Controlling of multidrug resistant Aeromonas hydrophila infected Nile tilapia (Oreochromis niloticus) using Ah01 and Ah02 virulent bacteriophages isolates

Abstract

The aim of this work is to investigate the efficiency Ah01 and Ah02 virulent bacteriophages isolated from fish in controlling Aeromonas hydrophila-infected Nile talipa fish. Aeromonas hydrophila strains were resistant to 64% of tested antibiotic, where the antibiotic sensitivity test revealed that among 28 different antibiotics, Aeromonas hydrophila (ATCC 13037) was resistant to 18 antibiotics. In this study, two lytic phages designated as Ah01 and Ah02 specific to Aeromonas hydrophila strain (ATCC 13037) were isolated from fish and agricultural irrigation water, respectively. The isolated phages exhibited tolerance to alkaline and acidic reactions (pH 3–12). The thermal inactivation point was found to be 84 °C for both phages (Ah01 and Ah02). Longevity in vitro of Ah01 and Ah02 phages was estimated to be 21 days at 22 ± 2 °C. The two phage isolates were polyvirulent, since, they were infectious to different species of genus Aeromonas. Transmission electron microscopy analysis revealed that Ah01 and Ah02 phages are of hexagonal head and contractile tail type. Therefore, these two phages belong to family Myoviridae. One step growth curve indicated that the latent periods were15 min for both phages (Ah01 and Ah02). The burst sizes were 102 and 209 PFU/ cell for Ah01 and Ah02 phages, respectively. The RAPD-PCR was used to study the genetic diversity and distances of DNA products of the two virulent phages (Ah01 & Ah02). Six primers with different sizes (P13, N8, B12, A01, S50 and S8) were succeeded to reproduce polymorphic DNA bands. Genetic distances relationship among the two phages were studied based on their similarity indices data of RAPD- PCR analysis. The RAPD primers achieved a rate of polymorphism ranged between 50% and 100%. Complete mortality was recorded in fish injected with Aeromonas hydrophila (ATCC 13037) within two days, but when adding phages, the phage isolates were combined with A. hydrophila strain (ATCC 13037) and intraperitoneal injected in fish, no mortality was detected until the end of experiment (15 days).