Aberrant proximal tubule DNA methylation underlies phenotypic changes related to kidney dysfunction in patients with diabetes.

Epigenetic mechanisms are considered to contribute to diabetic nephropathy by maintaining memory of poor glycemic control during the early stages of diabetes. However, DNA methylation changes in the human kidney are poorly characterized, because of the lack of cell type-specific analysis. We examined DNA methylation in proximal tubules (PTs) purified from patients with diabetic nephropathy and identified differentially methylated CpG sites, given the critical role of proximal tubules in the kidney injury. Hypermethylation was observed at CpG sites annotated to genes responsible for proximal tubule functions, including gluconeogenesis, nicotinamide adenine dinucleotide synthesis, transporters of glucose, water, phosphate, and drugs, in diabetic kidneys, whereas genes involved in oxidative stress and the cytoskeleton exhibited demethylation. Methylation levels of CpG sites annotated to ACTN1, BCAR1, MYH9, UBE4B, AFMID, TRAF2, TXNIP, FOXO3, and HNF4A were correlated with the estimated glomerular filtration rate, whereas methylation of the CpG site in RUNX1 was associated with interstitial fibrosis and tubular atrophy. Hypermethylation of G6PC and HNF4A was accompanied by decreased expression in diabetic kidneys. Proximal tubule-specific hypomethylation of metabolic genes related to HNF4A observed in control kidneys was compromised in diabetic kidneys, suggesting a role for aberrant DNA methylation in the dedifferentiation process. Multiple genes with aberrant DNA methylation in diabetes overlapped genes with altered expressions in maladaptive proximal tubule cells, including transcription factors PPARA and RREB1. In conclusion, DNA methylation derangement in the proximal tubules of patients with diabetes may drive phenotypic changes, characterized by inflammatory and fibrotic features, along with impaired function in metabolism and transport.NEW & NOTEWORTHY Cell type-specific DNA methylation patterns in the human kidney are not known. We examined DNA methylation in proximal tubules of patients with diabetic nephropathy and revealed that oxidative stress, cytoskeleton, and metabolism genes were aberrantly methylated. The results indicate that aberrant DNA methylation in proximal tubules underlies kidney dysfunction in diabetic nephropathy. Aberrant methylation could be a target for reversing memory of poor glycemic control.

Highly Efficient Blue Fluorescent Organic Light-Emitting Devices Based on λ5-Phosphinine Derivatives.

Although λ5-phosphinine derivatives are known as a promising class of blue fluorescent emitters, those photoluminescent quantum yield (PLQY) values have been reached up to 92 %, however, only a few examples have been explored as an emitter for blue organic light-emitting device (OLED), and the external quantum efficiency (EQE) has been below 2.4 % so far. In this study, we newly developed two types of blue λ5-phosphinine derivatives namely CN-COCF3 and CO2Me-CHO, and investigated the photophysical properties in the solid states. The photophysical analyses in solid state films suggested that the strong electron-accepting nature of these λ5-phosphinine derivatives caused the inferior PLQY values, and the exciplex formation with the host and neighboring materials should be avoided to improve the device efficiency. By choosing suitable host and neighboring materials with deep ionization potentials, we successfully realized efficient blue fluorescent OLEDs with EQE of over 4 % and CIE (0.14, 0.18). This is among the best in λ5-phosphinine-based blue OLEDs so far.

Diastereodivergent and Regiodivergent Total Synthesis of Princepin and Isoprincepin in Both (7″R,8″R) and (7″S,8″S) Isomers.

The first total synthesis of (7″R,8″R)-, (7″S,8″S)-isomers of princepin (1) and (7″R,8″R)-, (7″S,8″S)-isomers of isoprincepin (2) was accomplished in a highly stereoselective manner via para quinomethide-mediated construction of the furofuran and 1,4-benzodioxane rings. Structural confirmation methods of 1 and 2 were established by CD and HPLC analysis of each diastereomers with natural products.

Effect of Target Gene Silencing on Calcite Single Crystal Formation by Thermophilic Bacterium Geobacillus thermoglucosidasius NY05.

Geobacillus thermoglucosidasius NY05 catalyzes calcite single crystal formation at 60 °C by using acetate and calcium. Endospores are embedded at the central part of the calcite single crystal and carbon atoms in the calcite lattice are derived from acetate carbon. Here, we synthesized 21-mer antisense DNA oligonucleotides targeting sporulation transcription factor, acetate-CoA ligase, isocitrate lyase, and malate synthase G mRNAs and evaluated the effect of these oligonucleotides on calcite formation in G. thermoglucosidasius NY05. G. thermoglucosidasius NY05 cells containing antisense DNA oligonucleotides targeting sporulation transcription factor, acetate-CoA ligase, isocitrate lyase, and malate synthase G mRNAs had reduced calcite single crystal formation by 18.7, 50.6, 55.7, and 82.3%, respectively, compared with cells without antisense DNA oligonucleotides. These results support that calcite formation needs endospores as the nucleus to grow, and carbon dioxide generated from acetate, which is metabolized via the glyoxylate pathway and glucogenesis, is supplied to the crystal lattice.

Allexiviruses may have acquired inserted sequences between the CP and CRP genes to change the translation reinitiation strategy of CRP.

Allexiviruses are economically important garlic viruses that are involved in garlic mosaic diseases. In this study, we characterized the allexivirus cysteine-rich protein (CRP) gene located just downstream of the coat protein (CP) gene in the viral genome. We determined the nucleotide sequences of the CP and CRP genes from numerous allexivirus isolates and performed a phylogenetic analysis. According to the resulting phylogenetic tree, we found that allexiviruses were clearly divided into two major groups (group I and group II) based on the sequences of the CP and CRP genes. In addition, the allexiviruses in group II had distinct sequences just before the CRP gene, while group I isolates did not. The inserted sequence between the CP and CRP genes was partially complementary to garlic 18S rRNA. Using a potato virus X vector, we showed that the CRPs affected viral accumulation and symptom induction in Nicotiana benthamiana, suggesting that the allexivirus CRP is a pathogenicity determinant. We assume that the inserted sequences before the CRP gene may have been generated during viral evolution to alter the termination-reinitiation mechanism for coupled translation of CP and CRP.

Suvorexant is associated with a low incidence of delirium in critically ill patients: a retrospective cohort study.

BACKGROUND: Benzodiazepine use is a risk factor for the development of delirium in adult intensive care unit (ICU) patients. Suvorexant is an alternative to benzodiazepines to induce sleep, but the incidence of delirium in critically ill patients is unknown. We undertook this retrospective study to investigate the incidence of delirium in patients who receive suvorexant in the ICU. METHODS: This retrospective cohort study was conducted in a closed 12-bed ICU at a tertiary teaching hospital. Patients admitted to the ICU for 72 h or longer between January and June 2015 were evaluated for delirium using the Confusion Assessment Method for the Intensive Care Unit tool. We evaluated the incidence of delirium in patients who received suvorexant and those who did not. To adjust for confounding factors, multivariable logistic regression analysis was conducted. RESULTS: Study subjects included 118 patients, with a median age of 72 years and a median Acute Physiology and Chronic Health Evaluation II score of 18 points. Eighty-two patients (69.5%) were admitted after cardiovascular surgery. In the suvorexant group, there were fewer post-cardiovascular surgical patients and more medical patients. The duration of mechanical ventilation during ICU stay was longer in the suvorexant group, and sedatives and sleep inducers other than suvorexant were used more frequently in the suvorexant group. The incidence of delirium was 43.8% in the suvorexant group and 58.8% in the non-suvorexant group (P = 0.149). After adjustment for risk factors using multivariable logistic regression analysis, suvorexant was associated with a lower incidence of delirium (odds ratio = 0.23, 95% confidence interval: 0.07-0.73; P = 0.012). CONCLUSIONS: Suvorexant was associated with decreased odds of transitioning to delirium in critically ill patients. The use of suvorexant may lower the incidence of delirium in ICU patients. Future prospective studies are warranted.

Magnesium-Calcite Crystal Formation Mediated by the Thermophilic Bacterium Geobacillus thermoglucosidasius Requires Calcium and Endospores.

Fresh Geobacillus thermoglucosidasius cells grown on soybean-casein digest nutrient agar were inoculated as a parent colony 1 cm in diameter on the surface of an agar gel containing acetate and calcium ions (calcite-promoting hydrogel) and incubated at 60 °C for 4 days, after which magnesium-calcite single crystals of 50-130 µm in size formed within the parent colony. Addition of EDTA, polyacrylic acid or N,N-dicyclohexylcarbodiimide to the calcite-forming hydrogel inhibited the parent colony from forming magnesium-calcite crystals. Inoculation of G. thermoglucosidasius on calcite-forming hydrogel containing 5 µM cadmium and 20 µM zinc resulted in a decrease in the sporulation rate from 55 to 7-8 %. Magnesium-calcite synthesis decreased relative to the sporulation rate. G. thermoglucosidasius exhibited higher adsorption/absorbance of calcium than other Geobacillus sp. that do not mediate calcite formation and higher levels of magnesium accumulation. Calcium ions contained in the calcite-promoting hydrogel and magnesium ions concentrated in G. thermoglucosidasius cells serve as the elements for magnesium-calcite synthesis. The observed decreases in sporulation rate and magnesium-calcite formation support the hypothesis that endospores act as nuclei for the synthesis of magnesium-calcite single crystals.

Emergence of integrated behaviors through direct optimization for homeostasis.

Homeostasis is a self-regulatory process, wherein an organism maintains a specific internal physiological state. Homeostatic reinforcement learning (RL) is a framework recently proposed in computational neuroscience to explain animal behavior. Homeostatic RL organizes the behaviors of autonomous embodied agents according to the demands of the internal dynamics of their bodies, coupled with the external environment. Thus, it provides a basis for real-world autonomous agents, such as robots, to continually acquire and learn integrated behaviors for survival. However, prior studies have generally explored problems pertaining to limited size, as the agent must handle observations of such coupled dynamics. To overcome this restriction, we developed an advanced method to realize scaled-up homeostatic RL using deep RL. Furthermore, several rewards for homeostasis have been proposed in the literature. We identified that the reward definition that uses the difference in drive function yields the best results. We created two benchmark environments for homeostasis and performed a behavioral analysis. The analysis showed that the trained agents in each environment changed their behavior based on their internal physiological states. Finally, we extended our method to address vision using deep convolutional neural networks. The analysis of a trained agent revealed that it has visual saliency rooted in the survival environment and internal representations resulting from multimodal input.

Geobacillus thermoglucosidasius endospores function as nuclei for the formation of single calcite crystals.

Geobacillus thermoglucosidasius colonies were placed on an agar hydrogel containing acetate, calcium ions, and magnesium ions, resulting in the formation of single calcite crystals (calcites) within and peripheral to the plating area or parent colony. Microscopic observation of purified calcites placed on the surface of soybean casein digest (SCD) nutrient medium revealed interior crevices from which bacterial colonies originated. Calcites formed on the gel contained [1-(13)C]- and [2-(13)C]acetate, demonstrating that G. thermoglucosidasius utilizes carbon derived from acetate for calcite formation. During calcite formation, vegetative cells swam away from the parent colony in the hydrogel. Hard-agar hydrogel inhibited the formation of calcites peripheral to the parent colony. The calcite dissolved completely in 1 M HCl, with production of bubbles, and the remaining endospore-like particles were easily stained with Brilliant green dye. The presence of DNA and protein in calcites was demonstrated by electrophoresis. We propose that endospores initiate the nucleation of calcites. Endospores of G. thermoglucosidasius remain alive and encapsulated in calcites.

Effect of spo0A, sigE, sigG, and sigK disruption on butanol production and spore formation in Clostridium saccharoperbutylacetonicum strain N1-4 (ATCC13564).

Clostridium saccharoperbutylacetonicum strain N1-4 (ATCC13564) is a butanol-producing strain suitable for application to butanol production from cellulosic materials by co-culture with cellulolytic and thermophilic species, such as Hungateiclostridium thermocellum (synonym: Clostridium thermocellum). The optimal temperature for butanol production by strain N1-4 is 30 °C, and the strain is sensitive to a high culture temperature of 37 °C. Given that spore formation is observed at high frequency when strain N1-4 is cultivated at 37 °C, we assumed in a previous study that the initiation of sporulation is related to a decrease in butanol production. Therefore, to investigate the relationship between butanol production and spore formation, we generated strain N1-4 isolates in which genes related to spore formation were disrupted. The sporulation-related gene disruptants of spo0A, sigE, sigG, and sigK lost the ability to produce heat-resistant spores, irrespective of the culture temperature. Among the gene disruptants produced, only the spo0A disruptant lost butanol-producing ability when cultivated at 30 °C. Interestingly, the sigE disruptant maintained butanol productivity similar to that observed at 30 °C, even when cultivated at 37 °C. In addition, the sigE disruptant successfully produced butanol from Avicel cellulose by co-culture with H. thermocellum at a fermentation temperature of 37 °C.

Effect of Rj genotype and cultivation temperature on the community structure of soybean-nodulating bradyrhizobia.

The nodulation tendency and community structure of indigenous bradyrhizobia on Rj genotype soybean cultivars at cultivation temperatures of 33/28°C, 28/23°C, and 23/18°C for 16/8 h (day/night degrees, hours) were investigated using 780 bradyrhizobial DNA samples from an Andosol with 13 soybean cultivars of four Rj genotypes (non-Rj, Rj(2)Rj(3), Rj(4), and Rj(2)Rj(3)Rj(4)). A dendrogram was constructed based on restriction fragment length polymorphism of the PCR products (PCR-RFLP) of the 16S-23S rRNA gene internal transcribed spacer region. Eleven Bradyrhizobium U.S. Department of Agriculture strains were used as a reference. The dendrogram indicated seven clusters based on similarities among the reference strains. The occupancy rate of the Bj123 cluster decreased with increasing cultivation temperature, whereas the occupancy rates of the Bj110 cluster, Be76 cluster, and Be94 cluster increased with increasing cultivation temperature. In particular, the Rj(2)Rj(3)Rj(4) genotype soybeans were infected with a number of Bj110 clusters, regardless of the increasing cultivation temperature, compared to other Rj genotype soybean cultivars. The ratio of beta diversity to gamma diversity (H'(ß)/H'(γ)), which represents differences in the bradyrhizobial communities by pairwise comparison among cultivation temperature sets within the same soybean cultivar, indicated that the bradyrhizobial communities tended to be different among cultivation temperatures. Multidimensional scaling analysis indicated that the infection of the Bj110 cluster and the Bj123 cluster by host soybean genotype and the cultivation temperature affected the bradyrhizobial communities. These results suggested that the Rj genotypes and cultivation temperatures affected the nodulation tendency and community structures of soybean-nodulating bradyrhizobia.

Variability in the P1 gene helps to refine phylogenetic relationships among leek yellow stripe virus isolates from garlic.

Nucleotide sequences from the P1 gene and the 5' untranslated region of leek yellow stripe virus (LYSV), collected from several locations, were used to refine the phylogenetic relationships among the isolates. Multiple alignments revealed three distinct regions of insertions and deletions to classify LYSVs. In our phylogenetic analyses, the LYSV isolates separated into two major groups (N-type and S-type). S-type viruses had two large deletions compared to N-type viruses. Considering that the outgroup, onion yellow dwarf virus (OYDV) also has the sequences corresponding to the deletions in the S-type viruses, our study shows that the sequences missing in the S-type were present in the common ancestor of the N-type and S-type. In the phylogenetic trees, we found three distinct clades of isolates, from Uruguay (U), Okinawa (O) and Spain (Sp), suggesting that LYSVs have unique evolutionary histories depending on their garlic origins. The P1 gene of LYSV is thus quite suited to reflecting viral evolution, as recently suggested for other potyviruses.

Isolation of ammonia gas-tolerant extremophilic bacteria and their application to the elimination of malodorous gas emitted from outdoor heat-treated toilets.

Ammonia gas-tolerant extremophilic bacteria capable of growing in atmospheres containing up to 4000 ppm of gaseous ammonia were isolated. These bacteria were capable of growing in nutrient broth containing high concentrations of ammonia water, with growth in medium augmented with 0.1-0.2% ammonia exceeding that in medium without ammonia. The minimal inhibitory concentration of ammonia in the medium was 0.5%. The isolated ammonia gas-tolerant bacterium was moderately alkaliphilic, with optimum growth was observed at pH 9. DNA sequence analysis of the 16SrRNA gene revealed that the isolated bacterium was Bacillus lentus. Furthermore, extremophilic bacteria cultured in a 1300 ppm ammonia gas atmosphere on agar medium containing no nitrogen sources were observed to use ammonia gas for growth. These bacteria were identified as Paenibacillus lentus and Bacillus altitudinis based on 16SrRNA gene sequence analysis. The deodorizing effect of ammonia odor by the isolated bacteria immobilized on sawdust was evaluated. The findings showed that forcing ammonia gas through a column containing B. altitudinis immobilized on sawdust reduced the concentration of ammonia gas by 30% compared to columns containing sawdust only. The isolated bacteria immobilized on wood sawdust lost the capacity deodorization after drying, but this function could be restored with increased moisture. The ammonia gas-tolerant extremophilic bacteria immobilized on sawdust show considerable potential for use in ameliorating malodors associated with outdoor heat-treated toilets.

Identification of a Novel Quinvirus in the Family Betaflexiviridae That Infects Winter Wheat.

Yellow mosaic disease in winter wheat is usually attributed to the infection by bymoviruses or furoviruses; however, there is still limited information on whether other viral agents are also associated with this disease. To investigate the wheat viromes associated with yellow mosaic disease, we carried out de novo RNA sequencing (RNA-seq) analyses of symptomatic and asymptomatic wheat-leaf samples obtained from a field in Hokkaido, Japan, in 2018 and 2019. The analyses revealed the infection by a novel betaflexivirus, which tentatively named wheat virus Q (WVQ), together with wheat yellow mosaic virus (WYMV, a bymovirus) and northern cereal mosaic virus (a cytorhabdovirus). Basic local alignment search tool (BLAST) analyses showed that the WVQ strains (of which there are at least three) were related to the members of the genus Foveavirus in the subfamily Quinvirinae (family Betaflexiviridae). In the phylogenetic tree, they form a clade distant from that of the foveaviruses, suggesting that WVQ is a member of a novel genus in the Quinvirinae. Laboratory tests confirmed that WVQ, like WYMV, is potentially transmitted through the soil to wheat plants. WVQ was also found to infect rye plants grown in the same field. Moreover, WVQ-derived small interfering RNAs accumulated in the infected wheat plants, indicating that WVQ infection induces antiviral RNA silencing responses. Given its common coexistence with WYMV, the impact of WVQ infection on yellow mosaic disease in the field warrants detailed investigation.

Catalytic biomineralization of fluorescent calcite by the thermophilic bacterium Geobacillus thermoglucosidasius.

The thermophilic Geobacillus bacterium catalyzed the formation of 100-µm hexagonal crystals at 60°C in a hydrogel containing sodium acetate, calcium chloride, and magnesium sulfate. Under fluorescence microscopy, crystals fluoresced upon excitation at 365 ± 5, 480 ± 20, or 545 ± 15 nm. X-ray diffraction indicated that the crystals were magnesium-calcite in calcite-type calcium carbonate.

Loop-Mediated Isothermal Amplification Screening for COVID-19 in Asymptomatic Preoperative Orthopedic Patients in a General Hospital in Kanagawa, Japan.

The global coronavirus disease 2019 (COVID-19) pandemic has caused several million infections and hundreds of thousands of deaths. A large number of healthcare workers have died as a result of infection with this virus. Therefore, elective surgery was markedly reduced or stopped in our hospital's orthopedic department. The detection of asymptomatic COVID-19-positive patients became key to reducing the infection risk to physicians and staff to allow orthopedic surgery to be performed. A total of 21 patients were scheduled to undergo orthopedic surgery, including elective surgery, in Shonantobu General Hospital, Chigasaki City, Kanagawa, Japan. All 21 patients gave permission to undergo loop-mediated isothermal amplification (LAMP) screening the day before surgery. None of the 21 patients we tested was positive for COVID-19. All patients remained asymptomatic during the two to four weeks of postoperative follow-up. No physicians or medical staff developed COVID-19 symptoms. This was a very small study in a city with a relatively low incidence of COVID-19. We found that LAMP screening was accurate, in terms of its negative predictive value. Larger studies are needed.

Tylosin degradation during manure composting and the effect of the degradation byproducts on the growth of green algae.

In this study, we investigate the tylosin degradation in sheep feces during composting. The sheep feces containing tylosin were composted using the laboratory-scale composting units. Tylosin was degraded during composting, and the half-life of tylosin degradation decreased with increasing temperature from 40 °C to 65 °C. The tylosin degradation in sheep feces can be attributed to the microorganisms in the feces and not to heating because tylosin did not degrade over a period of 48 h at temperatures of 0 °C-65 °C in sterilized water. The artificial rainwater solution extracted from the composted sample did not inhibit the growth of Raphidocelis subcapitata, a type of green alga. Our results indicate that composting the feces containing tylosin is effective in degrading tylosin, which may result in the preservation of agricultural fields as well as nearby aquatic environments.

Plasmid uptake by bacteria: a comparison of methods and efficiencies.

The ability to introduce individual molecules of plasmid DNA into cells by transformation has been of central importance to the recent rapid advancement of plasmid biology and to the development of DNA cloning methods. Molecular genetic manipulation of bacteria requires the development of plasmid-mediated transformation systems that include (1) chemical transformation, (2) electro-transformation, (3) biolistic transformation, and (4) sonic transformation, leading to the introduction of exogenous plasmid DNA into bacterial cells. In this review, the manipulation properties and transformation efficiencies of these techniques are described. In addition to these methods, a conceptually novel transformation technique, namely the hydrogel exposure method, was developed. The hydrogel exposure method, based on the Yoshida effect, provides a significant advance over chemical means for transforming many strains of Escherichia coli and a variety of other bacterial species. The new term "tribos transformation" has been proposed for this novel technique. We also determined that, compared to conventional methods, the hydrogel exposure method is a novel and convenient method by which to transform bacteria.

Electrically Conductive Gels Prepared from Syndiotactic Polystyrene and an Ionic Liquid.

Syndiotactic polystyrene (SPS) physical gels containing a large amount of the ionic liquid 1-butylpyridinium bromide ([C4py]Br) were systematically prepared and their physical properties were examined in detail. The gels had stable forms for a long time, having storage elastic modulus values of normal gels. They showed nearly the same values of the electrical conductivity (∼7 mS/cm) as those of the mixed solutions of [C4py]Br, suggesting that the distribution of [C4py]Br was uniform in these gels and that the charge transportation in these SPS gels was not interrupted by a three-dimensional network of SPS fibrils consisting of the SPS δ crystalline phases.