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MB Sample ID: SA416006
| Local Sample ID: | Titration-10 |
| Subject ID: | SU003933 |
| Subject Type: | Bacteria |
| Subject Species: | Dorea longicatena |
| Taxonomy ID: | 88431 |
| Genotype Strain: | Flavonifractor plautii, Dorea longicatena, Turicibacter sanguinis, Clostridium bolteae, Clostridium symbiosum, Clostridium innocuum, Clostridium clostridioforme, Streptococcus parasanguinis |
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Collection:
| Collection ID: | CO003926 |
| Collection Summary: | All strains were cultured anaerobically at 37°C in different media types according to the experiment being performed. The media compositions used in this study include BHI-R, CHG-R, Gifu-R, and Mega-R, each formulated with specific components to support microbial growth. BHI-R was prepared by dissolving 18.5 g of Brain Heart Infusion (BHI) and 5 g of L-arginine in 500 mL of Milli-Q water, followed by sterilization via autoclaving (10 lbs, 115°C, 15 minutes). CHG-R was based on CHG media, which contained 18.5 g of BHI, 0.5 g each of D-(+)-cellobiose, D-(+)-maltose monohydrate, and D-(-)-fructose, and 0.25 g of cysteine in 500 mL of Milli-Q water. After filter sterilization, additional supplements, including 5 mL each of Vitamin K + Hematin (1%), trace mineral supplement, and vitamin supplement, were added in an anaerobic chamber. Gifu-R was derived from commercially available Gifu Anaerobic Media (GAM) (Himedia, Cat # M1801), prepared by suspending 59 g of GAM in 1000 mL of purified/distilled water and autoclaving under the same conditions as BHI-R. Mega-R was formulated with a complex mixture of components, including 5 g of Trypticase Peptone (BBL), 2.5 g each of Yeast Extract (Bacto) and Meat Extract, and multiple carbohydrate sources such as 1 g of D-(+)-glucose and 0.5 g each of D-(+)-cellobiose, D-(+)-maltose monohydrate, and D-(-)-fructose, all dissolved in 500 mL of Milli-Q water. Additional components included 50 mL of 1 M potassium phosphate buffer (pH 7.2), 20 mL of TYG salts solution, 1 mL of 25% Tween 80, and various supplements such as SCFA, CaCl₂, FeSO₄, and resazurin. Like CHG-R, Mega-R was supplemented with 5 mL each of Vitamin K + Hematin (1%), trace mineral supplement, and vitamin supplement. For CHG-R, Gifu-R, and Mega-R, an additional 5 g of L-arginine was incorporated into 500 mL of each respective medium to create their corresponding arginine-enriched formulations. All cultures were grown in an anaerobic chamber (Coy Laboratory Products) with an atmosphere of 5% CO2, 5% H2, and 90% N2 at 37°C. The isolates were streaked onto CHG plates and incubated for 48 hours. Once colonies were visible, a single colony from each plate was picked and processed for 16S rRNA Sanger sequencing using primer sequences 27F: AGAGTTTGATCMTGGCTCAG and 1492R: GGTTACCTTGTTACGACTT. Once the identity of the bacterial strains was confirmed, a single colony from each strain was inoculated in triplicates in 6 mL of the respective media types (CHG, Gifu, Mega) and incubated overnight. The overnight cultures were inoculated into fresh media and normalized to the OD600 of 0.05. At each time point after inoculation (6h, 24h, 48 h), 200 μL of the bacterial culture were collected. |
| Sample Type: | Bacterial cells |
