Study On The Antibacterial Activity And Mechanism Of Chicken Egg Yolk Antibodies Against Listeria Monocytogens

Listeria monocytogenes is a common pathogen in seafood. Because it can grow at a wide range of temperature and survive even at 4℃,it is difficult to effectively control the pollution and resulted hazards of Listeria monocytogenes in fresh products during their storage and transportation. Egg yolk antibody (IgY), a unique immunoglobulin in egg yolk, has a potent inhibitory activity to its antigen bacteria, and could be used as a potential antimicrobial agent to food micro-organisms. In this study, IgY antibodies against Listeria monocytogens were isolated, purified, characterized and validated to be used as an antimicrobial agent in sea food samples. The main results were summarized as follows:Four different methods were evaluated for extraction and purification of IgY, and the best outcoming was observed with ammonium sulfate precipitation, where the titer of anti-Listeria monocytogens antibodies was the highest and the SDS-PAGE analysis showed comparatively high purity of protein contents. The content of IgY in egg yolk reach to a high level after three times of booster immunization,-and-maintained stable for six weeks. The prepared IgY demonstrated excellent pH and thermal stabilities: more than 70% bioactivity could be retained in the range of pH 3-9, and only 33% decrease in the titer after 4-h incubation at 60℃.The inhibitory effect of the specific IgY on the growth of Listeria monocytogenes was tested in liquid medium, solid medium and food samples. After 8-h incubation with the specific IgY at 37℃,the growth of Listeria monocytogenes significantly decreased compared with the control ones, and positive correlation between inhibitory efficiency and IgY concentration is observed.Similar growth inhibitory effects were observed with incubation at 4℃for 120 h.The IgY also inhibited the growth of Listeria monocytogenes when coated onto solid medium, fresh or smoked salmon samples. More than 80% of Listeria monocytogenes were inhibited for growth when 200 mg/mL of the IgY was incubated with the solid medium for 30 min at room temperature. Compared with those of blanks, the amounts of Listeria monocytogenes in fresh or smoked salmon samples were reduced more than 2 log cycles after 15 days of storage at 6±1℃in the presence of 200 mg/mL of the specific IgY.Two different affinity chromatography methods were developed for the separation of the specific IgY from non-specific ones. After purification by either of the two methods, the titer of the IgY could increase more than 4 times, and the antibacterial efficiency could also improve more than 4 times when tested with milk stored at 6±1℃for 10 days as sample.The results of transmission electron microscopy showed that the binding of the specific IgY to Listeria Monocytogens resulted in significant destruction of cell wall and structural alterations on the bacterial surface. The products of the IgY and Listeria monocytogenes were collected and analysed by NMR and GC-MS.The result showed that the structure of those products was similar with that of teichoic acid, which is an important component of cell wall.The above data illustrated that the antibacterial mechanism of the IgY might be the destruction of teichoic acid.