Federal University of Paraiba, Brazil
Title: Identification of Lactic Acid Bacteria (LAB) in fruits using Matrix-Assisted Laser Desorption Ionization– Time Of Flight Mass Spectrometry (MALDI-TOF MS)
Estefania F Garcia has experience in the development of fermented foods and isolation of lactic bacteria to study probiotic potential and application in fermented foods. She has developed studies with isolation of bacteria from fruits and by-products of fruits and insertion in juices, smoothies, breads and cheese, beside evaluation of the quality these products. She recently initiated the study of freeze-dry of lactic bacteria for commercial insertion in foods.
Statement of the Problem: Matrix-Assisted Laser Desorption Ionization– Time Of Flight Mass Spectrometry (MALDI-TOF MS) can be a replacement for routine 16S rRNA gene sequencing for identification of organisms because it is a fast technique and low cost. This study aimed to isolate and then identify LAB in fruits using MALDI-TOF MS. Methodology & Theoretical Orientation: Samples of orange (Citrus sinensis (L.), grape (Vitis vinifera L.), apple (Malus domestica Borkh) and banana (Musa spp.) obtained from João Pessoa (Paraíba, Brazil). The samples were diluted, inoculated in MRS agar and incubated at 37ºC/48 hr. At least five colonies presenting different morphologies were randomly isolated from plates of each type of fruit and submitted to analyze. Isolates presumptively identified as LAB (catalase negative, gram-positive cocci or rods) were stored at -20°C. Initially, 1 μL of protein extract was deposited in four wells of the sample plate and overlaid with 1 μL of a saturated alpha-cyano-4-hydroxycinnamic acid solution in acetonitrile: water:Tri-Fluoroacetic Acid (TFA). The MALDI-TOF mass spectra measurements of samples were performed using a Bruker Biotyper 3.1. The identification results were expressed by BioTyper log (scores): ≥2.0 is considered accurate species-level identification; ≥1.7-2.0, a probable genus-level identification; and an isolate with a score <1.7 is considered to be unidentified. Findings: A total of 44 isolates of LAB were identified. Species belonging to Enterococcus genus were the most predominant (32/44 isolates; 73%) identified and the other isolates identified belonged to Lactobacillus genus (12/44; 27%). Lactobacillus are isolated in orange (L. brevis, L. plantarum, L. paraplantarum), banana (L. plantarum) and grape (L. brevis and L. plantarum). Enterococcus is isolated from apple (E. faecium and E. mundtii) and grape (E. faecium). From the identification of LABs isolated from fruits using the MALDI-TOF technique the selection of strains for future studies is possible.