sj-pdf-1-jnm-10.1177_10949968221102306 - Supplemental material for Do Handwritten Notes Benefit Online Retailers? A Field Experiment

Supplemental material, sj-pdf-1-jnm-10.1177_10949968221102306 for Do Handwritten Notes Benefit Online Retailers? A Field Experiment by Sanghwa Kim, Jeonghye Choi, and Seung Hyun Kim in Journal of Interactive Marketing</p

Data_Sheet_1_Lobophorin Producing Endophytic Streptomyces olivaceus JB1 Associated With Maesa japonica (Thunb.) Moritzi & Zoll..PDF

In this study, we focused on endophytes of Maesa japonica (Thunb.) Moritzi & Zoll. and the plant-microbe interaction at metabolite levels. We isolated seven endophytes associated with M. japonica (JB1−7), and focused on Streptomyces olivaceus JB1 because of antibacterial activities of its secondary metabolites. We confirmed lobophorin analogs production from the bacterial strain JB1 by using spectroscopic techniques such as NMR, UV, and LC/Q-TOF-MS. In the LC/MS system, thirteen reported lobophorin analogs and twelve unreported analogs were detected. Among metabolites, lobophorin A was clearly detected in the dried foliar residues of M. japonica which implies that JB1 resides in the host and accumulates its secondary metabolites likely interacting with the plant. Antimicrobial activity tests of the secondary metabolites against undesirable contaminants isolated from the external surface of M. japonica supported the host and microbe mutualistic relationship. In the meantime, lobophorin producing Streptomyces spp. were isolated from marine environments such as marine sediments, algae, corals, and sponges. As lobophorin producing Streptomyces is isolated commonly from marine environments, we conducted a saline water stress tolerance test with JB1 showing saline medium does not accelerate the growth of the bacterium.</p

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Multivariate statistical analysis plots of nine Lauraceae species based on LC-MS spectral data.

(A) PCA score plot. (B) PLS-DA score plot. LDE, Lindera erythrocarpa Makino; LJ, Litsea japonica (Thunb.) Jussieu; NS, Neolitsea sericea (Blume) Koidz.; MT, Machilus thunbergii Siebold & Zucc.; CC, Cinnamomum camphora (L.) J. Presl; CY, C. yabunikkei H. Ohba; NA, N. aciculata (Blume) Koidz.; MJ, M. japonica Siebold & Zucc.; and LC, L. coreana H.Lév.</p

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(CSV)</p

Phylogenetic relationship of Lauraceae samples.

Combined sequences of two universal barcoding regions, trnH-GUG and rbcL, from each sample were used to draw a neighbor-joining tree with 1000 bootstrap replicates. LDE, Lindera erythrocarpa Makino; LJ, Litsea japonica (Thunb.) Jussieu; NS, Neolitsea sericea (Blume) Koidz.; MT, Machilus thunbergii Siebold & Zucc.; CC, Cinnamomum camphora (L.) J. Presl; CY, C. yabunikkei H. Ohba; NA, N. aciculata (Blume) Koidz.; MJ, M. japonica Siebold & Zucc.; and LC, L. coreana H.Lév.</p

The networking analysis results of the nine Lauraceae species.

A network was generated using MS/MS spectra through classical molecular networking on the GNPS server and visualized with nodes and edges through Cytoscape 3.8.0. The nodes consist of pie charts based on the peak intensity proportion for each metabolite. The thickness of the edges was determined by the similarity between two connected nodes with edge widths ranging from 6.0 to 16.0. The blue, red, and green boxes indicate isoquinoline alkaloids, flavonoids, and lignin clusters, respectively. LDE, Lindera erythrocarpa Makino; LJ, Litsea japonica (Thunb.) Jussieu; NS, Neolitsea sericea (Blume) Koidz.; MT, Machilus thunbergii Siebold & Zucc.; CC, Cinnamomum camphora (L.) J. Presl; CY, C. yabunikkei H. Ohba; NA, N. aciculata (Blume) Koidz.; MJ, M. japonica Siebold & Zucc.; and LC, L. coreana H.Lév.</p

Pasakbumin A controls intracellular Mtb growth by increasing the production of NO and pro-inflammatory cytokine in H37Rv-infected macrophages.

Raw264.7 macrophages were stimulated with pasakbumin A (Pas A, 10 μM) for 48 h after infection with H37Rv (at MOIs of 1 or 5). (A) Intracellular bacterial survival was determined by counting the number of CFUs at 3-weeks after inoculation. (B) The cultures were grown in 7H9 medium supplemented with 10% ADC containing 0.2% glycerol at 37°C for 72 h with or without pasakbumin A (Pas A, 10 μM). Left, bacterial growth was measured as OD600 at 72 h after Mtb inoculation. Right, colony-forming units (CFUs) were measured by plating bacterial dilutions onto 7H10 agar supplemented with 10% OADC containing 0.5% glycerol at 72 h. Error bars represents the standard deviation of the mean. (C) Cell viability was assessed using trypan-blue exclusion assay. (D) Culture supernatants were measured for the production of TNF-α and IL-10 by ELISA in non-infected (NI) or H37Rv-infected cells. (E) Bar graph in the left panel represents NO production (as indicated by the nitrite level) by diazotization (Griess method) assay. Right panel represents NOS2 protein by western blot analysis. Actin served as a loading control. The full length of all western blots has been provided in the S1 Fig. Statistical significance is indicated as **, pp<0.001.</p

Representative chemical structures of eleven metabolites contributing to sample discrimination.

(1) neochlorogenic acid; (2) afzelin; (3) laurolitsine; (4) catechin; (6) chlorogenic acid; (7) coclaurine; (8) dihydrokaempferol; (9) epicatechin; (11) roemerine; (12) phenylalanine; and (13) quercitrin.</p

The chemical composition of Lauraceae samples based on the network analysis.

(A) Clusters annotated as isoquinoline alkaloids or flavonoids through classical molecular networking. The thickness of the edges was determined by the similarity between two connected nodes with edge widths ranging from 6.0 to 16.0. (B) The node composition ratio of isoquinoline alkaloids or flavonoids in each sample. LDE, Lindera erythrocarpa Makino; LJ, Litsea japonica (Thunb.) Jussieu; NS, Neolitsea sericea (Blume) Koidz.; MT, Machilus thunbergii Siebold & Zucc.; CC, Cinnamomum camphora (L.) J. Presl; CY, C. yabunikkei H. Ohba; NA, N. aciculata (Blume) Koidz.; MJ, M. japonica Siebold & Zucc.; and LC, L. coreana H.Lév.</p