Caroline Blaecher1, Eva Bauwens1, Alfred Tay2, Fanny Peters2, Sylvia Dobbs3,4,5, John Dobbs3,4,5, André Charlett3,6, Richard Ducatelle1, Freddy Haesebrouck1, Annemieke Smet1. 1. Department of Pathology, Bacteriology and Avian Diseases, Faculty of Veterinary Medicine, Ghent University, Merelbeke, Belgium. 2. The Marshall Centre for Infectious Diseases Research and Training, University of Western Australia, Perth, WA, Australia. 3. Institute of Pharmaceutical Science, King's College London, London, UK. 4. The Maudsley Hospital, London, UK. 5. Department of Gastroenterology, King's College Hospital, London, UK. 6. Statistics Unit, Centre for Infectious Disease Surveillance and Control, Public Health England, London, UK.
Abstract
BACKGROUND: Helicobacter suis is a very fastidious microorganism associated with gastritis, gastric ulcers, and mucosa-associated lymphoid tissue lymphoma in humans. In vitro isolation of this agent from human patients has so far been unsuccessful. MATERIALS AND METHODS: A probe-based real-time PCR (RT-PCR) for the rapid detection of H. suis in gastric biopsies was developed. Secondly, a mouse-passage-based protocol was optimized for isolation of low numbers of viable H. suis bacteria. Mice were inoculated with different numbers of viable H. suis (102 -108 ) and kept for 4 weeks to allow multiplication of this pathogen. RESULTS: The probe-based real-time PCR (RT-PCR) exhibited a high degree of diagnostic specificity and analytical sensitivity, high linear correlations (r2 between 0.995 and 0.999), and high amplification efficiencies (>90%) for H. suis. No cross-reactivity was detected with human, porcine, non-human primate, and murine DNA nor with DNA from other bacteria including Helicobacter spp. and Campylobacter spp. H. suis was successfully re-isolated from the stomach of mice inoculated with at least 104 viable H. suis, using a biphasic medium (pH 5), consisting of Brucella agar with Brucella broth on top, both supplemented with vitox supplement, Campylobacter-selective supplement, amphotericin (5 μg/mL), HCl (0.05%), fetal bovine serum (20%), and linezolid (5 μg/mL). Linezolid was necessary to inhibit proliferation of contaminants, including lactobacilli. CONCLUSION: The methods described above can be implemented for detection or isolation of H. suis from human gastric biopsies.
BACKGROUND:Helicobacter suis is a very fastidious microorganism associated with gastritis, gastric ulcers, and mucosa-associated lymphoid tissue lymphoma in humans. In vitro isolation of this agent from humanpatients has so far been unsuccessful. MATERIALS AND METHODS: A probe-based real-time PCR (RT-PCR) for the rapid detection of H. suis in gastric biopsies was developed. Secondly, a mouse-passage-based protocol was optimized for isolation of low numbers of viable H. suis bacteria. Mice were inoculated with different numbers of viable H. suis (102 -108 ) and kept for 4 weeks to allow multiplication of this pathogen. RESULTS: The probe-based real-time PCR (RT-PCR) exhibited a high degree of diagnostic specificity and analytical sensitivity, high linear correlations (r2 between 0.995 and 0.999), and high amplification efficiencies (>90%) for H. suis. No cross-reactivity was detected with human, porcine, non-human primate, and murine DNA nor with DNA from other bacteria including Helicobacter spp. and Campylobacter spp. H. suis was successfully re-isolated from the stomach of mice inoculated with at least 104 viable H. suis, using a biphasic medium (pH 5), consisting of Brucella agar with Brucella broth on top, both supplemented with vitox supplement, Campylobacter-selective supplement, amphotericin (5 μg/mL), HCl (0.05%), fetal bovine serum (20%), and linezolid (5 μg/mL). Linezolid was necessary to inhibit proliferation of contaminants, including lactobacilli. CONCLUSION: The methods described above can be implemented for detection or isolation of H. suis from human gastric biopsies.
Authors: Nicolas Feau; Stéphanie Beauseigle; Marie-Josée Bergeron; Guillaume J Bilodeau; Inanc Birol; Sandra Cervantes-Arango; Braham Dhillon; Angela L Dale; Padmini Herath; Steven J M Jones; Josyanne Lamarche; Dario I Ojeda; Monique L Sakalidis; Greg Taylor; Clement K M Tsui; Adnan Uzunovic; Hesther Yueh; Philippe Tanguay; Richard C Hamelin Journal: PeerJ Date: 2018-02-22 Impact factor: 2.984
Authors: Helena Berlamont; Chloë De Witte; Sofie De Bruyckere; James G Fox; Steffen Backert; Annemieke Smet; Filip Boyen; Freddy Haesebrouck Journal: Pathogens Date: 2021-03-18