pink, and purple. Further analysis using anthocyanin metabolomics and transcriptomics was conducted on five C. alismatifolia varieties with significant differences in coloration and total anthocyanin content. In addition to previously reported anthocyanins, delphinidin-3-O-glucoside and peonidin-3-O-rutinoside were identified for the first time as important contributors to the diverse bract coloration in C. alismatifolia. Fifty-three differentially expressed genes (DEGs) were identified in the anthocyanin biosynthesis pathway, and two significant gene modules were determined through WGCNA analysis. Correlation network analysis revealed two BZ1 genes that may be key terminal enzyme genes affecting anthocyanin synthesis in C. alismatifolia bracts. Multiple transcription factors, including MYB, NAC, WRKY, ERF, and bHLH, may be involved in regulating the accumulation of different anthocyanin contents in the bracts. This research sheds light on the genetic and metabolic factors that influence bract coloration in C. alismatifolia.

Cytotoxic Activity of Vancomycin-Resistant Enterococci Isolated from Hospitalised Patients.

Vancomycin-resistant enterococci (VRE) are considered one of the main nosocomial pathogens due to their increasing antibiotic resistance and ability to cause life-threatening infections in humans. This study included VRE isolates obtained from various specimens including urine, blood, faeces, wounds, sputum, and oral cavity wash. Of the 37 strains, 30 (81.1%) and 7 (18.9%) were identified by MALDI TOF as Enterococcus faecium and Enterococcus faecalis, respectively. The clinical vancomycin-resistant enterococci exhibited multi-drug resistance (MDR). Apart from vancomycin, the enterococci exhibited resistance to penicillins (89.1 to 100%), fluoroquinolones (100%), rifampicin (86.5%), tetracycline (27%), aminoglycosides (56.8 to 86.5%), quinupristin-dalfopristin (35.1%), and chloramphenicol (10.8%). Moreover, resistance to linezolid and tigecycline emerged among the tested vancomycin-resistant enterococci. The analysis of aminoglycoside modifying enzyme (AME) genes showed the presence of bifunctional aac(6')-Ie-aph(2″)-Ia genes contributed to high-level aminoglycoside resistance (HLAR) in the E. faecalis and E. faecium isolates. The other AME gene, i.e., aph(3')-IIIa, was also found in the VRE isolates. All strains carried the vanA gene. Enterococci from colonised gastrointestinal tracts (1/2.7%) and from infection (6/16.2%) showed cytotoxic activity against the human epithelial cell line HEp-2.

Composition and micromorphological determination of blue honeysuckle fruit (Lonicera caerulea L.) cuticular wax and its effects on fruit post-harvest quality

The chemical composition and structure of the cuticular wax in blue honeysuckle fruit were investigated using gas chromatography–mass spectrometry (GC–MS) and scanning electron microscopy (SEM). The results revealed that the cuticular wax was dense and uniform, taking on a tubular form. A total of 158 wax components were identified, including alkanes, terpenes, ketones, alcohols, fatty acids, and esters. The wax was found to be particularly rich in alkanes. After storage, the wax content decreased, whereas an increase in 1-undecanol. The destruction or reduction of cuticular wax resulted in a more rapid decline in storage quality, loss of nutrients, and a decrease in antioxidant phytochemicals. Meanwhile, wax metabolizing enzyme activity and gene expression increased. This study presents a deeper understanding of blue honeysuckle fruit cuticular wax composition and aids to developing effective measures to delay its postharvest fruit quality deterioration.

Exogenous Melatonin Alleviates NaCl Injury by Influencing Stomatal Morphology, Photosynthetic Performance, and Antioxidant Balance in Maize.

Maize (Zea mays L.) is sensitive to salt stress, especially during seed germination and seedling morphogenesis, which limits maize growth and productivity formation. As a novel recognized plant hormone, melatonin (MT) participates in multiple growth and developmental processes and mediates biotic/abiotic stress responses, yet the effects of salt stress on maize seedlings remain unclear. Herein, we investigated the effects of 150 μM exogenous MT on multiple phenotypes and physiologic metabolisms in three-leaf seedlings across eight maize inbred lines under 180 mM NaCl salt stress, including growth parameters, stomatal morphology, photosynthetic metabolisms, antioxidant enzyme activities, and reactive oxygen species (ROS). Meanwhile, the six gene expression levels controlling antioxidant enzyme activities and photosynthetic pigment biosynthesis in two materials with contrasting salt resistance were examined for all treatments to explore the possible molecular mechanism of exogenous MT alleviating salt injury in maize. The results showed that 150 μM exogenous MT application protected membrane integrity and reduced ROS accumulation by activating the antioxidant system in leaves of maize seedlings under salt stress, their relative conductivity and H2O2 level average reduced by 20.91% and 17.22%, while the activities of superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), and ascorbate peroxidase (APX) averaged increased by 13.90%, 17.02%, 22.00%, and 14.24% relative to salt stress alone. The improvement of stomatal size and the deposition of photosynthetic pigments were more favorable to enhancing photosynthesis in leaves when these seedlings treated with MT application under salt stress, their stomatal size, chlorophyll content, and net photosynthetic rate averaged increased by 11.60%, 19.64%, and 27.62%. Additionally, Gene expression analysis showed that MT stimulation significantly increased the expression of antioxidant enzyme genes (Zm00001d009990, Zm00001d047479, Zm00001d014848, and Zm00001d007234) and photosynthetic pigment biosynthesis genes (Zm00001d011819 and Zm00001d017766) under salt stress. At the same time, 150 μM MT significantly promoted seedling growth and biomass accumulation. In conclusion, our study may unravel crucial evidence of the role of MT in maize seedlings against salt stress, which can provide a novel strategy for improving maize salt stress resistance.

Association of Angiotensin Converting Enzyme (insertion\deletion) and Angiotensin II Type 1 Receptor (A1166C) gene polymorphisms with diabetic nephropathy in Iraqi type 2 diabetic patients

Renin-angiotensin-aldosterone system abnormalities are the most prevalent cause of renal hemodynamic abnormalities, and candidate genes in this system are involved in the etiology of diabetic nephropathy (DN). A polymorphism in the angiotensin converting enzyme (ACE) gene I(insertion)\D(deletion) has been correlated to plasma ACE levels. Furthermore, the Angiotensin II Type 1 Receptor AGT1R (A1166C) expression pattern is highly related to nephropathy. The objectives of this study involved evaluating the frequency of the ACE (I/D) and AGT1R (A1166C) gene polymorphisms and investigating the association of these polymorphism with the development of DN in Iraqi patients with Type 2 diabetes mellitus (T2DM) and evaluating the levels of several urinary and serum markers in relation to studied polymorphisms. This is a cross-sectional study that included 161 T2DM patients whom were divided into two groups: T2DM with DN(included 98 patients) and normalbuminuric T2DM (included 63 patients). ACE gene polymorphism analysis revealed that the D allele was far more prevalent in DN patients compared to normalbuminuric patients (60.2% vs. 50.8%), while the I allele frequency was 39.8% in DN and 49.2% in normoalbuminuric patients. In addition, DN patients carrying the DD genotype had higher serum kidney injury molecule 1 (KIM1), serum cystatin C (CysC), and HbA1C and lower glomerular filtration rate (GFR) compared to ID+II genotypes. For AGT1R (A1166C), both DN and normalbuminuric patients had a comparable high prevalence of the AA genotype, followed by the AC genotype, while the CC genotype had been seen only in 3 patients. In conclusion, among the studied T2DM patients, individuals with DD genotype had higher frequency of DN and had 2-fold risk for DN compared to II genotype which had the lowest risk for DN. Also the current study shows that the A1166C polymorphisms of AGT1R distribution frequency were similar for both study groups with higher frequency for A allele compared to C allele and not associated with risk of DN in Iraqi T2DM Keywords: Angiotensin Converting Enzyme (I\D) gene polymorphism, Angiotensin II Type 1 Receptor (A166C) gene polymorphisms, Cystatin C, Diabetic nephropathy, Kidney injury molecule1.

Abstract C047: Autophagy-mediated proline-supply is important for cell survival in pancreatic cancers

Abstract Oncogenic KRAS mutations present in 90–95% of pancreatic cancer patients. KRAS mutation in pancreatic cancer increased the dependency on glutamine and consumption of proline. Proline is a non-essential amino acid but it is important for the synthesis of proteins such as collagen, and also for precursors of antioxidant molecules. Several enzymes such as P5CS(ALDH18A1),PYCR1 and PYCR2 are involving proline synthesis. The effects of proline synthesis enzymes on pancreatic cancer cells are relatively less studied than that of enzymes catalyzing proline. Here, we examined the impact of proline synthesis enzymes on pancreatic cancer cell survival and proliferation. We first examined that exogenous proline promotes cell survival under low glutamine conditions in various pancreatic cancer cells. Exogenous proline supplementation increased some pancreatic cell survival even when glutamine was not sufficiently available, whereas proline supplement is dispensible for certain cell types of pancreatic cancer. Next, to investigate the importance of proline synthesis enzymes in cell survival, we measured the cell proliferation of pancreatic cancer with knock-down the proline synthesis enzymes. The knockdown of ALDH18A1 and PYCR1,2, which are proline biosynthesis enzyme genes, reduced cell survival in pancreatic cancer cells, suggesting that in PDAC cells, the enzymes that synthesize proline from glutamine are crucial for cancer cell survival and growth. To test the effect of proline-sysnthesis enzymes on autophagy, we measured autophagy flux in the cell harboring GFP-RFP-LC3 or GFP-LC3 as a reporter. Knockdown of proline synthesis enzymes, particularly ALDH18A1 or PYCR1 led to an increase in autophagy levels, indicating that proline-independent cells can bypass proline synthesis by autophagy induction in the absence of proline synthesis genes. Furthermore gemcitabine-resistant cells showed reduced levels of proline synthesis enzymes. In pancreatic cancer cell lines, proline promotes cancer cell survival, generated by both biosynthesis from glutamine and starvation-induced autophagy, which also critically important for supporting cellular drug resistance. Taken together, these results indicate that proline synthesis is mechanistically associated with autophagy induced by glutamine-starvation, which is pivotal for supplying proline in pancreatic cancer cells. Citation Format: Ji hyeon Kim, Heesun Cheong. Autophagy-mediated proline-supply is important for cell survival in pancreatic cancers [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr C047.

Enhanced indigenous bacteria-mediated bioremediation of aged petroleum-contaminated soil by Fenton pretreatment: Synergistic effect of soil environmental improvement and microbial community response

Due to the harsh soil environment and low microbial activity, the efficacy of indigenous bacteria-mediated bioremediation is often limited in aged petroleum-contaminated soils. In this study, Fenton pretreatment (H2O2: 200 ∼ 600 mmol/kg, H2O2: Fe2+ = 100: 1) was applied to improve the aged soil environment and rebuild the microbial community to enhance subsequent bioremediation. After pretreated by 400 mmol/kg H2O2 and 4 mmol/kg Fe2+ (T2), the removal efficiency of total petroleum hydrocarbons (TPHs) was elevated to 93.27 %, equivalent to 1.96-fold of the natural attenuation (CK). Additionally, the half-life of biodegradation was shorted to 45.57 d, representing a decrease to 0.29-fold that of CK. Correlation analysis revealed a radar plot comprising five indicators, including bacterial quantification (BQ), dehydrogenase (DHA), polyphenol oxidase (PPO), permeability coefficient (K), and bulk density coefficient (γ), all of which exhibited positive correlations with TPHs degradation. Compared to CK, Fenton pretreatment increased the richness and evenness of the microbial community during incubation, thus facilitating TPHs biodegradation. Moreover, Fenton pretreatment accelerated the shift of the dominant genus in T2 from Pseudomonas (0.599) to Pseudomonas (0.258), Nocardioides (0.109), Dietzia (0.157), and Stenotrophomonas (0.120) throughout the incubation. Based on the analysis of Kyoto Encyclopedia of Genes and Genomes (KEGG), the metabolism and enzymatic functional genes related to the biodegradation of petroleum hydrocarbons in T2 were enhanced during incubation, accounting for the high removal efficiency of TPHs. These findings demonstrated that moderate Fenton pretreatment was an effective approach to stimulate indigenous bacteria for the enhanced biodegradation of aged petroleum-contaminated soil.

356 Exploring the manganese requirement of feedlot steers

Abstract Angus-cross steers [n = 144; body weight (BW) = 359 kg ± 13.4] were used to assess the effect of dietary Mn and steroidal implants on trace mineral (TM) status, and liver and serum metabolites related to N metabolism. Steers (n = 6/pen) were stratified by BW in a 3 × 2 factorial. GrowSafe bunks recorded individual feed intake (experimental unit = steer; n = 24/treatment). Dietary treatments included (MANG; 8 pens per treatment; Mn as MnSO4): 1) no supplemental Mn (analyzed 14 mg Mn/kg DM; Mn0); 2) 20 mg supplemental Mn/kg DM (Mn20); 3) 50 mg supplemental Mn/kg DM (Mn50). Within MANG, steers received a steroidal implant treatment (IMP) on d 0: 1) no implant; NO; or 2) combination implant (Revalor-200; REV). Liver biopsies for TM analysis and qPCR, and blood for serum urea-N (SUN) analysis were collected on d 0, 20, 40, and 77. Liver TM, serum metabolite, enzyme activity, and gene expression data were analyzed as repeated measures using PROC MIXED of SAS 9.4. The Correlation Procedure of SAS 9.4 was utilized to assess correlations between liver arginase activity, SUN concentrations, and liver Mn concentrations. No MANG × IMP effects were noted (P ≥ 0.12) for growth or carcass measures. Steroidal implants increased final BW (FBW), overall average daily gain (ADG), and gain to feed (G:F; P ≤ 0.01). Dietary Mn did not influence FBW, overall ADG, or overall G:F (P ≥ 0.14). Liver Mn concentration increased with supplemental Mn (P = 0.01). An IMP × DAY effect was noted for liver Mn (P = 0.01) where NO and REV were similar on d 0 but NO cattle increased liver Mn from d 0 to 20 while REV liver Mn decreased. Relative expression of liver Mn transporter ZnT10 tended to be least (MANG; P = 0.10) in Mn20 while Mn0 and Mn100 were similar. Relative expression of liver Mn transporter ZIP8 was not influenced by MANG, IMP, or MANG × IMP (P ≥ 0.23). Liver arginase activity was not influenced by MANG, IMP, MANG × DAY, IMP × DAY, or MANG × IMP (P ≥ 0.15) but tended to decrease over time (DAY; P = 0.10). On d 0, 20, and 40, arginase activity was correlated (r ≤ 0.33; P ≤ 0.01) with SUN and liver Mn. Relative expression of MnSOD in the liver was greater in REV (P = 0.03) compared with NO and within a MANG × IMP effect (P = 0.01) REV increased liver MnSOD activity. This study indicates current NASEM Mn recommendations are adequate to meet demands of finishing beef cattle given a steroidal implant. This study also reveals how implants and supplemental Mn influence liver enzyme activity and genes related to arginine metabolism, urea synthesis, antioxidant capacity, and TM homeostasis.

Effect of creatine administration on locomotor activity and stress response in olive flounder (Paralichthys olivaceus)

The creatine kinase system is crucial for maintaining cellular energy homeostasis and plays a role in regulating locomotor behavior in organisms, but its significance in the regulating the motionless behavior in olive flounder is limited. In the first experiment of this study, elevated levels of creatine kinase (CK) activity in the spinal cord were detected in the juvenile group (JG) flounder compared to the adult group (AG) flounder. In the second experiment, to further confirm the involvement of CK in the locomotor behavior, the adult flounder was given an intraperitoneal injection of creatine (150 mg/kg), while the flounder in the control group received a saline solution. After one week post-injection, the behavioral analysis revealed that the flounder in the creatine-treated group displayed higher levels of locomotor activity and a greater number of escape attempts in response to external stimuli when compared to the control group. However, the acute stress response, induced by intraperitoneal injection and characterized by tail beating, was significantly alleviated in the flounder in the creatine-treated group. Additionally, there was an upregulation of the UII and AchR genes in the spinal cord, as well as increased levels of UII and AchR in the muscle tissues of the creatine-treated flounder. However, a reduction in UI mRNA levels was observed in the brain of the flounder. Collectively, our data provide the evidence that the elevated enzyme activity and gene expression of creatine kinase play important roles in off-bottom swimming behavior in the JG flounder. Furthermore, administration of creatine improved the locomotor activity and alleviated the stress response in flounder, which is associated with regulation of the locomotor- and stress-related gene in the brain, spinal cord, and muscle.

Regulatory mechanism of antioxidant enzymes on microbial metabolism and NADH in anaerobic fermentation of food waste for hydrogen production

Anaerobic fermentation is frequently hampered by toxicity arising from oxidative stress, and antioxidant enzymes play a crucial role in combating oxidative stress. In this study, the mechanism of microbial consortium and metabolic pathways regulated by antioxidant enzyme genes in anaerobic fermentation with different pH values was revealed. The results showed that antioxidant enzyme genes, such as glutathione peroxidase, peroxidase, and superoxide dismutase, were 5 times, 3 times, and 2 times higher at pH 7 than at pH 5, respectively. This allowed Clostridium to effectively resist oxidative stress, with substrate metabolism dominated by glycolysis, leading to an increase in the NADH/NAD+ ratio. Furthermore, the relative abundance of hydrogenase and electron transfer efficiency increased by 4.5 times and 2.71 times, respectively, resulting in a hydrogen production of 21.48 L/L. When antioxidant enzyme genes were inhibited at pH 5, the substrate mainly produced biomolecules for combating oxidative stress through the pentose phosphate and glycerophospholipid pathways, and led to the transformation of dominant genus into Lactobacillus. With an increased lactate dehydrogenase activity of 5.34 times, the final lactate production reached 65.17 g/L, which was 19.69 times higher than at pH 7. These results elucidate the regulatory mechanism of pH mediated antioxidant enzyme genes on hydrogen production and improve the controllability of anaerobic fermentation.

Integrated Metabolome, Transcriptome and Long Non-Coding RNA Analysis Reveals Potential Molecular Mechanisms of Sweet Cherry Fruit Ripening.

Long non-coding RNAs (lncRNAs), a class of important regulatory factors for many biological processes in plants, have received much attention in recent years. To explore the molecular roles of lncRNAs in sweet cherry fruit ripening, we conducted widely targeted metabolome, transcriptome and lncRNA analyses of sweet cherry fruit at three ripening stages (yellow stage, pink stage, and dark red stage). The results show that the ripening of sweet cherry fruit involves substantial metabolic changes, and the rapid accumulation of anthocyanins (cyanidin 3-rutinoside, cyanidin 3-O-galactoside, and cyanidin 3-O-glucoside) is the main cause of fruit coloration. These ripening-related alterations in the metabolic profile are driven by specific enzyme genes related to the synthesis and decomposition of abscisic acid (ABA), cell wall disintegration, and anthocyanin biosynthesis, as well as transcription factor genes, such as MYBs, bHLHs, and WD40s. LncRNAs can target these ripening-related genes to form regulatory modules, incorporated into the sweet cherry fruit ripening regulatory network. Our study reveals that the lncRNA-mRNA module is an important component of the sweet cherry fruit ripening regulatory network. During sweet cherry fruit ripening, the differential expression of lncRNAs will meditate the spatio-temporal specific expression of ripening-related target genes (encoding enzymes and transcription factors related to ABA metabolism, cell wall metabolism and anthocyanin metabolism), thus driving fruit ripening.

Methanogenic degradation of long-chain fatty acids associated with syntrophic microbial dynamics during thermophilic AnMBR treatment of lipid-rich dairy wastes

Anaerobic membrane reactors (AnMBRs) have been applied to effectively treat dairy wastes because they can address the drawbacks of sludge flotation and biomass washout. However, when dairy wastes contain considerable proportion of lipids, any anaerobic systems will likely encounter challenges with long-chain fatty acid (LCFA) inhibition. In this study, a thermophilic AnMBR was operated to treat dairy processing wastewater (DPW) plus ice cream waste (ICW). To evaluate the impact of LCFAs, the lipid-rich ICW was gradually introduced at different levels (0, 5, 10, and 20 %) into the AnMBR, along with a stepwise increase in lipid loading rate (LLR) from 0.22 to 11.24 g lipid/L/d. The results of biomethanation revealed that the AnMBR could be successfully operated when the LLR was maintained at 5.73 g lipid/L/d with a lipid/VS ratio of 50.22 % in the feedstock. However, at ultra-high LLR of 11.24 g lipid/L/d (20 % ICW added), a significant inhibitory effect on methane yields was observed, resulting in a reduction of 56.90 %. Dynamic analysis of the LCFAs profiled that a lower degradation rate was an important factor contributing to LCFA accumulation, and the inhibition concentration of various LCFAs was identified. Based on microbial evolution and co-occurrence network results, potential syntrophic partnerships between LCFA-degrading bacteria and methanogens were proposed. The responsible enzyme genes in the LCFA-degrading, acetoclastic, and hydrogenotrophic methanogenic pathways played crucial roles in LCFA inhibition and degradation. These findings offer practical insights for the efficient methanogenic treatment of lipid-rich wastes.

Retrospective analysis of arginase 1 deficiency progression in adults over 5 years at a single metabolic centre

AbstractBackgroundThe clinical presentation of ARG1‐D is characterised by elevated arginine levels leading to neurological and mobility impairments. Information about long‐term outcomes in adults is lacking, which prompted us to undertake a retrospective observational study.MethodsWe extracted ARG1‐D patient data spanning a 5‐year period from electronic health records. Ethical approval was not required for the study. Informed consent was obtained.ResultsWe identified nine ARG1‐D patients from consanguineous backgrounds. Age of symptom onset ranged from infancy to age 7 years, age of diagnosis from infancy to 20 years. Patients had paraparesis or altered gait of varying degree and had experienced early ARG1‐D onset. Over 5 years, mobility declined in six (6/9, 67%) patients. Three patients (3/9, 33%) were fully dependent and hoisted. Two (2/9, 22%) reached adulthood before experiencing hyperammonaemia, another one (1/9, 11%) first experienced hyperammonaemia at age 15 years. One patient (1/9, 11%) started on ammonia scavenger therapy in adulthood, one (1/9, 11%) required a second scavenger to be added to their treatment regimen. Two patients (2/9, 22%) had gastrostomy tubes inserted for nutrition and supplements at age 9 years and 15 years. Six patients (6/9, 67%) had raised levels of ALT; of these, four (4/9, 44%) also had elevated AFP. Heterogeneity of ARG1‐D symptoms was evident, suggesting complex genetic and environmental interactions.ConclusionARG1‐D presents significant lifelong challenges with deteriorating mobility and more frequent metabolic crises. Current management strategies are insufficient for preventing progression, highlighting the need for innovative treatments like enzyme replacement and gene therapy.

TRNAVal allows four-way decoding with unmodified uridine at the wobble position in Lactobacillus casei.

Modifications at the wobble position (position 34) of tRNA facilitate interactions that enable or stabilize non-Watson-Crick basepairs. In bacterial tRNA, 5-hydroxyuridine (ho5U) derivatives xo5U [x: methyl (mo5U), carboxymethyl (cmo5U), and methoxycarbonylmethyl (mcmo5U)] present at the wobble positions of tRNAs are responsible for recognition of NYN codon families. These modifications of U34 allow basepairing not only with A and G but also with U and in some cases C. mo5U was originally found in Gram-positive bacteria, and cmo5U and mcmo5U were found in Gram-negative bacteria. tRNAs of Mycoplasma species, mitochondria, and chloroplasts adopt four-way decoding in which unmodified U34 recognizes codons ending in A, G, C, and U. Lactobacillus casei, Gram-positive bacteria and lactic acid bacteria, lacks the modification enzyme genes for xo5U biosynthesis. Nevertheless, L. casei has only one type of tRNAVal with the anticodon UAC [tRNAVal(UAC)]. However, the genome of L. casei encodes an undetermined tRNA (tRNAUnd) gene, and the sequence corresponding to the anticodon region is GAC. Here, we confirm that U34 in L. casei tRNAVal is unmodified and that there is no tRNAUnd expression in the cells. In addition, in vitro transcribed tRNAUnd was not aminoacylated by L. casei valyl-tRNA synthetase suggesting that tRNAUnd is not able to accept valine, even if expressed in cells. Correspondingly, native tRNAVal(UAC) with unmodified U34 bound to all four valine codons in the ribosome A site. This suggests that L. casei tRNAVal decodes all valine codons by four-way decoding, similarly to tRNAs from Mycoplasma species, mitochondria, and chloroplasts.

Isolation and characterization of koji mold (Aspergillus oryzae) from nature in Niigata

Koji mold (Aspergillus oryzae) is a key microorganism in brewing and fermentation in Japan. We isolated koji molds from the environment in Niigata Prefecture. Eighty-one environmental samples were placed on isolation medium made from steamed rice with wood ash and 36 Aspergillus section Flavi-like strains were obtained. Of those, 26 strains did not produce aflatoxin. We studied their morphology, sequence of ITS region, calmodulin gene, aflatoxin biosynthetic homologous gene cluster and α-amylase gene and fermentation-related enzyme activities. Furthermore, DNA-seq analysis of 14 strains from 26 non-aflatoxin producing strains were conducted and compared the three mycotoxin biosynthetic gene clusters (aflatoxin, cyclopiazonic acid, and aflatrem) and fermentation-related genes against those of reference strain A.oryzae RIB40. In some strains, gene sequences confirmed the absence of mycotoxin production, but differences in fermentation-related enzyme activities could not be explained well by amino acid substitutions. We classified the 26 isolates into 6 morphology types based on the appearance of colonies and mating types, and it was found that strains of the same morphology type had similar enzymatic profiles and gene sequences. Our results show that koji molds with various properties occur in the environment, and it will expand the possibilities of koji mold in industrial use.