PMC

(A) % invasion and (B) fold proliferation of STM WT and STM ΔchiA strains, and (C) % invasion and (D) fold proliferation of STY WT, STY ΔchiA, STY ΔchiA:chiA and STY ΔchiA:pQE60 strains in U937 derived monocytes by gentamicin protection assay. (N = 3, n = 3). (E) % invasion and (F) fold proliferation of STM WT and STM ΔchiA strains, and (G) % invasion and (H) fold proliferation of STY WT, STY ΔchiA, STY ΔchiA:chiA and STY ΔchiA:pQE60 strains in BMDCs by gentamicin protection assay. (N = 3, n = 3). (I) Extracellular NO was estimated by Greiss assay from spent media obtained from STM WT, STM ΔchiA, STY WT, STY ΔchiA and STY ΔchiA:chiA infected BMDCs. (UI- Uninfected). (N = 3, n = 3). Two-way ANOVA was used to analyze the data. Intracellular survival of (J) STM WT and STM ΔchiA and (K) STY WT and STY ΔchiA strains was calculated in WT (NOS2^+/+) and NOS2^-/- BMDCs by gentamicin protection assay. (N = 3, n = 3). (L) Representative flow cytometry plot for ROS estimation by DCFDA assay from STM WT, STM ΔchiA, STY WT, STY ΔchiA and STY ΔchiA:chiA infected and lipopolysaccharide (LPS; control) treated PMs. (UI US- Unstained and uninfected, UI- Uninfected). (M) ³H thymidine incorporation assay to assess CD8⁺ T cell proliferation 20 hpi with STM WT, STM ΔchiA, STY WT, STY ΔchiA and STY ΔchiA:chiA (UI- Uninfected, OVA- Ovalbumin, ConA- Concanavalin A). (N = 3, n = 6). (N) Quantification of the MHC-II density per unit length of the cell membrane of STM WT, STM ΔchiA, STY WT, STY ΔchiA and STY ΔchiA:chiA infected PMs after the indicated time (UI- Uninfected). (N = 2). (O) Representative flow cytometry plot showing surface MHC-II level on PMs infected with STM WT and STM ΔchiA for 20 hours (UI US- Unstained and uninfected, UI- Uninfected). Unpaired Student’s t test was used to analyze the data for A, B, E, F, J, K and one-way ANOVA was used to analyze the data for C, D, G, H, M.

(A) Representative image of Caco-2 cells infected with STM WT, STM ΔchiA, STY WT and STY ΔchiA strains to visualize the intracellular niche of the bacteria. The SCVs were stained for LAMP1 (UI- Uninfected; Yellow arrows- LAMP1⁺ SCVs, white arrowheads- LAMP1^- SCVs). (B) % of STM WT and STM ΔchiA, (C) % of STY WT and STY ΔchiA bacteria inside and outside the LAMP1⁺ SCVs 16 hpi was calculated. (N = 3). (D) Representative image of Caco-2 cells infected with STM WT, STM ΔchiA, STY WT and STY ΔchiA strains at MOI 50 to visualize EEA1 and LAMP1 recruitment on the SCVs (scale bars- 10μm; insets show SCVs). Absolute CFU/ml values of (E) STM WT and STM ΔchiA, and (F) STY WT, STY ΔchiA and STY ΔchiA:chiA in Caco-2 cells in chloroquine resistance assay 16 hpi. (N = 3, n = 3). One-way ANOVA was used to analyze the data.

GISH on somatic metaphase cells from root tips of Elymus sinosubmuticus and Elymus breviaristatus. a-c, E. sinosubmuticus. a, 14 chromosomes showed H genome single which from Hordeum bogdanii when blocked with StY genome which from Roegneria ciliaris. b, 28 chromosomes showed StY singles when blocked with H genome. c, 14 chromosomes showed H genome singles and 28 chromosomes showed StY singles when StY genome and H genome as probes. d-f, E. breviaristatus. d, 14 chromosomes showed H genome singles when blocked with StY genome. e, 28 chromosomes showed StY singles when blocked with H genome. f, 14 chromosomes showed H genome singles and 28 chromosomes showed StY singles when StY genome and H genome as probes. Scale bar equal 10 μm

Inside the enterocyte: STY localization and survival.
(A) Immunostaining of STY-infected biopsies with cytoskeletal protein tubulin, intracellular early endosome marker Rab5, STY and genomic material (DAPI). STY colocalizes with tubulin during biopsy invasion but exists independent of Rab5. Inset images demonstrating co-localization are marked with a blue arrow; independent STY are marked with a pink arrow.
(B) Loss of intracellular early endosome marker indicates masking of cellular localization by STY. RNAseq data predicts inhibition of phagosome formation and maturation; downregulated genes are in red, upregulated genes are in green.
(C) Inhibition of phagosome maturation critically impairs antigen harvest and bacterial clearance, which are overarching themes in disease pathways predicted to be downregulated in response to STY infection of terminal ileum biopsies.
(D) Predicted deactivated disease states related to the immune response deciphered from downregulation of genes during STY infection. The negative Z-score indicates a downregulation of pathways; the number of genes refers to the number of genes downregulated during STY infection that corresponds to the pathway listed. Notable deactivated disease states include quantity of phagocytes, activation of lymphocytes, localization of mononuclear cells, activation of lymphocytes and leukocyte migration.