Anais do Congresso Brasileiro de Virologia & Encontro de Virologia do Mercosul

ISOLATION AND CHARACTERIZATION OF A MICROCOCCUS LUTEUS-SPECIFIC BACTERIOPHAGE FOR CONTROLLING BACTERIAL BIOFILM

Published in 04/01/2021 - ISBN: 978-65-5941-175-7

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DOI
10.29327/cbv.297866  
Paper Title
ISOLATION AND CHARACTERIZATION OF A MICROCOCCUS LUTEUS-SPECIFIC BACTERIOPHAGE FOR CONTROLLING BACTERIAL BIOFILM
Authors
  • Paloma Cavalcante Cunha
  • Ariel Ribeiro Siqueira
  • Jéssica Duarte
  • Marcella Silva Vieira
  • Maraisa Marques Marcelino
  • Mirelly Jady Fernandes e Silva
  • Sergio Oliveira de Paula
  • Roberto Sousa Dias
Modality
Resumo
Subject area
Ambiental
Publishing Date
04/01/2021
Country of Publishing
Brasil
Language of Publishing
Inglês
Paper Page
https://www.even3.com.br/anais/cbv/297866-isolation-and-characterization-of-a-micrococcus-luteus-specific-bacteriophage-for-controlling-bacterial-biofilm
ISBN
978-65-5941-175-7
Keywords
Gram-positive bacteria, biological control, biofilm, bacteriophage
Summary
Micrococcus luteus is a gram-positive bacterium found in several habitats and is part of the natural microbiota of mammalian skin. Despite being considered a harmless commensal organism, M. luteus can be an opportunistic pathogen associated with the occurrence of various infections in hospital environments. M. luteus also plays a role as a bridging organism in the biofilm consortium, making it an important target in the study of clinically relevant polymicrobial biofilms. Among the promising strategies for controlling bacterial pathogens and biofilms are the use of bacteriophages. In this work, we isolated and evaluated the performance of a bacteriophage specific to M. luteus (named UFV015) to control bacterial biofilm. The phage isolation was done by the enrichment method, adding a water sample collected from São Bartolomeu creek (Viçosa - Minas Gerais, Brazil) to the culture of M. luteus, which was previously grown overnight. The mixture was incubated for 24 hours at 37 ° C and then centrifuged and filtered. The filtrate was used for two more rounds of enrichment with the bacteria as previously described. The final propagate was plated by the double-layer method and incubated overnight at 37 ° C. A lysis plate was selected, propagated, and then plated to determine the titer in PFU/ml. The phage was also tested against four gram-negative (i.e., Escherichia coli 30, E. coli SAN3, Pseudomonas fluorescens, and Proteus vulgaris) and one gram-positive (i.e., Bacillus cereus) bacteria for specificity. For the microplate biofilm quantification assay, a culture of M. luteus was inoculated in DO600 0.1 (control) and M. luteus + phage UFV015 MOI 0.001 (treatment). The plate was then incubated for 24 hours and, subsequently, the liquid was discarded, fixed with 99% methanol and stained with 0.1% crystal violet solution. The crystal violet was discarded and the dye attached to the biofilm was recovered by adding an alcohol: acetone solution (80:20 v/v). The absorbance (590 nm) of each microplate well was measured in a plate reader for indirect quantification of the biomass present in the biofilm. The same procedure was repeated with the bacteria used in the specificity test. The phage M. luteus UFV015 presented lysis plates with discrete morphology, with a small, translucent center and a halo that became more evident a few days after plating. Using a low phage titer (around 2.5x105 PFU/mL), the biofilm of M. luteus was practically eradicated (DO600 0.1), while the control had a DO600 of approximately 1.5. The phage showed no activity against any of the other bacteria tested, both in the host spectrum and in the biofilm assay. These results indicate that this phage is specific for M. luteus and highly effective in eliminating its host's biofilm. Future tests using other bacteria should be conducted to ensure the high specificity of the phage. In addition, morphological and genomic analyzes should be performed in order to better classify and characterize the phage, enabling future clinical trials to control M. luteus infections. Financial support: CAPES and Petrobras.
Title of the Event
XXXI Congresso Brasileiro de Virologia & XV Encontro de Virologia do Mercosul
Title of the Proceedings of the event
Anais do Congresso Brasileiro de Virologia & Encontro de Virologia do Mercosul
Name of the Publisher
Even3
Means of Dissemination
Meio Digital
DOI

How to cite

CUNHA, Paloma Cavalcante et al.. ISOLATION AND CHARACTERIZATION OF A MICROCOCCUS LUTEUS-SPECIFIC BACTERIOPHAGE FOR CONTROLLING BACTERIAL BIOFILM.. In: Congresso Brasileiro de Virologia & Encontro de Virologia do Mercosul. Anais...Porto Alegre(RS) Online, 2020. Available in: https//www.even3.com.br/anais/cbv/297866-ISOLATION-AND-CHARACTERIZATION-OF-A-MICROCOCCUS-LUTEUS-SPECIFIC-BACTERIOPHAGE-FOR-CONTROLLING-BACTERIAL-BIOFILM. Access in: 28/04/2025

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