Species Identification and Strain Typing of Staphylococcus agnetis and Staphylococcus hyicus Isolates from Bovine Milk by Use of a Novel Multiplex PCR Assay and Pulsed-Field Gel Electrophoresis.
Staphylococcus hyicusandStaphylococcus agnetisare two coagulase-variable staphylococcal species that can be isolated from bovine milk and are difficult to differentiate. The objectives of this study were to characterize isolates of bovine milk origin from a collection that had previously been characterized as coagulase-positiveS. hyicusbased on phenotypic species identification methods and to develop a PCR-based method for differentiatingS. hyicus,S. agnetis, andStaphylococcus aureusIsolates (n= 62) were selected from a previous study in which milk samples were collected from cows on 15 dairy herds. Isolates were coagulase tested and identified to the species level using housekeeping gene sequencing. A multiplex PCR to differentiateS. hyicus,S. agnetis, andS. aureuswas developed. Pulsed-field gel electrophoresis was conducted to strain type the isolates. Based on gene sequencing, 44/62 of the isolates were determined to be eitherS. agnetis(n= 43) orS. hyicus(n= 1). Overall, 88% (37/42) of coagulase-positiveS. agnetisisolates were found to be coagulase positive at 4 h. The herd-level prevalence of coagulase-positiveS. agnetisranged from 0 to 2.17%. Strain typing identified 23 different strains. Six strains were identified more than once and from multiple cows within the herd. Three strains were isolated from cows at more than one time point, with 41 to 264 days between samplings. These data suggest thatS. agnetisis likely more prevalent on dairy farms thanS. hyicusAlso, someS. agnetisisolates in this study appeared to be contagious and associated with persistent infections.
Department of Veterinary Medicine and Surgery, University of Missouri, Columbia, Missouri, USA.
P R F
PRF
Adkins
Department of Veterinary Medicine and Surgery, University of Missouri, Columbia, Missouri, USA middletonjr@missouri.edu.
J R
JR
Middleton
Department of Veterinary Pathobiology, University of Missouri, Columbia, Missouri, USA.
M J
MJ
Calcutt
Department of Veterinary Pathobiology, University of Missouri, Columbia, Missouri, USA.|Bond Life Sciences Center, University of Missouri, Columbia, Missouri, USA.
G C
GC
Stewart
Department of Veterinary Clinical Sciences, Washington State University, Pullman, Washington, USA.
L K
LK
Fox
D000818:Animals; D002417:Cattle; D002418:Cattle Diseases / Q000382:microbiology; D016521:Electrophoresis, Gel, Pulsed-Field / Q000379:methods*; D008892:Milk / Q000382:microbiology*; D058889:Molecular Typing / Q000379:methods*; D060885:Multiplex Polymerase Chain Reaction / Q000379:methods*; D013203:Staphylococcal Infections / Q000382:microbiology / Q000662:veterinary; D013210:Staphylococcus / Q000145:classification* / Q000235:genetics* / Q000302:isolation & purification
Staphylococcus; cattle; milk