Tuft cells mediate commensal remodeling of the small intestinal antimicrobial landscape.

Succinate produced by the commensal protist Tritrichomonas musculis (T. mu) stimulates chemosensory tuft cells, resulting in intestinal type 2 immunity. Tuft cells express the succinate receptor SUCNR1, yet this receptor does not mediate antihelminth immunity nor alter protist colonization. Here, we report that microbial-derived succinate increases Paneth cell numbers and profoundly alters the antimicrobial peptide (AMP) landscape in the small intestine. Succinate was sufficient to drive this epithelial remodeling, but not in mice lacking tuft cell chemosensory components required to detect this metabolite. Tuft cells respond to succinate by stimulating type 2 immunity, leading to interleukin-13-mediated epithelial and AMP expression changes. Moreover, type 2 immunity decreases the total number of mucosa-associated bacteria and alters the small intestinal microbiota composition. Finally, tuft cells can detect short-term bacterial dysbiosis that leads to a spike in luminal succinate levels and modulate AMP production in response. These findings demonstrate that a single metabolite produced by commensals can markedly shift the intestinal AMP profile and suggest that tuft cells utilize SUCNR1 and succinate sensing to modulate bacterial homeostasis.

Release of Small RNA-containing Exosome-like Vesicles from the Human Filarial Parasite Brugia malayi.

Lymphatic filariasis (LF) is a socio-economically devastating mosquito-borne Neglected Tropical Disease caused by parasitic filarial nematodes. The interaction between the parasite and host, both mosquito and human, during infection, development and persistence is dynamic and delicately balanced. Manipulation of this interface to the detriment of the parasite is a promising potential avenue to develop disease therapies but is prevented by our very limited understanding of the host-parasite relationship. Exosomes are bioactive small vesicles (30-120 nm) secreted by a wide range of cell types and involved in a wide range of physiological processes. Here, we report the identification and partial characterization of exosome-like vesicles (ELVs) released from the infective L3 stage of the human filarial parasite Brugia malayi. Exosome-like vesicles were isolated from parasites in culture media and electron microscopy and nanoparticle tracking analysis were used to confirm that vesicles produced by juvenile B. malayi are exosome-like based on size and morphology. We show that loss of parasite viability correlates with a time-dependent decay in vesicle size specificity and rate of release. The protein cargo of these vesicles is shown to include common exosomal protein markers and putative effector proteins. These Brugia-derived vesicles contain small RNA species that include microRNAs with host homology, suggesting a potential role in host manipulation. Confocal microscopy shows J774A.1, a murine macrophage cell line, internalize purified ELVs, and we demonstrate that these ELVs effectively stimulate a classically activated macrophage phenotype in J774A.1. To our knowledge, this is the first report of exosome-like vesicle release by a human parasitic nematode and our data suggest a novel mechanism by which human parasitic nematodes may actively direct the host responses to infection. Further interrogation of the makeup and function of these bioactive vesicles could seed new therapeutic strategies and unearth stage-specific diagnostic biomarkers.

Proliferation and copy number variation of BEL-like long terminal repeat retrotransposons within the Diabrotica virgifera virgifera genome.

The proliferation of retrotransposons within a genome can contribute to increased size and affect the function of eukaryotic genes. BEL/Pao-like long-terminal repeat (LTR) retrotransposons were annotated from the highly adaptable insect species Diabrotica virgifera virgifera, the Western corn rootworm, using survey sequences from bacterial artificial chromosome (BAC) inserts and contigs derived from a low coverage next-generation genome sequence assembly. Eleven unique D. v. virgifera BEL elements were identified that contained full-length gag-pol coding sequences, whereas 88 different partial coding regions were characterized from partially assembled elements. Estimated genome copy number for full and partial BEL-like elements ranged from ~8 to 1582 among individual contigs using a normalized depth of coverage (DOC) among Illumina HiSeq reads (total genome copy number ~8821). BEL element copy number was correlated among different D. v. virgifera populations (R2=0.9846), but individual element numbers varied ≤ 1.68-fold and the total number varied by ~527 copies. These data indicate that BEL element proliferation likely contributed to a large genome size, and suggest that differences in copy number are a source of genetic variability among D. v. virgifera.

A single major QTL controls expression of larval Cry1F resistance trait in Ostrinia nubilalis (Lepidoptera: Crambidae) and is independent of midgut receptor genes.

The European corn borer, Ostrinia nubilalis (Lepidoptera: Crambidae), is an introduced crop pest in North America that causes major damage to corn and reduces yield of food, feed, and biofuel materials. The Cry1F toxin from Bacillus thuringiensis (Bt) expressed in transgenic hybrid corn is highly toxic to O. nubilalis larvae and effective in minimizing feeding damage. A laboratory colony of O. nubilalis was selected for high levels of Cry1F resistance (>12,000-fold compared to susceptible larvae) and is capable of survival on transgenic hybrid corn. Genetic linkage maps with segregating AFLP markers show that the Cry1F resistance trait is controlled by a single quantitative trait locus (QTL) on linkage group 12. The map position of single nucleotide polymorphism (SNP) markers indicated that midgut Bt toxin-receptor genes, alkaline phosphatase, aminopeptidase N, and cadherin, are not linked with the Cry1F QTL. Evidence suggests that genes within this genome interval may give rise to a novel Bt toxin resistance trait for Lepidoptera that appears independent of known receptor-based mechanisms of resistance.

A rearrangement of the Z chromosome topology influences the sex-linked gene display in the European corn borer, Ostrinia nubilalis.

Males are homogametic (ZZ) and females are heterogametic (WZ) with respect to the sex chromosomes in many species of butterflies and moths (insect order Lepidoptera). Genes on the Z chromosome influence traits involved in larval development, environmental adaptation, and reproductive isolation. To facilitate the investigation of these traits across Lepidoptera, we developed 43 degenerate primer pairs to PCR amplify orthologs of 43 Bombyx mori Z chromosome-linked genes. Of the 34 orthologs that amplified by PCR in Ostrinia nubilalis, 6 co-segregated with the Z chromosome anchor markers kettin (ket) and lactate dehydrogenase (ldh), and produced a consensus genetic linkage map of ~89 cM in combination with 5 AFLP markers. The O. nubilalis and B. mori Z chromosomes are comparatively co-linear, although potential gene inversions alter terminal gene orders and a translocation event disrupted synteny at one chromosome end. Compared to B. mori orthologs, O. nubilalis Z chromosome-linked genes showed conservation of tissue-specific and growth-stage-specific expression, although some genes exhibited species-specific expression across developmental stages or tissues. The O. nubilalis Z chromosome linkage map provides new tools for isolating quantitative trait loci (QTL) involved in sex-linked traits that drive speciation and it exposes genome rearrangements as a possible mechanism for differential gene regulation in Lepidoptera.

Thyroid hormone action on skin: diverging effects of topical versus intraperitoneal administration.

Previously, we demonstrated stimulation of epidermal proliferation and hair growth in triiodothyronine (T(3)) treated mice. To distinguish skin effects of directly applied T(3) from those of systemic hyperthyroidism, we treated CD-1 mice with either intraperitoneally (IP) or topically administered T(3). Relative to controls, mice receiving T(3) IP had 10% thinner epidermis (p < 0.01) and 48% fewer hairs (p < 0.001). By contrast, mice receiving T(3) topically had 78% thicker epidermis (p < 0.01) and 160% more hairs (p < 0.01). To gain insight into factors responsible for the diverging effects, we contrasted T(3) effect on proliferation of isolated keratinocyte cultures versus keratinocytes cocultured with dermal fibroblasts. For keratinocytes grown in the absence of fibroblasts, T(3) stimulated proliferation in a dose-dependent, biphasic pattern with the peak at 0.5 nM T(3) (84 +/- 30%, p < 0.05). Paradoxically, T(3) inhibited proliferation of keratinocytes cocultured with fibroblasts, the nadir at 0.1 nM T(3) (34% +/- 4%, p < 0.001). These studies are the first describing divergent effects of IP and topically administered thyroid hormone. The data suggest that while T(3) stimulated keratinocyte proliferation, T(3) also stimulated proliferation inhibitory factor(s) from skin fibroblasts. Insight into the interplay among the competing factors will be important in understanding thyroid hormone regulation of skin physiology.