The sessile biofilms of Vibrio parahaemolyticus and Listeria monocytogenes have increasingly become a critical threat in seafood safety. This study aimed to evaluate the effects of modified atmospheres on the formation ability of V. parahaemolyticus and L. monocytogenes biofilms. The stress responses of bacterial biofilm formation to modified atmospheres including anaerobiosis (20% carbon dioxide, 80% nitrogen), micro-aerobiosis (20% oxygen, 80% nitrogen), and aerobiosis (60% oxygen, 40% nitrogen) were illuminated by determining the live cells, chemical composition analysis, textural parameter changes, expression of regulatory genes, etc. Results showed that the biofilm formation ability of V. parahaemolyticus was efficiently decreased, supported by the fact that the modified atmospheres significantly reduced the key chemical composition [extracellular DNA (eDNA) and extracellular proteins] of the extracellular polymeric substance (EPS) and negatively altered the textural parameters (biovolume, thickness, and bio-roughness) of biofilms during the physiological conversion from anaerobiosis to aerobiosis, while the modified atmosphere treatment increased the key chemical composition of EPS and the textural parameters of L. monocytogenes biofilms from anaerobiosis to aerobiosis. Meanwhile, the expression of biofilm formation genes (luxS, aphA, mshA, oxyR, and opaR), EPS production genes (cpsA, cpsC, and cpsR), and virulence genes (vopS, vopD1, vcrD1, vopP2β, and vcrD2β) of V. parahaemolyticus was downregulated. For the L. monocytogenes cells, the expression of biofilm formation genes (flgA, flgU, and degU), EPS production genes (Imo2554, Imo2504, inlA, rmlB), and virulence genes (vopS, vopD1, vcrD1, vopP2β, and vcrD2β) was upregulated during the physiological conversion. All these results indicated that the modified atmospheres possessed significantly different regulation on the biofilm formation of Gram-negative V. parahaemolyticus and Gram-positive L. monocytogenes, which will provide a novel insight to unlock the efficient control of Gram-negative and Gram-positive bacteria in modified-atmosphere packaged food.

温度是影响细菌生长与失活的关键因素,在食品生产中常用来控制致病微生物潜在的风险.但由于不同菌株间个体的差异,细菌在相同温度作用下呈现出不同的失活趋势,这种行为方式称为菌株的失活异质性,容易导致微生物风险控制的不确定性和变异性.比较了19株副溶血性弧菌(16株临床菌株和3株环境菌株)在巴氏消毒温度(65℃)及冷链温度(10℃)作用下的失活情况,并结合Weibull模型,拟合相应的失活参数(tR值),探究了不同菌株间的失活异质性.在65℃处理条件下,19株副溶血性弧菌的tR值介于22.62～67.23 s,VPC-1为耐热性最强菌株,而VPC-10为耐热性最弱菌株,热失活参数tR值最适的概率分布为Normal(44.82,12.27).在10℃条件下,tR值介于113.96～371.38 h,VPC-3为耐冷性最强菌株,VPC-2为耐冷性最弱菌株,冷失活参数tR值最适的概率分布为Loglogistic(51.45,148.88,4.67).结果表明,副溶血性弧菌的热失活和冷失活间没有显著的相关性,菌株的失活异质性广泛存在于副溶血性弧菌之中,仅基于单一菌株进行失活模型的拟合,很难描述其整体的失活趋势.同时,初步构建了菌株失活异质性的随机模型,并使用概率分布代替了传统的失活参数.

The extracellular polymeric substances (EPS) construct the three-dimensional (3-D) structure of biofilms, but their respective roles are still not clear. Therefore, this study aimed to illuminate the role of key chemical components [extracellular DNA (eDNA), extracellular proteins, and carbohydrates] of EPS in biofilm formation of Vibrio parahaemolyticus. The correlations between each key chemical component and biofilm formation were first determined, showing that the biofilm formation of V. parahaemolyticus was strongly positively correlated with both eDNA and protein content (P < 0.01), but not with carbohydrates. Subsequently, individual DNase I or protease K treatment markedly reduced the initial adhesion and structural stability of the formed biofilms by hydrolyzing the eDNA or extracellular proteins, but did not induce significant dispersion of mature biofilms. However, the combination of DNase I and protease K treatment induced the obvious dispersion of the mature biofilms through the concurrent destruction of eDNA and extracellular proteins. The analysis at a structural level showed that the collapse of biofilms was mainly attributed to the great damage of the loop configuration of eDNA and the secondary structure of proteins caused by the enzyme treatment. Therefore, this study provides a deep understanding of the role of key chemical components of EPS in biofilm development of V. parahaemolyticus, which may give a new strategy to develop environmentally friendly methods to eradicate the biofilms in food industry.