Partial characterization of an enzymatic extract from Bentong ginger (Zingiber officinale var. Bentong).

Extraction of protease from a local ginger rhizome (Zingiber officinale var. Bentong) was carried out. The effect of extraction pH (6.4, 6.8, 7.0, 7.2, 7.6, 8.0, 8.4, and 8.8) and stabilizers (0.2% ascorbic acid, 0.2% ascorbic acid and 5 mM EDTA, or 10 mM cysteine and 5 mM EDTA) on protease activity during extraction was examined. pH 7.0 potassium phosphate buffer and 10 mM cysteine in combination with 5 mM EDTA as stabilizer were found to be the most effective conditions. The extraction procedure yielded 0.73% of Bentong ginger protease (BGP) with a specific activity of 24.8±0.2 U/mg protein. Inhibitory tests with some protease inhibitors classified the enzyme as a cysteine protease. The protease showed optimum activity at 60 °C and pH 6-8, respectively. The enzyme was completely inhibited by heavy metal cations such as Cu2+, and Hg2+. SDS stimulated the activity of enzyme, while emulsifiers (Tween 80 and Tween 20) slightly reduced its activity. The kinetic analysis showed that the protease has Km and Vmax values of 0.21 mg mL-1 and 34.48 mg mL-1 min-1, respectively. The dried enzyme retained its activity for 22 months when stored at -20 °C.

Characterization of pR18, a novel rolling-circle replication plasmid from Lactobacillus plantarum.

Lactobacillus plantarum PA18, a strain originally isolated from the leaves of Pandanus amaryllifolius, contains a pR18 plasmid. The pR18 plasmid is a 3211bp circular molecule with a G+C content of 35.8%. Nucleotide sequence analysis revealed two putative open reading frames, ORF1 and ORF2, in which ORF2 was predicted (317 amino acids) to be a replication protein and shared 99% similarity with the Rep proteins of pLR1, pLD1, pC30il, and pLP2000, which belong to the RCR pC194/pUB110 family. Sequence analysis also indicated that ORF1 was predicted to encode linA, an enzyme that enzymatically inactivates lincomycin. The result of Southern hybridization and mung bean nuclease treatment confirmed that pR18 replicated via the RCR mechanism. Phylogenetic tree analysis of pR18 plasmid proteins suggested that horizontal transfer of antibiotic resistance determinants without genes encoding mobilization has not only occurred between Bacillus and Lactobacillus but also between unrelated bacteria. Understanding this type of transfer could possibly play a key role in facilitating the study of the origin and evolution of lactobacillus plasmids. Quantitative PCR showed that the relative copy number of pR18 was approximately 39 copies per chromosome equivalent.

Inhibitory effects of lactic acid and lauricidin on spoilage organisms of chicken breast during storage at chilled temperature.

Different concentrations of lauricidin (LU, containing 1% lactic acid) and lactic acid alone (LA) were evaluated for their effectiveness in reducing naturally occurring microflora of raw chicken breasts. Chicken breasts were dipped in 0 (control), 0.5, 1.0, 1.5, and 2.0% solutions of LU (w/v) or LA (v/v) for 10, 20, and 30 min and stored at 4°C for 14 d. Total Plate Counts (TPC) and populations of Pseudomonas spp. and Enterobacteriaceae were determined before and after dipping and after storing for 1, 3, 7, 10, and 14 d. Additionally, Hunter L, a, and b values and pH of the chicken breast were also determined. From the obtained results, TPC on chicken breast treated with LU was found to be decreased by 0.92 to 1.2 log CFU/g from a mean initial log 5.69 CFU/g, while those dipped in LA decreased by 0.53 to 2.36 log CFU/g. Pseudomonas population on chicken breast dipped in LU decreased by 0.79 to 1.77 log CFU/g from an initial 3.90 log CFU/g, while in LA treated it decreased by 0.39 to 1.82 log CFU/g. Enterobacteriaceae counts were also found to be reduced by 0.14 to 1.14 log CFU/g on chicken breast dipped in LU, while the reduction was from 0.59 to 2.18 log CFU/g in chicken breast dipped in LA. The major bacterial types isolated from LU treated chicken breast belonged to the Enterobacteriaceae group, which included: Enterobacter, E. coli and Citrobacter. Whereas, in the LA treated breast it belonged to: Pseudomonas, E. coli, and Kocuria rhizophila (formerly Micrococcus luteus). Dipping chicken breast in LU and LA caused a significant decrease (p ≤ 0.05) in their pH values. Also, treatment with LU and LA caused a slight darkening in color (decreased Hunter L value), increase in redness (increased Hunter a value), and increase in yellowness (increased Hunter b value). Based on the results obtained in the present study, Lactic acid and Lauricidin showed high potential to be used as a sanitizer in reducing the population of spoilage microorganisms naturally occurring on raw chicken, and can be explored commercially for extension of their shelf life.

Prevalence and resistance to antibiotics for Aeromonas species from retail fish in Malaysia.

A total of 87 market fish samples representing five types of fish were evaluated for the presence of Aeromonas spp. Of the samples examined, 69%, 55%, 11.5% and 2.3% harbored Aeromonas spp., A. veronii biovar sobria, A. hydrophila and A. caviae, respectively. The 60 isolated Aeromonas spp. strains were further examined for hemolytic activity, resistance to antimicrobial agents and presence of plasmids. Hemolytic activity varied widely among the isolated strains. Though all the isolates demonstrated resistance to three or more of the antibiotics tested, all were susceptible to ceptazidime. Thirty-four (56.7%) of the sixty isolates harbored plasmids, with sizes ranging from 2.3 to 15.7 kb. These results indicate that hemolytic, multiple antibiotic resistant and genetically diverse aeromonads are easily recovered from fish in this region.