Enterobacter sakazakii is a gram-negative rod bacterium that was formerly known as "yellow-pigmented Enterobacter cloacae" until 1980. It is an emerging opportunistic pathogen associated with bacterial meningitis in immunocompromised neonates who ingested contaminated powdered infant formula. In several E. sakazakii-related outbreaks and sporadic cases, powdered infant formula was epidemiologically or microbiologically established as the source of infection. The International Commission on Microbiological Specifications for Foods has ranked E. sakazakii as a "severe hazard for restricted populations." However, the organism was isolated at very low levels from commercial powdered infant formula and dry environmental samples collected from infant formula factories. In most cases, the contamination did not exceed 1 cfu per 100 g. Progress in understanding the epidemiology of E. sakazakii outbreaks and instituting effective control measures has been poor to date. To clarify the mechanisms involved in infectivity and enterotoxin production, we performed two-dimensional gel electrophoresis (2-DE) coupled with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and isotope-coded affinity tags (ICATs)-LC -MS to identify and quantify differences in protein expression between the E. sakazakii strains. We found 23 spots in 2-DE and 37 proteins in ICAT analysis that were differentially expressed in E. sakazakii. These two methods gave us closely related but different information, suggesting that the two methods are complementary or at least supplementary. Enterobacter sakazakii has been implicated in meningitis or enteritis with 20% to >50% mortality of infants who contract the disease. Our research has been used in verification of Enterobacter sakazakii isolated from infant formula and other milk and milk related foods. We have established standard maps of two Enterobacter sakazakii strains which were isolated from infant formula. Through appearance or absence of specific spots and their isoelectric point or molecular weight comparison, a two dimensional electrophoresis standard map can be developed and used as a powerful tool to differentiate Enterobacter sakazakii from other Enterobacteriaceae. This offers microbiologists powerful tools for Enterobacter sakazakii detection and provides more basic information to investigate the pathogenesis of Enterobacter sakazakii and contamination sources in infant formula production.
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