High-parametric evaluation of human invariant natural killer T cells to delineate heterogeneity in allo- and autoimmunity.

Human invariant natural killer T (iNKT) cells are a rare innate-like lymphocyte population that recognizes glycolipids presented on CD1d. Studies in mice have shown that these cells are heterogeneous and are capable of enacting diverse functions, and the composition of iNKT cell subsets can alter disease outcomes. In contrast, far less is known about how heterogeneity in human iNKT cells relates to disease. To address this, we used a high-dimensional, data-driven approach to devise a framework for parsing human iNKT heterogeneity. Our data revealed novel and previously described iNKT cell phenotypes with distinct functions. In particular, we found 2 phenotypes of interest: (1) a population with T helper 1 function that was increased with iNKT activation characterized by HLA-II+CD161- expression, and (2) a population with enhanced cytotoxic function characterized by CD4-CD94+ expression. These populations correlate with acute graft-versus-host disease after allogeneic hematopoietic stem cell transplantation and with new onset type 1 diabetes, respectively. Our study identifies human iNKT cell phenotypes associated with human disease that could aid in the development of biomarkers or therapeutics targeting iNKT cells.

Habitat visualization and genomic analysis of "Candidatus Pantoea carbekii," the primary symbiont of the brown marmorated stink bug.

Phytophagous pentatomid insects can negatively impact agricultural productivity and the brown marmorated stink bug (Halyomorpha halys) is an emerging invasive pest responsible for damage to many fruit crops and ornamental plants in North America. Many phytophagous stink bugs, including H. halys, harbor gammaproteobacterial symbionts that likely contribute to host development, and characterization of symbiont transmission/acquisition and their contribution to host fitness may offer alternative strategies for managing pest species. "Candidatus Pantoea carbekii" is the primary occupant of gastric ceca lumina flanking the distal midgut of H. halys insects and it is acquired each generation when nymphs feed on maternal extrachorion secretions following hatching. Insects prevented from symbiont uptake exhibit developmental delays and aberrant behaviors. To infer contributions of Ca. P. carbekii to H. halys, the complete genome was sequenced and annotated from a North American H. halys population. Overall, the Ca. P. carbekii genome is nearly one-fourth (1.2 Mb) that of free-living congenerics, and retains genes encoding many functions that are potentially host-supportive. Gene content reflects patterns of gene loss/retention typical of intracellular mutualists of plant-feeding insects. Electron and fluorescence in situ microscopic imaging of H. halys egg surfaces revealed that maternal extrachorion secretions were populated with Ca. P. carbekii cells. The reported findings detail a transgenerational mode of symbiont transmission distinct from that observed for intracellular insect mutualists and illustrate the potential additive functions contributed by the bacterial symbiont to this important agricultural pest.

Evolutionary basis of mitonuclear discordance between sister species of mole salamanders (Ambystoma sp.).

Distinct genetic markers should show similar patterns of differentiation between species reflecting their common evolutionary histories, yet there are increasing examples of differences in the biogeographic distribution of species-specific nuclear (nuDNA) and mitochondrial DNA (mtDNA) variants within and between species. Identifying the evolutionary processes that underlie these anomalous patterns of genetic differentiation is an important goal. Here, we analyse the putative mitonuclear discordance observed between sister species of mole salamanders (Ambystoma barbouri and A. texanum) in which A. barbouri-specific mtDNA is found in animals located within the range of A. texanum. We test three hypotheses for this discordance (undetected range expansion, mtDNA introgression, and hybridization) using nuDNA and mtDNA data analysed with methods that varied in the parameters estimated and the timescales measured. Results from a Bayesian clustering technique (structure), bidirectional estimates of gene flow (migrate-n and IMa2) and phylogeny-based methods (*beast, bucky) all support the conclusion that the discordance is due to geographically restricted mtDNA introgression from A. barbouri into A. texanum. Limited data on species-specific tooth morphology match this conclusion. Significant differences in environmental conditions exist between sites where A. texanum with and without A. barbouri-like mtDNA occur, suggesting a possible role for selection in the process of introgression. Overall, our study provides a general example of the value of using complimentary analyses to make inferences of the directionality, timescale, and source of mtDNA introgression in animals.

Obligate insect endosymbionts exhibit increased ortholog length variation and loss of large accessory proteins concurrent with genome shrinkage.

Extreme genome reduction has been observed in obligate intracellular insect mutualists and is an assumed consequence of fixed, long-term host isolation. Rapid accumulation of mutations and pseudogenization of genes no longer vital for an intracellular lifestyle, followed by deletion of many genes, are factors that lead to genome reduction. Size reductions in individual genes due to small-scale deletions have also been implicated in contributing to overall genome shrinkage. Conserved protein functional domains are expected to exhibit low tolerance for mutations and therefore remain relatively unchanged throughout protein length reduction while nondomain regions, presumably under less selective pressures, would shorten. This hypothesis was tested using orthologous protein sets from the Flavobacteriaceae (phylum: Bacteroidetes) and Enterobacteriaceae (subphylum: Gammaproteobacteria) families, each of which includes some of the smallest known genomes. Upon examination of protein, functional domain, and nondomain region lengths, we found that proteins were not uniformly shrinking with genome reduction, but instead increased length variability and variability was observed in both the functional domain and nondomain regions. Additionally, as complete gene loss also contributes to overall genome shrinkage, we found that the largest proteins in the proteomes of nonhost-restricted bacteroidetial and gammaproteobacterial species often were inferred to be involved in secondary metabolic processes, extracellular sensing, or of unknown function. These proteins were absent in the proteomes of obligate insect endosymbionts. Therefore, loss of genes encoding large proteins not required for host-restricted lifestyles in obligate endosymbiont proteomes likely contributes to extreme genome reduction to a greater degree than gene shrinkage.