Biodiversity of Bacteriophages Specific to ESBL-Producing Klebsiella pneumoniae Isolated from Wastewater
Nuridha Audinia Safitri (a,e*), Meity Mardiana (e), Leli Saptawati (c), Dewi Anggraini (d), Iftita Rahmatika (b), Cindy Rianti Priadi (b), Ruby Setiawan (f), Yustinus Maladan (e), Andriansjah Rukmana (a), Dodi Safari (e), Yulia Rosa Saharman (a), Rosantia Sarassari (e)

a) Department of Microbiology, Faculty of Medicine, Universitas Indonesia, Jakarta, Indonesia
*nuridhaaudinias[at]gmail.com
b) Environmental Engineering Study Program, Faculty of Engineering, Universitas Indonesia, Jakarta, Indonesia
c) Department of Microbiology, Faculty of Medicine, Universitas Sebelas Maret, Surakarta, Indonesia
d) Department of Microbiology, Faculty of Medicine, Universitas Riau, Riau, Indonesia
e) Eijkman Research Center for Molecular Biology, National Research and Innovation Agency, Bogor, Indonesia
f) Biosystematics and Evolution Research Center, National Research and Innovation Agency, Bogor, Indonesia

Abstract

Extended-spectrum beta-lactamase-producing Klebsiella pneumoniae (ESBL-KP) is an antimicrobial-resistant (AMR) bacteria that poses a major challenge in infection treatment. Bacteriophages are viruses that infect bacteria. Phage plaque morphology can serve as an initial screening to identify lytic or lysogenic bacteriophages that can be potential alternative for antimicrobial teraphy. This study aims to characterize and analyze the diversity of bacteriophages infecting ESBL-KP from wastewater based on plaque morphology. The study was conducted from April to May 2024. Wastewater samples were collected from seven locations. Samples were filtered using a 0.22 Millipore membrane filter. The resulting filtrate was mixed with suspensions of 29 ESBL-KP isolates. Phage plaque morphology was tested using the double overlay plaque assay and spot test. The formed plaques were observed and analyzed based on turbidity and the presence or absence of halos. This study isolated a total 160 bacteriophages from seven wastewater sample points againts 29 ESBL-KP isolates. Of these, 101 phages formed turbid plaques and 59 formed clear plaques. Among the 59 clear plaques, 30 formed with halos (semi-transparent zones around the plaques). Clear plaques indicate lytic (virulent) activity, whereas turbid plaques indicate lysogenic (temperate) activity. This study is an initial investigation into bacteriophages. Morphological analysis reported of diversity among bacteriophages isolated from wastewater. In order to know which phage is suitable for future teraphy, genome analysis, and interaction between host and phages are needed to be explored.

Keywords: bacteriophage, Klebsiella pneumoniae, ESBL, wastewater, plaque turbidity

Topic: Genetic and Molecular Biology