msgg medium  (Difco)

Journal: BMC Microbiology

Article Title: The role of nitric-oxide-synthase-derived nitric oxide in multicellular traits of Bacillus subtilis 3610: biofilm formation, swarming, and dispersal

doi: 10.1186/1471-2180-11-111

Figure Lengend Snippet: Influence of NO and NO synthase (NOS) on colony morphology and fruiting body formation of B. subtilis 3610 . (A-E) Colonies were grown for 4 d on MSgg agar and images were captured with a digital camera. (F-K) Colonies were grown for 3 d on MSgg agar and images were captured with a CCD camera mounted on a microscope. NO scavenger (c-PTIO), NOS inhibitor (L-NAME) and NO donor (Noc-18) were added to biofilm incubations of B. subtilis wild-type. Scale bars are 1 cm (A-E) and 200 μm (F-K) .

Article Snippet: Biofilms were homogenized in the MSgg medium by sonication (Labsonic U, B. Braun, Melsungen, Germany) for 10 min at ~ 40 W on ice.

Techniques: Microscopy

Journal: BMC Microbiology

Article Title: The role of nitric-oxide-synthase-derived nitric oxide in multicellular traits of Bacillus subtilis 3610: biofilm formation, swarming, and dispersal

doi: 10.1186/1471-2180-11-111

Figure Lengend Snippet: Influence of NO and NO synthase (A) on the cell concentration and (B) the percentage of spores per cell during the development of biofilms of B. subtilis 3610 and 3610Δ nos at the liquid-air interface as determined by plate counting . Biofilms of wild-type 3610 were grown in 25 mL MSgg medium in glass tubes without supplementation (control), supplemented with 100 μM L-NAME (NOS inhibitor), 75 μM c-PTIO (NO scavenger), and 130 μM Noc-18 (NO donor). Error bars indicate standard deviation (N = 3).

Article Snippet: Biofilms were homogenized in the MSgg medium by sonication (Labsonic U, B. Braun, Melsungen, Germany) for 10 min at ~ 40 W on ice.

Techniques: Concentration Assay, Standard Deviation

Journal: BMC Microbiology

Article Title: The role of nitric-oxide-synthase-derived nitric oxide in multicellular traits of Bacillus subtilis 3610: biofilm formation, swarming, and dispersal

doi: 10.1186/1471-2180-11-111

Figure Lengend Snippet: Influence of NO and NO synthase on (A) dispersal and (B) germination of B. subtilis 3610 . (A) The dispersal assay was conducted with 3610 wild-type (white bars) and 3610Δ nos (gray bars). Colonies grew for 4 d on MSgg agar and were mounted with a drop of 100 μL MSgg medium. The NOS inhibitor L-NAME and the NO scavenger c-PTIO were supplemented to agar and drop, while the NO donor SNAP was only supplemented to the drop. Vegetative cells that dispersed within 2 h into the drop liquid were quantified with flow cytometry. Error bars indicate standard error (N = 10). (B) The germination assay was conducted in a separate experiment, employing a similar set-up and the same treatments as for the dispersal assay. MSgg medium (including supplements) was mixed with B. subtilis spores, placed as a 100 μL drop on a sterile polystyrene surface and incubated for 2 h. Spores only (open bars in panel B) and total cells (hatched bars in panel B) were determined by plating and counting the colony forming units (cfu). The results are normalized to the spore concentration. Error bars indicate standard deviation (N = 5). The results show that any difference in the dispersal assay is caused by effects of NO and NOS on active dispersal of vegetative biofilm cells and not on germination of spores.

Article Snippet: Biofilms were homogenized in the MSgg medium by sonication (Labsonic U, B. Braun, Melsungen, Germany) for 10 min at ~ 40 W on ice.

Techniques: Flow Cytometry, Germination Assay, Incubation, Concentration Assay, Standard Deviation

Journal: BMC Microbiology

Article Title: The role of nitric-oxide-synthase-derived nitric oxide in multicellular traits of Bacillus subtilis 3610: biofilm formation, swarming, and dispersal

doi: 10.1186/1471-2180-11-111

Figure Lengend Snippet: Nitric oxide microprofiles measured during the dispersal assay . The y-axis shows the biofilm depth with 0 (dashed line) denoting the surface of the biofilm. Positive values are inside the spot colony biofilm and negative values are above the biofilm in the MSgg medium drop. MSgg medium was supplemented with 300 μM of the NO donor SNAP (closed symbols) or supplied without supplementation of SNAP (open symbols). Wild-type B. subtilis 3610 was incubated with a drop of MSgg (A) without further supplementation and (B) further supplemented with 100 μM NOS inhibitor L-NAME. ( C) shows B. subtilis 3610 Δ nos supplied with MSgg without further supplementation. Error bars depict the standard deviation (N = 10) between repeated measurements at the same position in the sample reflecting the precision of the measurement.

Article Snippet: Biofilms were homogenized in the MSgg medium by sonication (Labsonic U, B. Braun, Melsungen, Germany) for 10 min at ~ 40 W on ice.

Techniques: Incubation, Standard Deviation

Journal: bioRxiv

Article Title: Arabidopsis thaliana induces multigenerational stress tolerance against biotic and abiotic stressors and memorization of host colonization in Bacillus subtilis

doi: 10.1101/2022.05.29.493878

Figure Lengend Snippet: (A) B. subtilis bacteria were cultured in liquid MSgg (“untreated”) or in A. thaliana secretions and then washed in PBS. These bacteria were termed “ancestors”. Some of the bacteria were transferred to a shaking culture in liquid LB and grown for four hours. These bacteria were termed “descendants”. The bacteria were then regrown in MSgg and monitored for planktonic growth -/+ 600 mM NaCl in a plate reader. Alternatively ancestors and descendants were cultured with A. thaliana plants -/ + 600 mM NaCl, after which root colonization was examined with a confocal microscope, CFU counts and FACS analysis. (B-C) Growth curves and lag time of secretion treated and untreated bacteria (+sec and -sec respectively) and their descendants in the absence and presence of NaCl. Bacteria were grown in a microplate reader as described above and their lag time was calculated (D) Lag time of bacteria with and without plant secretions in the absence and presence of salicylic acid in “ancestor” bacteria. (E) Growth curves of bacteria that were treated or untreated with secretions, re-streaked separately on LB plates and grown overnight. Single colonies were picked up, transferred to liquid shaking culture for 4 hours at 37°C and then grown in a microplate reader as described above. All experiments were performed in triplicates at least three separate and independent times. Data is expressed as average values ± standard deviations of the means from a representative experiment, performed with replicates. Data was analyzed by two tailed student’s t-test (alpha = 0.05).

Article Snippet: 10-day-old seedlings were washed in PBS (Biological Industries), transferred to 6 ml liquid MSgg medium of each well of a 6-well microplate (Thermo Scientific), and then grown for additional four days.

Techniques: Cell Culture, Microscopy, Two Tailed Test

Journal: bioRxiv

Article Title: Arabidopsis thaliana induces multigenerational stress tolerance against biotic and abiotic stressors and memorization of host colonization in Bacillus subtilis

doi: 10.1101/2022.05.29.493878

Figure Lengend Snippet: B. subtilis colonization of A. thaliana and tomato roots after 24h with and without secretion treatment (A-B). Secretion-treated and untreated WT bacteria (+sec and -sec respectively) and their descendants were cultured separately in liquid MSgg with A. thaliana seedlings in the absence or presence of NaCl. After 24h, the bacteria colonizing the roots were quantified with CFU counts. Data is expressed as average values ± standard deviations of the means from all independent experiments.

Article Snippet: 10-day-old seedlings were washed in PBS (Biological Industries), transferred to 6 ml liquid MSgg medium of each well of a 6-well microplate (Thermo Scientific), and then grown for additional four days.

Techniques: Cell Culture

Journal: bioRxiv

Article Title: Arabidopsis thaliana induces multigenerational stress tolerance against biotic and abiotic stressors and memorization of host colonization in Bacillus subtilis

doi: 10.1101/2022.05.29.493878

Figure Lengend Snippet: (A) Fluorescently-labeled bacteria treated or untreated with secretions were cultured together in liquid MSgg with A. thaliana seedlings in the absence or presence of NaCl. The percentage of secretion-treated and untreated ancestor bacteria on the root was quantified with FACS. Representative bright field and fluorescent images of secretion-treated and untreated ancestor bacteria on A. thaliana roots in the absence (B) and presence (C) of NaCl. (D) The percentage of secretion-treated versus untreated bacteria on tomato roots without NaCl quantified with FACS (as described above). (E) The percentage of secretion-treated versus untreated descendant bacteria on A. thaliana roots without NaCl. Descendant bacteria were grown and treated as described in and quantified with FACS (as described above). All experiments were performed in triplicates at least three separate and independent times. Data is expressed as average values ± standard deviations of the means from representative experiment, done in technical triplicates. Data was analyzed by two tailed student’s t-test (alpha = 0.05).

Article Snippet: 10-day-old seedlings were washed in PBS (Biological Industries), transferred to 6 ml liquid MSgg medium of each well of a 6-well microplate (Thermo Scientific), and then grown for additional four days.

Techniques: Labeling, Cell Culture, Two Tailed Test