Identification of major pathogens isolated from subclinical intramammary infection by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

Abstract

The hypothesis of this study is that methodology known as MALDI-TOF MS 'matrix-assisted laser desorption/ionization time-of-flight mass spectrometry' effectively identifies the major pathogens causing subclinical intramammary infection (IMI) compared to microbiological culture. This study will aim to identify the major pathogens (Staphylococcus aureus, environmental Streptococcus, Streptococcus agalactiae, Escherichia coli and Klebsiella spp.) causing subclinical IMI by MALDI-TOF MS. For this, six dairy herds (600 cows) will be envolved, where lactating cows will be selected (estimated 330 animals) with subclinical IMI caused by major pathogens distributed in 5 groups (1: Staphylococcus aureus; 2: Streptococcus environmental; 3: Streptococcus agalactiae; 4: Escherichia coli and 5: Klebsiella spp). Mammary quarters (n = 150) from dairy cows diagnosed with IMI caused by major pathogens will be selected. In addition, a second sampling step of milk at mammary quarters level will be performed considering the mammary quarters previously selected and from these somatic cell count and microbiological culture will also be performed. After identifying the pathogens by microbiological culture, ribosomal proteins extraction will be carried out from tube extraction technique. These extraction ribosomal proteins step will be necessary to read the protein profiles (fingerprint) and identify pathogens in the species level by MALDI-TOF MS. The results of the two techniques (MALDI-TOF MS and microbiology culture) will be compared using the McNemar test for paired proportions using SAS ® version 9.2 statistical program. It is hoped that MALDI-TOF MS can be adopted as a routine laboratory technique for identification of pathogens causing subclinical bovine mastitis. The MALDI-TOF MS will provide rapid identification of mastitis pathogens enabling quick results to farmers. (AU)