| Vacuum cooling,as an emerging preservation technique,can quickly evaporate moisture under low pressure conditions to achieve product cooling,which plays an important role in the cold chain transportation system.Cross-contamination caused by microorganisms during post-harvest processing and transportation of products will cause significant loss in product quality.As one of the important pathogenic bacteria causing vegetable spoilage,Pectobacterium carotovorum subsp.carotovorum(Pcc)can attach and then biofilm form on the contact surface,which has a protective effect on bacterial cells.It is currently a difficult point to control microbial contamination of products.The indirect contamination of vegetables by the biofilm on the contact surface after vacuum cooling is an important reason for its spoilage.The study of this process is conducive to the prevention and control of the infection of Pcc biofilm during the vacuum cooling process of post-harvest vegetables according to the change of the growth characteristics of Pcc biofilm at different formation stages.The main research contents and conclusions of this article are as follows:(1)The growth process and its influencing factors of the biofilm of Pectobacterium carotovorum subsp.Carotovorum under normal temperature(30?).The crystal violet staining method,ultrasonic sampling coating plate method and field emission scanning electron microscope were used to characterize the biofilm at various formation stages at normal temperature.The results showed that 0-24 hours was the attachment stage,24-60 hours was the development stage,and 60-120 hours was the mature stage.The effects of four factors,including concentration of bacteria suspension,mass concentration of sodium chloride,type of culture medium,and the p H value of the culture medium,on the growth of Pcc biofilm at the normal temperature were studied.The optimal growth conditions were the concentration of bacterial suspension was 1×10~7 CFU/m L,the mass concentration of sodium chloride was 4%,the medium was LB liquid culture medium,and the p H of the medium was 7.0.(2)The changes of biofilm production at different formation stages under the accumulative effect of vacuum cooling were studied.Based on the optimal growth conditions of the biofilm at normal temperature,the study found that with the increase of the cumulative treatments number of vacuum cooling,the amount of biofilm production decreased.The bacterial concentration of biofilm after three times vacuum cooling treatment decreased from 10~6CFU/cm~2 to 10~3 CFU/cm~2.The amount of biofilm production in the development stage increased,and the biofilm bacterial concentration per unit contact area increased by 1.2logarithmic gradients after three cumulative effects.However,the amount of biofilm production in the mature stage was not affected by the cumulative effect of vacuum cooling,and the corresponding bacterial concentration was maintained at 1×10~7 CFU/cm~2.The bacterial characteristics of the biofilms with different formation stages under the effect of vacuum cooling were further studied,including its motility,hydrophobicity and surface charge.The results showed that the cluster capability of bacterial colonies was strong during the attachment stage and development stage,and the bacterial hydrophobicity was decreased in the attachment stage.The hydrophobicity ratio of development stage and mature stage was 15%H and 14%H,respectively.The surface acid charge content of the strains in all three strains was above 90%h.Based on the characteristics of the bacterial strains in three states,the bacterial strains of the biofilm in the development stage exhibit strong ability of biofilm formation.(3)Monitoring of polysaccharides and protein of Pcc biofilm by SERS.Firstly,chemical method was used to extract and detect changes in polysaccharides and proteins in extracellular polymeric substances(EPS)of biofilm at different growth stages at room temperature condition and after vacuum cooling.Secondly,silver-coated gold nanorods Au@Ag NRs substrate was prepared.The length of gold rod was 86 nm and the width was 21 nm.The average thickness on both sides of the silver shell was 7.52 nm,and the average thickness on both ends was 5.02nm.The results showed that after the vacuum cooling treatment,the bacterial cells of biofilm in adhesion stage were greatly reduced and damaged,resulting in an increase in the content of polysaccharides and protein components.Increasing the consumption of the biofilm bacteria reached a certain level in the mature stage,maintaining a relative balance between nutrients and metabolites limits the growth of the biofilm,thereby keeping the content of these two components constant. |
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