C. jejuni strain 81-176 was thawed from frozen stocks and grown on columbia agar (with 5% sheep blood) and selective karmali agar plates (both from Oxoid, Wesel, Germany). Age- and sex-matched microbiota-depleted IL-10-/- mice (4-month-old littermates) were infected perorally with 109 colony forming units (CFU) of the pathogen in 0.3 mL sterile phosphate-buffered saline (PBS, Thermo Fisher Scientific, Waltham, MA, USA) on days 0 and 1 by gavage.
Selective karmali agar plates
Manufactured by Thermo Fisher Scientific
Sourced in Germany
Selective karmali agar plates are a microbiological culture medium used for the isolation and identification of Campylobacter species from clinical and environmental samples. The agar contains selective agents that inhibit the growth of non-target organisms, allowing for the selective growth of Campylobacter species.
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Lab products found in correlation
Sterile pbs, by Thermo Fisher Scientific (4 mentions)
Phosphate buffered saline (pbs), by Thermo Fisher Scientific (2 mentions)
Columbia agar, by Thermo Fisher Scientific (2 mentions)
Sterile phosphate buffered saline pbs, by Thermo Fisher Scientific (1 mentions)
Clove eo, by Merck Group (1 mentions)
Campygas packs, by Thermo Fisher Scientific (1 mentions)
11 protocols using selective karmali agar plates
1
Infection of Microbiota-Depleted Mice with C. jejuni
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C. jejuni strain 81-176 was thawed from frozen stocks and grown on columbia agar (with 5% sheep blood) and selective karmali agar plates (both from Oxoid, Wesel, Germany). Age- and sex-matched microbiota-depleted IL-10-/- mice (4-month-old littermates) were infected perorally with 109 colony forming units (CFU) of the pathogen in 0.3 mL sterile phosphate-buffered saline (PBS, Thermo Fisher Scientific, Waltham, MA, USA) on days 0 and 1 by gavage.
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C. jejuni strain 81-176 was thawed from frozen stocks and grown on columbia agar (with 5% sheep blood) and selective karmali agar plates (both from Oxoid, Wesel, Germany). Age- and sex-matched microbiota-depleted IL-10-/- mice (4-month-old littermates) were infected perorally with 109 colony forming units (CFU) of the pathogen in 0.3 mL sterile phosphate-buffered saline (PBS, Thermo Fisher Scientific, Waltham, MA, USA) on days 0 and 1 by gavage.
2
Campylobacter Infection in IL-10 Knockout Mice
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On days 0 and 1, 4-month-old secondary abiotic IL-10-/- mice (sex-matched littermates; balanced gender ratio) were infected perorally with 109 colony forming units (CFU) of the C. jejuni strain 81–176 grown on Columbia agar (with 5% sheep blood) and selective karmali agar plates (both from Oxoid, Wesel, Germany). Therefore, bacterial colonies were harvested with a sterile swab from respective agar plates after 48-h incubation under microaerophilic conditions and transferred to sterile phosphate-buffered saline (PBS, Thermo Fisher Scientific, Waltham, MA, USA). Immediately thereafter, mice received 0.3 mL of the bacterial suspension by gavage.
As described earlier, the daily pathogen loads were determined in fecal samples after C. jejuni infection and upon necropsy in luminal samples from the gastrointestinal tract (stomach, duodenum, ileum, and colon) by culture [16 (link),38 (link)]. Briefly, serial dilutions of each sample (in sterile PBS, Thermo Fisher Scientific, Waltham, MA, USA) were plated onto karmali agar and incubated under microaerophilic conditions for at least 48 h and at 37 °C. The detection limit of viable pathogens was 100 CFU per g (CFU/g).
As described earlier, the daily pathogen loads were determined in fecal samples after C. jejuni infection and upon necropsy in luminal samples from the gastrointestinal tract (stomach, duodenum, ileum, and colon) by culture [16 (link),38 (link)]. Briefly, serial dilutions of each sample (in sterile PBS, Thermo Fisher Scientific, Waltham, MA, USA) were plated onto karmali agar and incubated under microaerophilic conditions for at least 48 h and at 37 °C. The detection limit of viable pathogens was 100 CFU per g (CFU/g).
3
Clove Oil Mitigates C. jejuni Infection in Mice
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C. jejuni strain 81-176 was thawed from frozen stocks and grown on Columbia agar (with 5% sheep blood) and selective Karmali agar plates (both from Oxoid, Wesel, Germany). Age- and sex-matched microbiota-depleted IL-10−/− mice (4-month-old littermates) were infected perorally with 109 colony forming units (CFU) of the pathogen on days 0 and 1 by gavage. For one experiment, mice were assigned to three groups: uninfected und untreated (i.e., naive) group (n = 4); C. jejuni infected group treated with placebo (n = 4); and the infected group treated with clove EO (100 mg per kg body weight per day, n = 4). Treatment with clove EO (purchased from Sigma-Aldrich, Munich, Germany) was performed from day 2 post-infection (p.i.) until the end of the observation period and applied to autoclaved tap water (final concentration of 5 g/L; ad libitum). The placebo control mice received autoclaved tap water only.
4
Murine Model of Campylobacter Infection
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The C. jejuni strain 81-176 used for the experiments was grown on selective karmali agar plates (from Oxoid, Wesel, Germany). Age- and sex-matched microbiota-depleted IL-10−/− mice (3-month-old littermates) were infected perorally with 109 colony forming units (CFU) of the pathogen on days 0 and 1 by gavage. Mice were treated with DESF (purchased from Sigma-Aldrich, Munich, Germany) added to the autoclaved drinking water (final concentration of 500 mg/L; ad libitum) starting 7 days prior infection. This DESF concentration was chosen to exceed the iron concentration of ≈160 mg/kg in the mice feeds (standard food pellets: sniff R/M-H, V1534-300; Sniff, Soest, Germany; ad libitum). Mice from the placebo (PLC) cohort received autoclaved tap water only.
5
Establishing C. jejuni Infection in Mice
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C. jejuni strain 81-176 was thawed from frozen stocks and grown on selective karmali agar plates (Oxoid, Wesel, Germany). Age- and sex-matched hma and SAB IL-10−/− mice (3-month-old littermates) were infected perorally with 109 colony-forming units (CFU) of the pathogen on days 0 and 1 by gavage (0.3 mL total volume).
6
Infection of Humanized Mice with C. jejuni
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C. jejuni strain 81-176 bacterial stocks were stored at −80 °C. After thawing, the bacteria were streaked out and incubated on selective karmali agar plates (purchased from Oxoid, Wesel, Germany) at 37 °C for 48 h in a jar under microaerophilic conditions (CampyGas Packs; Oxoid, Wesel, Germany). Grown C. jejuni bacteria were harvested in sterile PBS (Thermo Fisher Scientific, Waltham, MA, USA) immediately before infection. Age- and sex-matched human intestinal microbiota-associated (hma) IL-10−/− mice (3-month-old littermates) were then infected perorally with 109 colony forming units (CFU) of the pathogen (in 0.3 mL) on d0 and d1 (Fig. 1A ) by gavage as reported earlier [48 (link)].
7
Colonization of IL-10 Deficient Mice with C. jejuni
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Viable C. jejuni strain 81-176 bacteria stored at −80 °C were thawed, streaked out, and incubated on selective karmali agar plates (purchased from Oxoid, Wesel, Germany) at 37 °C for 48 h under micro-aerophilic conditions. In order to generate an inoculum of 109 bacterial cells, bacteria were harvested in sterile PBS (Thermo Fisher Scientific, Waltham, MA, USA). Age- and sex-matched human-gut-microbiota-associated IL-10−/− mice (3-month-old littermates) were then infected perorally with 109 colony forming units (CFU) of the pathogen on two consecutive days (i.e., on days 0 and 1; Figure 1 ) by gavage as stated in detail elsewhere [57 (link)].
8
Colonization of IL-10-/- Mice by C. jejuni
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Viable C. jejuni strain 81-176 bacteria stored at −80 °C were thawed, streaked out, and incubated on selective karmali agar plates (purchased from Oxoid, Wesel, Germany) at 37 °C for 48 h under microaerophilic conditions. In order to generate an inoculum of 109 bacterial cells, bacteria were harvested in sterile PBS. Age- and sex-matched human intestinal microbiota-associated (hma) IL-10−/− mice (3-month-old littermates) were then infected perorally with 109 colony forming units (CFU) of the pathogen on two consecutive days (i.e., on d0 and d1; Fig. 1 ) by gavage as state in detail elsewhere [25 (link)].
9
Infecting Microbiota-Depleted IL-10 Knockout Mice with Campylobacter jejuni
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After thawing from a stock, C. jejuni strain 81–176 was grown on columbia agar (with 5% sheep blood) and selective karmali agar plates (both from Oxoid, Wesel, Germany) in a microaerophilic atmosphere for 48 h (37 °C). Colonies were harvested from a densely grown bacterial cell layer with a sterile cotton swab and transferred to sterile phosphate buffered saline (PBS; Thermo Fisher Scientific, Waltham, MA, USA). Microbiota-depleted IL-10−/− mice (four-month-old sex- and age-matched littermates) were infected perorally with 109 colony forming units (CFU) of the pathogen on days 0 and 1 by gavage.
10
Murine Campylobacter jejuni Infection
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After thawing from frozen stocks kept at −80 °C, C. jejuni strain 81–176 bacteria were cultivated for a minimum of 2 days at 37 °C in a microaerophilic environment on selective karmali agar plates (Oxoid, Wesel, Germany). As previously described [33 (link)], grown bacteria were transferred to sterile PBS (Thermo Fisher Scientific, Waltham, MA, USA) with sterile cotton swabs to create an inoculum of 109 bacterial cells. On days 0 and 1, hma IL-10−/− mice (12-week-old age- and sex-matched littermates) were gavaged for infection with 109 colony-forming units (CFU) of the pathogen (in 0.3 mL volume).
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