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Comparison of standard and on-plate extraction protocols for identification of mastitis-causing bacteria by MALDI-TOF

M M Barcelos1, L Martins1, R C Grenfell2

  • 1Department of Animal Sciences, School of Veterinary Medicine and Animal Sciences, University of São Paulo (USP), 225 Duque de Caxias Norte Avenue, Zip code, Pirassununga, SP, 13635-900, Brazil.

Brazilian Journal of Microbiology : [Publication of the Brazilian Society for Microbiology]
|July 1, 2019
PubMed

Insights

Comparing standard and on-plate sample preparation for mastitis bacteria identification using MALDI-TOF MS revealed similar overall bacterial identification rates. The standard protocol showed slightly higher accuracy, suggesting its use when the on-plate method fails.

Area of Science:

  • Veterinary Microbiology
  • Analytical Chemistry
  • Bacterial Identification

Background:

  • Subclinical mastitis in cows poses significant economic challenges in the dairy industry.
  • Accurate and rapid identification of mastitis-causing bacteria is crucial for effective treatment and herd management.
  • Matrix-Assisted Laser Desorption/Ionization-Time of Flight Mass Spectrometry (MALDI-TOF MS) offers a promising alternative to conventional microbiological methods.

Purpose of the Study:

  • To compare the efficacy of standard versus on-plate sample preparation protocols for bacterial identification in subclinical mastitis.
  • To evaluate the identification rates and reliability of both protocols using MALDI-TOF MS.
  • To determine the optimal protocol for routine identification of mastitis pathogens.

Main Methods:

  • Two hundred and eighty-six bacterial isolates from cows with subclinical mastitis were analyzed.
  • Samples were prepared using either a standard reagent-based lysis protocol or a simplified on-plate protocol.
  • Bacterial identification was performed using MALDI-TOF MS, with analysis of both species-level (score ≥ 2.0) and genus-level (score ≥ 1.7) identification.

Main Results:

  • The on-plate protocol achieved a bacterial identification rate of 91.4% (170/186), comparable to the standard protocol's 94.6% (176/186).
  • Reducing the identification score threshold to genus-level improved identification rates for both protocols.
  • The standard protocol demonstrated higher genus-level identification accuracy (100%) compared to the on-plate protocol (94.6%) when compared against a gold standard.

Conclusions:

  • Both standard and on-plate protocols are viable alternatives to conventional methods for identifying subclinical mastitis pathogens via MALDI-TOF MS.
  • The standard protocol may be preferred when the on-plate method yields inconclusive results, serving as a more robust reference.
  • Optimizing the on-plate protocol, potentially through enhanced reference database libraries, could further improve its diagnostic utility.
Keywords:
MALDI-TOF MSMastitisStandard and on-plate extractionSubclinical case

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