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Abstract Bovine leukemia virus (BLV) is the etiological agent of enzootic bovine leukosis and causes a persistent infection that can leave cattle with no symptoms. Many countries have been able to successfully eradicate BLV through improved detection and management methods. However, with the increasing novel molecular detection methods there have been few efforts to standardize these results at global scale. This study aimed to determine the interlaboratory accuracy and agreement of 11 molecular tests in detecting BLV. Each qPCR/ddPCR method varied by target gene, primer design, DNA input and chemistries. DNA samples were extracted from blood of BLV-seropositive cattle and lyophilized to grant a better preservation during shipping to all participants around the globe. Twenty nine out of 44 samples were correctly identified by the 11 labs and all methods exhibited a diagnostic sensitivity between 74 and 100%. Agreement amongst different assays was linked to BLV copy numbers present in samples and the characteristics of each assay (i.e., BLV target sequence). Finally, the mean correlation value for all assays was within the range of strong correlation. This study highlights the importance of continuous need for standardization and harmonization amongst assays and the different participants. The results underscore the need of an international calibrator to estimate the efficiency (standard curve) of the different assays and improve quantitation accuracy. Additionally, this will inform future participants about the variability associated with emerging chemistries, methods, and technologies used to study BLV. Altogether, by improving tests performance worldwide it will positively aid in the eradication efforts.

To identify a biomarker for the early diagnosis of enzootic bovine leukosis (EBL) caused by bovine leukemia virus (BLV), we investigated the expression of a microRNA, bta-miR-375, in cattle serum. Using quantitative reverse-transcriptase PCR analysis, we measured bta-miR-375 levels in 27 samples from cattle with EBL (EBL cattle), 45 samples from animals infected with BLV but showing no clinical signs (NS cattle), and 30 samples from cattle uninfected with BLV (BLV negative cattle). In this study, we also compared the kinetics of bta-miR-375 with those of the conventional biomarkers of proviral load (PVL), lactate dehydrogenase (LDH), and thymidine kinase (TK) from the no-clinical-sign phase until EBL onset in three BLV-infected Japanese black (JB) cattle. Bta-miR-375 expression was higher in NS cattle than in BLV negative cattle (P < 0.05) and greater in EBL cattle than in BLV negative and NS cattle (P < 0.0001 for both comparisons). Receiver operating characteristic curves demonstrated that bta-miR-375 levels distinguished EBL cattle from NS cattle with high sensitivity and specificity. In NS cattle, bta-miR-375 expression was increased as early as at 2 months before EBL onset-earlier than the expression of PVL, TK, or LDH isoenzymes 2 and 3. These results suggest that serum miR-375 is a promising biomarker for the early diagnosis of EBL.

Fascioliasis is a neglected tropical zoonotic disease caused by liver flukes belonging to the genus Fasciola. The emergence of resistance to triclabendazole, the only World Health Organization-recommended drug for this disease, highlights the need for the development of new drugs. Helminths possess an anaerobic mitochondrial respiratory chain (fumarate respiration) which is considered a potential drug target. This study aimed to evaluate the occurrence of fumarate respiration in Fasciola flukes. We analyzed the properties of the respiratory chain of Fasciola flukes in both adults and newly excysted juveniles (NEJs). Fasciola flukes travel and mature through the stomach, bowel, and abdominal cavity to the liver, where oxygen levels gradually decline. High fumarate reductase activity was observed in the mitochondrial fraction of adult Fasciola flukes. Furthermore, rhodoquinone-10 (RQ10 Em’= −63 mV), a low-potential electron mediator used in fumarate respiration was found to be predominant in adults. In contrast, the activity of oxygen respiration was low in adults. Rotenone, atpenin A5, and ascochlorin, typical inhibitors of mitochondrial enzymes in complexes I, II, and III, respectively, inhibit the activity of each enzyme in the adult mitochondrial fraction. These inhibitors were then used for in vitro viability tests of NEJs. Under aerobic conditions, NEJs were killed by rotenone or ascochlorin, which inhibit aerobic respiration (complex I–III), whereas atpenin A5, which inhibits complex II involved in fumarate respiration, did not affect NEJs. Moreover, ubiquinone-10 (UQ10 Em’= +110 mV), which is used in oxidative respiration, was detected in NEJs, in addition to RQ10. In contrast, under anaerobic conditions, rotenone and atpenin A5, which inhibit fumarate respiration (complex I–II), were crucial for NEJs. These findings demonstrate that NEJs have active hybrid respiration, in which they can properly use both oxygen and fumarate respiration, depending on oxygen availability. Thus, fumarate respiration is a promising drug target for Fasciola flukes, because it plays an essential role in both adults and NEJs.

A degradative solvent extraction method for upgrading low-rank coal was performed at 200–350 °C for 90 min to obtain a substance dissolved in the solvent at room temperature (Soluble). Because the resulting mixture exhibited a high carbon content without ash, it could be readily used as a fuel. Furthermore, deep eutectic solvents (DESs) have attracted attention for improving the Soluble yield and decreasing the oxygen content in Soluble. DES is known to cleave oxygen-containing functional groups in biomass and is considered effective for deoxidizing low-rank coal. Herein, DES was prepared by mixing choline chloride, FeCl3·6H2O, and Adaro subbituminous coal (AD) and then added to 1-methylnaphthalene (1-MN) in a non-polar solvent, followed by degradative solvent extraction in the range of 200–350 °C. The effects of reaction temperature and added DES amount on the product yield and the composition were evaluated. As the reaction temperature and amount of DES added increased, the Soluble yield and carbon content increased. It was also found that the thermal decomposition temperature and oxygen content decreased with the increasing DES amounts. This decrease indicates that DES promotes the deoxygenation and decomposition of AD and increases the soluble yield of the fuel source.

ABSTRACT Bovine leukemia virus (BLV), a retrovirus, infects B cells of ruminants and is integrated into the host genome as a provirus for lifelong infection. After a long latent period, 1% to 5% of BLV-infected cattle develop aggressive lymphoma, enzootic bovine leukosis (EBL). Since the clonal expansion of BLV-infected cells is essential for the development of EBL, the clonality of proviral integration sites could be a molecular marker for diagnosis and early prediction of EBL. Recently, we developed Rapid Amplification of the Integration Site without Interference by Genomic DNA Contamination (RAISING) and an analysis software of clonality value (CLOVA) to analyze the clonality of transgene-integrated cells. RAISING-CLOVA is capable of assessing the risk of adult T-cell leukemia/lymphoma development in human T-cell leukemia virus-I-infected individuals through the clonality analysis of proviral integration sites. Thus, we herein examined the performance of RAISING-CLOVA for the clonality analysis of BLV-infected cells and conducted a comprehensive clonality analysis by RAISING-CLOVA in EBL and non-EBL cattle. RAISING-CLOVA targeting BLV was a highly accurate and reproducible method for measuring the clonality value. The comprehensive clonality analysis successfully distinguished EBL from non-EBL specimens with high sensitivity and specificity. A longitudinal clonality analysis in BLV-infected sheep, an experimental model of lymphoma, also confirmed the effectiveness of RAISING-CLOVA for early detection of EBL development. Therefore, our study emphasizes the usefulness of RAISING-CLOVA as a routine clinical test for monitoring virus-related cancers. IMPORTANCE Bovine leukemia virus (BLV) infection causes aggressive B-cell lymphoma in cattle and sheep. The virus has spread to farms around the world, causing significant economic damage to the livestock industry. Thus, the identification of high-risk asymptomatic cattle before they develop lymphoma can be effective in reducing the economic damage. Clonal expansion of BLV-infected cells is a promising marker for the development of lymphoma. Recently, we have developed a high-throughput method to amplify random integration sites of transgenes in host genomes and analyze their clonality, named as RAISING-CLOVA. As a new application of our technology, in this study, we demonstrate the value of the RAISING-CLOVA method for the diagnosis and early prediction of lymphoma development by BLV infection in cattle. RAISING-CLOVA is a reliable technology for monitoring the clonality of BLV-infected cells and would contribute to reduce the economic losses by EBL development.

Substrate channeling can improve biosynthetic efficiency and has been implicated in the reactions of fusicoccadiene synthase. Here, the authors analyze this bifunctional enzyme complex by cryoEM, cross-linking MS and integrative modeling, providing structural insights into how substrate channeling is achieved.

The conserved eukaryotic nucleotide excision repair (NER) pathway protects the genome from a wide variety of environmentally induced DNA lesions. Here, the authors provide insights into how NER is initiated on lesions by determining the cryo-EM structure of the yeast TFIIH/Rad4–Rad23-Rad33 complex bound to a DNA containing a single carcinogen-DNA adduct.

Respiratory chains generate the proton motive force used for ATP synthesis. Cryo-EM structures of functional respiratory CIII2CIV supercomplex and native CIII2 from Rhodobacter capsulatus provide insight into CIII2CIV assembly and respiratory electron transport pathways in Gram-negative bacteria.

Rough-surfaced nanocone structures are preferred for use as transparent conductive oxides due to their high optical transparency and electrical conductivity. Structural properties of Ga-doped ZnO terrace-truncated nanocones, which were grown by advanced spray pyrolysis deposition technique, vastly changes with the spraying angle. In the present study, the effect of the spray angle on terrace-truncated nanocone structure formation was investigated. Spray pyrolysis deposition technique was used to grow the nanostructure as the growth rate can be controlled easily. The prepared samples were characterized using X-ray diffraction spectroscopy (XRD) and scanning electron microscopy (SEM) techniques. Optical and electrical properties were investigated by the UV-visible spectrum and four-probe method. The lowest spray angle of 15º showed homogeneous and hexagonal shaped nanocone structure with an average top diameter of 22.8 nm and an average height of 240 nm. An excellent transparent conductive oxide behavior was obtained from the sample synthesized at the lowest spray angle of 15o with high conductivity of 2.5×103 ?-1 .cm-1 and high transparency of 82% in the visible range.

A novel measurement system consisting of scanning electron microscopy and infrared (IR) thermography has been constructed to measure the thermal conductivity and the thermal diffusivity on a micro-/nanometer scale. In this study, we measured them for a stainless-steel wire with a diameter of 50 μm to demonstrate the validity of our system. The time evolution of temperature distribution on the whole of the stainless-steel wire was recorded by the IR thermograph when the wire was irradiated by the electron beam (EB). A time response of the wire temperature corresponding to the on/off of the EB irradiation was clearly observed, indicating that local EB heating was realized. In addition, the experimental time evolution of temperature at its transient phenomena was in good agreement with the theoretical calculation based on a one-dimensional (1D) equivalent-circuit model of heat flow coupled with electric current, where the thermal parameters, such as thermal conductivity and heat capacitance, were set to bulk values. Therefore, it can be concluded that our system is valid to evaluate the thermal conductivity of the 1D wire sample. For evaluating the thermal diffusivity of the wire sample, we applied a periodic EB irradiation by a rectangular wave with a frequency of 0.25 Hz. The thermal diffusivity was evaluated to be (2.8 ± 0.6) × 10−5 m2 s−1, which was seven times as large as that of bulk stainless steel. Our system can be applied to an AC calorimetry method for a 1D system sample after the analysis procedure is further modified.

In this study, vertically aligned zinc oxide (ZnO) nanorod (NR) arrays were grown with the 2-step method. Spin coating was used to apply the seed layer, and by increasing the number of repetitions (n), higher thickness of the seed layer was achieved. The effects of different seed layer thicknesses, spin coated at different rotational speeds (v) and how the variables (v and n) influence the morphological and crystal properties of the resulting ZnO NRs were analyzed. The effects of 1, 3, and 5 (n), with (v) of 3000 or 4000 rpm were investigated, and the formed seed layers were characterized by using scanning electron microscopy (SEM) and X-ray diffraction (XRD) profiles. The ZnO NR arrays were grown by a chemical bath deposition method because of its simplicity and ease of use. Full width at half maximum (FWHM) analysis was conducted on the XRD chart and Raman spectrum to analyze the grain size and Wurtzite distortion of the ZnO NRs. A lower FWHM value was attributed to the lower number of structural disorders in the ZnO NRs, which equated to the most stable or structured Wurtzite structures in the ZnO NRs, achieved at the lowest FWHM value. The lowest FWHM for (002) XRD peak was obtained at 0.35° for ZnO NRs grown with a seed layer spin coated at 4000 rpm once. The crystallite size calculated by Scherrer’s equation gave a size of 25.14 nm. The lowest FWHM for E2(High) Raman peak was obtained at 6.00 cm–1 for ZnO NRs grown with a seed layer spin coated at 4000 rpm three times.

Abstract Background Bovine leukemia virus (BLV) is the causative agent of enzootic bovine leukosis (EBL). The incidence of EBL in Japan is increasing annually; and the cases of EBL in cattle younger than 2 years old has been reported. Therefore, it is vital to find a method to control BLV infection, especially in young calves. In this study, to evaluate the protective ability of colostral antibodies against BLV infection, as well as the potential for BLV infection mediated by colostrum/milk, we investigated temporal fluctuations in the anti-BLV antibody titer and BLV proviral load (PVL) in colostrum/milk and peripheral blood of six infected dams during lactation. The association between PVL and antibody titer in colostrum and peripheral blood was then investigated using samples from a further twenty-seven cattle. Antibody concentrations were measured with a Syncytium-induction Inhibition Assay using colostral/milk whey and serum. PVL in peripheral blood and colostrum was measured by real-time PCR. Results Colostral antibodies showed high inhibitory activity until day 3 of lactation. The antibody titer and PVL in peripheral blood showed lesser changes than those in colostrum/milk throughout lactation. The colostral antibody titer was significantly higher than the serum antibody titer in all samples, whereas the colostrum PVL was significantly lower than the blood PVL. The blood PVL showed a significant correlation with serum antibody titer, colostrum PVL, and colostral antibody titer. However, there were no major correlations between the serum and colostral antibody titers. Conclusions This is the first report investigating the temporal changes in colostral antibody titer in terms of inhibiting BLV infection in vitro. The results of antibody detection by Syncytium-induction Inhibition Assay suggested that the protective activity of the colostral antibodies against BLV infection would be conferred by anti-BLV gp51 antibody. The high antibody titer of colostral whey suggests that colostral whey could be a potential source of antibodies with a low risk of infection in neonatal calves.

Bovine leukemia virus (BLV) is the causative agent of enzootic bovine leukosis (EBL), a malignant B cell lymphoma. However, the mechanisms of BLV-associated lymphomagenesis remain poorly understood. Here, after deep sequencing, we performed comparative analyses of B cell microRNAs (miRNAs) in cattle infected with BLV and those without BLV. In BLV-infected cattle, BLV-derived miRNAs (blv-miRNAs) accounted for 38% of all miRNAs in B cells. Four of these blv-miRNAs (blv-miR-B1-5p, blv-miR-B2-5p, blv-miR-B4-3p, and blv-miR-B5-5p) had highly significant positive correlations with BLV proviral load (PVL). The read counts of 90 host-derived miRNAs (bta-miRNAs) were significantly down-regulated in BLV-infected cattle compared to those in uninfected cattle. Only bta-miR-375 had a positive correlation with PVL in BLV-infected cattle and was highly expressed in the B cell lymphoma tissue of EBL cattle. There were a few bta-miRNAs that correlated with BLV tax/rex gene expression; however, BLV AS1 expression had a significant negative correlation with many of the down-regulated bta-miRNAs that are important for tumor development and/or tumor suppression. These results suggest that BLV promotes lymphomagenesis via AS1 and blv-miRNAs, rather than tax/rex, by down-regulating the expression of bta-miRNAs that have a tumor-suppressing function, and this downregulation is linked to increased PVL.

Bovine leukemia virus (BLV) is an oncogenic virus belonging to the genus Deltaretrovirus and is the causative agent of enzootic bovine leukosis. Proviral load (PVL) determined by real-time quantitative PCR (qPCR) is now widely used as an indicator of not only BLV infection, but also BLV disease progression. To interpret PVLs determined by different qPCRs used in Japan, we compared a chimeric cycling probe-based qPCR, CY415, targeting the BLV tax region; a TaqMan probe-based qPCR, RC202, targeting the BLV pol region; and a TaqMan probe-based qPCR, CoCoMo, targeting the BLV long terminal repeat (LTR) region. Whole-blood samples collected from 317 naturally BLV-infected cattle (165 Holstein–Friesian and 152 Japanese Black) and tumor tissue samples collected from 32 cattle at a meat inspection center were used. The PVLs determined by each qPCR were strongly correlated. However, the PVL and the proportion of BLV-infected cells determined by RC202 or CoCoMo were significantly higher than those determined by CY415. Genetic analysis of three tumor tissue samples revealed that LTR region mutations or a deletion affected the PVL determined by CoCoMo. These results suggest that the TaqMan-based RC202 or CoCoMo qPCR is better than CY415 for BLV PVL analysis. However, qPCR target region mutations were not rare in tumors and could hamper PVL analysis by using qPCR.

We aimed to evaluate choroid structural changes using swept-source optical coherence tomography (SS-OCT) following hemodialysis initiation in diabetic and nondiabetic patients with end-stage kidney disease (ESKD). In this multicenter, prospective, cross-sectional study, diabetic (DM group; 30 eyes; 16 patients) and nondiabetic patients (NDM group; 30 eyes; 15 patients) with ESKD were evaluated after hemodialysis initiation. SS-OCT findings were analyzed using a manual delineation technique and binarization method before the first and last hemodialysis sessions, conducted approximately 2 weeks apart. Subfoveal choroidal thickness changes and mean large choroidal vessel layer thickness were significantly greater in the DM group (-13.3% ± 2.5% and -14.5% ± 5.2%, respectively) than the NDM group (-9.5% ± 3.1% and -9.2% ± 3.4%, respectively; p = 0.049 and p = 0.02, respectively). Binarized SS-OCT analysis revealed that the mean subfoveal choroidal area was significantly larger in the DM group (-21.9% ± 6.5%) than the NDM group (-17.2% ± 5.9%; p = 0.032). The change ratio in mean luminal area values was significantly greater in the DM group (-27.7% ± 8.7%) than the NDM group (-17.7% ± 5.8%; p = 0.007). The DM group exhibited substantial changes in the choroidal layer, possibly reflecting choroidal vascular disorders caused by diabetes.

Abstract This study investigated the steam gasification of Indonesian Adaro coal and Japanese cedar mixed in a 1:1 ratio with the physical addition of 10 wt% Fe2O3 in 50 vol% H2O at ambient atmospheric pressure and 800°C for 1 h. The primary objective of this work was to examine the effectiveness of an iron catalyst on the interaction between Indonesian Adaro coal and Japanese cedar. The study demonstrated that the H2 evolution amount for co‐gasification of Japanese cedar and Adaro coal (with 1:1 ratio in weight) without a Fe catalyst was 100 mmol/g‐char. However, the H2 evolution amount for co‐gasification with the addition of Fe2O3 was 152 mmol/g‐char. The increase in the co‐gasification for H2 evolution was based on a change in the char structure during pyrolysis and gasification.

Different morphologies of zinc oxide (ZnO) can be obtained through various synthesizing methods, such as that of the water bath. By synthesizing under various conditions, different ZnO morphologies can be seen as the result of the water bath method. Replacing ZnO nanoparticles with vertically aligned ZnO nanorods results in a much higher energy conversion efficiency. Yet vertically aligned nanorods can only be grown through difficult and expensive methods. Several researchers have studied the growth of one-dimensional (1D) nanorods on homogeneous film with various growth conditions. However, there has been little in the way of research on ZnO nanorods grown on ZnO seed layers using the water bath method. In this research, vertically aligned nanorods with an optimum size ratio were formed through a simple water bath method. This method reveals that the ZnO nanorods are well aligned and grown with a high density and uniformity on the substrate. Their X-ray diffraction patterns reveal that the nanorods are grow in the [001] direction. The density, diameter, and length of the ZnO nanorods can be altered by changing the growing condition. All of the samples were characterized using a scanning electron microscope, X-ray diffraction, and micro Raman spectroscopy. To investigate crystal growth, zinc nitrate and zinc acetate were used when preparing the solution. The results demonstrate that the morphology and alignment of ZnO nanorods are determined by the precursor’s type and deposition time.

Arrays of zinc oxide (ZnO) nanorod (NR) were fabricated in a vertical axis direction through the two-step method of seed layer’s deposition and growth of the NR. The seed layer was applied by spin coating with a three-time repetition (n) and rotational speed (v) at 3000 rpm. After the seed layer had grown, ZnO NRs were grown with a growth solution made by combining one zinc source with one hydroxide source. There were two different zinc sources, i.e., zinc acetate dehydrate and zinc nitrate hexahydrate and, for comparison, zinc acetate (ZA) and zinc nitrate (ZN) were each combined with the same hydroxide source, hexamethylenetetramine (HMT). Later, the growth solutions were processed by the chemical bath deposition (CBD) method using a waterbath machine. The CBD method was started at room temperature until it reached the designated temperature at 85°C. At that point, the growth time was calculated from the zero-minute condition. It was found that ZnO NRs had already grown at a thickness of about 100 nm for both ZA and ZN sources. The growth time varied at 15, 60, 90, and 120 minutes after the zero-minute point. By using two separate and independent zinc sources while growing ZnO NRs at various growth periods, several ZnO NRs’ thicknesses were controlled. According to a paper by Lee et al., the lower thickness of ZnO NRs boosted the charge transfer properties of perovskite solar cells (PSCs) because the series resistance between ZnO/perovskite interfaces was lessened. Scanning electron microscopy (SEM) images were observed to analyze the morphological shape of the ZnO NRs. X-ray diffraction (XRD) profiles were characterized to obtain the data for ZnO NR crystallinity. Full width at half maximum (FWHM) analysis was performed at the (002) ZnO peak to calculate the crystal size of the peak. From the results, the smallest crystallite sizes for ZnO NRs grown from ZA and ZN sources were 10.70 nm and 19.29 nm, respectively, which would be the most suitable condition for PSC application.

Cattle are major reservoirs of the provisionally named influenza D virus, which is potentially involved in the bovine respiratory disease complex. Here, we conducted a serological survey for the influenza D virus in Japan, using archived bovine serum samples collected during 2010-2016 from several herds of apparently healthy cattle in various regions of the country. We found sero-positive cattle across all years and in all the prefectural regions tested, with a total positivity rate of 30.5%, although the positivity rates varied among regions (13.5-50.0%). There was no significant difference in positivity rates for Holstein and Japanese Black cattle. Positivity rates tended to increase with cattle age. The herds were clearly divided into two groups: those with a high positive rate and those with a low (or no) positive rate, indicating that horizontal transmission of the virus occurs readily within a herd. These data demonstrate that bovine influenza D viruses have been in circulation for at least 5 years countrywide, emphasizing its ubiquitous distribution in the cattle population of Japan.

A 36-year-old woman who had been diagnosed with ulcerative colitis at the age of 17 years was referred to our hospital because of severe abdominal pain and repeated bloody diarrhea that persisted during pregnancy despite combination therapy with high-dose corticosteroids and weekly granulocyte and monocyte adsorptive apheresis (GMA). She underwent combination therapy consisting of high-dose corticosteroids, intensive GMA (two sessions per week) and vancomycin, which was used to eradicate Clostridium difficile, under total parenteral nutrition control until the estimated weight of her fetus reached 1,000 g. This combination therapy was partially successful, resulting in almost complete disappearance of abdominal pain and a marked decrease in stool frequency. However bloody diarrhea persisted and the patient developed anemia and hypoalbuminemia and was unable to prolong her gestation time. Cesarean section was conducted at 28 weeks of gestation without any congenital abnormalities or neurological defects. Oral administration of tacrolimus was begun 7 days after cesarean section, which was followed by rapid induction of remission. Corticosteroids were then gradually tapered off. Tacrolimus is one therapeutic option after cesarean section in pregnant patients who do not respond well to GMA and high-dose corticosteroids for persistent active ulcerative colitis.

The aims of this study were to automate irrigation in a conventional, bottom-irrigated cabbage seedling bed and to record the irrigation with a volumetric water content sensor on a water-absorbing mat. Two crops were grown in summer, with healthy seedling establishment rates of 91.3% and 88.6%. Growth was normal in the first crop, in which the soil moisture content was always maintained above 20% during germination, but some plants died in the second crop, in which the content dropped to 0% at night owing to excessive drought. Standard practice was to irrigate the beds from sunrise to 10:00 to -35% water content and then let the soil dry out in the afternoon. Irrigation was also applied in the afternoon on hot days and just before planting. In the second crop, plant death was attributed to insufficient irrigation frequency under increased evaporation due to high temperatures, low humidity, and a high humidity deficit. These results make it clear that simply setting a certain number of irrigations per day is insufficient to account for changes in environment. Instead, it is effective to set a target moisture content for individual time periods and for irrigation to be applied as needed when moisture content is insufficient. [ABSTRACT FROM AUTHOR]

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