Gas-pak

Published in Ronald M. Atlas, James W. Snyder, Handbook Of Media for Clinical Microbiology, 2006

Ronald M. Atlas, James W. Snyder

Preparation of Medium: Add components, exhemin solution and vitamin K1 solution, to distilled/ deionized water and bring volume to 1.0L. Mix thoroughly. Gently heat and bring to boiling. Adjust pH to 7.0 ± 0.2 at 25°C. Autoclave for 15 min at 15 psi pressure-121°C. Cool to 45°-50°C. Aseptically add 0.5mL of sterile hemin solution and 1.0mL of sterile vitamin K1 solution. Mix thoroughly. Pour into sterile Petri dishes or distribute into sterile tubes. Reduce medium for 24h by incubation in an anaerobic glove box of GasPak jar prior to use.

Mucin degrader Akkermansia muciniphila accelerates intestinal stem cell-mediated epithelial development

Published in Gut Microbes, 2021

Seungil Kim, Yun-Chan Shin, Tae-Young Kim, Yeji Kim, Yong-Soo Lee, Su-Hyun Lee, Mi-Na Kim, Eunju O, Kwang Soon Kim, Mi-Na Kweon

A total of 32 fecal samples were obtained from the human dock center of the Asan Medical Center. The samples were collected from fresh residual samples after fecal occult blood and parasitic examination on the same date. Fecal samples were suspended in PBS and then seeded onto brain heart infusion agar without dextrose (Kisan-Bio, Seoul, South Korea) supplemented with 0.4% mucin (BHI-M). The fecal cultures were maintained at 37°C under anaerobic conditions generated using a GasPak 100 system (BD Bioscience). Approximately 50 colonies were selected from BHI-M plates and tested with PCR for species-specific sequences with the primer set, 5′-CAGCACGTGAAGGTGGGGAC-3′ and 5′-CCTTGCGGTTGGCTTCAGAT-3′. A. muciniphila was isolated from 11 samples. One strain was established per sample by sub-culturing for future experimentation.

Multi-targeted properties of the probiotic saccharomyces cerevisiae CNCM I-3856 against enterotoxigenic escherichia coli (ETEC) H10407 pathogenesis across human gut models

Published in Gut Microbes, 2021

Charlène Roussel, Kim De Paepe, Wessam Galia, Jana de Bodt, Sandrine Chalancon, Sylvain Denis, Françoise Leriche, Pascal Vandekerkove, Nathalie Ballet, Stéphanie Blanquet-Diot, Tom Van de Wiele

The M-SHIME® consists of a series of connected double-jacketed reactors (Pierreglas, Vilvoorde, Belgium) mimicking conditions of the upper and lower part of the human gastrointestinal tract, operated in a semi-continuous mode to mimic gastrointestinal transit.66,67 Three successive compartments simulating the combined stomach/duodenum-jejunum, the ileum and the ascending colon were run in parallel for a probiotic vs control condition (Figure 1b). Only ileum and ascending colon were inoculated with fecal microbiota. To capture interindividual variability in ETEC behavior and probiotic effects, six healthy adults were inoculated the M-SHIME (3 women and 3 men aged between 25 and 36 years old, including Belgian, African, Turkish, and French origins with two vegetarians). Fresh fecal samples were collected in sterile airtight containers comprising anaerogen bags (BD GasPak™, Erembodegem, Belgium). Consent for fecal collection was obtained under registration number BE670201836318 (Gent University). A 20% (w/v) fecal slurry was prepared as previously described.67 All vessels were flushed with N2 immediately after inoculation to generate anaerobic conditions. In addition, the mucosal environment was mimicked in both ileum and ascending colon compartments, through the incorporation of microcosms (AnoxKaldnes K1 carrier, Lund, Sweden) coated with type III porcine mucin-agar (Sigma͐–aldrich, St. Louis, US), instead of type II mucin, as described by Van den Abbeele et al.68 Functioning of the system, mucin carrier replacement and media composition have been presented in Roussel et al.65

Motility and biofilm formation of the emerging gastrointestinal pathogen Campylobacter concisus differs under microaerophilic and anaerobic environments

Published in Gut Microbes, 2019

Sandra Ovesen, Juliana Durack, Karina Frahm Kirk, Hans Linde Nielsen, Henrik Nielsen, Susan V. Lynch

A total of 46 clinical isolates of C. concisus were cultured on 5% Horse Blood Agar plates with 1% added yeast extract for 48 hours under microaerophilic conditions (80% N2, 10% CO2, 5% H2, 5% O2). Subsequently, these isolates were cultured in Brain Heart Infusion broth (BHI) under either anaerobic (85% N2, 5% CO2, 10% H2, 0.05%< O2) or microaerophilic conditions. Anaerobic (AN) atmosphere was maintained in an anaerobic chamber (Coy Laboratory Products). Microaerophilic (MA) atmosphere was generated using CampyGen and CO2Gen gas generating kits (BD GasPaK™ EZ) and 0.2g sodium borohydride powder (Aldrich) dissolved in 10ml of sterile milli-Q water.