Dewi, Nabilah Evelina (2025) Rapid and Specific Detection Development of Aeromonas hydrophila and Aeromonas veronii in Nile Tilapia Fish Using Single and Duplex PCR. Undergraduate thesis, Fakultas Sains dan Matematika.

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Abstract

Aeromonas hydrophila and Aeromonas veronii are pathogens capable of coinfecting Nile tilapia simultaneously. This study aims to develop Single PCR and
Duplex PCR methods for the rapid and specific detection of Aeromonas hydrophila
carrying the aerolysin gene and Aeromonas veronii with the rpoB gene in Nile
tilapia (Oreochromis niloticus). These study was conducted to evaluate the
efficiency of Duplex PCR compared to Single PCR in simultaneously detecting
both bacteria, making it an ideal tool for aquaculture pathogen surveillance, with
target bands at 417 bp for Aerolysin gene and 224 bp for rpoB gene. The results
showed that A. hydrophila and A. veronii isolates exhibited distinct colony
morphologies on TSA, GSP, RS, and MacConkey media and were confirmed as
Gram-negative, β-hemolytic bacteria, catalase and oxidase positive. Single PCR
successfully detected the aer gene (417 bp) for A. hydrophila and the rpoB gene
(224 bp) for A. veronii, with clear bands at optimal annealing temperature of 58°C
in 30 cycle. The Limit of Detection (LOD) for duplex PCR was 10⁻⁵ dilution, while
single PCR reached up to 10⁻¹⁰. Koch's postulates confirmed the pathogenicity of
both bacteria, with co-infected fish showing clinical signs such as hemorrhage,
exophthalmia, and internal organ damage, highlighting the importance of early
detection for disease management in aquaculture

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