Judul: Penggunaan Mutan C. sakazakii GPFuv Untuk Kajian Pembentukan Biofilm Multispesies
Abstrak: Biofilms are defined as sessile communities of bacterial cells attached to a surface or to each other and followed by growth and biofilm formation, usually enclosed in the extracellular polymeric substances produced by the bacteria. The biofilms formed in food processing environtment may potentially become sources of contamination resulting in food spoilage or foodborne illness. Biofilm cells has been reported are more resistant to antimicrobial agents than their planktonic counterparts. Cronobacter sakazakii is a Gram-negative, rod-shaped, non-spore-forming pathogenic bacteria. C. sakazakii has been reported to be able to attach to and form biofilms on silicon, latex, polycarbonate, stainless steel, glass, and polyvinyl chloride. In observation of wild-type biofilm bacteria in a complex environment or the conditions of competition between bacteria often found difficulty in distinguishing the bacteria target with other bacteria. Green Fluorescent Protein (GFPuv) has become an especially valuable marker for nondestructively visualizing cells, particularly in biofilms. Labeling bacteria using GFPuv can be an alternative to observe biofilm-forming bacteria in complex environments or multispecies. C. sakazakii labeled GFPuv can become fluorescent under UV light to indicate the specific green fluorescent colonies that can be distinguished by other bacteria. The purpose of this research is to investigate the behavior of C. sakazakii mutant GFPuv in the formation of biofilm on several food contact surfaces and compare it to those by its wild-type as well as the formation of biofilm of this mutant in the presence of S. aureus and/or L. rhamnosus and the performance of direct microscopic count and plating method for enumeration of the adherent cells. This research was carried out into two stages. The first stage was prelimenary research consist of : (a) preparatory bacteria cultures C. sakazakii mutant GFPuv and wild-type FWHd16 and YRt2a, S. aureus SA25923 and L. rhamnosus R23 ; (b) preparatory food surface contact plates consist of SS, buna-N and PTFE in the growth media of TSB and mRSB. The second stage was compare GFPuv labeled and wild-type C. sakazakii strains pertaining their ability to biofilm formation on several food contact surfaces, i.e. SS, buna-N, and PTFE under incubated at three different times, i.e 24, 48 and 72 h. Mutant C. sakazakii is also used for the formation of biofilms multispesies using three different bacteria, i.e C. sakazakii mutant GFPuv FWHd16 and YRt2a, S. aureus SA25923 and L. rhamnosus R23 on stainless steel which also incubated at 24, 48 and 72 h. The use of bacteria that have been inserted with marker plasmid GFPuv can be applied to see the ability of bacteria to form biofilms on food contact surfaces. C. sakazakii mutant GFPuv able to form biofilms with the number of population and patterns of biofilm formation are relatively similar to wild-type C. sakazakii on all food contact surfaces, ie, SS, Buna-N and PTFE. The statistical results showed that C. sakazakii can attach highest numbers on in PTFE, followed SS and Buna-N. The treatment effect of incubation time providing the highest biofilm density was formed at 48 h by wild-type and GFPuv labeled C. sakazakii FWH16 as well as wild-type C. sakazakii YRt2a. The GFPuv
Keyword: Biofilm, C. sakazakii, GFPuv, Multispecies, Competition