Production of Bacteriocin-Like Inhibitory Compounds by Human Fecal Bifidobacterium Strains

Abstract

Acid- and bile-resistant Bifidobacterium strains were isolated from human feces and identified by genus-specific PCR and randomly amplified polymorphic DNA PCR. Twenty-four different strains were screened for possible production of proteinaceous antimicrobial compounds by assaying the inhibitory effects of their neutralized culture supernatants. Six Bifidobacterium strains (BIR-0304, BIR-0307, BIR-0312, BIR-0324, BIR-0326, and BIR-0349) were selected on the basis of their broad inhibitory spectra. These strains were active against gram-positive and gram-negative bacteria and yeasts relevant to food safety and human health. The antagonistic effects of the six selected Bifidobacterium strains were related to bacteriocin-like compounds, which were active at pH values between 3 and 10, stable at 100 degrees C for 10 min, resistant to alpha-amylase and lipase A, but sensitive to proteinases (trypsin, proteinase K, protease A, pepsin, and cathepsin B). The molecular masses of the antimicrobial compounds produced by Bifidobacterium BIR-0312 and BIR-0324 were in the range of 10 to 30 kDa, and those of the compounds produced by Bifidobacterium BIR-0304, BIR-0307, BIR-0326, and BIR-0349 were less than 10 kDa. All Bifidobacterium strains produced maximum antimicrobial activities in the late logarithmic phase of growth and in the presence of Tween 80. These results confirm that the synthesis of bacteriocin-like inhibitory compounds is a key factor in the in vitro inhibition of pathogen and spoilage bacteria by Bifidobacterium strains.

Probiotics Applications in Autoimmune Diseases

“…The balance of different bacteria stabilises as people age but is still affected by factors including diet, location, antibiotic use and radiation exposure in adults. Gut composition seems to become more unstable again as people age, as the faecal microbial profiles of those 65 years and older show considerably more variability between individuals [89].…”

Produção de bacteriocina por Bifidobacterium lactis a partir de soro de leite

“…Additional antimicrobials from six Bifidobacterium strains were found to exhibit broad inhibitory spectra against both Gram-negative and Gram-positive bacteria, namely Clostridium difficile, Brochothrix thermosphacta, L. monocytogenes, S. aureus, Helicobacter pylori, S. typhimurium, Arcobacter butzleri, and some pathogenic yeasts. These heat-stable compounds were sensitive to proteinases and resistant to pH in the range from 3 to 10 (Collado et al, 2005b), but were neither purified nor subject to further investigation. Finally, von Ah (2006) identified, recovered by methanol/acetone extraction and reversed-phase HPLC and partly characterized thermophilicin B67, a bacteriocin produced by Bifidobacterium thermophilum RBL67 that exhibited a narrow inhibition spectrum towards three Listeria strains and Lactobacillus acidophilus.…”

“…was able to stimulate bacteriocin production by the latter and, in some cases, the addition of surfactants such as Tween 80 increased the concentration of bacteriocins produced as a consequence of cell growth acceleration. Surfactants may also enhance the sensitivity of the indicator strain and form micelles with proteinaceous compounds, thus stabilizing the bacteriocins (Carolissen-Mackay et al, 1997;Cheikhyoussef et al, 2008;Collado et al, 2005b).…”

“…Other alternatives include the use of acid or organic solvents as precipitating agents (Gibson and Wang, 1994a). Dialysis and ultrafiltration can also be used to further concentrate and purify bacteriocins, even though some Bifidobacterium-associated studies pointed out large product losses due to incomplete precipitation (Collado et al, 2005b;Liévin et al, 2000). The final purification step generally consists of reversed phase high-performance liquid chromatography (RP-HPLC) with acetonitrile gradient, which can be followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) or mass spectrometry for molecular size determination.…”

“…Nonetheless, several investigations have demonstrated that the use of complex growth media with a high peptide content is necessary to ensure high bacteriocin productivity (Carolissen-Mackay et al, 1997), and thus a tradeoff is required. Similarly, if from one hand the addition of Tween 80 can enhance bacteriocin production, from the other it can affect the purification process through, for example, the formation of precipitates in the culture supernatants (Collado et al, 2005b).…”