Congresso Brasileiro de Microbiologia 2023

Congresso Brasileiro de Microbiologia 2023

Resumo: 1073-1

1073-1

ISOLATION AND PRELIMINARY CHARACTERIZATION OF A LYTIC BACTERIOPHAGE FOR KLEBSIELLA PNEUMONIAE.

Autores:

Bruna Guerrero (UNISO - PHAGELAB – BIOFILMS AND BACTERIOPHAGES LABORATORY) ; Luan Mota (UNISO - PHAGELAB – BIOFILMS AND BACTERIOPHAGES LABORATORY) ; Stephen Emencheta (UNISO - PHAGELAB – BIOFILMS AND BACTERIOPHAGES LABORATORY, UNN NSUKKA - DEPARTMENT OF PHARMACEUTICAL MICROBIOLOGY AND BIOTECHNOLOGY) ; Arthur Pereira (UNISO - PHAGELAB – BIOFILMS AND BACTERIOPHAGES LABORATORY) ; Erica Silva (UNISO - PHAGELAB – BIOFILMS AND BACTERIOPHAGES LABORATORY) ; Marta Vila (UNISO - PHAGELAB – BIOFILMS AND BACTERIOPHAGES LABORATORY) ; Victor Balcão (UNISO - PHAGELAB – BIOFILMS AND BACTERIOPHAGES LABORATORY, UNIVERSIDADE AVEIRO - DEPARTMENT OF BIOLOGY AND CESAM, UNIVERSITY OF AVEIRO,)

Resumo:

ABSTRACT:Klebsiella pneumoniae is an enterobacteria that causes opportunistic infections and is on the list of critical priority pathogens of the World Health Organization (WHO), often associated with hospital infections because it remains in surgical instruments and affects neonates and immunosuppressed patients. Currently, bacteria have become increasingly resistant to antibiotics due to their exacerbated use, generating persistent infections and high cost of treatment. In view of the lack of new antibiotics, the use of bacteriophages, which are highly specific viruses that have the ability to eliminate only target bacteria, without compromising the microbiota, reappears as a therapeutic alternative. Therefore, this work aimed to isolate and characterize in a preliminary way a lytic bacteriophage for Klebsiella pneumoniae. Initially the bacteriophage was isolated from nature, propagated and purified. It was then characterized by biological tests such as: Spot-test, Phage Titre, Curve of a Single Synchronous Cycle (OSGC) and physical-chemical tests: Spectrophotometric analysis in the UV-VIS zone, Electrophoresis in Polyacrylamide Gel with Sodium Dodecylsulfate (SDSPage ) and Transmission Electron Microscopy (TEM). The bacteriophage was isolated from a sample collected from wastewater at the Veterinary Hospital of the University of Sorocaba (Sorocaba/SP, Brazil), being named vB_KpnS_Uniso1E, it was purified to remove cellular debris and propagated to obtain a larger amount of the solution. containing the bacteriophage. After the isolation, a Spot test is performed in a Petri dish with double layer agar containing a mat of the host bacteria, the test indicated a high lytic capacity. The phage titer was 1.37x1014 Plaque Forming Units per mL (PFU/mL). In the one step growth curve, a burst size of 17.67 virions/host cell was observed. The maximum absorbance observed through the spectrophotometric analysis in the UV-VIS zone was 256 nm and the minimum was 244 nm, which allowed obtaining the molar extinction coefficient, which was 1,278x10-13 (UFP/mL)-1 •cm -1. SDS PAGE electrophoresis indicated molecular weights of structural proteins between 34 -169 kDa. By electronic microscopy, it was observed that the isolated phage belongs to the Caudoviricetes order and, probably, to the Siphoviridae family. It was found that the phage suspension obtained containing phage particles against Klebsiella pneumoniae has a large amount of virions and a high lytic potential. Based on the results obtained, the phage demonstrated viability for application in phage therapy to combat infections caused by Klebsiella pneumoniae.

Palavras-chave:

Biotechnology, Characaterization, Infections, Klebsiella pneumoniae, Phage