小龙虾微冻贮藏过程中特定腐败菌的分离鉴定及致腐产胺能力分析

Isolation and identification of specific spoilage organisms and analysis of saprogenic amine production capacity during micro-frozen storage of Procambarus clarkii

目的探究小龙虾微冻贮藏过程中特定腐败菌(specific spoilage organisms,SSOs)种类及其致腐产胺能力。方法本研究利用选择性培养基从微冻至腐败小龙虾中分离、纯化优势腐败菌,通过16S rDNA技术结合细菌生化鉴定法进行鉴定;将鉴定出的SSOs接种至无菌虾肉中,通过监测pH、挥发性盐基氮(total volatile basic nitrogen,TVB-N)、菌落总数等指标的变化,计算腐败代谢产物产量因子(YTVB-N/CFU)以评价各SSOs的致腐能力,并比较各SSOs接种组的产生物胺能力。结果小龙虾微冻过程中的SSOs为荧光假单胞菌(Pseudomonas fluorescens)、腐败希瓦氏菌(Shewanella putrefaciens)、温和气单胞菌(Aeromonas sobria)和蜂房哈夫尼亚菌(Hafnia alvei);在贮藏15 d后,P.fluorescens、S.putrefaciens、A.sobria和H.alvei接种组TVB-N含量分别为205.88、54.01、60.60、24.24 mg/100 g,菌落总数分别为9.33、8.92、9.02、9.11 log10 CFU/g,YTVB-N/CFU大小顺序为P.fluorescens>A.sobria>S.putrefaciens>H.alvei;小龙虾腐败过程主要生物胺为腐胺和尸胺,各SSOs产胺能力为P.fluorescens>S.putrefaciens≈A.sobria>H.alvei。结论本研究明确了小龙虾微冻贮藏过程中的特定腐败菌种类及其致腐产胺能力,为针对性地筛选小龙虾高效抑菌保鲜剂提供了理论基础。

Objective To explore the types of specific spoilage organisms(SSOs)during micro-frozen storage of Procambarus clarkii and study their saprogenic amine production capacity.Methods Dominant spoilage organisms were isolated and purified from spoilage Procambarus clarkii under micro-frozen storage by using selective culture medium,and were identified by 16S rDNA technique combined with bacterial biochemical identification.Then,the identified SSOs were inoculated into sterile Procambarus clarkii meat,and the pH,total volatile basic nitrogen(TVB-N)and total colonies counts were monitored.According to the results of TVB-N and total colonies count,the spoilage metabolite yield factor(YTVB-N/CFU)was calculated to evaluate the spoilage potential of each SSOs,and the amine production capacity were compared among each SSOs by determining biogenic amine concentration.Results The SSOs of spoilage Procambarus clarkii under micro-frozen storage were identified as Pseudomonas fluorescens,Shewanella putrefaciens,Aeromonas sobria and Hafnia alvei.After 15 days storage,the TVB-N content and total colonies counts in samples inoculated with P.fluorescens,S.putrefaciens,A.sobria and H.alvei were 205.88,54.01,60.60 and 24.24 mg/100 g and 9.33,8.92,9.02,9.11 log10 CFU/g,respectively.The order of YTVB-N/CFU was P.fluorescens>A.sobria>S.putrefaciens>H.alvei.The main amine during the spoilage process of Procambarus clarkii were putrescine and cadaverine,and the order of amine production capacity was P fluorescens>S.putrefaciens≈A.sobria>H.alvei.Conclusion This study clarifies the species,spoilage potential and amine production capacity of SSOs,and provides a theoretical basis for the targeted screening of efficient bacterial preservatives for Procambarus clarkii.