A Comprehensive Study of the Immunophenotype and its Clinicopathologic Significance in Adult T-Cell Leukemia/Lymphoma.

Adult T-cell leukemia/lymphoma (ATLL) is a mature T-cell tumor caused by human T-lymphotropic virus type 1 (HTLV-1). The typical ATLL immunophenotypes are described in the 2017 World Health Organization Classification of Tumours of Haematopoietic and Lymphoid Tissues (positive: CD2, CD3, CD5, CD4, and CD25; negative: CD7, CD8, and cytotoxic markers; and partially positive: CD30, CCR4, and FOXP3). However, limited studies are available on the expression of these markers, and their mutual relationship remains unknown. Furthermore, the expression status of novel markers associated with T-cell lymphomas, including Th1 markers (T-bet and CXCR3), Th2 markers (GATA3 and CCR4), T follicular helper markers (BCL6, PD1, and ICOS), and T-cell receptor (TCR) markers, and their clinicopathologic significance is unclear. In this study, we performed >20 immunohistochemical stains in 117 ATLL cases to determine the comprehensive immunophenotypic profile of ATLL, which were compared on the basis of clinicopathologic factors, including morphologic variants (pleomorphic vs anaplastic), biopsy locations, treatments, Shimoyama classification-based clinical subtype, and overall survival. CD3+/CD4+/CD25+/CCR4+ was considered a typical immunophenotype of ATLL, but approximately 20% of cases did not conform to this pattern. Simultaneously, the following new findings were obtained: (1) most cases were negative for TCR-ß and TCR-δ (104 cases, 88.9%), indicating the usefulness of negative conversion of TCR expression to provide differentiation from other T-cell tumors; (2) the positivity of CD30 and CD15 and the negativity of FOXP3 and CD3 were significantly associated with anaplastic morphology; and (3) atypical cases, such as T follicular helper marker-positive (12 cases, 10.3%) and cytotoxic molecule-positive cases (3 cases, 2.6%), were identified. No single markers could predict the overall survival among patients with acute/lymphoma subtypes of ATLL. The results of this study illustrate the diversity of ATLL phenotypes. In T-cell tumors occurring in HTLV-1 carriers, the possibility of ATLL should not be eliminated even when the tumor exhibits an atypical phenotype, and the confirmation of HTLV-1 in the tissue is recommended.

Electrostatic Adsorption Behaviors of Polymer Plates to a Droplet.

Electrostatic transfer and adsorption of electrically conductive polymer-coated poly(ethylene terephthalate) plates from a particle bed to a water droplet were studied, with the influence of plate thickness and shape observed. After synthesis and confirmation of the particles' properties using stereo and scanning electron microscopies, elemental microanalysis, and water contact angle measurement, the electric field strength and droplet-bed separation distance required for transfer were measured. An electrometer and high-speed video footage were used to measure the charge transferred by each particle, and its orientation and adsorption behavior during transfer and at the droplet interface. The use of plates of consistent square cross section allowed the impact of contact-area-dependent particle cohesion and gravity on the electrostatic transfer of particles to be decoupled for the first time. The electrostatic force required to extract a plate was directly proportional to the plate mass (thickness), a trend very different from that previously observed for spherical particles of varied diameter (mass). This reflected the different relationship between mass, surface area, and cohesive forces for spherical and plate-shaped particles of different sizes. Thicker plates transferred more charge to the droplet, probably due to their remaining at the bed at higher field strengths. The impact of plate cross-sectional geometry was also assessed. Differences in the ease of transfer of square, hexagonal, and circular plates seemed to depend only on their mass, while other aspects of their comparative behavior are attributed to the more concentrated charge distribution present on particles with sharper vertices.

Genetic profile of adult T-cell leukemia/lymphoma in Okinawa: Association with prognosis, ethnicity, and HTLV-1 strains.

Genetic alterations in adult T-cell leukemia/lymphoma (ATLL), a T-cell malignancy associated with HTLV-1, and their clinical impacts, especially from the perspective of viral strains, are not fully elucidated. We employed targeted next-generation sequencing and single nucleotide polymorphism array for 89 patients with ATLL in Okinawa, the southernmost islands in Japan, where the frequency of HTLV-1 tax subgroup-A (HTLV-1-taxA) is notably higher than that in mainland Japan, where most ATLL cases have HTLV-1-taxB, and compared the results with previously reported genomic landscapes of ATLL in mainland Japan and the USA. Okinawan patients exhibited similar mutation profiles to mainland Japanese patients, with frequent alterations in TCR/NF-ĸB (eg, PRKCB, PLCG1, and CARD11) and T-cell trafficking pathways (CCR4 and CCR7), in contrast with North American patients who exhibited a predominance of epigenome-associated gene mutations. Some mutations, especially GATA3 and RHOA, were detected more frequently in Okinawan patients than in mainland Japanese patients. Compared to HTLV-1-taxB, HTLV-1-taxA was significantly dominant in Okinawan patients with these mutations (GATA3, 34.1% vs 14.6%, P = .044; RHOA, 24.4% vs 6.3%, P = .032), suggesting the contribution of viral strains to these mutation frequencies. From a clinical viewpoint, we identified a significant negative impact of biallelic inactivation of PRDM1 (P = .027) in addition to the previously reported PRKCB mutations, indicating the importance of integrated genetic analysis. This study suggests that heterogeneous genetic abnormalities in ATLL depend on the viral strain as well as on the ethnic background. This warrants the need to develop therapeutic interventions considering regional characteristics.

A new diagnostic algorithm using biopsy specimens in adult T-cell leukemia/lymphoma: combination of RNA in situ hybridization and quantitative PCR for HTLV-1.

Histopathological distinction between adult T-cell leukemia/lymphoma (ATLL) and other T-cell neoplasms is often challenging. The current gold standard for the accurate diagnosis of ATLL is the Southern blot hybridization (SBH) assay, which detects clonal integration of human T-cell leukemia virus type I (HTLV-1) provirus. However, SBH cannot be performed with small biopsy or formalin-fixed paraffin-embedded (FFPE) tissue samples because this assay requires a large amount of DNA without degradation. Here we developed a new diagnostic algorithm for the accurate diagnosis of ATLL using FFPE samples. This method combines two HTLV-1 detection assays, namely, ultrasensitive RNA in situ hybridization using RNAscope for HTLV-1 bZIP factor (HBZ-RNAscope), and quantitative PCR targeting the tax gene (tax-qPCR). We analyzed 119 FFPE tissue specimens (62 ATLL, and 57 non-ATLL, including 41 HTLV-1 carriers) and compared them with the SBH results using the corresponding fresh-frozen samples. As a result, tax-qPCR had a higher ATLL identification rate than HBZ-RNAscope (88% [52/59], and 63% [39/62], respectively). However, HBZ-RNAscope clearly visualized the localization of HTLV-1-infected tumor cells and its identification rate increased to 94% (17/18) when the analysis was limited to samples up to 2 years old, indicating its usefulness in the daily diagnosis. The diagnostic algorithm combining these two assays successfully evaluated 94% (112/119) of samples and distinguished ATLL from non-ATLL cases including HTLV-1 carriers with 100% sensitivity and specificity. This method is expected to replace SBH and increase the accuracy of the diagnosis of ATLL.

Rock glaciers and related cold rocky landforms: Overlooked climate refugia for mountain biodiversity.

Mountains are global biodiversity hotspots where cold environments and their associated ecological communities are threatened by climate warming. Considerable research attention has been devoted to understanding the ecological effects of alpine glacier and snowfield recession. However, much less attention has been given to identifying climate refugia in mountain ecosystems where present-day environmental conditions will be maintained, at least in the near-term, as other habitats change. Around the world, montane communities of microbes, animals, and plants live on, adjacent to, and downstream of rock glaciers and related cold rocky landforms (CRL). These geomorphological features have been overlooked in the ecological literature despite being extremely common in mountain ranges worldwide with a propensity to support cold and stable habitats for aquatic and terrestrial biodiversity. CRLs are less responsive to atmospheric warming than alpine glaciers and snowfields due to the insulating nature and thermal inertia of their debris cover paired with their internal ventilation patterns. Thus, CRLs are likely to remain on the landscape after adjacent glaciers and snowfields have melted, thereby providing longer-term cold habitat for biodiversity living on and downstream of them. Here, we show that CRLs will likely act as key climate refugia for terrestrial and aquatic biodiversity in mountain ecosystems, offer guidelines for incorporating CRLs into conservation practices, and identify areas for future research.

First molecular detection of Hemolivia and Hepatozoon parasites in reptile-associated ticks on Iriomote Island, Japan.

Hepatozoon and Hemolivia are members of the haemogregarines and are reported in reptiles and reptile-associated ticks. However, no studies have reported on Hepatozoon and Hemolivia in Japanese reptile-associated ticks. This study aimed to molecularly identify and to characterize Hepatozoon and Hemolivia in Japanese reptile-associated ticks, Amblyomma geoemydae (Cantor, 1847) and Amblyomma nitidum (Hirst & Hirst, 1910). A total of 41 and 75 DNA samples from A. geoemydae and A. nitidum ticks, respectively, were used for screening of Hepatozoon and Hemolivia with polymerase chain reaction targeting 18S rDNA. As a result, Hemolivia and Hepatozoon were detected in two A. geoemydae and one A. nitidum, respectively. The sequences of Hemolivia spp. showed a 99.5% (1,050/1,055 bp) identity with Hemolivia parvula (KR069083), and the Hemolivia spp. were located in the same clade as H. parvula in the phylogenetic tree. The sequences of Hepatozoon sp. showed a 98.4% (1,521/1,545 bp) identity with Hepatozoon colubri (MN723844), and the Hepatozoon sp. was distinct from validated Hepatozoon species in the tree. Our findings highlight the first molecular record of Hemolivia in Japan and present the first detection of Hepatozoon in A. nitidum. Further investigations on these tick-borne protozoa are required to understand their life cycle and pathogenicity.

Allogeneic hematopoietic stem cell transplantation at the first remission for younger adults with FLT3-internal tandem duplication AML: The JALSG AML209-FLT3-SCT study.

In this phase II multicenter study (JALSG AML209-FLT3-SCT), we aimed to prospectively elucidate the role of allogeneic hematopoietic stem cell transplantation (allo-HSCT) at first complete remission (CR1) for FLT3-internal tandem duplication (ITD)-positive AML. Newly diagnosed de novo AML patients with FLT3-ITD were enrolled at the achievement of CR1 and received allo-HSCT as soon as possible after the first consolidation therapy. Mutations of 57 genes in AML cells at diagnosis were also analyzed. Among 48 eligible patients with a median age of 38.5 (17-49) years, 36 (75%) received allo-HSCT at a median of 108 days after CR1. The median follow-up was 1726 days. The primary end-point, 3-year disease-free survival (DFS) based on an intent to treat analysis, was 43.8% (95% confidence interval [CI], 30%-57%), suggesting the efficacy of this treatment because the lower limit of the 95% CI exceeded the threshold response rate of 20%. The 3-year overall survival, post-transplant DFS, and non-relapse mortality rates were 54.2% (95% CI, 39%-67%), 58.3% (95% CI, 41%-72%), and 25.0% (95% CI, 12%-40%), respectively. The median ITD allelic ratio (AR) was 0.344 (0.006-4.099). Neither FLT3-ITD AR nor cooccurring genetic alterations was associated with a poor DFS. This prospective study indicated the efficacy and safety of allo-HSCT for FLT3-ITD AML patients in CR1. This study was registered at: www.umin.ac.jp/ctr/ as #UMIN000003433.

Hydrogeological characterization of an alpine aquifer system in the Canadian Rocky Mountains.

Groundwater storage in alpine regions is essential for maintaining baseflows in mountain streams. Recent studies have shown that common alpine landforms (e.g., talus and moraine) have substantial groundwater storage capacity, but the hydrogeological connectivity between individual landforms has not been understood. This study characterizes the hydrogeology of an alpine cirque basin in the Canadian Rocky Mountains that contains typical alpine landforms (talus, meadow, moraines) and hydrological features (tarn, streams, and springs). Geological, hydrological, and hydrochemical observations were used to understand the overall hydrogeological setting of the study basin, and three different geophysical methods (electrical resistivity tomography, seismic refraction tomography, and ground penetrating radar) were used to characterize the subsurface structure and connectivity, and to develop a hydrogeological conceptual model. Geophysical imaging shows that the talus is typically 20-40 m thick and highly heterogeneous. The meadow sediments are only up to 11 m thick but are part of a 30-40-m-thick accumulation of unconsolidated material that fills a bedrock overdeepening (i.e. a closed, subglacial basin). A minor, shallow groundwater system feeds springs on the talus and streams on the meadow, whereas a deep system in the moraine supplies most of the water to the basin outlet springs, thereby serving as a 'gate keeper' of the basin. Although the hydrologic functions of the talus in this study are substantially different from other locations, primarily due to differences in bedrock lithology and geomorphic processes, the general conceptual framework developed in this study is expected to be applicable to other alpine regions.

Dans les régions alpines, le stockage des eaux souterraines est essentiel au maintien du débit de base des cours d'eau de montagne. De récentes études ont montré que les formes de relief alpin courants (par exemple le talus et la moraine) ont une capacité de stockage des eaux souterraines substantielle, mais la connectivité hydrogéologique entre les formes particulières de relief n'a pas été comprise. La présente étude caractérise l'hydrogéologie d'un bassin d'un cirque alpin des Montagnes Rocheuses Canadiennes qui présente des formes typiques de relief (talus, prairie, moraines) et des caractéristiques hydrogéologiques (petits lacs, cours d'eau et sources). Des observations géologiques, hydrologiques et hydrochimiques ont été utilisées pour comprendre le contexte hydrogéologique général du bassin étudié, et trois méthodes géophysiques différentes (tomographie par résistivité électrique, tomographie par sismique réfraction et géoradar) ont été utilisées pour caractériser la structure et la connectivité du sous-sol et pour développer un modèle conceptuel hydrogéologique. L'imagerie géophysique montre que le talus a généralement une épaisseur de 20­40 m et est fortement hétérogène. Les sédiments de la prairie n'ont que 11 m d'épaisseur mais font partie d'une accumulation épaisse de 30­40 m de matériel non consolidé, qui remplit un surcreusement profond dans le substratum (c-à-d, un bassin fermé sous-glaciaire). Un petit système aquifère phréatique nourrit des sources sur le talus et des cours d'eau sur la prairie, tandis que le système profond de la moraine fournit la plus grande partie de l'eau aux sources sortant du bassin, servant ainsi de « portier ¼ du bassin. Bien que les fonctions hydrologiques du talus soient dans cette étude substantiellement différentes de celles d'autres localisations, surtout en raison des différences de lithologie du socle et des processus géomorphologiques, on escompte que le schéma conceptuel général développé ici est applicable à d'autres régions alpines.

El almacenamiento de aguas subterráneas en las regiones alpinas es esencial para mantener los flujos de base en los cursos de agua de las montañas. Estudios recientes han demostrado que las formas de relieve alpino comunes (por ejemplo, el talud y la morena) tienen una capacidad importante de almacenamiento de aguas subterráneas, pero no se ha comprendido la interrelación hidrogeológica entre las distintas formas de relieve. Este estudio caracteriza la hidrogeología de una cuenca de circo alpino en las Rocky Mountains del Canadá que contiene las típicas formas de relieve alpino (talud, pradera, morenas) y características hidrológicas (lagunas, arroyos y manantiales). Se utilizaron observaciones geológicas, hidrológicas e hidroquímicas para comprender el contexto hidrogeológico general de la cuenca de estudio, y se emplearon tres métodos geofísicos diferentes (tomografía de resistividad eléctrica, tomografía de refracción sísmica y georradar) para caracterizar la estructura y la conectividad en el subsuelo, y para elaborar un modelo conceptual hidrogeológico. Las imágenes geofísicas muestran que el talud suele tener un espesor de 20 a 40 m y es muy heterogéneo. Los sedimentos de la pradera sólo tienen un espesor de hasta 11 m, pero forman parte de una acumulación de material no consolidado de 30­40 m de espesor que rellena un basamento sobreexcavado (es decir, una cuenca subglacial cerrada). Un sistema menor de aguas subterráneas poco profundas alimenta los manantiales en el talud y los arroyos en la pradera, mientras que un sistema profundo en la morena suministra la mayor parte del agua a los manantiales de salida de la cuenca, sirviendo, así como "portadora" de la cuenca. Aunque las funciones hidrológicas del talud en este estudio son sustancialmente diferentes de las de otros lugares, principalmente debido a las diferencias en la litología del basamento y los procesos geomórficos, se espera que el marco conceptual general desarrollado en este estudio sea aplicable a otras regiones alpinas.

O armazenamento de águas subterrâneas em regiões alpinas é essencial para manter os fluxos de base nos córregos das montanhas. Estudos recentes mostraram que formas de relevo alpinas comuns (por exemplo, tálus e morena) têm capacidade substancial de armazenamento de águas subterrâneas, mas a conectividade hidrogeológica entre formas de relevo individuais não foi compreendida. Este estudo caracteriza a hidrogeologia de uma bacia de circo alpino nas Montanhas Rochosas Canadenses que contém formas de relevo alpinas típicas (talus, prados, morenas) e características hidrológicas (tarn, córregos e nascentes). Observações geológicas, hidrológicas e hidroquímicas foram usadas para entender o cenário hidrogeológico geral da bacia em estudo e três métodos geofísicos diferentes (tomografia de resistividade elétrica, tomografia de refração sísmica e radar de penetração no solo) foram usados ​​para caracterizar a estrutura e a conectividade do subsolo, e desenvolver um modelo conceitual hidrogeológico. As imagens geofísicas mostram que o tálus tem tipicamente 20­40 m de espessura e é altamente heterogêneo. Os sedimentos dos prados têm apenas 11 m de espessura, mas fazem parte de um acúmulo de material não consolidado com 30­40 m de espessura que preenche um afundamento no leito de rocha (isto é, uma bacia subglacial fechada). Um sistema menor de águas subterrâneas rasas alimenta fontes no tálus e córregos no prado, enquanto um sistema profundo na morena fornece a maior parte da água às fontes de saída da bacia, servindo assim como um 'guardião' da bacia. Embora as funções hidrológicas do tálus neste estudo sejam substancialmente diferentes de outros locais, principalmente devido a diferenças na litologia dos leitos de rocha e nos processos geomórficos, espera-se que o quadro conceitual geral desenvolvido neste estudo seja aplicável a outras regiões alpinas.

[Determination of Tributyltin and Triphenyltin in Fish and Shellfish Using Accelerated Solvent Extraction and Liquid Chromatography with Tandem Mass Spectrometry].

A determination method for tributyltin (TBT) and triphenyltin (TPT) in fish and shellfish using an accelerated solvent extractor (ASE) and LC-MS/MS was developed. The chromatographic separation was conducted on a Poroshell 120 EC-C18 column using an isocratic mobile phase of 0.1% formic acid in 70% methanol. Sample preparation was performed using ASE at 125℃ with n-hexane and a cleanup using a Florisil cartridge. Internal calibration curves using deuterium-labeled TBT and TPT were employed for quantification. For both TBT and TPT, the calibration curves were linear in the range of 0.2-250 ng/mL, and the method quantification limits were 0.8 ng/g for both TBT and TPT. A National Institute for Environmental Studies certified reference material, No. 15 (adductor muscle of scallop), was analyzed to assess the performance of the developed method. The trueness, relative standard deviations of repeatability, and within laboratory reproducibility of this method, evaluated using a recovery test with four spiked fish species and one shellfish, ranged from 89.3 to 105.3%, 1.0 to 4.5%, and 1.3 to 7.6%, respectively.

[Improvement of Analytical Method of PCBs by GC-MS/MS Using an Accelerated Solvent Extractor and Expansion of Target Food Products].

We examined the expansion of target food products for the analysis of polychlorinated biphenyls (PCBs). From our studies in fish using an accelerated solvent extractor (ASE) and GC-MS/MS, we found that recovery of low-chlorinated PCBs (some isomer of 3 or 4 chlorinated PCBs) in specific food products was low. Therefore, we attempted to improve the analytical method. In Japanese sea perch (Suzuki) and milk, freeze drying before extracting with ASE did not sufficiently improve the recovery of low-chlorinated PCBs; however, it was significantly improved by changing the column to a silica gel/H2SO4 silica gel multilayer column. To evaluate this method for other food products, PCBs in fish, meat, eggs, milk, and dairy products were analyzed. The samples were extracted using ASE at 125℃ with n-hexane, cleaned up using a silica gel/H2SO4 silica gel multilayer column, and analyzed by GC-MS/MS for 3-7 chlorinated PCBs. The limit of quantification for this method was was 0.03-0.16 µg/kg for the 3-7 chlorinated PCBs. The recovery and the coefficient of variation of 3-7 chlorinated PCBs from each of the food products (n=5) was in the range of 84%-112% and 1%-9%, respectively.

Palladium-Catalyzed Construction of Quaternary Stereocenters by Enantioselective Arylation of γ-Lactams with Aryl Chlorides and Bromides.

Herein, we report the first Pd-catalyzed enantioselective arylation of α-substituted γ-lactams. Two sets of conditions were developed for this transformation, allowing for the use of either aryl chlorides or bromides as electrophiles. Utilizing a highly electron-rich dialkylphosphine ligand we have been able to construct α-quaternary centers in good yields (up to 91 % yield) and high enantioselectivities (up to 97 % ee).

Evaluation of two prognostic indices for adult T-cell leukemia/lymphoma in the subtropical endemic area, Okinawa, Japan.

Aggressive adult T-cell leukemia/lymphoma (ATL) has an extremely poor prognosis and is hyperendemic in Okinawa, Japan. This study evaluated two prognostic indices (PIs) for aggressive ATL, the ATL-PI and Japan Clinical Oncology Group (JCOG)-PI, in a cohort from Okinawa. The PIs were originally developed using two different Japanese cohorts that included few patients from Okinawa. The endpoint was overall survival (OS). Multivariable Cox regression analyses in the cohort of 433 patients revealed that all seven factors for calculating each PI were statistically significant prognostic predictors. Three-year OS rates for ATL-PI were 35.9% (low-risk, n = 66), 10.4% (intermediate-risk, n = 256), and 1.6% (high-risk, n = 111), and those for JCOG-PI were 22.4% (moderate-risk, n = 176) and 5.3% (high-risk, n = 257). The JCOG-PI moderate-risk group included both the ATL-PI low- and intermediate-risk groups. ATL-PI more clearly identified the low-risk patient subgroup than JCOG-PI. To evaluate the external validity of the two PIs, we also assessed prognostic discriminability among 159 patients who loosely met the eligibility criteria of a previous clinical trial. Three-year OS rates for ATL-PI were 34.5% (low-risk, n = 42), 9.2% (intermediate-risk, n = 109), and 12.5% (high-risk, n = 8). Those for JCOG-PI were 22.4% (moderate-risk, n = 95) and 7.6% (high-risk, n = 64). The low-risk ATL-PI group had a better prognosis than the JCOG-PI moderate-risk group, suggesting that ATL-PI would be more useful than JCOG-PI for establishing and examining novel treatment strategies for ATL patients with a better prognosis. In addition, strongyloidiasis, previously suggested to be associated with ATL-related deaths in Okinawa, was not a prognostic factor in this study.

[Analytical Method of PCBs in Fishes by GC-MS/MS Using an Accelerated Solvent Extractor].

An analytical method for PCBs in fishes using an accelerated solvent extractor (ASE) and GC-MS/MS was evaluated. After the extraction of ASE at 125℃ with n-hexane and clean-up with an AgNO3 silica gel/H2SO4 silica gel multilayer column, samples were analyzed by GC-MS/MS. This method was fast, effective and easy to operate. The limit of quantitation of the method was calculated to be 0.78 µg/kg for total PCBs. The recovery and the coefficient of variation of PCBs (n=5) from 6 fishes (Japanese sea perch, chub mackerel, yellowtail, salmon, pacific saury, and sardine) of total PCBs were 91-108% and 1-3%, respectively.

Ni-Catalyzed Enantioselective C-Acylation of α-Substituted Lactams.

A new strategy for catalytic enantioselective C-acylation to generate α-quaternary-substituted lactams is reported. Ni-catalyzed three-component coupling of lactam enolates, benzonitriles, and aryl halides produces ß-imino lactams that then afford ß-keto lactams by acid hydrolysis. Use of a readily available Mandyphos-type ligand and addition of LiBr enable the construction of quaternary stereocenters on α-substituted lactams to form ß-keto lactams in up to 94% ee.

A thaumatin-like protein, Rj4, controls nodule symbiotic specificity in soybean.

Soybeans exhibit a nitrogen-fixing symbiosis with soil bacteria of the genera Bradyrhizobium and Ensifer/Sinorhizobium in a unique organ, the root nodule. It is well known that nodulation of soybean is controlled by several host genes referred to as Rj (rj) genes. Among these genes, a dominant allele, Rj4, restricts nodulation with specific bacterial strains such as B. elkanii USDA61 and B. japonicum Is-34. These incompatible strains fail to invade the host epidermal cells as revealed by observations using DsRed-labeled bacteria. Here, we describe the molecular identification of the Rj4 gene by using map-based cloning with several mapping populations. The Rj4 gene encoded a thaumatin-like protein (TLP) that belongs to pathogenesis-related (PR) protein family 5. In rj4/rj4 genotype soybeans and wild soybeans, we found six missense mutations and two consecutive amino acid deletions in the rj4 gene as compared with the Rj4 allele. We also found, using hairy root transformation, that the rj4/rj4 genotype soybeans were fully complemented by the expression of the Rj4 gene. Whereas the expression of many TLPs and other PR proteins is induced by biotic/abiotic stress, Rj4 gene expression appears to be constitutive in roots including root nodules.

Rare case of peritoneal complete hydatidiform mole.

A 23-year-old woman, gravida 1, para 1, was transferred to our hospital with acute lower abdominal pain and vital signs consistent with shock. Her urine concentration of human chorionic gonadotrophin was 8000 mIU/mL. Transvaginal ultrasound revealed an echo-free space with mosaic echo pattern in the right adnexal area and no gestational sac in the uterus. With a preoperative diagnosis of ruptured ectopic pregnancy, emergency laparotomy was performed. The rectouterine pouch was filled with many clots containing small amounts of villous tissue. After removal of the conceptus, which was infiltrating into the peritoneum of the Pouch of Douglas, bleeding was controlled by Argon laser. Histological examination of the conceptus by immunohistochemical staining with p57(kip2) showed features of complete hydatidiform mole. This case demonstrates that the peritoneum in the Pouch of Douglas is a possible site of ectopic complete hydatidiform mole occurrence and that immunohistochemical stain is useful to confirm the diagnosis of ectopic complete hydatidiform mole.

The receptor-like kinase KLAVIER mediates systemic regulation of nodulation and non-symbiotic shoot development in Lotus japonicus.

In legumes, the number of symbiotic root nodules is controlled by long-distance communication between the shoot and the root. Mutants defective in this feedback mechanism exhibit a hypernodulating phenotype. Here, we report the identification of a novel leucine-rich repeat receptor-like kinase (LRR-RLK), KLAVIER (KLV), which mediates the systemic negative regulation of nodulation in Lotus japonicus. In leaf, KLV is predominantly expressed in the vascular tissues, as with another LRR-RLK gene, HAR1, which also regulates nodule number. A double-mutant analysis indicated that KLV and HAR1 function in the same genetic pathway that governs the negative regulation of nodulation. LjCLE-RS1 and LjCLE-RS2 represent potential root-derived mobile signals for the HAR1-mediated systemic regulation of nodulation. Overexpression of LjCLE-RS1 or LjCLE-RS2 did not suppress the hypernodulation phenotype of the klv mutant, indicating that KLV is required and acts downstream of LjCLE-RS1 and LjCLE-RS2. In addition to the role of KLV in symbiosis, complementation tests and expression analyses indicated that KLV plays multiple roles in shoot development, including maintenance of shoot apical meristem, vascular continuity, shoot growth and promotion of flowering. Biochemical analyses using transient expression in Nicotiana benthamiana revealed that KLV has the ability to interact with HAR1 and with itself. Together, these results suggest that the potential KLV-HAR1 receptor complex regulates symbiotic nodule development and that KLV is also a key component in other signal transduction pathways that mediate non-symbiotic shoot development.

Genetic diversity and geographical distribution of indigenous soybean-nodulating bradyrhizobia in the United States.

We investigated the relationship between the genetic diversity of indigenous soybean-nodulating bradyrhizobia and their geographical distribution in the United States using nine soil isolates from eight states. The bradyrhizobia were inoculated on three soybean Rj genotypes (non-Rj, Rj(2)Rj(3), and Rj(4)). We analyzed their genetic diversity and community structure by means of restriction fragment length polymorphisms of PCR amplicons to target the 16S-23S rRNA gene internal transcribed spacer region, using 11 USDA Bradyrhizobium strains as reference strains. We also performed diversity analysis, multidimensional scaling analysis based on the Bray-Curtis index, and polar ordination analysis to describe the structure and geographical distribution of the soybean-nodulating bradyrhizobial community. The major clusters were Bradyrhizobium japonicum Bj123, in the northern United States, and Bradyrhizobium elkanii, in the middle to southern regions. Dominance of bradyrhizobia in a community was generally larger for the cluster belonging to B. elkanii than for the cluster belonging to B. japonicum. The indigenous American soybean-nodulating bradyrhizobial community structure was strongly correlated with latitude. Our results suggest that this community varies geographically.

24-Hour Rest-Activity Rhythm in Middle-Aged and Older Persons with Depression.

Changes in rest or active states were clinically observed in persons with depression. However, the association between symptoms of depression and 24 h rest-activity rhythm (RAR) components that can be measured using wearable devices was not clarified. This preliminary cross-sectional study aimed to clarify the 24 h RAR components associated with symptoms of depression in middle-aged and older persons. Participants were recruited from among inpatients and outpatients requiring medical treatment at Akita University Hospital for the group with depression and from among healthy volunteers living in Akita prefecture, Japan, for the healthy control group. To assess RAR parameters including inter-daily stability (IS), intra-daily variability (IV), relative amplitude (RA), and average physical activity level for the most active 10 h span (M10) or for the least active 5 h span (L5), all the participants were instructed to wear an Actiwatch Spectrum Plus device on their non-dominant wrist for seven days. Twenty-nine persons with depression and 30 controls were included in the analysis. The results of a binomial regression analysis showed that symptoms of depression were significantly associated with a high IS value (odds ratio [OR], 1.20; 95% confidence interval [95% CI], 1.01-1.44; p = 0.04) and a low M10 value (OR, 0.85; 95% CI, 0.74-0.96; p = 0.01). Our findings suggest potential components of 24 h RAR are associated with depression.

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.