title: High-Resolution Melting Curve Analysis for Identification of Pasteurellaceae Species in Experimental Animal Facilities.
abstract: Pasteurellaceae are among the most prevalent bacterial pathogens isolated from mice housed in experimental animal facilities. Reliable detection and differentiation of Pasteurellaceae are essential for high-quality health monitoring. In this study, we combined a real-time PCR assay amplifying a variable region in the 16S rRNA sequence with high-resolution melting curve analysis (HRM) to identify and differentiate among the commonly isolated species Pasteurella pneumotropica biotypes "Jawetz" and "Heyl", Actinobacillus muris, and Haemophilus influenzaemurium. We used a set of six reference strains for assay development, with the melting profiles of these strains clearly distinguishable due to DNA sequence variations in the amplicon. For evaluation, we used real-time PCR/HRM to test 25 unknown Pasteurellaceae isolates obtained from an external diagnostic laboratory and found the results to be consistent with those of partial 16S rRNA sequencing. The real-time PCR/HRM method provides a sensitive, rapid, and closed-tube approach for Pasteurellaceae species identification for health monitoring of laboratory mice.
authors.affiliation: Research Unit Comparative Medicine, German Research Center for Environmental Health, Neuherberg, Germany.
authors.forename: Manuel
authors.initials: M
authors.lastname: Miller
authors.affiliation: Research Unit Comparative Medicine, German Research Center for Environmental Health, Neuherberg, Germany.
authors.forename: Julia
authors.initials: J
authors.lastname: Zorn
authors.affiliation: Research Unit Comparative Medicine, German Research Center for Environmental Health, Neuherberg, Germany.
authors.forename: Markus
authors.initials: M
authors.lastname: Brielmeier
mesh_terms: D000822:Animal Husbandry; D000818:Animals; D004269:DNA, Bacterial / Q000235:genetics; D006799:Housing, Animal; D051379:Mice; D015661:Pasteurellaceae / Q000302:isolation & purification*; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D012376:Rodent Diseases / Q000382:microbiology*
chemical_list: D004269:DNA, Bacterial; D012336:RNA, Ribosomal, 16S