Purification and characterization of enterocin FH 99 produced by a faecal isolate Enterococcus faecium FH 99

Abstract

Enterococcus faecium FH 99 was isolated from human faeces and selected because of its broad spectrum of inhibitory activity against several Gram-positive foodborne spoilage and pathogenic bacteria. Ent. faecium FH 99 accumulates enterocin in large number in early stationary phase of the growth. The enterocin FH 99 was stable over a wide pH range (2-10) and recovered activity even after treatment at high temperatures (10 min at 100°C). The enterocin was subjected to different purifi cation techniques viz., gel fi lteration, cation exchange chromatography and reverse-phase high-performance liquid chromatography. The activity was eluted as one individual active fraction. SDS-PAGE revealed a molecular weight of less than 6.5 kDa. Studies carried out to identify the genetic determinants for bacteriocin production showed that this trait may be plasmid encoded as loss in both of the plasmids (size>chromosomal DNA) led to loss in bacteriocin production by Ent. faecium FH 99. Ent. faecium strain FH 99 is a newly discovered high bacteriocin producer with Activity Units 1.8 × 10 5 AU ml -1 and its characteristics indicate that it may have strong potential for application as a protective agent against pathogens and spoilage bacteria in foods.

Antibacterial efficacy of Nisin, Pediocin 34 and Enterocin FH99 against Listeria monocytogenes and cross resistance of its bacteriocin resistant variants to common food preservatives

“…Enterococcus faecium FH99, bacteriocinogenic strain was a isolate from human faeces (Gupta et al , 2010). Pediococcus pentosaceous 34, a bacteriocinogenic strain was an isolate from cheddar cheese.…”

Antibacterial efficacy of nisin, pediocin 34 and enterocin FH99 against L. monocytogenes, E. faecium and E. faecalis and bacteriocin cross resistance and antibiotic susceptibility of their bacteriocin resistant variants

“…Bacterial strains and culture conditions E. faecium FH99, bacteriocinogenic strain was an isolate from human feces (Gupta et al 2010). Pediococcus pentosaceous 34 (Rao and Malik 2003), a bacteriocinogenic strain was an isolate from cheddar cheese, Pediococcus acidilactici LB 42 (a sensitive strain used for detection of bacteriocin producers), was obtained from , Department of Animal Science, University of Wyoming, Laramie Wyoming, USA.…”

“…Crude enterocin FH99 and pediocin 34 were precipitated from broth media by 60% ammonium sulphate precipitation and the precipitates were dissolved in sterilized Milli Q water. The bacteriocins were purified by the method earlier described by Gupta et al (2010). Nisin A (Nisaplin ®) was procured from Danisco (Gurgaon, India).…”

Mechanism of Nisin, Pediocin 34, and Enterocin FH99 Resistance in Listeria monocytogenes

“…Enterococcus faecium FH99 bacteriocinogenic strain was an isolate from human feces [14]. Pediococcus pentosaceous 34, a bacteriocinogenic strain isolated from cheddar cheese and Pediococcus acidilactici LB 42 (a sensitive strain used for detection of bacteriocin producers), was obtained from Prof Bibek Ray, Department of Animal Science, University of Wyoming, Laramie, Wyoming, USA.…”

“…One liter aliquots of MRS broth (pH 6.5) (HiMedia, Mumbai) were inoculated with active cultures of E. faecium FH 99 [14] and P. pentosaceus 34 (1% v/v) and incubated at 37°C for 24 h. Cell-free culture supernatants (CFCS) were prepared by centrifugation of the cultures in refrigerated centrifuge (HANIL, Supra-30 K) at 7,000g for 10 min, filter sterilized by passing through a 0.2 lm, 45 mm diameter membrane filter and used for partial purification after neutralization. To 1,000 mL of cell-free culture supernatant, solid ammonium sulfate was added slowly with constant stirring to achieve 60% saturation and stirring was continued for another 1 h in a cold room at 5-7°C.…”