24 research outputs found

    Antibacterial potential components of Bacillus species and antibiotics residues in branded and unbranded honey samples from Nigeria

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    Honey is a sweet viscous liquid produced by honey bee, Apis mellifera from the nectar of plants. Honey is a natural product that has been used from ancient times till now as food and for medicinal purpose. This study was carried out to determine the mode of action of Bacillus species and antibiotics residues in branded and unbranded honey samples from Nigeria. Bacilli spp. count was carried out by initially heating diluted honey samples in water bath set at 80°C for 15 min, while total bacterial count was carried out using the pour plate method. Antibacterial activity of identified Bacillus spp on Micrococcus was determined using well-in agar method while the mode of action was carried out by reporter assay method. Detection of tetracycline, gentamycin and streptomycin was analyzed using high performance liquid chromatography (HPLC) column oven L-2300 and Column intensil ODS-3C18 (250 x 4.6 mm). Honey samples (2 g) were extracted for HPLC by deprotenizing using acetonitrile and methanol with flow rate of 1 mL/min and RID detector was used to detect antibiotic residue. Bacilli from honey were characterized physiologically, morphologically and biochemically, they were tentatively identified as Bacilli licheniformis, Bacilli subtilis, Bacilli coagulans, Bacilli cirulans, Bacilli pumilis and Bacilli badius. The most prevalent Bacillus spp. were B. licheniformis and B. subtilis. Total bacteria count for branded honey ranged from 2.2 x 102 to 5.5 x 103 cfu/g, while Bacilli count ranged from nil to 6.2 x 102 cfu/g. For unbranded honey samples, total bacteria count ranged from 7.0 x 103 to 3.5 x 102 cfu/g, while Bacilli count ranged from 5 x 101 to 1.6 x 103 cfu/g. Four of the isolates representing branded (SF2 and RW2) and unbranded honey samples (EH2 and TC2) exhibited antibacterial activity against Micrococcus; one isolate (SF2) showed cell wall causing antibacterial activity. Tetracycline was detected more in the unbranded honey samples while gentamycin and streptomycin were detected in just two unbranded honey samples, indicating that tetracycline is used frequently for the treatment of bee diseases that is why it is detected as residue in the finished honey product.Key words: Antibiotics, Bacillus, health benefit honey, high performance liquid chromatography (HPLC), residues

    Heterologous expression, purification and refolding of an anti-listerial peptide produced by Pediococcus acidilactici K7

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    The fusion protein, 6XHis-Xpress-PedA was constructed and expressed in Escherichia coli BL21 (DE3). The presence of a 12.8 kDa recombinant protein, localized in inclusion bodies (IBs) at high concentration, was confirmed by SDS-PAGE analysis and by western blotting using anti-His antibody. The rec-pediocin was purified by Nickel-nitrilotriacetic acid beads and refolded using 5 mM of \u3b2-mercaptoethanol along with 1 M glycine. Results indicated that the refolded rec-pediocin had an early elution profile in the RP-HPLC when compared to the unfolded protein and it exhibited biological activity against Listeria monocytogenes V7 which was approximately 25 times less active compared to native counterpart. The final yield of purified rec-pediocin was 3 mg/l of the culture and is estimated to be 8-10 times higher than the purification by conventional methods

    Molecular characterization of class IIa, heat-stable enterocin<i style=""> </i>produced by <i style="">Enterococcus faecium</i> MTCC 5153

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    307-315 The objective of this study was to provide molecular evidences for enterocin biosynthesis by Enterococcus faecium MTCC 5153. The culture filtrate (CF) of E. faecium MTCC 5153 exhibited a broad inhibitory spectrum against several enterococci, food-spoilage lactic acid bacteria (LAB) as well as pathogenic Gram-positive bacteria. The antimicrobial compound present in the CF showed the properties of class IIa bacteriocin. PCR was employed for taxonomic identification and detection of genes coding for virulence factors and different enterocins. Based on amplification of enterocin A (entA) gene by PCR, additional characterization of enterocin A operon was carried out. Southern hybridization confirmed the presence of chromosomally encoded enterocin A (entA) gene. These results suggest distribution of conserved enterocin A operon among different strains of E. faecium. Tricine SDS-PAGE activity gel assay in combination with mass spectral analysis indicated the production of single enterocin of 4.8 kDa in size. Production of enterocin was carried out using different sugars in tryptone, yeast extract (TYE) medium, under microaerophilic condition and the antimicrobial activity in the range of 10-50 x103 arbitrary units per milliliter (AU mL-1) was observed. This study indicates the potentiality of native isolate to produce broad spectrum bacteriocin which can be used to deal with spoilage bacteria and food-borne pathogens like listeria

    Cloning of pediocin PA-1 and its immunity genes from Pediococcus acidilactici K7 using pAMJ shuttle vector into Lactococcus lactis MG1363

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    550-553The matured pediocin PA-1 encoding gene, pedA and its immunity counterpart, pedB from Pediococcus acidilactici K7 were cloned by PCR technique and ligated into the E. coli-lactic shuttle, protein expression vector pAMJ2008. The recombinant pAMJAB was constructed and sequenced to confirm the intactness of reading frame of the deduced pediocin PA-1 fusion protein. Lactococcus lactis MG1363 was electroporated with the plasmids and the transformation efficiency of ~ 2-3 10³ g⁻¹ was observed. This was further confirmed by plasmid DNA isolation and analysis

    Redalyc.Heterologous expression, purification and refolding of an anti-listerial peptide produced by Pediococcus acidilactici K7

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    Chile Halami, Prakash M.; Chandrashekar, Arun Heterologous expression, purification and refolding of an anti-listerial peptide produced by Pediococcus acidilactici K

    Conjugal transfer of erm(B) and multiple tet genes from Lactobacillus spp. to bacterial pathogens in animal gut, in vitro and during food fermentation

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    Three strains of Lactobacillus comprising Lactobacillus salivarius (CHS-1E and CH7-1E) and Lactobacillus reuteri (CH2-2) previously isolated from chicken meat were analyzed for their transferability of antibiotic resistance (AR) genes to pathogenic strains under in vivo, in vitro, and during food fermentation. For in vivo model, Albino Wistar rats were inoculated with 10(10) CFU/g/ml of Enterococcus faecalis JH2-2 (recipient). After 7 days, either of two donors L. salivarius CH7-1E or L. reuteri harbouring erythromycin and tetracycline resistance genes] were introduced at a concentration of 10(9) CFU/ml daily for 1 week. Two days after donor introduction, there was a stable increase in the number of transconjugants in the animal faeces from 10(2) to 10(3)CFU/g and presented erm (B), tet(M), tet(L) and tet(W) in their genome like donor strains. Similar observations were made with in vitro filter mating between CHS-1E, CH2-2 and CH7-1E and E. faecalis JH2-2 with transfer frequencies of 1 x 10(-4), 3.8 x 10(-3) and 2 x 10(-3) per donor cell respectively. With the results obtained in vivo and in vitro, the AR transferability of donor strains was estimated during food fermentation (chicken sausage, fermented milk or idii batter) with pathogenic recipient strains added as contaminants. At the end of mating period, phenotypic resistance to erythromycin and tetracycline in Listeria monocytogenes and Yersinia enterocolitica strains was observed. This study showed the ability of food borne Lactobacillus in diffusing their AR traits in diverse natural environments increasing their concern of AR dissemination in the food chain when used as food additives and/or probiotics

    Heterogeneity of macrolide-lincosamide-streptogramin phenotype & conjugal transfer of erm(B) in Pediococcus pentosaceus

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    Background & objectives: Pediococcus pentosaceus has been reported to cause clinical infections while it is being promoted as probiotic in food formulations. Antibiotic resistance (AR) genes in this species are a matter of concern for treating clinical infections. The present study was aimed at understanding the phenotypic resistance of P. pentosaceus to macrolide-lincosamide-streptogramin B (MLSB) antibiotics and the transfer of AR to pathogens. Methods: P. pentosacues isolates (n=15) recovered from fermented foods were screened for phenotypic resistance to MLSB antibiotics using disc diffusion and microbroth dilution methods. Localization and transferability of the identified resistance genes, erm(B) and msr(C) were evaluated through Southern hybridization and in vitro conjugation methods. Results: Four different phenotypes; sensitive (S) (n=5), macrolide (M) (n=7), lincosamide (L) (n=2) and constitutive (cMLS(B)) (n=1) were observed among the 15 P. pentosaceus isolates. High-level resistance (>256 mu g/ml) to MLSB was observed with one cMLS(B) phenotypic isolate IB6-2A. Intermediate resistance (8-16 mu g/ml) to macrolides and lincosamides was observed among M and L phenotype isolates, respectively. Cultures with S phenotype were susceptible to all other antibiotics but showed unusual minimum inhibitory concentration (MIC) values of 8-16 mu g/ml for azithromycin. Southern hybridization studies revealed that resistance genes localized on the plasmids could be conjugally transferred to Enterococcus faecalis JH2-2. Interpretation & conclusions: The study provides insights into the emerging novel resistance patterns in P. pentosaceus and their ability to disseminate AR. Monitoring their resistance phenotypes before use of MLS antibiotics can help in successful treatment of Pediococcal infections in humans

    Emerging Microbial Concerns in Food Safety and New Control Measures

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    The editorial collects a brief summary of the topics discussed in the articles that are published in Emerging Microbial Concerns in Food Safety and New ControlMeasures. We hope that readers of this special issue will find some information of interest in order to expand their knowledge in this field and to increase their level of attention on matters here reported
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