Genomic and GWAS-Based Insights into Antimicrobial Resistance in <i>Shewanella algae</i> Isolated from <i>Penaeus monodon</i>

<b>Background/Objectives:</b> The emergence of antimicrobial-resistant (AMR) pathogens in aquaculture ecosystems poses a significant risk to both food security and human health. <i>Shewanella</i> species are recognized as significant AMR reservoirs, yet their prevalence and resistance mechanisms within a shrimp-related ecosystem remain poorly characterized. This study aimed to perform a genotypic and phenotypic characterization of <i>S. algae</i> VK101, isolated from wild-caught black tiger shrimp (<i>Penaeus monodon</i>) broodstock. <b>Methods:</b> A complete 5.21 Mb genome was generated using a hybrid Illumina and Oxford Nanopore sequencing approach. Antimicrobial susceptibility was evaluated for 21 antibiotics via Minimum Inhibitory Concentration (MIC) testing. Comparative pangenomics and genome-wide association studies (GWAS) across 125 <i>S. algae</i> genomes were conducted to identify novel resistance determinants. <b>Results:</b> MIC analysis revealed that VK101 was resistant to ampicillin (>16 µg/mL) and colistin (8 µg/mL), while showing intermediate susceptibility to imipenem and ciprofloxacin. In silico analysis identified 205 antimicrobial resistance genes (ARGs), including a perfect hit for the fluoroquinolone resistance gene <i>qnrA3</i>. Notably, no <i>mcr</i> genes were detected. Although VK101 exhibited moderate resistance (8 µg/mL), GWAS across the broader <i>S. algae</i> population linked a specific <i>lptA</i> mutation (K140N) to high-level resistance (64 µg/mL). Other GWAS-identified genes (e.g., <i>czcA</i>, <i>ampC</i>, and <i>oprM</i>) likely represent indirect associations driven by genetic linkage or clade-specific markers rather than direct causal factors. <b>Conclusions:</b> These findings highlighted the presence of multidrug-resistant <i>S. algae</i> in wild-caught <i>P. monodon</i> broodstock, reflecting the occurrence of antimicrobial resistance in aquatic environments. Colistin resistance in these isolates was primarily mediated by chromosomal variants rather than mobile <i>mcr</i> elements, indicating the need for integrated genomic surveillance within the aquaculture value chain.