An Assessment by In Situ Hybridization Method for Pathogens of Severe Respiratory Infection

It has been shown that the in situ hybridization (ISH)method for the detection of phagocyted bacteria in polymorphonuclear neutrophils is more sensitive than conventional blood culture in patients with sepsis.The present study was designed to further evaluate the clinical utility of the ISH method, using a commercially available kit (Hybrizep(R)), for detecting pathogens in patients with severe respiratory infections. Peripheral blood was taken from patients with severe respiratory infections for both routine blood culture and the ISH method,respectively.In addition,focal samples including sputum,bronchoalveolar lavage,central and thoracic catheter, etc. were simultaneously examined for bacterial culture. A total number of 22 specimens was examined.The positive cases by ISH were 50.0% in the respiratory infections,which were significantly higher than those in blood cultures (9.1%).However, identical pathogens in the ISH method were not isolated from blood and/or other sources in respiratory sites. These findings suggest a possibility of high frequency of bacteremia or multifactorious pathogens in severe respiratory infections.The ISH method may provide additional information on serious respiratory infections for the detection of bacteremia. However, the clinical utility by the ISH method for patients with severe respiratory infection remains undetermined. Shinshu Med J 59 : 223―228, 2011Article信州医学雑誌 59(4): 223-228(2011)departmental bulletin pape

Conformational alterations in unidirectional ion transport of a light-driven chloride pump revealed using X-ray free electron lasers

光でイオンを輸送する膜タンパク質の巧妙な仕組み --XFELが捉えた光駆動型イオンポンプロドプシンの構造変化--. 京都大学プレスリリース. 2022-02-28.Light-driven chloride-pumping rhodopsins actively transport anions, including various halide ions, across cell membranes. Recent studies using time-resolved serial femtosecond crystallography (TR-SFX) have uncovered the structural changes and ion transfer mechanisms in light-driven cation-pumping rhodopsins. However, the mechanism by which the conformational changes pump an anion to achieve unidirectional ion transport, from the extracellular side to the cytoplasmic side, in anion-pumping rhodopsins remains enigmatic. We have collected TR-SFX data of Nonlabens marinus rhodopsin-3 (NM-R3), derived from a marine flavobacterium, at 10-µs and 1-ms time points after photoexcitation. Our structural analysis reveals the conformational alterations during ion transfer and after ion release. Movements of the retinal chromophore initially displace a conserved tryptophan to the cytoplasmic side of NM-R3, accompanied by a slight shift of the halide ion bound to the retinal. After ion release, the inward movements of helix C and helix G and the lateral displacements of the retinal block access to the extracellular side of NM-R3. Anomalous signal data have also been obtained from NM-R3 crystals containing iodide ions. The anomalous density maps provide insight into the halide binding site for ion transfer in NM-R3

Cell type-specific transcriptome of Brassicaceae stigmatic papilla cells from a combination of laser microdissection and RNA sequencing

Pollination is an early and critical step in plant reproduction, leading to successful fertilization. It consists of many sequential processes, including adhesion of pollen grains onto the surface of stigmatic papilla cells, foot formation to strengthen pollen-stigma interaction, pollen hydration and germination, and pollen tube elongation and penetration. We have focused on an examination of the expressed genes in papilla cells, to increase understanding of the molecular systems of pollination. From three representative species of Brassicaceae (Arabidopsis thaliana, A. halleri and Brassica rapa) stigmatic papilla cells were isolated precisely by laser microdissection, and cell type-specific gene expression in papilla cells was determined by RNA sequencing. As a result, 17,240, 19,260 and 21,026 unigenes were defined in papilla cells of A. thaliana, A. halleri and B. rapa, respectively, and among these 12,311 genes were common to all three species. Among the17,240 genes predicted in A. thaliana, one-third were papilla specific while approximately half of the genes were detected in all tissues examined. Bioinformatics analysis revealed that genes related to a wide range of reproduction and development functions are expressed in papilla cells, particularly metabolism, transcription, and membrane-mediated information exchange. These results reflect the conserved features of general cellular function and also the specific reproductive role of papilla cells, highlighting a complex cellular system regulated by a diverse range of molecules in these cells. This study provides fundamental biological knowledge to dissect the molecular mechanisms of pollination in papilla cells and will shed light on our understanding of plant reproduction mechanism