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2011 - 20153
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Subject: Microbiology × Author: GUPTA, ANUPAMA ×
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    ThesisItemOpen Access
    PCR DETECTION OF BACTERIA ISOLATE NO. A75 AND APPLICATION OF BACTERIOCIN PRODUCED FROM IT AS FOOD BIOPRESERVATIVE
    (2011) GUPTA, ANUPAMA; SHARMA, NIVEDITA
    ABSTRACT The present investigation was carried out to purify, characterize and apply bacteriocin produced from Bacillus sp. A75 as food biopreservative. Bacillus sp. A75 was tested for antimicrobial activity against ten indicator strains viz. Listeria monocytogenes, Bacillus subtilis, Staphylococcus aureus, Escherichia coli, Bacillus cereus, Clostridium perfringens, Aeromonas hydrophila, Leuconostoc mesenteroides, Enterococcus faecalis and Lactobaciilus plantarum by bit/disk and well diffusion method. The antimicrobial activity was seen against 6 tested indicators viz. L. monocytogenes, B. subtilis, C. perfringens, S. aureus, L. plantarum and L. mesenteroides. On the basis of zone size three indicator strains were selected viz. L. monocytogenes, B. subtilis and C. perfringens for further studies. Isolate A75 was identified as Bacillus sp. at molecular level using 16S r RNA gene technique. Increase in specific activity of Bacillus sp. A75 in case of partially purified bacteriocin was 3.5 fold while in case of purified bacteriocin it was 41 fold. The recovery in case of partially purified bacteriocin was 82% and in case of purified bacteriocin it was 9%. The zone sizes, noted by well diffusion assay, in case of purified bacteriocin of Bacillus sp. A75 increased against the respective indicators. Bacteriocin of Bacillus sp. A75 was characterized by studying the effect of temperature, pH, proteolytic enzyme and stability. Bacteriocin was found to be thermostable upto 100oC, showed sensitivity to trypsin and was found to be active at neutral pH. The bacteriocin of Bacillus sp. A75 showed better resistance to the spoilage causing microorganisms in milk, cheese, orange juice and tomato paste as compared to chemical preservative-sodium benzoate while it showed almost as effective results as shown by nisin- commercial biopreservative at refrigeration and room temperature (15-18oC). The use of bacteriocin as food biopreservative has been found effective for shelf stability of food products at 4oC and room temperature as compared to chemical preservative.
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    ThesisItemOpen Access
    CHARACTERIZATION OF POTENTIALLY ACTIVE NEW PROBIOTIC STRAINSISOLATEDFROM DIFFERENT SOURCES AND TO STUDY THEIR PROSPECTS AS NUTRACEUTICAL AGENTS
    (2015) GUPTA, ANUPAMA; SHARMA, NIVEDITA
    ABSTRACT The present investigation was carried out to isolate most efficient potential probiotic lactic acid bacteria from rare and unexplored food sources, their screening, identification, safety assessment, evaluationof probiotic attributes, metabolic fingerprinting as well the development of novel nutraceutical products. In total 27 bacterial isolates were obtained from fermented/non fermented food sources. Out of all, 6 bacterial isolates were screened by bit/disc as well as well diffusion method on the basis of their wide inhibitory spectrum against tested pathogens. Isolates C-1, LB-CC, LB-WC, Ch-1, Ch-2 and 107 were selected for further studies having widest antagonistic potential, which were isolated fromkodo millet flour, lasoda bari, chuli and salori, respectively that are traditional food products of Himachal Pradesh. Isolates C-1, LB-CC and LB-WC were identifies as Pediococcus pentosaceus, Ch-1 as Enterococcus faecium, Ch-2 as Pediococcus acidilactici while 107 identified as Pediococcus damnosus by 16S rRNA gene technique and registered in NCBI under accession no. KM251461, KM251460, KJ489415, KJ541885, KJ541886 and KJ577588, respectively. Safety assessment of isolates was done by evaluating antibiotic susceptibility, haemolytic, DNase and gelatinase activities. All the isolates exhibited 80-100% antibiotic sensitivity, non-heamolytic, non-DNase and non-gelatinase activities, thereby proving their safe status. These screened isolates were further examined for their probiotic potential viz. acidity tolerance, bile tolerance, simulated gastrointestinal transit, auto-, co-aggregation, hydrophobicity, adhesion to gastric as well as mammalian cell lines, competitive exclusion of pathogens, antimicrobial potential, exopolysaccharides and β-galactosidase production, compatibility and cumulative probiotic potential. All the six screened isolates were found to be highly acidity tolerant strains with 93.65-99.41 % survival rate at pH 3 for 3 h. These six isolates were able to resist high bile salt concentration i.e. 2.0% with 93.12-98.22 % survival rate for 8 h. Resistance to gastrointestinal transit as found to be in a range from 43.09-90.12 %. All the six isolates exhibited good autoaggregation capacity i.e. greater than 40% after 5 h and showed string hydrophobicity towards xylene i.e. > 40%. All the isolates were able to adhere to gastric mucin with 14.7-49.5% adhesion and all showed adhesion potential to mammalian epithelial cell lines. These screened probiotic isolates showed broad and strong inhibitory spectrum against both gram positive as well as gram negative pathogenic bacteria through secretion of antimicrobial metabolites viz. lactic acid (7.18-8.46 mg/l), H2O2 (0.24-0.38g/l) and bacteriocins (77.33-100.0 AU/ml). Metabolic fingerprinting was done to elucidate the total metabolic profile of isolates and P. pentosaceus LB-CC and P. acidilactici Ch-2 were being first lactic acid bacteria to be reported for the production of squalene with anticancerous properties. The entire screened six isolates were highly qualified with cumulative score of 95.83% and are being used to prepare probiotic nutraceutical products with high antioxidant content viz. novel cereal based probiotic acidulant, probiotic enriched mango fortified shrikhand and probiotic enriched pulse fortified yogurtand are successfully accepted in their sensory evaluation. Hence, this study affirms the use of P. pentosaceus C-1, P. pentosaceus LB-CC, P. pentosaceus LB-WC, E. faecium Ch-1, P. acidilactici Ch-2 and P. damnosus 107 in the development of new functional foods and nutraceutical preparations to impart betterment to the public health as these six strains isolated in the present study have been proved safe as well as highly effective probiotic candidates
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    ThesisItemOpen Access
    CHARACTERIZATION OF POTENTIALLY ACTIVE NEW PROBIOTIC STRAINS ISOLATED FROM DIFFERENT SOURCES AND TO STUDY THEIR PROSPECTS AS NUTRACEUTICAL AGENTS
    (2015) GUPTA, ANUPAMA; SHARMA, NIVEDITA
    ABSTRACT The present investigation was carried out to isolate most efficient potential probiotic lactic acid bacteria from rare and unexplored food sources, their screening, identification, safety assessment, evaluation of probiotic attributes, metabolic fingerprinting as well the development of novel nutraceutical products. In total 27 bacterial isolates were obtained from fermented/non fermented food sources. Out of all, 6 bacterial isolates were screened by bit/disc as well as well diffusion method on the basis of their wide inhibitory spectrum against tested pathogens. Isolates C-1, LB-CC, LB-WC, Ch-1, Ch-2 and 107 were selected for further studies having widest antagonistic potential, which were isolated from kodo millet flour, lasoda bari, chuli and salori, respectively that are traditional food products of Himachal Pradesh. Isolates C-1, LB-CC and LB-WC were identifies as Pediococcus pentosaceus, Ch-1 as Enterococcus faecium, Ch-2 as Pediococcus acidilactici while 107 identified as Pediococcus damnosus by 16S rRNA gene technique and registered in NCBI under accession no. KM251461, KM251460, KJ489415, KJ541885, KJ541886 and KJ577588, respectively. Safety assessment of isolates was done by evaluating antibiotic susceptibility, haemolytic, DNase and gelatinase activities. All the isolates exhibited 80-100% antibiotic sensitivity, non-heamolytic, non-DNase and non-gelatinase activities, thereby proving their safe status. These screened isolates were further examined for their probiotic potential viz. acidity tolerance, bile tolerance, simulated gastrointestinal transit, auto-, co-aggregation, hydrophobicity, adhesion to gastric as well as mammalian cell lines, competitive exclusion of pathogens, antimicrobial potential, exopolysaccharides and β-galactosidase production, compatibility and cumulative probiotic potential. All the six screened isolates were found to be highly acidity tolerant strains with 93.65-99.41 % survival rate at pH 3 for 3 h. These six isolates were able to resist high bile salt concentration i.e. 2.0% with 93.12-98.22 % survival rate for 8 h. Resistance to gastrointestinal transit as found to be in a range from 43.09-90.12 %. All the six isolates exhibited good autoaggregation capacity i.e. greater than 40% after 5 h and showed string hydrophobicity towards xylene i.e. > 40%. All the isolates were able to adhere to gastric mucin with 14.7-49.5% adhesion and all showed adhesion potential to mammalian epithelial cell lines. These screened probiotic isolates showed broad and strong inhibitory spectrum against both gram positive as well as gram negative pathogenic bacteria through secretion of antimicrobial metabolites viz. lactic acid (7.18-8.46 mg/l), H2O2 (0.24-0.38g/l) and bacteriocins (77.33-100.0 AU/ml). Metabolic fingerprinting was done to elucidate the total metabolic profile of isolates and P. pentosaceus LB-CC and P. acidilactici Ch-2 were being first lactic acid bacteria to be reported for the production of squalene with anticancerous properties. The entire screened six isolates were highly qualified with cumulative score of 95.83% and are being used to prepare probiotic nutraceutical products with high antioxidant content viz. novel cereal based probiotic acidulant, probiotic enriched mango fortified shrikhand and probiotic enriched pulse fortified yogurt and are successfully accepted in their sensory evaluation. Hence, this study affirms the use of P. pentosaceus C-1, P. pentosaceus LB-CC, P. pentosaceus LB-WC, E. faecium Ch-1, P. acidilactici Ch-2 and P. damnosus 107 in the development of new functional foods and nutraceutical preparations to impart betterment to the public health as these six strains isolated in the present study have been proved safe as well as highly effective probiotic candidates.