Effect of chitosan, nisin and storage time on the growth of Listeria innocua and Shewanella putrefaciens in fish homogenates

L. I. SCHELEGUEDA, M. F. GLIEMMO AND C. A. CAMPOS

Valladolid, España

Conferencia; I International Conference on Antimicrobial Research (ICAR2010); 2010

Fish and other fishery products are highly perishable food. The process of degradation starts immediately after the fish’s death and it often develops more quickly than in other meats. Therefore, the use of a preservation method is essential when it comes to the mentioned products. The use of natural antimicrobials together with other preservation measures has proved to be a good option to extend the shelf life of this kind of food. The aim of this study was to evaluate the effect of chitosan, nisin and storage time on the inhibition of Shewanella putrefaciens and Listeria innocua growth in fish homogenates. For such purpose a factorial design in three blocks was performed, on which the concentration of chitosan and nisin, and the storage time were the factors studied. The selected levels were: chitosan (0, 2000 ppm), nisin (0, 5000 ppm) and storage time (0, 48 hours). A central point was included (chitosan 1000 ppm, nisin 2500 ppm, storage time 24 hours). L. innocua was selected as a representative microorganism of Gram-positive flora and as an alternative to the pathogen Listeria monocytogenes. S. putrefaciens, which is the typical spoilage microorganism of fishery products when they are stored in cold and aerobic conditions, was selected as the Gram-negative bacterium representative. Fish homogenate was prepared processing Argentine hake fillets (Merluccius hubbsi) and distilled water in a ratio of 1:1; pH was adjusted to 5.5 using citric acid 10%m/m and homogenate aliquots were put into screw-cap flasks. They were sterilized for 15 minutes at 100ºC. Preservatives were added and then microorganisms were inoculated reaching a level of 105 CFU g-1. The microorganisms had been previously incubated in Mueller-Hinton broth, overnight at 30ºC. The inoculated homogenates were stored at 30ºC for 48 h.  S. putrefaciens and L. innocua populations were enumerated  by puor-plating in TSA agar and Mueller-Hinton agar, respectively. The plates were incubated for 48 hours at 30ºC. As it was expected, in all cases, time exerted a significant effect on the growth of both microorganisms. When antimicrobials were added, counts were significantly reduced (P < 0.05). A significant interaction between antimicrobials and time was noted; however, this effect depended on the microorganism being tested. In the case of S. putrefaciens, both preservatives reduced growth during storage; on the other hand, the development of L. innocua was reduced by chitosan during storage, but it increased in the presence of nisin. There was no significant interaction between the two preservatives. However, different trends in respect of time are noted. At time zero, nisin alone or together with chitosan reduced the count of both microorganisms between 2 and 3 log cycles. After 48 hours of storage, both antimicrobials showed an additive effect reducing the development of both microorganisms between 4 and 5 log cycles. To sum up, nisin and chitosan were able to inhibit the development of S. putrefaciens and L. innocua in fish homogenate at pH 5.5. It is clear that nisin controlled the growth of S. putrefaciens, Gram-negative bacterium. Although the interaction was not significant, the joint use of the studied antimicrobials could be promising, since the nisin´s effects on counts are shown immediately after its application, while chitosan´s effects can be noticed throughout the storage.