Journal: eLife

Article Title: Respiratory tissue-associated commensal bacteria offer therapeutic potential against pneumococcal colonization

doi: 10.7554/eLife.53581

Figure Lengend Snippet: ( A ) S. pneumoniae cultures were grown in presence of fresh (FM) or L. murinus ( Lm CM) or E. coli ( Ec CM) conditioned media. Culture growth was followed hourly by optical density at 600 nm (OD600) for 12 hr. Pooled data from two independent experiments are given as mean ± SD. Student t-test was applied for significance test. ( B ) S. pneumonia cultures were grown for 6 hr in presence of FM, Lm CM, or Ec CM. Culture growth was measured by OD600. Representative data from two independent experiments are depicted as mean ± sd (n = 3). Student t-test was applied for significance test in comparison to fresh media treated group.( C ) Following incubation in presence of FM, Lm CM, or Ec CM for 6 hr (note, the same OD was taken for the efficiency of plating at the 6 hr time point), S. pneumonia cultures are serially diluted and growth on fresh TSB agar plate with 5% sheep blood for 24 hr. Representative images are shown. ( D ) Different S. pneumonia strains, that is the virulent encapsulated strain (ATCC6303), non-encapsulated clinical isolate (110.58) and a derivate of the non-encapsulated avirulent lab strain R6 (R-6 derivative), or ( E ) Staphylococcus aureus (USA300) cultures were grown in presence of FM, Lm CM, or Ec CM for 6 hr. Culture growth was measured by OD600. Representative data from two independent experiments are depicted as mean ± SD. Student t-test was applied for significance test in comparison to fresh media group of each bacterial strain.

Article Snippet: Streptococcus pneumoniae strains, i.e. antigenic type 3 (ATCC-6303, LGC, Germany), Swiss non-encapsulated nasopharyngeal pneumococcal isolate 110.58 (MLST 344) and R6-derivative strain, were cultivated on Trypticase soy agar plates (bioMerieux, France) with 5% sheep blood (bioMerieux, France) at 37°C with 5% CO 2 .

Techniques: Comparison, Incubation

Journal: eLife

Article Title: Respiratory tissue-associated commensal bacteria offer therapeutic potential against pneumococcal colonization

doi: 10.7554/eLife.53581

Figure Lengend Snippet: S. pneumonia cultures were grown to 0.6 OD600 in presence of FM, Lm CM or Ec CM. Cellular chain lengths of bacterial cultures are quantified using light microscopy. Each symbol represents an individual multicellular S. pneumoniae chain. Data was pooled from three independent experiments. Number of chains evaluated for each group indicated on the graph ( n ). Student t-test is applied for statistical analysis. Representative images are shown.

Article Snippet: Streptococcus pneumoniae strains, i.e. antigenic type 3 (ATCC-6303, LGC, Germany), Swiss non-encapsulated nasopharyngeal pneumococcal isolate 110.58 (MLST 344) and R6-derivative strain, were cultivated on Trypticase soy agar plates (bioMerieux, France) with 5% sheep blood (bioMerieux, France) at 37°C with 5% CO 2 .

Techniques: Light Microscopy

Journal: eLife

Article Title: Respiratory tissue-associated commensal bacteria offer therapeutic potential against pneumococcal colonization

doi: 10.7554/eLife.53581

Figure Lengend Snippet: Hydrogen peroxide, causing similar growth inhibition on S. pneumoniae cultures, is not the active substance in Lm CM. ( A ) S. pneumoniae cultures grown for 6 hr in presence of FM or media conditioned by L. murinus in aerobic (+O 2 ) or hypoxic environment (-O 2 ). Culture growth was measured by OD600. Representative data from two independent experiments are depicted as mean ± SD (n = 3). Student t-test was applied for significance test. ( B ) S. pneumoniae cultures grown for 6 hr in presence of mock or catalase treated FM, Lm CM or Hydrogen peroxide solution (H 2 O 2 , 3.26 mM). Culture growth was measured by OD600. Mean ± SD are depicted in the graph (n = 3). Student t-test was applied for significance test in comparison to fresh media treated group unless otherwise stated by the arrows.

Article Snippet: Streptococcus pneumoniae strains, i.e. antigenic type 3 (ATCC-6303, LGC, Germany), Swiss non-encapsulated nasopharyngeal pneumococcal isolate 110.58 (MLST 344) and R6-derivative strain, were cultivated on Trypticase soy agar plates (bioMerieux, France) with 5% sheep blood (bioMerieux, France) at 37°C with 5% CO 2 .

Techniques: Inhibition, Comparison

Journal: eLife

Article Title: Respiratory tissue-associated commensal bacteria offer therapeutic potential against pneumococcal colonization

doi: 10.7554/eLife.53581

Figure Lengend Snippet: Lactic acid present in Lm CM is responsible for growth inhibition in S. pneumoniae cultures. ( A ) pH measurements of fresh (FM) or L. murinus conditioned (LmCM) media. Data is pooled from three independent experiments and depicted as mean ± SD (n = 3). ( B ) S. pneumonia cultures were grown for 6 hr in presence of FM, Lm CM or pH adjusted Lm CM (pH:~6.2, NaOH). Culture growth was measured by OD600. Representative data from two independent experiments are depicted as mean ± SD (n = 3). Student t-test was applied for significance test in comparison to fresh media treated group unless otherwise stated by the arrows. ( C ) Representative GC-MS total ion chromatograms of FM (dashed blue line) and Lm CM (straight red line) are shown. Concentrations of Lactate in each media, calculated based on a standard curve acquired using DL-Lactic acid, depicted in the graph (Mean ± SD, n = 3). Student t-test was applied for significance test. ( D ) S. pneumonia cultures were grown for 6 hr in presence of FM, LmCM, DL-Lactic acid (DLL, 100 mM) or pH adjusted LmCM (pH:~6.2, NaOH) and pH adjusted DLL (pH:~6.2, NaOH). Culture growth was measured by OD600. Representative data from two independent experiments are depicted as mean ± SD (n = 3). Student t-test was applied for significance test in comparison to fresh media treated group unless otherwise stated by the arrows.

Article Snippet: Streptococcus pneumoniae strains, i.e. antigenic type 3 (ATCC-6303, LGC, Germany), Swiss non-encapsulated nasopharyngeal pneumococcal isolate 110.58 (MLST 344) and R6-derivative strain, were cultivated on Trypticase soy agar plates (bioMerieux, France) with 5% sheep blood (bioMerieux, France) at 37°C with 5% CO 2 .

Techniques: Inhibition, Comparison, Gas Chromatography-Mass Spectrometry

Journal: eLife

Article Title: Respiratory tissue-associated commensal bacteria offer therapeutic potential against pneumococcal colonization

doi: 10.7554/eLife.53581

Figure Lengend Snippet: S. pneumonia cultures were grown to 0.6 OD600 in presence of ( A ) FM, Lm CM or DL-Lactic acid (DLL,~100 mM), ( B ) FM, Lm CM or pH adjusted Lm CM (pH:~6.2, NaOH) and ( C ) mock or proteinase K (proK) treated FM and Lm CM. Cellular chain lengths of bacterial cultures are quantified using light microscopy. Each symbol represents an individual multicellular S. pneumoniae chain. Data was pooled from three independent experiments. Number of chains evaluated for each group indicated on the graph (n). Student t-test is applied for statistical analysis.

Article Snippet: Streptococcus pneumoniae strains, i.e. antigenic type 3 (ATCC-6303, LGC, Germany), Swiss non-encapsulated nasopharyngeal pneumococcal isolate 110.58 (MLST 344) and R6-derivative strain, were cultivated on Trypticase soy agar plates (bioMerieux, France) with 5% sheep blood (bioMerieux, France) at 37°C with 5% CO 2 .

Techniques: Light Microscopy

Journal: eLife

Article Title: Respiratory tissue-associated commensal bacteria offer therapeutic potential against pneumococcal colonization

doi: 10.7554/eLife.53581

Figure Lengend Snippet: ( A ) Germ-free (GF) or specific pathogen-free (SPF) mice were intranasally administered with PBS or L. murinus (~10 5 cfu/animal). Three days post-administration, mice were challenged with S. pneumoniae (~1500 cfu/animal). S. pneumoniae lung titers (cfu/organ) of L. murinus (half black/half white) or PBS (plain black) administered GF (circles) or SPF (squares) are quantified 24 hr post infection (n = 10). Medians are depicted by a black line for each group. Pooled data from two independent experiments are shown. BALF pH does not significantly differ between colonized and GF mice (right panel). Median pH values for indicated mice are shown. Each dot represents one animal. Mann-Whitney test is applied for statistical analysis.( B ) Average relative initial body weight (%)± SD is depicted for mock treated (black line with circles) and IAV infected (red line with squares) mice (n = 5). Mice were challenged with S. pneumoniae (~1500 cfu/animal) 10 days post IAV infection. S. pneumoniae lung titers (cfu/organ) of mock treated (black circles) or IAV infected (red squares) are quantified 24 hr post infection (n = 5). Medians are depicted by a black line for each group. Student t-test was applied for significance test. ( C ) All mice are infected with IAV (40 pfu). Seven days post IAV infection, PBS (red line with red squares) or L. murinus (~10 5 cfu/animal, black line with red squares) is intranasally administered to the animals. Average relative initial body weight (%)± SD is depicted for both group (n = 10). Mice were challenged with S. pneumoniae (~1500 cfu/animal) 10 days post IAV challenge. S. pneumoniae lung titers (cfu/organ) of IAV infected, PBS (red squares) or L. murinus administered (black rounded red squares) are quantified 24 hr post infection (n = 10). Medians are depicted by a black line for each group. Pooled data from two independent experiments are shown. Student t-test was applied for significance test. ( D ) pH of BALF from mock treated or L. murinus colonized mice 3 days post colonization and 10 days post IAV vaccination (n = 7–8 animals/group).

Article Snippet: Streptococcus pneumoniae strains, i.e. antigenic type 3 (ATCC-6303, LGC, Germany), Swiss non-encapsulated nasopharyngeal pneumococcal isolate 110.58 (MLST 344) and R6-derivative strain, were cultivated on Trypticase soy agar plates (bioMerieux, France) with 5% sheep blood (bioMerieux, France) at 37°C with 5% CO 2 .

Techniques: Infection, MANN-WHITNEY