This study investigated changes in the microbial compositions of crayfish tails during storage at 4 °C (for 0–12 days) as measured using high-throughput sequencing (HTS). The specific spoilage organisms (SSOs) in the crayfish tails were isolated using culture-dependent cultivation methods, and they were identified by 16S rRNA and characterized for their enzymatic spoilage potentials (e.g., protease, lipase, phospholipase, and amylase). The spoilage abilities of the selected strains in the crayfish tails were assessed by inoculating them into real food. Moreover, the microbial growth and the volatile basic nitrogen (TVB-N) changes were monitored during the storage period. The results from the HTS showed that the dominant genus of shrimp tails evolved from Streptococcus (D0) to Pseudomonas (D4) and, finally, to Paenisporosarcina (D12) during storage. Seven bacterial species (Acinetobacter lwoffii, Aeromonas veronii, Kurthia gibsonii, Pseudomonas sp., Exiguobacterium aurantiacum, Lelliottia amnigena, and Citrobacter freundii) were screened from the spoiled shrimp tails by the culture-dependent method, among which Aeromonas veronii had the strongest spoilage ability.
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