First screening for Brachyspira hampsonii in Swiss pigs applying a new high resolution melting assay.
A new High Resolution Melting (HRM) assay was developed for the rapid detection of Brachyspira (B.) hampsonii. B. hampsonii occurs in different European countries, however, until today it has not been encountered in Switzerland. Four B. hampsonii reference strains were used to develop the HRM assay: B. hampsonii clade I ATCC BAA2463 and clade II ATCC BAA2464 strain, as well as two isolated strains P280/1 from the UK and the German isolate 5369-1x/12. A conserved region of the nox gene was used to design B. hampsonii-specific primers. The HRM melting curves for the four reference strains showed reproducible difference graphs with distinct differences between the four strains based on a slight variation between the four amplicon sequences. In addition, DNA from 22 B. hampsonii strains representing four genetic B. hampsonii groups was used to validate the method. Melting temperatures in the interval between 73.1 and 74°C were obtained for all B. hampsonii strains and allow differentiating B. hampsonii from other Brachyspira species. In total 897 Swiss porcine fecal Brachyspira isolates, cultured between 2009 and 2015, were analysed by the HRM protocol. B. hampsonii was not detected among these Swiss Brachyspira isolates. In conclusion, the rapid and low-cost HRM approach allows a sensitive and specific identification of B. hampsonii.
Institute of Veterinary Bacteriology, Vetsuisse Faculty, University of Zurich, Winterthurerstrasse 270, CH 8057 Zurich, Switzerland. Electronic address: simone.scherrer@vetbakt.uzh.ch.
Simone
S
Scherrer
Institute of Veterinary Bacteriology, Vetsuisse Faculty, University of Zurich, Winterthurerstrasse 270, CH 8057 Zurich, Switzerland.
Anna
A
Borgström
Institute of Veterinary Bacteriology, Vetsuisse Faculty, University of Zurich, Winterthurerstrasse 270, CH 8057 Zurich, Switzerland.
Daniel
D
Frei
Institute of Veterinary Bacteriology, Vetsuisse Faculty, University of Zurich, Winterthurerstrasse 270, CH 8057 Zurich, Switzerland.
Max M
MM
Wittenbrink
D000818:Animals; D017400:Brachyspira / Q000235:genetics / Q000302:isolation & purification*; D005243:Feces / Q000382:microbiology; D016905:Gram-Negative Bacterial Infections / Q000175:diagnosis / Q000382:microbiology / Q000662:veterinary*; D016415:Sequence Alignment / Q000662:veterinary; D017422:Sequence Analysis, DNA / Q000662:veterinary; D013552:Swine; D013553:Swine Diseases / Q000175:diagnosis / Q000382:microbiology*; D013557:Switzerland
Brachyspira hampsonii; High resolution melting assay; Swine dysentery; nox gene