Comparison of media and growth conditions for culturing enterohepaticHelicobacterspecies

Abstract

This research aims to compare the culturing conditions for enterohepatic Helicobacter, evaluating culture media, incubation atmosphere and susceptibility to antimicrobials used to generate selective conditions. Four common media for the closely related genus Campylobacter (Columbia, Bolton, Brucella and CCDA agar), as well as the need for hydrogen in the microaerobic incubation atmosphere, were evaluated. Serial dilutions of 13 strains belonging to six species (H. apodemus, H. bilis, H. canicola, H. canis, H. equorum and Helicobacter sp.) were inoculated in each media and incubated at 37°C for 48 to 96 h using CampyGen (OXOID) and gaseous exchange (including hydrogen) in parallel. Columbia or Brucella agars were the most appropriate for culturing EHH (P < 0·05). However, there was no significant difference between the atmospheres evaluated (P = 0·13). In addition, minimal inhibitory concentration for six antibiotics showed that all isolates were resistant to trimethoprim, whereas for the rest of the antibiotics (cephalothin, cefoperazone, cefsulodin, teicoplanin and vancomycin) the inhibition range was between 8 and 64 μg ml−1. Our findings suggest that Columbia or Brucella media, regardless of the use of hydrogen, can be used for the EHH isolation. In addition, the concentration of antibiotics included in commercial campylobacteria supplements is suitable for EHH species recovery. Significance and Impact of the Study Enterohepatic Helicobacter (EHH) infections have been associated with several diseases in humans such as acute gastroenteritis, inflammatory bowel disease and hepatobiliary diseases. Although they are frequently detected in clinical samples by molecular methods, only occasionally they are isolated using culture conditions described for the taxonomic related pathogen Campylobacter sp. This is because the optimal conditions for the isolation of EHH have not yet been described, which results in an underestimation of the prevalence and clinical importance of these emerging pathogens. Therefore, this study provides insight for culturing EHH species.

Systematic Culturomics Shows that Half of Chicken Caecal Microbiota Members can be Grown in Vitro Except for Two Lineages of Clostridiales and a Single Lineage of Bacteroidetes

“…This number is close to similar studies [13,14]. In fact, the real number of chicken gut microbiota which can be grown in vitro is likely higher since some bacterial genera like Helicobacter and Campylobacter did not grow under culture conditions which were tested in this study though specific culture conditions for these genera are known [3,23]. Moreover, the ability to culture decreased with the decreasing abundance of the OTU in the initial caecal sample.…”

Exploring the role of healthy dogs as hosts of enterohepatic Helicobacter species using cultivation‐dependent and ‐independent approaches

“…Faecal samples were cultivated on Columbia agar (Oxoid) prepared with the addition of 5% sheep blood (CBA) and the selective supplement CAT (cefoperazone, teicoplanin and amphotericin B, Oxoid) by direct inoculation (Ochoa et al, 2019). Also, the samples were homogenized with physiological saline and 400 μl of samples were filtered through 0.45μm pore nitrocellulose filters (Millipore) on antibiotic-free CBA.…”

“…Detection of Helicobacter species was performed in the present study using the methodology described by Ochoa et al, 2019. Greater cultivation of EHH was seen using CBA supplemented with CAT (7.4%, 29/390) compared to filtration on non-selective CBA…”

Review: Other Helicobacter species

“…Although they are frequently detected in clinical samples by molecular methods, these fastidious organisms are rarely isolated by traditional culture technique, resulting in an underestimation of the prevalence and clinical importance of these pathogens. A study compared media and growth conditions for culturing EHH, ie Helicobacter canicola , Helicobacter apodemus , Helicobacter bilis , Helicobacter canis , Helicobacter equorum and other Helicobacter species 6 . Regardless of the use of hydrogen, Columbia or Brucella media can be used for the EHH isolation.…”

“…Regardless of the use of hydrogen, Columbia or Brucella media can be used for the EHH isolation. All isolates were resistant to trimethoprim but the antibiotic concentrations included in commercial supplements for Campylobacter culture are suitable for EHH culture 6 . Among EHH, Helicobacter cinaedi is most frequently associated with human diseases.…”