Gaspak em

Phocaeicola dorei 5_1_36 and Lachnoclostridium symbiosum WAL-14673 were obtained from BEI Resources (HM-29 and HM-319). For inoculations and initial tests, both microorganisms were cultured in Reinforced Clostridium Media (RCM; Becton, Dickinson, Franklin Lakes, NJ) supplemented with 0.5 g/liter of l-cysteine (RCM-cys; Loba Chemie, India). All incubations were performed at 37°C for 24 to 48 h in an anaerobic jar (Anaerocult; Merck, Darmstadt, Germany) with anaerobic packs (Gaspak EM; Becton, Dickinson, Franklin Lakes, NJ, USA). mZMB medium was prepared as in reference 48 (link).

Strains used in this study were obtained from the UC Davis Viticulture & Enology Culture Collection (Bifidobacterium longum subsp. infantis ATCC 15697, Lactobacillus acidophilus ATCC 4356, Escherichia coli K12), and the American Type Culture Collection (Bacteroides vulgatus ATCC 8482; Manassas, VA, USA). For routine experiments, bifidobacteria were grown on de Mann-Rogose-Sharp (MRS) broth supplemented with 0.05% w/vl-cysteine (Loba Chemie, Maharashtra, India) and incubated for 48 h at 37 °C in an anaerobic jar (Anaerocult, Merck, Darmstadt, Germany) with anaerobic packs (Gaspak EM, Becton-Dickinson, Franklin Lakes, NJ, USA). Lactobacilli were grown under the same conditions but without l-cysteine. Bacteroides strains were grown anaerobically using Reinforced Clostridium Medium (Becton-Dickinson) supplemented with 1 g·L⁻¹l-cysteine. All media were pre-reduced in an anaerobic jar overnight before inoculation. E. coli was routinely grown on Standards Methods Broth (Becton-Dickinson). Prior to each assay all bacteria were subcultured twice. All chemicals were acquired from Merck, Sigma or Calbiochem.

C. baratii C3 was routinely cultured in Reinforced Clostridium Medium (RCM, Becton-Dickinson, Franklin Lakes, NJ, USA) supplemented with 0.5 g/L of L-cysteine (RCM-cys; Loba Chemie, India). All incubations were performed at 37 °C for 24–48 h in an anaerobic jar (Anaerocult, Merck, Darmstadt, Germany) with anaerobic packs (Gaspak EM, Becton-Dickinson, Franklin Lakes, NJ, USA). Two overnight cultures of C. baratii were made on Blood Agar and Starch Agar plates (Winkler, Santiago, Chile). The bacteria were also inoculated in API-20A strips (Biomerieux, Craponne, France) following the manufacturer’s instructions. Finally, antibiotic resistance patterns were evaluated using antibiotic discs in RCM agar plates incubated for 48 h (Lilfilchem, Roseto degli Abruzzi TE, Italy) for 48 h. Cefazolin (30 µg), ciprofloxacin (5 µg), chloramphenicol (30 µg), erythromycin (15 µg), penicillin G (10 µg), piperacill/tazobactam (110 µg), trimethoprim-sulfamethoxazole (25 µg), and vancomycin (30 µg) were used.