Single Cell Analysis of Lymph Node Tissue from HIV-1 Infected Patients Reveals that the Majority of CD4+ T-cells Contain One HIV-1 DNA Molecule

Genetic recombination contributes to the diversity of human immunodeficiency virus (HIV-1). Productive HIV-1 recombination is, however, dependent on both the number of HIV-1 genomes per infected cell and the genetic relationship between these viral genomes. A detailed analysis of the number of proviruses and their genetic relationship in infected cells isolated from peripheral blood and tissue compartments is therefore important for understanding HIV-1 recombination, genetic diversity and the dynamics of HIV-1 infection. To address these issues, we used a previously developed single-cell sequencing technique to quantify and genetically characterize individual HIV-1 DNA molecules from single cells in lymph node tissue and peripheral blood. Analysis of memory and naïve CD4⁺ T cells from paired lymph node and peripheral blood samples from five untreated chronically infected patients revealed that the majority of these HIV-1-infected cells (>90%) contain only one copy of HIV-1 DNA, implying a limited potential for productive recombination in virus produced by these cells in these two compartments. Phylogenetic analysis revealed genetic similarity of HIV-1 DNA in memory and naïve CD4⁺ T-cells from lymph node, peripheral blood and HIV-1 RNA from plasma, implying exchange of virus and/or infected cells between these compartments in untreated chronic infection.     

Patterns of Root Colonization in Epacridaceous Plants Collected from Different Sites

Root colonization was studied in ten species of the Epacridaceaeat three sites in Victoria by morphological and cross-inoculationexperiments. The sites and genera chosen were Cranbourne [Epacrisimpressa Labill. andLeucopogon ericoides(Smith) R. Br.] andRye [L. parviflorus(Andrews) Lindley] on the Mornington Peninsula,and the Grampians[Astroloma conostephioides(Sond.) Benth.,A.humifusum(Cav.) R. Br.,A pinifolium(R. Br.) Benth,Brachylomadaphnoides(Smith) Benth.,E. impressa, E. impressavar.grandifloraBenth.andStyphelia adscendensR. Br.] in western Victoria. For morphologicalstudies, samples of roots from each species at each site werecleared and stained and examined microscopically. For cross-inoculationstudies, cuttings from each site were struck in potting mediuminoculated with soil from the same and other sites. The ericoidmycorrhizae in the roots of plants found at or grown in Cranbourneand Rye soils were similar. Both were significantly differentfrom the internal hyphae found in the roots of plants foundat or grown in Grampians soils, which were three times largerin diameter and formed dense coils which filled the host celland invaded adjacent epidermal cells. This suggests that morethan one fungus is involved in the relationships, that the MorningtonPeninsula sites had a different fungus from the Grampians siteand that host specificity is low. Vesicular structures werealso found commonly on plants at the Grampians site, in contrastwith other sites. Epacridaceae; root; fungus; mycorrhiza; morphology; inoculation     

Autoimmune lymphoproliferative syndrome with defective Fas: genotype influences penetrance

Autoimmune lymphoproliferative syndrome (ALPS) is a disorder of lymphocyte homeostasis and immunological tolerance. Most patients have a heterozygous mutation in the APT1 gene, which encodes Fas (CD95, APO-1), mediator of an apoptotic pathway crucial to lymphocyte homeostasis. Of 17 unique APT1 mutations in unrelated ALPS probands, 12 (71%) occurred in exons 7-9, which encode the intracellular portion of Fas. In vitro, activated lymphocytes from all 17 patients showed apoptotic defects when exposed to an anti-Fas agonist monoclonal antibody. Similar defects were found in a Fas-negative cell line transfected with cDNAs bearing each of the mutations. In cotransfection experiments, Fas constructs with either intra- or extracellular mutations caused dominant inhibition of apoptosis mediated by wild-type Fas. Two missense Fas variants, not restricted to patients with ALPS, were identified. Variant A(-1)T at the Fas signal-sequence cleavage site, which mediates apoptosis less well than wild-type Fas and is partially inhibitory, was present in 13% of African American alleles. Among the ALPS-associated Fas mutants, dominant inhibition of apoptosis was much more pronounced in mutants affecting the intracellular, versus extracellular, portion of the Fas receptor. Mutations causing disruption of the intracellular Fas death domain also showed a higher penetrance of ALPS phenotype features in mutation-bearing relatives. Significant ALPS-related morbidity occurred in 44% of relatives with intracellular mutations, versus 0% of relatives with extracellular mutations. Thus, the location of mutations within APT1 strongly influences the development and the severity of ALPS.     

Ultrasound imaging in an experimental model of fatty liver disease and cirrhosis in rats

Background

Atypical scrapie was first identified in Norwegian sheep in 1998 and has subsequently been identified in many countries. Retrospective studies have identified cases predating the initial identification of this form of scrapie, and epidemiological studies have indicated that it does not conform to the behaviour of an infectious disease, giving rise to the hypothesis that it represents spontaneous disease. However, atypical scrapie isolates have been shown to be infectious experimentally, through intracerebral inoculation in transgenic mice and sheep. The first successful challenge of a sheep with 'field' atypical scrapie from an homologous donor sheep was reported in 2007.

Results

This study demonstrates that atypical scrapie has distinct clinical, pathological and biochemical characteristics which are maintained on transmission and sub-passage, and which are distinct from other strains of transmissible spongiform encephalopathies in the same host genotype.

Conclusions

Atypical scrapie is consistently transmissible within AHQ homozygous sheep, and the disease phenotype is preserved on sub-passage.     

Mapping of citrullinated fibrinogen B-cell epitopes in rheumatoid arthritis by imaging surface plasmon resonance

Introduction  

Rheumatoid arthritis (RA) frequently involves the loss of tolerance to citrullinated antigens, which may play a role in pathogenicity. Citrullinated fibrinogen is commonly found in inflamed synovial tissue and is a frequent target of autoantibodies in RA patients. To obtain insight into the B-cell response to citrullinated fibrinogen in RA, its autoepitopes were systematically mapped using a new methodology.     

Quantification of global transcription patterns in prokaryotes using spotted microarrays

We describe an analysis, applicable to any spotted microarray dataset produced using genomic DNA as a reference, that quantifies prokaryotic levels of mRNA on a genome-wide scale. Applying this to Mycobacterium tuberculosis, we validate the technique, show a correlation between level of expression and biological importance, define the complement of invariant genes and analyze absolute levels of expression by functional class to develop ways of understanding an organism's biology without comparison to another growth condition.     

An annotated catalogue of salivary gland transcripts in the adult female mosquito, Ædes ægypti*

Background

The number of completely sequenced plastid genomes available is growing rapidly. This array of sequences presents new opportunities to perform comParative analyses. In comParative studies, it is often useful to compare across wide phylogenetic spans and, within angiosperms, to include representatives from basally diverging lineages such as the genomes reported here: Nuphar advena (from a basal-most lineage) and Ranunculus macranthus (a basal eudicot). We report these two new plastid genome sequences and make comparisons (within angiosperms, seed plants, or all photosynthetic lineages) to evaluate features such as the status of ycf15 and ycf68 as protein coding genes, the distribution of simple sequence repeats (SSRs) and longer dispersed repeats (SDR), and patterns of nucleotide composition.

Results

The Nuphar [GenBank:NC_008788] and Ranunculus [GenBank:NC_008796] plastid genomes share characteristics of gene content and organization with many other chloroplast genomes. Like other plastid genomes, these genomes are A+T-rich, except for rRNA and tRNA genes. Detailed comparisons of Nuphar with Nymphaea, another Nymphaeaceae, show that more than two-thirds of these genomes exhibit at least 95% sequence identity and that most SSRs are shared. In broader comparisons, SSRs vary among genomes in s of abundance and length and most contain repeat motifs based on A and T nucleotides.

Conclusion

SSR and SDR abundance varies by genome and, for SSRs, is proportional to genome size. Long SDRs are rare in the genomes assessed. SSRs occur less frequently than predicted and, although the majority of the repeat motifs do include A and T nucleotides, the A+T bias in SSRs is less than that predicted from the underlying genomic nucleotide composition. In codon usage third positions show an A+T bias, however variation in codon usage does not correlate with differences in A+T-richness. Thus, although plastome nucleotide composition shows "A+T richness", an A+T bias is not apparent upon more in-depth analysis, at least in these aspects. The pattern of evolution in the sequences identified as ycf15 and ycf68 is not consistent with them being protein-coding genes. In fact, these regions show no evidence of sequence conservation beyond what is normal for non-coding regions of the IR.