Metagenomic analysis exploring soil microbial communities associated with Antarctic vascular plants and functional analysis. Supplementary figures & tables

Rhizosphere core microbiome and relative abundance eggNOG categories heatmap for all analyzed rhizospheric soil samples from Antarctic vascular plants. Raw reads were trimmed and aligned using DIAMOND agains NR database (NCBI). Taxonomical and functional classification was performed using MEGAN

Rodriguez et al 2018

Raw data for publicatio

Mice exposed to uninfected <i>P. duboscqi</i> bites are protected against vector-transmitted <i>L. major</i>.

<p>(<b>A</b>) Representative parasite load and percent metacyclics in the gut of infected <i>P. duboscqi</i> sand flies the day of transmission. The mean ± SEM are shown. (<b>B–C</b>) Mice exposed to uninfected bites in the right ear (▪) or are naïve () were challenged with 10 <i>L. major</i>-infected <i>P. duboscqi</i> sand flies in the left ear two weeks after the last exposure. (<b>B</b>) Weekly measurement of ear lesions after transmission. The mean ± SEM for 10 mice per group are shown. (<b>C</b>) The number of parasites per ear for five mice at four weeks post-transmission determined by limiting dilution assay. The bar represents the mean parasite burden per ear. Data are representative of three independent experiments. *<i>p</i><0.02; **<i>p</i> = 0.008.</p

IFN- γ expression by T cells and NK cells following infected sand fly bites.

<p>Ear cells were recovered from naïve and exposed mice at 6 h, 24 h, 48 h and one week after transmission and cultured for 16 h. Brefeldin, PMA and ionomycin were added during the last four hours. The percent and absolute number of CD4+ T cells (<b>A</b>) or NK cells (<b>B</b>) expressing IFN-γ. Percentages shown are representative of three independent experiments; absolute numbers shown represent the mean ± SEM from three independent experiments. Five mice were used per group per experiment. *<i>p</i><0.05.</p

Characterization of the early leukocyte infiltrate following vector-transmission of <i>L. major</i>.

<p>Ear cells were recovered from naïve and exposed mice at 6 h, 24 h, 48 h and one week after transmission. At the indicated times after challenge, cells were stained <i>ex vivo</i> and analyzed by FACS for surface expression of TCR-β, CD4, NK1.1, F4/80, CD11b, and Ly-6G to identify specific leukocyte populations. (<b>A</b>) Absolute number of leukocytes per ear; (<b>B</b>) The gating strategy used to identify cells (NK1.1^posTCR-β^neg); CD4⁺ T cells (TCR-β^posCD4^pos), neutrophils (F4/80^neg CD11b^highLy6G^high), Gr1⁺ inflammatory monocytes (F4/80^posCD11b^highLy6G^high) and macrophages (F4/80^highLy6G^negCD11b^high); (<b>C</b>) Absolute number of neutrophils, NK cells, Gr1⁺ monocytes, macrophages and CD4⁺ T cells per ear. The numbers shown represent mean ± SEM from 3 independent experiments. Five mice were used per group per experiment. *<i>p</i><0.05.</p

Intradermal Immunization of <i>Leishmania donovani</i> Centrin Knock-Out Parasites in Combination with Salivary Protein LJM19 from Sand Fly Vector Induces a Durable Protective Immune Response in Hamsters

<div><p>Background</p><p>Visceral leishmaniasis (VL) is a neglected tropical disease and is fatal if untreated. There is no vaccine available against leishmaniasis. The majority of patients with cutaneous leishmaniasis (CL) or VL develop a long-term protective immunity after cure from infection, which indicates that development of an effective vaccine against leishmaniasis is possible. Such protection may also be achieved by immunization with live attenuated parasites that do not cause disease. We have previously reported a protective response in mice, hamsters and dogs with <i>Leishmania donovani</i> centrin gene knock-out parasites (<i>LdCen</i>^<i>-/-</i>), a live attenuated parasite with a cell division specific centrin1 gene deletion. In this study we have explored the effects of salivary protein LJM19 as an adjuvant and intradermal (ID) route of immunization on the efficacy of <i>LdCen</i>^<i>-/-</i> parasites as a vaccine against virulent <i>L</i>. <i>donovani</i>.</p><p>Methodology/Principal Findings</p><p>To explore the potential of a combination of <i>LdCen</i>^-/- parasites and salivary protein LJM19 as vaccine antigens, <i>LdCen</i>^-/- ID immunization followed by ID challenge with virulent <i>L</i>. <i>donovani</i> were performed in hamsters in a 9-month follow up study. We determined parasite burden (serial dilution), antibody production (ELISA) and cytokine expression (qPCR) in these animals. Compared to controls, animals immunized with <i>LdCen</i>^<i>-/-</i> + LJM19 induced a strong antibody response, a reduction in spleen and liver parasite burden and a higher expression of pro-inflammatory cytokines after immunization and one month post-challenge. Additionally, a low parasite load in lymph nodes, spleen and liver, and a non-inflamed spleen was observed in immunized animals 9 months after the challenge infection.</p><p>Conclusions</p><p>Our results demonstrate that an ID vaccination using <i>LdCen</i>^<i>-/-</i>parasites in combination with sand fly salivary protein LJM19 has the capability to confer long lasting protection against visceral leishmaniasis that is comparable to intravenous or intracardial immunization.</p></div

Most relevant biological processes involved during table grape berry ripening.

<p>In this schematic resume, the biological processes which show an up-regulated transcriptional profile are presented in red. In contrast, the processes presented a decrease of its related transcripts are presented in green.</p

Additional file 1: Table S1. of mRNA-seq reveals skeletal muscle atrophy in response to handling stress in a marine teleost, the red cusk-eel (Genypterus chilensis)

Complete list of up-regulated and down-regulated transcripts in response to handling stress. (XLSX 66 kb

Transcriptome validation by qRT-PCR.

<p>Nine genes were selected according their biological role in the cell metabolism belong from different profile of expression in order to confirm the expression values obtained by RNASeq. For each gene, the FPKM value (continue line) and relative expression to A1 of qRT-PCR (dashed line) was evaluated in all selected points, using the expression of MVK gene for normalization. Vertical bars represent standard error of the mean of three biological replicates for qRT-PCR.</p

Cytokine responses 5 weeks after immunization with <i>LdCen</i>^<i>-/-</i> parasites and LJM19.

<p>Immune response in the ear tissue of immunized hamsters was measured. The expression of mRNA encoding IFN-γ (A), iNOS (B), IL-12 (C), IL-4 (D) and IL-10 (E) was evaluated by qPCR. The data were normalized to β-Actin expression. (F) Fold increase of the cytokines compared to BSA (G4) group. Statistical differences (<i>p</i><0.05) are indicated in letters (a: G1).</p