Pak-Lam Yu

Enterocins in food preservation.

The Enterococcus genus, a member of the Lactic Acid Bacteria (LAB) is found in various environments, but more particularly in the intestines of humans and other animals. Although sometimes associated with pathogenicity these bacteria have many benefits. They have been found in traditional artisanal fermented products, are used as probiotic cultures and nowadays extensively studied for the production of bacteriocins--the enterocins. Many of these enterocins have been found to be active against Listeria monocytogenes, and a few have also been reported to be active even against Gram negative bacteria, an unusual property for the bacteriocins produced by LAB. These properties have resulted in many studies describing the use of enterocins as preservatives in foods of animal and vegetable origin. This review covers the most recent information on the use of enterocins as food preservatives, either produced in-situ by the addition of enterocin producing strains or as external preservatives in the form of purified or semi-purified extracts, to prevent the growth of spoilage and pathogenic microorganisms.

Antimicrobial Activity and Bacterial-Membrane Interaction of Ovine-Derived Cathelicidins

ABSTRACT Three ovine-derived cathelicidins, SMAP29, OaBac5mini, and OaBac7.5mini, were compared with respect to their antibacterial activities and interactions with membranes. SMAP29 was confirmed to be α-helical, broad spectrum, and able to disrupt both the outer and the cytoplasmic membranes at relatively low concentrations. In contrast, the two proline- and arginine-rich OaBac peptides had more-modest antibacterial activities, reduced levels of lipopolysaccharide binding, and a lesser ability to depolarize the cytoplasmic membrane, consistent with a cytoplasmic target.

Genetic analysis of a lactococcal plasmid replicon

SummaryThe sequence and genetic organization was determined of the 2508 by lactococcal portion of pFX2, which was derived from a crypticLactococcus lactis subsp.lactis plasmid and used as the basis for construction of a series of lactococcal vectors. A lactococcal plasmid plus origin and two replication protein-coding regions (repA andrepB) were located. RepA has a helix-turn-helix motif, a geometry typical of DNA-binding proteins. RepB shows a high degree of homology to the plasmid replication initiation proteins from other gram-positive bacteria andMycoplasma. The transcribed inverted repeat sequence betweenrepA andrepB could form an attenuator to regulate pFX2 replication. Upstream of theori site, and in a region which was non-essential for replication, a 215 by sequence identical to the staphylococcal plasmid pE194 and carrying the RSA site was identified. The genetic organization of this lactococcal plasmid replicon shares significant similarity with pE194 group plasmids.

Rapid and molecular selective electrochemical sensing of phthalates in aqueous solution

Reported research work presents real time non-invasive detection of phthalates in spiked aqueous samples by employing electrochemical impedance spectroscopy (EIS) technique incorporating a novel interdigital capacitive sensor with multiple sensing thin film gold micro-electrodes fabricated on native silicon dioxide layer grown on semiconducting single crystal silicon wafer. The sensing surface was functionalized by a self-assembled monolayer of 3-aminopropyltrietoxysilane (APTES) with embedded molecular imprinted polymer (MIP) to introduce selectivity for the di(2-ethylhexyl) phthalate (DEHP) molecule. Various concentrations (1-100 ppm) of DEHP in deionized MilliQ water were tested using the functionalized sensing surface to capture the analyte. Frequency response analyzer (FRA) algorithm was used to obtain impedance spectra so as to determine sample conductance and capacitance for evaluation of phthalate concentration in the sample solution. Spectrum analysis algorithm interpreted the experimentally obtained impedance spectra by applying complex nonlinear least square (CNLS) curve fitting in order to obtain electrochemical equivalent circuit and corresponding circuit parameters describing the kinetics of the electrochemical cell. Principal component analysis was applied to deduce the effects of surface immobilized molecular imprinted polymer layer on the evaluated circuit parameters and its electrical response. The results obtained by the testing system were validated using commercially available high performance liquid chromatography diode array detector system.

Differential induction of β-galactosidase and phospho-β-galactosidase activities in the fermentation of whey permeate by Clostridium acetobutylicum

SummaryStrains of Clostridium acetobutylicum were tested for the presence of β-galactosidase and phospho-β-galactosidase activities when grown on lactose. All strains, except C. acetobutylicum ATCC 824, showed both enzyme activities. Only phospho-β-galactosidase activity was detected with C. acetobutylicum ATCC 824. C. acetobutylicum strains P262 and ATCC 824 showed no detectable β-galactosidase or phospho-β-galactosidase activities when grown on glucose. In the fermentation of whey permeate C. acetobutylicum P262 showed an early induction of phospho-β-galactosidase associated with the acidogenic phase. The β-galactosidase activity peaked at a later stage of the fermentation (22 h) coinciding with the solvent production phase. Similar induction of phospho-β-galactosidase at the early stages (13 h) of fermentation of whey permeate by C. acetobutylicum ATCC 824 was also shown. No β-galactosidase activity was detected during the entire course of fermentation by strain ATCC 824.

Conjugal transfer of Streptococcal antibiotic resistance plasmids intoClostridiumacetobutylicum

SummaryStreptococcal plasmids pAMβ1, pVA797, pVA797∶∶Tn917 and pAM610 were transferred or mobilized toClostridiumacetobútylicum fromStreptococcussanguis,S.faecalis andS.lactis donors.Clostridum transconjugants were able to retransfer pAMβ1 and pVA797 to a plasmid-freeS.lactis recipient.

Synergistic effects of ovine-derived cathelicidins and other antimicrobials against Escherichia coli O157:H7 and Staphylococcus aureus 1056 MRSA

Synergistic effects of ovine-derived cathelicidins SMAP29 and OaBac5mini with the antimicrobials polymyxin B, lysozyme, nisin and lactoferrin were investigated against E. coli O157:H7 and S. aureus 1056 MRSA. Lysozyme showed synergy against E. coli O157:H7 with SMAP29, polymyxin B and lactoferrin. Synergy was also found between SMAP29 and lactoferrin against this host. Against S. aureus 1056 MRSA, lysozyme showed synergy with OaBac5mini, polymyxin B and nisin, while synergy was also found between nisin and OaBac5mini and polymyxin B. Other combinations of the antimicrobials were either additive or non-synergistic.

Molecular cloning and expression of a proteinase gene from Lactococcus lactis subsp. cremoris H2 and construction of a new lactococcal vector pFX1

The 6.5 kb HindIII DNA fragment of the Lactococcus lactis subsp. cremoris H2 plasmid pDI21 was cloned into Escherichia coli POP13 with λNM1149, and also directly into Lactococcus lactis subsp. lactis 4125 using a newly-constructed broad host-range vector pFX1. Proteinase was experessed in both transformed organisms. The proteinase resembles a PI type since it preferentially degraded β-casein. The restriction map of the 6.5 kb proteinase gene fragment has minor differences from those of published plamid proteinase genes. High-efficiency electroporation with pFX1 provides a direct approach for gene cloning in lactococci.

Molecular cloning and expression of multiple cellulase genes of Ruminococcus flavefaciens strain 186 in Escherichia coli

SummaryCellulase genes of the ruminant micro-organism Ruminococcus flavefaciens strain 186 have been cloned and expressed in Escherichia coli using the bacteriophage vector λNM1149. Twenty-six clones showed expression of endo-β-1,4-d-glucanases and were divided into four groups according to their insert sizes of approximately 2, 3, 4 or 9 kilobases (kb). Two of the clones with 4 kb inserts also showed exo-β-1,4-d glucanase activity while two clones with 9 kb inserts showed β-glucosidase activity. One of the clones with 9 kb inserts (λM903) showed the activities of all three cellulase activities. In addition, two of the 4 kb-insert clones and one 9 kb-insert clone degraded Avicel (PH101).

Antimicrobial fragments of the pro-region of cathelicidins and other immune peptides

In addition to the numerous cathelicidin peptides that are associated with the antimicrobial activity exhibited by a crude extract from ovine blood, a further three peptides with antimicrobial activity have been identified. These were part of the precursor cathelin domain of cathelicidins, a large fragment of platelet factor 4 and a small peptide similar to signal peptide of the T-cell glycoprotein CD4 precursor. Fragments of proteins that are involved in protecting the host from infection may have a secondary purpose as antimicrobial agents once they have carried out their primary purpose and are cleaved the main protein.