Fine mapping and DNA fiber FISH analysis locates the tobamovirus resistance gene L3 of Capsicum chinense in a 400-kb region of R-like genes cluster embedded in highly repetitive sequences

The tobamovirus resistance gene L3 of Capsicum chinense was mapped using an intra-specific F2 population (2,016 individuals) of Capsicum annuum cultivars, into one of which had been introduced the C. chinenseL3 gene, and an inter-specific F2 population (3,391 individuals) between C. chinense and Capsicum frutescence. Analysis of a BAC library with an AFLP marker closely linked to L3-resistance revealed the presence of homologs of the tomato disease resistance gene I2. Partial or full-length coding sequences were cloned by degenerate PCR from 35 different pepper I2 homologs and 17 genetic markers were generated in the inter-specific combination. The L3 gene was mapped between I2 homolog marker IH1-04 and BAC-end marker 189D23M, and located within a region encompassing two different BAC contigs consisting of four and one clones, respectively. DNA fiber FISH analysis revealed that these two contigs are separated from each other by about 30 kb. DNA fiber FISH results and Southern blotting of the BAC clones suggested that the L3 locus-containing region is rich in highly repetitive sequences. Southern blot analysis indicated that the two BAC contigs contain more than ten copies of the I2 homologs. In contrast to the inter-specific F2 population, no recombinant progeny were identified to have a crossover point within two BAC contigs consisting of seven and two clones in the intra-specific F2 population. Moreover, distribution of the crossover points differed between the two populations, suggesting linkage disequilibrium in the region containing the L locus

Neutron diffraction and thermal studies of amorphous

We have succeeded in preparing amorphous carbon disulphide (\chem{CS_2}) by depositing its vapour on a cold substrate at 10\un{K}. Complete formation of the amorphous state has been confirmed by neutron diffraction and differential thermal analysis (DTA). The amorphous sample crystallised at ca. 70\un{K}, which is lower than the hypothetical glass transition temperature (92\un{K}) estimated from the DTA data of the (\chem{CS_2})x(\chem{S_2Cl_2})$_{1-x}$ binary mixture. \chem{CS_2}, a symmetric linear tri-atomic molecule, is the simplest of the amorphised molecular substances whose structural and thermal information has been reported so far. Comparison of the static structure factors $S(Q)$ has shown that the orientational correlation of \chem{CS_2} molecules may be much stronger in the amorphous state than in the liquid state at higher temperature

CD4+ and CD8+ cytotoxic T lymphocytes may induce mesenchymal cell apoptosis in IgG4-related disease

Background: IgG4-related disease (IgG4-RD) is an immune-mediated fibrotic disorder that has been linked to CD4+ cytotoxic T lymphocytes (CD4+CTLs). The effector phenotype of CD4+CTLs and the relevance of both CD8+ cytotoxic T lymphocytes (CD8+CTLs) and apoptotic cell death remain undefined in IgG4-RD. Objective: We sought to define CD4+CTL heterogeneity, characterize the CD8+CTL response in the blood and in lesions, and determine whether enhanced apoptosis may contribute to the pathogenesis of IgG4-RD. Methods: Blood analyses were undertaken using flow cytometry, cell sorting, transcriptomic analyses at the population and single-cell levels, and next-generation sequencing for the TCR repertoire. Tissues were interrogated using multicolor immunofluorescence. Results were correlated with clinical data. Results: We establish that among circulating CD4+CTLs in IgG4-RD, CD27loCD28loCD57hi cells are the dominant effector subset, exhibit marked clonal expansion, and differentially express genes relevant to cytotoxicity, activation, and enhanced metabolism. We also observed prominent infiltration of granzyme A–expressing CD8+CTLs in disease tissues and clonal expansion in the blood of effector/memory CD8+ T cells with an activated and cytotoxic phenotype. Tissue studies revealed an abundance of cells undergoing apoptotic cell death disproportionately involving nonimmune, nonendothelial cells of mesenchymal origin. Apoptotic cells showed significant upregulation of HLA-DR. Conclusions: CD4+CTLs and CD8+CTLs may induce apoptotic cell death in tissues of patients with IgG4-RD with preferential targeting of nonendothelial, nonimmune cells of mesenchymal origin

Comprehensive full-length sequence analyses of human parechoviruses: diversity and recombination

Human parechoviruses (HPeVs) are highly prevalent pathogens among very young children. Although originally classified into two serologically distinct types, HPeV1 and -2, recent analyses of variants collected worldwide have revealed the existence of 12 further types classified genetically by sequence comparisons of complete genome sequences or the capsid (VP1) gene. To investigate the nature of HPeV evolution, its population dynamics and recombination breakpoints, this study generated 18 full-length genomic sequences of the most commonly circulating genotypes, HPeV1 and -3, collected over a time span of 14 years from The Netherlands. By inclusion of previously published full-length sequences, 35 sequences were analysed in total. Analysis of contemporary strains of HPeV1 and those most similar to the prototype strain (Harris) showed that HPeV1 variants fall into two genetically distinct clusters that are much more divergent from each other than those observed within other HPeV types. Future classification criteria for HPeVs may require modification to accommodate the occurrence of variants with intermediate degrees of diversity within types. Recombination was frequently observed among HPeV1, -4, -5 and -6, but was much more restricted among HPeV3 strains. Favoured sites for recombination were found to flank the capsid region, and further sites were found within the non-structural region, P2. In contrast to other HPeV types, the majority of the HPeV3 sequences remained monophyletic across the genome, a possible reflection of its lower diversity and potentially more recent emergence than other HPeV types, or biological and/or epidemiological constraints that limit opportunities for co-infections with potential recombination partners