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ThesisItem Restricted Use of bioinsecticides in compost for button mushroom cultivation(Punjab Agricultural University, Ludhiana, 2020) Simran; Sharma, ShivaniAgaricus bisporus is the most commonly cultivated edible variety in the world which possesses an ability to grow on agricultural wastes. Therefore, the present study has dealt with the use of pre botanical insecticides such as neem powder, vinegar, castor powder and two biological insecticides such as Bacillus thuringiensis and Beauveria bassiana to study its effect on mycelial growth of A. bisporus on PDA medium and wheat straw based compost. Maximum growth of 55mm and 29mm was recorded with neem powder on PDA and compost respectively. However, vinegar and Bacillus thuringiensis also showed significant mycelial run on PDA as well as compost. The natural insecticides were supplemented @0.5 to 4.5% (w/w) in ready compost and spawned @1.0% in compost. The supplementation of neem powder @1.5% showed significantly improved yield potential 43.40% and increased number of fruit bodies 37.55%. In case of castor powder (0.5 to 4.5% v/w), supplementation @ 4.5% (v/w) showed an increase of 23.22% in yield of button mushroom. An increase upto 20.23% and 46.42% in yield and fruit bodies was observed when vinegar was supplemented @2.5%. Bacillus thuringiensis supplementation @2.5% showed maximum yield potential 33.96%. Similarly, Beauveria bassiana was supplemented at a population of 3.4×107 cfu/ml @0.54.5% (v/w) in compost which significantly improved the yield upto 14.44% and 12.62% when Beauveria bassiana was supplemented @1.5% and 0.5% at the time of spawning. Thus, it was concluded from our study that plant-derived products can be recommended to substitute for synthetic chemicals in the commercial production of edible mushrooms.ThesisItem Open Access Evaluation of antioxidant activity in selenium biofortified oyster mushrooms; Pleurotus spp.(Punjab Agricultural University, Ludhiana, 2020) Sravani, Anduri; Sharma, ShivaniPleurotus florida, P. eryngii, P. cornucopiae and P. djamor were studied for comparing the effect of selenium biofortification on antioxidant activity. Maximum radial growth, biomass production and total soluble protein content of mycelium were observed @ 5 mg L-1 in P. florida (9.00 mm/day, 88.0 g L-1 and 2.28 mg g-1 respectively), @ 10 mg L-1 in P. cornucopiae (8.02 mm/day, 58.3g L-1 and 1.94 mg g-1 respectively), P. djamor (7.11 mm/day, 88.5g L-1and 1.97 mg g-1 respectively) and @ 20 mg L-1 sodium selenate supplementation in P. eryngii (8.56 mm/day, 77.0 g L-1 and 2.55 mg g-1 respectively). The highest Se content was observed at 20 mg L-1 of Se concentration in P. djamor (920.32 µg g-1dw), followed by P. florida (880.61 µg g-1dw), P. eryngii (850.27 µg g-1dw) and P. cornucopiae (792.72 µg g-1dw). SEM studies showed that 10 mg L-1 Se concentration was best for the growth of P. florida, P. eryngii, P. djamor and P. cornucopiae and the mycelial texture became coarser and fragile in response to increased concentration of Se. EDS analysis showed that the % weight and atomic % of Se were highest in 10 mg L-1 Se supplementation treatment in all the four Pleurotus spp. There was no significant difference in the biological efficiency (%) of P. florida (66.50 vs 64.57), P. cornucopiae (36.74 vs 33.11), P. eryngii (30.41 vs 27.16) and P. djamor (30.14 vs 29.16) cultivated on Se-enriched and non-enriched wheat straw respectively. The total soluble protein content, total phenolic content, flavonoid content and free radical scavenging activity (%) were found to be highest in Se enriched fruit bodies when compared to control samples in all the four Pleurotus spp. Elemental analysis of Se biofortified Pleurotus mushrooms using SEM-EDS showed signals characteristic for selenium on the surface of P. florida and P. djamor conforming that the selenium was incorporated into the cell wall of these mushroom fruiting bodies and were absent in P. eryngii and P. cornucopiae. The Se content was found to be 22.34 µg g-1dw in Se enriched wheat straw and 0.059 µg g-1dw in respective non-enriched wheat straw. Se content of Se-enriched fruit bodies were found to be higher as compared to non-enriched fruit bodies of P. florida (124.00 vs 2.48 µg g-1dw), P. eryngii (57.25 vs 0.49 µg g-1dw), P. cornucopiae (46.25 vs 0.161 µg g-1dw) and P. djamor (156.6 vs 4.33 µg g-1dw). FT-IR spectra of proteins from all the four Pleurotus spp. indicated an increase in flexibility, unfolding and hydrophilicity upon Se supplementation. Therefore, Pleurotus spp. can be used as an excellent nutraceutical with numerous health benefits.ThesisItem Open Access Use of bioinsecticides in compost for button mushroom cultivation(Punjab Agricultural University, Ludhiana, 2020) Simran; Sharma, ShivaniAgaricus bisporus is the most commonly cultivated edible variety in the world which possesses an ability to grow on agricultural wastes. Therefore, the present study has dealt with the use of pre botanical insecticides such as neem powder, vinegar, castor powder and two biological insecticides such as Bacillus thuringiensis and Beauveria bassiana to study its effect on mycelial growth of A. bisporus on PDA medium and wheat straw based compost. Maximum growth of 55mm and 29mm was recorded with neem powder on PDA and compost respectively. However, vinegar and Bacillus thuringiensis also showed significant mycelial run on PDA as well as compost. The natural insecticides were supplemented @0.5 to 4.5% (w/w) in ready compost and spawned @1.0% in compost. The supplementation of neem powder @1.5% showed significantly improved yield potential 43.40% and increased number of fruit bodies 37.55%. In case of castor powder (0.5 to 4.5% v/w), supplementation @ 4.5% (v/w) showed an increase of 23.22% in yield of button mushroom. An increase upto 20.23% and 46.42% in yield and fruit bodies was observed when vinegar was supplemented @2.5%. Bacillus thuringiensis supplementation @2.5% showed maximum yield potential 33.96%. Similarly, Beauveria bassiana was supplemented at a population of 3.4×107 cfu/ml @0.5- 4.5% (v/w) in compost which significantly improved the yield upto 14.44% and 12.62% when Beauveria bassiana was supplemented @1.5% and 0.5% at the time of spawning. Thus, it was concluded from our study that plant-derived products can be recommended to substitute for synthetic chemicals in the commercial production of edible mushrooms.ThesisItem Restricted Upscaling and comparison of beer production by Saccharomyces cerevisiae fermentation of sweet sorghum(Punjab Agricultural University, Ludhiana, 2020) Jasmine Kaur; KeshaniBeer is the third most popular beverage in the world. It is generally made from malted barley. Tropical countries need to import barley from temperate countries for brewing which makes it an expensive process. So, brewers are searching for alternate substrates for beer production to meet the increasing demand of beer and to reduce its cost. Sweet sorghum is one of the potential substrate for beer production. In the present research, fifteen varieties of sweet sorghum grains viz. SPV 2402, SPV 2530, SPV 2596, SPV 2597, SPV 2599, SPV 2600, SPV 2601, SPV 2603, SPV 2606, SPV 2610, SPV 2611, CSV 19 SS, CSV 24SS, CSH 22 SS and SSV 84 were evaluated for their potential as a substrate for beer production. Beer was produced using sweet sorghum and pilsner malt blend in the ratio of 40:60 respectively. Out of all the sweet sorghum varieties assessed, beer made from CSV 24SS using Saccharomyces cerevisiae GP4 had highest alcohol content of 5.8% (v/v) and the highest sensorial acceptance. The CSV 24SS variety was then used for upscaled production (20 litres) of beer using S. cerevisiae GP4 fermentation. The beer produced was pale gold in color and had a pH of 4.2 and titrable acidity of 0.36%. The beer was evaluated by a panel of judges for its sensorial profile and received a mean score of 31.5 out of 36 on the Hedonic scale. The beer was subjected for shelf life studies. Few bottles of beer were force carbonated and others were kept flat and stored at 4°C. The flat beer showed greater stability during storage as compared to carbonated beer.ThesisItem Open Access Seasonal emergence of pathogenic bacteria in drinking water(Punjab Agricultural University, Ludhiana, 2020) Pandey, Swati; Sahota, Param PalThe impact of seasonal variations on microbial diversity of drinking water poses an imminent threat to public health. As such the seasonal dynamics of drinking water quality characteristics were investigated during the year 2019-2020 in Punjab. A total of N=600 drinking water samples from various sources and utilities were analysed for bacteriological and physicochemical attributes; viz heterotrophic plate counts (HPC), total coliforms (TC), faecal coliform E. coli, emerging pathogens, pH, conductivity (EC), and TDS. A total of 66.3% (n=398) samples were tested positive for the presence of coliforms. The mean HPC and TCC were found to differ significantly during seasons (summer, monsoon, and winter) of the year (P < 0.05). There were considerable seasonal variations in prevalence of indicator E. coli (16.7-60%) as well as the emerging pathogens in contaminated drinking water samples. Statistical analysis of the dataset showed significant positive correlation trends between temperature and TCC (r = 0.785), E. coli (r = 0.690) and HPC (r = 0.678). The prevalence of emerging pathogens in contaminated samples and temperature showed a significant positive correlation for Enterococcus spp., Campylobacter spp., Vibrio spp. and inverse significant correlation for Aeromonas spp., Yersinia spp., Pseudomonas spp. and Listeria spp. The bacterial pathogens Klebsiella spp., Shigella spp., Staphylococcus spp., Salmonella spp. were detected from contaminated samples throughout the year in low percentages regardless of temperature influences. The concurrence of faecal indicator E. coli and emerging bacterial pathogens in contaminated samples was weak and variable. The bacterial isolates from drinking water tested for antibiotic sensitivity assay showed MAR index range of 0.12-0.87. The commercially available UV filter was explored as a remedial technology for bacteriologically contaminated water. Its effectiveness under applicable conditions debunks the common belief about the advanced disinfection potential. The presence of injured coliforms and bacterial pathogens showing marked phenotypic and antibiotic sensitivity changes post-treatment raises the alarming concern of consumer safety. Lastly collection of a large dataset at high frequency over a period of one calendar year has enabled the characterization of previously undocumented seasonal dynamics in the drinking water characteristics and highlights the need for reframed standard methods to enhance pathogenspecific assessment for better risk management.ThesisItem Open Access Ester Profile of indigenous Saccharomyces cerevisiae strains isolated from traditional alcoholic beverages(Punjab Agricultural University, Ludhiana, 2020) Harmanpreet Kaur; KeshaniRed wine was prepared from the red grapes var. Zinfandel by the action of twenty-three yeast cultures isolated from traditional alcoholic beverages from Lahaul & Spiti and Bada Bhangal region. The yeasts were identified on the basis of morphological studies with respect to colony characteristics, characteristics in broth culture, and cell shape as well as biochemical characteristics. Extracted juice was subjected to skin treatment for 16h to improve the color intensity of juice. The later was fermented by twenty-three different Saccharomyces cerevisiae strains under optimized conditions viz. 5% (v/v) inoculum size, 50 mg/100 ml DAHP was supplemented at 28 ˚C. The prepared wine was compared for their alcoholic content as well as their overall acceptance on the basis of appearance, mouthfeel and flavor by sensory evaluation. The ester analysis of prepared wine was done by GC-MS technique. Esters such as ethyl acetate, isoamyl acetate, phenyl ethyl acetate, ethyl caprylate and ethyl caproate as well as the other volatile compounds were reported out of which ethyl acetate and isoamyl acetate were predominant. Maximum alcohol content (11.03% v/v) and fermentation efficiency (86.16%) was noticed in the wine produced by strain code AB. It also produces enough esters viz. ethyl acetate (36.55 mg/l) and isoamyl acetate (940 mg/l) to provide suitable flavor to the wine which leads to score maximum points (7) on the “Hedonic-scale” and was considered best among all.ThesisItem Open Access Bioprocessing of Indian gooseberry for the production of lactic acid beverage and pickles(Punjab Agricultural University, Ludhiana, 2019) Kirandeep Kaur; Sahota, Param PalThe plant-based fermented beverages have received considerable interest for their rich nutrition, high content of soluble and insoluble fibers and for their fermentability by lactic acid bacteria (LAB). A set of ten allochthonous lactic acid bacterial strains phenotypically and genotypically characterized for their probiotic potential were used as starter culture for the optimization and production of Indian gooseberry-based functional beverage and lacto-pickle. The optimized bioprocess comprised of gooseberry juice blend [gooseberry juice 2: ginger extract 1: guava juice 2], dilution ratio (1:3), condiment concentration (0.7%), pasteurized at 82ºC for 10 seconds and 5.0% (v/v) (106 CFU/mL) active starter culture for fermentation at 37ºC for 24 hr. The microbiological and physicochemical parameters of stored beverage (90 days) were; 1ºBrix, Titrable acidity 0.30-0.89 %, pH 6.0 -2.6, Total Sugars 44.3-35.3 mg/100mL, Reducing sugars 5.4-10.7 mg/100mL and antioxidant activity (85-67%), Total polyphenolic content (42.5-32.6 mg GAE/100mL),Total flavanoids (14.5-28.7 mg/100mL) with LAB capable of maintaining cell counts of 7.39 log CFU/mL. The bioprocess optimized for the Indian gooseberry lacto pickle as 5.0% (v/v) (106 CFU/mL) active starter culture and 5% brine solution for fermentation at 37ºC for 24 hr. The microbiological and physicochemical parameters of stored lacto-pickle (90 days) were; LAB count-6.22-7.27 log CFU/mL, 1ºBrix, Titrable acidity 0.24-0.64 %, pH 7.0-2.8, Total Sugars 20.1-11.6 mg/100g, Reducing sugars 2.60-14.6 mg/100g, viz. nutraceuticals Total antioxidant activity 86.1- 90.2%, Total polyphenolic content 16.6-37.8 mg GAE/100g and Total flavanoids 4.007-55.7 mg/100g. The overall acceptability of beverage and pickle was 7±0.2 on nine point hedonic scale. The bio-interventions: Gooseberry beverage and pickle endowed with organic acids, metabolites, antioxidants, polyphenols and flavonoids showed antimicrobial activity against food borne pathogens Staphylococcus aureus MTCC3906, Listeria monocytogenes MTCC657, Klebsiella pneumoniae MTCC109, Escherichia coli MTCC443, Aeromonas hydrophila MTCC173 as well as arrested the initial phase of Molt-4 cancer cell lines, down regulating the expression of proto-oncogenes and up regulating the tumor suppressor gene exhibiting the antitumorigenic effect.ThesisItem Open Access Bioutilization of spent mushroom compost for remediation of heavy metal rich effluents(Punjab Agricultural University, Ludhiana, 2020) Avneet Kaur; Katyal, PriyaThe present study was conducted to use spent mushroom compost (SMC) as a low cost effective biosorbent combining the advantage of biosorption and bioaccumulation approach for remediation of heavy metal rich effluents. Effluent samples were collected from two Common Effluent Treatment Plants (CETPs) located at Ludhiana (L-CETP) and Jalandhar (JCETP). Samples were analysed for various physico-chemical parameters, whereby DO (43.92 mg/l) and BOD values (13.2 mg/l) were low in L-CETP whereas COD value (320 mg/l) was higher than permissible limit (250 mg/l) indicating that sample was polluted with organic contaminants. In J-CETP, DO (92.0 mg/l) and BOD (71.5 mg/l) values were high while COD value was low (240 mg/l) indicating that sample has indigenous microflora. Total solids (TS) and Total dissolved solids (TDS) were found to be high in J-CETP. Metal profile of effluent samples was estimated using Inductively Coupled Argon Plasma-Atomic Emission Spectroscopy (ICAP-AES). In Ludhiana CETP, Cr was dominant heavy metal with a concentration of 226.62 ppm, followed by Ni 28.12 ppm and Pb 0.77 ppm. In Jalandhar CETP, the concentration of Cr was 1.83 ppm, followed by Pb 0.39 ppm and Ni 0.10 ppm. Heavy metal tolerance was determined by growing cultures of Pleurotus florida, Agaricus bisporus, Volvariella volvacea and Calocybe indica in Potato Dextrose Agar (PDA) plates amended with heavy metals. Cultures of white rot fungi were most sensitive to Hg and least sensitive to Cr. Among the four selected strains, Volvariella volvacea was found to be highly resistant to Cr with 22.5% inhibition of growth at 250 ppm. SMC of selected white rot fungi was dried, grinded and analysed for various physico-chemical parameters whereby low values of cellulose, hemicellulose and lignin indicated their degradation during the process of composting and mushroom growth. Batch experiments were carried out in erlenmeyer flasks to study the effect of various parameters like pH, temperature and initial metal concentration on biosorption. Maximum removal of Cr was obtained at Ci-50 mg/l, pH-4-6 and temperature-25°C. Experimental results were well fitted to Langmuir isotherm than to Freundlich isotherm. These standardized conditions were used to study metal uptake from effluent samples by SMC packed in a bioreactor. Maximum removal of Cr was achieved with contact time of 2 hrs with A. bisporus SMC, 3hrs for P. florida SMC and 1.5 hrs for C. indica SMC. FT-IR spectrum analysis of SMC showed shift in peaks at 2918 cm-1 to 2980 cm-1 due to stretching of CH and CH2 group, 1744.7 cm-1 to 1739.6 cm-1 because of masking of protonated carboxyl group and peak at 1500 cm-1 showed the presence of lignin group in treated and untreated spent compost.ThesisItem Open Access Development of plant growth promoting microbial consortium for alleviating abiotic stress in soybean(Punjab Agricultural University, Ludhiana, 2020) Rajveer Kaur; Sharma, PoonamThe present study was aimed to screen consortium of authenticated Bradyrhizobium sp. (LSBR-3) and rhizobacteria (LSE-1, LSE-2, LSE-3 and LSE-4) possessing plant growth promoting traits to mitigate concerted abiotic stresses (salinity, drought and temperature) in soybean. Single and consortium inoculants were screened for abiotic stress tolerance viz. salinity (0-25% NaCl), drought (Polyethylene glycol (PEG 0-30%) and temperature (28, 37°& 45°C) in terms of growth by spectrophotometer and agar plate assay. Maximum growth of all treatments was observed at 5% each of NaCl and PEG (6000). Single and consortium inoculants exhibited luxuriant growth at 28° and 37°C.Whereas, sharp decline was noticed at 45°C. In concerted stress, dual inoculants revealed higher growth over single inoculant and maximum growth was observed in LSBR-3 with LSE-3 (OD600nm 0.737 and 0.858) and LSE- 1 (OD600nm 0.676 and 0.688) at 45°C under normal vs stressed conditions respectively. Further, consortium of LSBR-3 with LSE-3 and LSE-1 also possessed multiple plant growth promoting traits viz. indole acetic acid (12.62, 6.65 and 6.96 μgml-1) and (13.95, 6.34 and 7.78 μgml-1), phosphate solubilization (10.09, 6.91 and 7.83 mg/100ml) and (10.33, 7.01 and 10.36 mg/100ml); 1-aminocyclopropane-1-carboxylate deaminase (OD600nm 0.3062, 0.6449 and 0.6081) and (OD600nm 0.3197, 0.6271and0.5871) respectively under normal, induced salt and drought stress at 45°C. At 45°C highest biofilm formation and total soluble sugars was noticed with NaCl and PEG treated dual treatments upto 24 h of incubation. Induced salinity and drought enhanced proline content of dual inoculants over control treatment. Greenhouse experiment conducted for normal and induced drought stress conditions with different abiotic stress tolerant consortium inoculants for growth, symbiosis, nutrient acquisition, antioxidants and yield in soybean revealed improvement in all parameters over un-inoculated control. Higher amount of antioxidant enzymes viz .superoxide dismutase (26.21μmol O2 mg-1 protein-1), catalase (12.01H2O2 mg-1 protein), peroxidase (0.315min-1mg-1) and osmolyteproline (7.81μgml-1) were noticed with LSBR-3+LSE-3 in induced drought stress over control treatment. Under stressed conditions, 1.5 and 2.9 folds enhancement in grain yield was observed with consortium of Bradyrhizobium sp. LSBR-3 with rhizobacteria LSE- 3, as compared to rhizobacterium single inoculants LSE-3 and control respectively. Present study highlights the role of consortium treatment of Bradyrhizobium sp. and rhizobacteria in ameliorating the oxidative damage induced by drought stress in soybean.
