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Li Z.,Ocean University of China | Lu Z.,Ocean University of China | Khan M.N.,Ocean University of China | Khan M.N.,Microbiological Analytical Center | And 2 more authors.
Food and Chemical Toxicology | Year: 2014

The present study aimed to investigate the relationship between protein carbonylation and changes of the IgE reactivity of turbot parvalbumin (PV) following electron beam (EB) irradiation. The concentration of protein carbonyls, specific IgE binding, and IgE binding inhibition between irradiated and oxidized PV were assessed. Irradiation resulted in a 3-fold enhancement in the protein carbonyl content. In purified PV irradiated with a 10-kGy dose, specific IgE binding was reduced by 91.2±6.2%. When raw PV was treated with reactive oxygen species (ROS), the protein carbonyl content increased 17.6-fold, with the specific IgE binding being reduced by 87.9±6.5% at an ROS concentration of 10nmol/mL. The IgE binding inhibition between irradiated and oxidized PV was investigated using an inhibition ELISA. Results showed that oxidized PV can inhibit the binding between irradiated PV and specific IgE with an IC50 of 8.2-58ng according to different doses of irradiation. These findings suggest that EB irradiation reduces specific IgE binding, probably by the induction of protein carbonylation. © 2014 Elsevier Ltd.

Rong R.,Ocean University of China | Lin H.,Ocean University of China | Wang J.,Ocean University of China | Khan M.N.,Ocean University of China | And 2 more authors.
Aquaculture | Year: 2014

Vibrio parahaemolyticus is closely associated with oysters in China, which are normally consumed raw or lightly cooked. Bacteriophages are a safe bio-controlling agent, and have been recognized in aquaculture for their pathogen reduction properties. This study investigated the potential application of the bacteriophage VPp1 during depuration to reduce V. parahaemolyticus in oysters at different multiplicity of infection (MOI) and temperature levels. Oysters were infected with 105, 106, and 107 colony-forming units (CFU)/mL of V. parahaemolyticus in the seawater and each infected group was treated with three different MOI values: 10, 1, and 0.1. Infected oysters were depurated in non-recirculating seawater at 22°C, 20°C, 16°C, and 12°C for 36h. Results revealed that temperatures <20°C were safe for oyster rearing. Depuration at 16°C with 0.1 MOI was the best condition for reducing V. parahaemolyticus in oysters, which decreased by 2.35-2.76logCFU/g within 36h. This study provides the basis for the use of bacteriophages in depuration techniques to eliminate V. parahaemolyticus in oysters. © 2013 Elsevier B.V.

Duan C.,Ocean University of China | Chen C.,Ocean University of China | Khan M.N.,Ocean University of China | Khan M.N.,Microbiological Analytical Center | And 4 more authors.
Food Control | Year: 2014

There is an emerging trend of non-destructive and onsite analysis of microbial contaminations for better food safety. A new strategy for determination of total bacterial in fish products (flounder fillets) was established using a portable near infrared spectrometer. Results revealed that the pretreatment of near infrared spectrum by the wavelet transform could significantly improve the accuracy and precision of the analysis. In comparison to usually exploited partial least squares regression (PLS), a combination of genetic algorithm (GA) and back-propagation artificial neural network (BP-ANN) exhibited much better efficiency, and the correlation coefficient (R) and root mean square error (RMSE) of the prediction model were calculated as 0.985 and 0.095, respectively, and validated as 0.966 and 0.083, respectively. These results allowed us to suggest a promising potential of the established technique for non-destructive and onsite monitoring of total bacteria in fishery products. © 2014 Elsevier Ltd.

Mirani Z.A.,Microbiological Analytical Center | Jamil N.,University of Karachi
Journal of the College of Physicians and Surgeons Pakistan | Year: 2013

Objective: To study the genomic organization of vancomycin resistance in a local isolate of vancomycin resistant Staphylococcus aureus (VRSA). Study Design: Experimental study. Place and Duration of Study: Department of Microbiology, University of Karachi, January 2008 through December 2010. Methodology: A vancomycin-resistant Staphylococcus aureus (VRSA-CP2) isolate (MIC 16 μg/ml) was isolated from a local hospital of Karachi. Species identification was confirmed by Gram staining, standard biochemical tests and PCR amplification of the nuc gene. The vancomycin MIC was re-confirmed by E-test. For the genetic determination of vancomycin resistance, in-vitro amplification of vanA cassette was performed by using plasmid DNA of CP2, CP2's transformant as template on MWG Thermo-Cycler. Amplified products of vanR, vanS, vanH, vanA, vanY, orf2, orf1D, orf2E, orf-Rev and IS element genes were subjected to Sanger's electrophoresis based sequence determination using specific primers. The Basic Local Alignment Search Tool (BLAST) algorithm was used to identify sequences in GenBank with similarities to the vanA cassette genes. Results: The vancomycin-resistant isolate CP2 was found to be resistant to oxacillin, chloramphenicol, erythromycin, rifampicin, gentamicin, tetracycline and ciprofloxacin, as well. The isolate CP2 revealed four bands: one of large molecular size ~56.4 kb and three of small size ~6.5 kb, ~6.1 kb and ~1.5 kb by agarose gel electrophoresis indicating the presence of 3 plasmids. The plasmid DNA of isolate CP2 was analyzed by PCR for the presence of the van cassettes with each of the vanA, vanB and vanC specific primers. It carried vanA cassette, which comprises of vanR, vanS, vanH, vanA, vanY, and orf2. The vanA cassette of isolate CP2 also carried an insertion element (IS). However, it did not show the PCR product for orf1. Vancomycin resistance was successfully transferred from the donor CP2 to a vancomycin-sensitive recipient S. aureus. The MIC of vancomycin for the transformant was 16 μg/ml, similar to the parent isolate CP2. Nucleotide sequencing of the PCR product showed similarity with van genes of enterococci and other VRSA reported from different parts of the world. Conclusion: Sequence of vanA cassette of CP2 showed partial homology with vancomycin resistant enterococci, VRSA vanA cassette element recorded in gene bank NCBI.

Han F.,Ocean University of China | Li M.,Ocean University of China | Lin H.,Ocean University of China | Wang J.,Ocean University of China | And 3 more authors.
Journal of Industrial Microbiology and Biotechnology | Year: 2014

Shewanella putrefaciens has been identified as a specific spoilage organism commonly found in chilled fresh fish, which contributes to the spoilage of fish products. Limiting S. putrefaciens growth can extend the shelf-life of chilled fish. Endolysins, which are lytic enzymes produced by bacteriophages, have been considered an alternative to control bacterial growth, and have been useful in various applications, including food preservation. We report here, for the first time, the complete genome sequence of a novel phage Spp001, which lyses S. putrefaciens Sp225. The Spp001 genome comprises a 54,789-bp DNA molecule with 67 open reading frames and an average total G + C content of 49.42%. In silico analysis revealed that the Spp001 open reading frames encode various putative functional proteins, including an endolysin (ORF 62); however, no sequence for genes encoding the holin polypeptides, which work in concert with endolysins, was identified. To examine further the lytic activity of Spp001, we analyzed the lytic enzyme-containing fraction from phages released at the end of the phage lytic cycle in S. putrefaciens, using diffusion and turbidimetric assays. The results show that the partially purified extract contained endolysin, as indicated by a high hydrolytic activity towards bacterial peptidoglycan decrease in the OD590 value by 0.160 in 15 min. The results will allow further investigation of the purification of natural Spp001 endolysin, the extension of Spp001 host range, and the applications of the phage-encoded enzymes. © Society for Industrial Microbiology and Biotechnology 2014.

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