Non-culture-based identification of mastitis-causing bacteria by MALDI-TOF mass spectrometry.
The purpose of this study was to evaluate the detection limit of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) for direct identification, without previous microbiological culture, of bovine mastitis-causing bacteria from milk samples. Milk samples (n = 15) were experimentally contaminated with Staphylococcus aureus, Streptococcus uberis, Streptococcus agalactiae, Streptococcus dysgalactiae, and Escherichia coli to have bacterial counts ranging from 103to 109cfu/mL. These contaminated milk samples were subjected to a preparation protocol for bacterial ribosomal protein extraction using the MALDI Sepsityper kit (Bruker Daltonik, Bremen, Germany), which allowed MALDI-TOF MS coupled with Biotyper software (Bruker Daltonik) to identify bacterial fingerprints based on intact ribosomal proteins. The ability of MALDI-TOF MS to correctly identify bacterial strains from experimentally contaminated milk (without previous microbiological culture) depended on the bacterial count of the samples and on the species of the bacteria evaluated. Adequate identification at the bacterial species level (score ≥2.0) directly from milk samples required bacterial counts in the following ranges: ≥106cfu/mL of Staph. aureus, ≥107cfu/mL of E. coli, and ≥108cfu/mL of Strep. agalactiae, Strep. dysgalactiae, and Strep. uberis. We concluded that direct identification of mastitis-causing pathogens is possible for Staph. aureus, E. coli, Strep. agalactiae, Strep. dysgalactiae, and Strep. uberis, but correct identification depended on the bacterial count in the milk samples.
Department of Animal Sciences, School of Veterinary Medicine and Animal Sciences, University of São Paulo, Pirassununga, SP, Brazil, 13.635-900.
Juliana Regina
JR
Barreiro
Department of Animal Sciences, School of Veterinary Medicine and Animal Sciences, University of São Paulo, Pirassununga, SP, Brazil, 13.635-900.
Juliano Leonel
JL
Gonçalves
ThoMSon Mass Spectrometry Laboratory, Institute of Chemistry, University of Campinas (UNICAMP), Campinas, SP, Brazil, 13.083-970.
Patrícia Aparecida Campos
PAC
Braga
Department of Animal Sciences, School of Veterinary Medicine and Animal Sciences, University of São Paulo, Pirassununga, SP, Brazil, 13.635-900.
Aline Gerato
AG
Dibbern
ThoMSon Mass Spectrometry Laboratory, Institute of Chemistry, University of Campinas (UNICAMP), Campinas, SP, Brazil, 13.083-970.
Marcos Nogueira
MN
Eberlin
Department of Animal Sciences, School of Veterinary Medicine and Animal Sciences, University of São Paulo, Pirassununga, SP, Brazil, 13.635-900. Electronic address: mveiga@usp.br.
Marcos
M
Veiga Dos Santos
D000818:Animals; D001419:Bacteria; D002417:Cattle; D004926:Escherichia coli; D005260:Female; D008414:Mastitis, Bovine / Q000175:diagnosis / Q000382:microbiology; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013211:Staphylococcus aureus
MALDI-TOF mass spectrometry; bacteria; mastitis; milk