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Open Access
Characterization of psychrotolerant rhizobacteria for Zn biofortification in lentil (Lens culinaris L. Medikus)
(Punjab Agricultural University, Ludhiana, 2018) Jaskiran Kaur; Khanna, Veena
Seventy psychrotolerant rhizobacterial isolates were evaluated for their PGP traits at 10°, 20° and 30°C. Highest IAA production was recorded with isolates PRh-9, PRh-14, PRh- 15, PRh-60, PRh-61 and that was confirmed by HPLC (8.98, 4.91, 5.66, 2.73 and 1.62 μg/ml respectively). Isolate PRh-14 recorded maximum Zn-solubilization potential of 354, 304.8 and 284.5 ppm at 10°, 20° and 30°C. It also recorded the maximum P-solubilization efficiency, gibberellins, EPS production and biofilm formation at 10°, 20° and 30°C. Eleven isolates amplified acdS gene which confirmed the presence of ACCD activity. Maximum naringin and quercetin production was shown by isolate PRh-61 and PRh-9, further confirmed by TLC studies. Highest antagonistic activity was recorded by PRh-9 against Rhizoctonia bataticola and Fusarium oxysporum (33.3% and 48.1% respectively) and was also established by SEM analysis. Potent siderophore, salicyclic acid and HCN producers were PRh-9, PRh- 61 and PRh-60. The SEM analysis of root surface revealed that coinoculation with Rhiobium and PRh-14 helped in maintaining the root integrity owing to biofilm formation on the root surface. Under field conditions coinoculation of Rhizobium with PRh-15, PRh-14, PRh-9, PRh-61, PRh-60 and PRh-30 further increased by grain yield by 1.7, 4.4, 3.7, 2.5, 2.09, 1.5% respectively, over Rhizobium alone. A significant increase in the grain and straw zinc content was recorded with the treatment R+PRh-14 (105.9 and 141.0 ppm respectively). The DGGE analysis of soil DNA depicted a composite banding pattern that reflected a high microbial diversity. The promising rhizobacterial isolates were identified as PRh-9 (Pseudomonas flourescens), PRh-14 (Aeromonas hydrophila), PRh-60 (Lysinibacillus fusiformis) and PRh-61 (Pseudomonas korensis) based on 16S rDNA sequencing.
Open Access
Impact of long term organic and inorganic fertilization on microbial community and screening for cellulose degrading bacteria
(Punjab Agricultural University, Ludhiana, 2018) Bhagat, Poonam
Impact of long term incorporation of rice straw and inorganic nitrogen was studied on microbial communities, soil enzymatic activities and on soil parameters at different time intervals (0, 45, 90 and 120 DAS) for two consecutive years. The different doses of rice straw (0, 5, 7.5 and 10 t/ha) along with different doses of inorganic nitrogen (0, 90, 120 and 150 kg/ha) were incorporated in the wheat field. The first year results revealed maximum total bacterial count (8.31 log 10 cfu/g of soil), fungal count (4.89 log 10 cfu/g of soil), celllulse degrading bacteria count (7.11 log 10 cfu/g of soil) with 7.5 t/ha RS + 120 kg N/ha while actinomycetes count (7.14 log 10 cfu/g of soil), diazotrophic count (6.07 log 10 cfu/g of soil) with 10 t/ha RS alone at 45 DAS. The maximum dehydrogenase activity (23.96 µg TPF/g soil/hr), alkaline phosphatase activity (14.77 µg pNP/g soil/hr) and urease activity (330.0 µg urea/hr/g soil) observed at 45 DAS with 7.5 t/ha RS plus 120 kg N/ha. The microbial population and enzymatic activities found higher in second year over first year. The soil pH was found to be altered from 7.82 to 6.99 and EC from 0.219 to 0.201 dSm-1 with treatment 10 t/ha RS +120 kg N/ha whereas OC from 0.26% to 0.49% with 10 t/ha RS + 150 kg N/ha in two years at 120 DAS. The maximum soil available nutrients viz. nitrogen (136.7 kg/ha), phosphorous (29.94 kg/ha) observed with treatment10 t/ha RS + 150 kg N/ha, potassium (119.78 kg/ha) with treatment 7.5 t/ha RS + 120 kg N/ha at 45 DAS in the first year. The maximum straw and grain nitrogen (0.46%) and (1.37%), phosphorus (0.15%) and (0.38%), potassium (1.72%) and (0.53%) contents respectively in first year found with 10 t/ha RS + 150 kg N/ha. The maximum straw and grain yield (9.65 t/ha) and (6.28 t/ha) respectively observed in first year with 7.5 t/ha RS + 120 kg N/ha. The available nutrients and yield found higher in second year than first year. The cellulose degrading bacteria were isolated and halozone formation confirmed their cellulolytic activity. Seventeen isolates were characterized biochemically and most were positive for oxidase and catalase while negative for H2S test, indole test and MR-VP test and few were positive for citrate utilization, gelatin solubilization and starch hydrolysis. The functional characterization of isolates showed highest cellulolytic activity (1.44µg/ml), IAA production (34.85µg/ml), ammonia excretion (2.048µg/ml) and phosphate solubility (28.56 µg/ml) was exhibited by isolate SKPB3. Isolates were analysed phylogenetically by 16S rDNA sequencing. Isolates clustered in phylogentic trees indicated high similarity and the abundance of particular cellulolytic strains. Identification of representative cultures using parial sequencing of 16S rDNA revealed presence of Acinetobacter sp, Pseudomonas sp, Stenotrophomonas sp, Bacillus sp.
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Synergistic effect of Mesorhizobium and non rhizobial endophytes on plant growth promotion in chickpea (Cicer arietinum L.)
(Punjab Agricultural University, Ludhiana, 2018) Chhabra, Deepika; Sharma, Poonam
The present investigation was carried out to exploit non rhizobial endophytic bacteria from root and nodules of chickpea. Of 263 non rhizobial endophytic bacteria 74.4 %, 87.4%, 12.6%, 36.6%, 37.4% and 55.7% were found to be positive for catalase, oxidase, citrate utilization, nitrate reduction, methyl red and Voges Proskauer‟s, respectively. 71 were found to be good for phosphate solubilization and IAA production. Significantly high P solubilization was observed in root isolate RBR20 (20.60 mg100ml-1) whereas, among nodule‟s isolates with RBN17 (21.0 mg100ml-1). In the presence of tryptophan, the isolate LCRE 9 produced the maximum amount of IAA (39.60 μgml-1) whereas in the absence of tryptophan the isolate RBR164 produced the maximum amount of IAA (19.93 μgml-1). RBN16 isolate showed highest growth in DF medium with ACC (OD 1.04) followed by RBR164 (OD 0.9985). High amount of GA production was observed in RBR19, RBR127, RBR136, RBR164 and LCNE6 (112.15 μgml-1). Highest amount of EPS production was observed LCRE8 (105.01 μgml-1). Out of 75 isolates 38.7%, 82.7%, 9.3% and 21.3% isolates were able to produce cellulase, protease, HCN and fluorescent pigment, respectively. On the basis of overall PGP traits and compatibility studies 3 potential non rhizobial endophytic bacteria (RBR164, LCRE9 and RBN17) were selected for colonization study in different combinations with ampicillin (6μgml-1) an antibiotic marker. Colonization was maximum in RB1+LGR33+ RBN164 treatment (1.16 and 1.60) at 15th and 50th respectively. Triple inoculant treatment LGR33+RB1+RBR164 resulted into maximum increase in plant height, dry weight of shoot and total bacterial count at 90 DAS as compared to dual, single and uninoculated control under field conditions. Maximum increase in dry weight of root, root shoot ratio, nodule dry weight and leghaemoglobin was observed with same treatment at 60 and 90 DAS. Similar trend was observed for total NPK content of shoot and seed. Grain yield was increased with triple inoculants treatment LGR33+RB1+RBR164 by 8.9% and 3.6% over uninoculated control and recommended consortium of chickpea, respectively. It seems from the foregoing study that consortium of Mesorhizobium with multifarious plant growth promoting endophytic bacteria can be developed and used as biofertilizer for chickpea.
Open Access
Development of microbial consortia for pretreatment of paddy straw and its utilization for ethanol production
(Punjab Agricultural University, Ludhiana, 2017) Lota, Pardeep; Kocher, Gurvinder Singh
The present study was conducted with the objective of standardization of a fungal consortium from selected ligninolytic cultures for biological pretreatment of paddy straw and its subsequent conversion to bioethnaol. The proximate analysis of paddy straw revealed that 30 mesh straw had highest proportion of cellulose (39.5%) besides, 23.3% hemicellulose and 11.5% lignin. Among different chemical pretreatment methods, acid (1.0% H2SO4) –autoclave (15 psi for 90 min) followed by alkali (2.0% NaOH) pretreatment resulted in 82.61% and 59.23% decrease in lignin and hemicellulose, respectively and 36.60% increase in relative proportion of cellulose. Four fungal strains (viz., LS1, LS2, LCu1 and LCu2) from a total of fifty nine strains isolated from diverse natural habitats, tested positive for the production of laccase (Lacc), lignin peroxidase (LiP) and manganese peroxidase (MnP). Out of three consortia developed on the basis of ligninolytic activities, consortium III comprising Pleurotus ostreatus and Phanerochaete chrysosporium, resulted in Lacc, LiP and MnP activities of 2.40, 37.92 and 62.50 nkat/gds, respectively, significantly higher than the activities of individual fungal strains. Biological pretreatment of paddy straw under Response surface methodology optimized conditions (moisture, 121.0 %; temperature, 31.3°C and log spore count, 8.0 spores/ml) resulted in cellulose, 43.0%; hemicellulose, 12.7%; lignin, 7.0%; total sugars,13.65 mg/gds and reducing sugars, 4.00 mg/gds. The pretreatment of paddy straw with concentrated (10x) ligninolytic enzyme under shake flask conditions using optimized physico-chemical parameters viz. paddy straw concentration, 2.5 g; enzyme volume, 8.0 ml; Ca2+ ions (10 mM) and incubation temperature, 45°C in 72 h of enzymatic action resulted in 80.87% and 47.64% decrease in lignin and hemicellulose, respectively and 29.96% increase in relative proportion of cellulose. For saccharification studies, from a total of forty seven fungal strains isolated from diverse natural sources, two strains (Aspergillus sp. CTS1 and Aspergillus sp. CTS2) tested positive for thermophilic and cellulolytic nature. The two strains were used for the development of fungal consortium which revealed filter paper, carboxymethyl cellulase and cellobiase activities of 10.2, 30.0 and 7.9 U/gds, respectively, significantly higher than the activities of individual fungal strains. Under the RSM optimized conditions [temperature, 55°C; pH,4.8 and substrate concentration, 5% (w/v)] and 30FPU/g dry substrate Arrowzyme yielded 476.0 mg/gds reducing sugars from biological pretreated paddy straw which were further increased to 492.0 mg/gds by the addition of Tween 20 at 0.2% (v/v). Enzymatic saccharification of biological pretreated straw by in-house concentrated cellulase (CC1) from thermophilic fungal consortium resulted in 229.0 mg/gds reducing sugars. The saccharification of chemical pretreated straw with Arrowzyme and CC1 resulted in 566.0 mg/gds and 250.0 mg/gds reducing sugar, respectively. Optimization of fermentation under separate hydrolysis and fermentation (SHF) conditions using RSM [S.cerevisiae inoculum size 10.0 % (v/v) and DAHP concentration, 0.2% (w/v)] resulted in 0.172 g/g and 0.193g/g ethanol from biological and chemical pretreated hydrolysate, respectively. Similarly, the CC1 saccharified hydrolysate resulted in an ethanol content of 0.076 g/g (biological pretreated) and 0.085 g/g (chemical pretreated). Fermentation of non-enzymatically saccharified biological and chemical hydrolysate by S.cerevisiae produced 0.013 g/g and 0.041 g/g ethanol, respectively, whereas sequential fermentation of chemical hydrolysate with S.cerevisiae and Pachysolen tannophilus produced 0.047g/g ethanol. The combined ethanol production from enzymatically and non-enzymatically saccharified biological and chemical pretreated paddy straw hydrolyaste was observed to be 0.185 g/g and 0.240 g/g, respectively.
Open Access
Bioprospects of microalgal isolates from water logged area of Punjab for biogas production
(Punjab Agricultural University, Ludhiana, 2017) Dar, Rouf Ahmad; Phutela, Urmila Gupta
The present study was aimed at isolation, identification, screening and characterization of microalgae from the waterlogged area of south-west Punjab, India. Optimization of cultural conditions of potential microalgal strains, their biogas production potential and anaerobic co-digestion studies were also conducted. Nineteen microalgal cultures (BGLR1-BGLR19) were isolated and were screened using different culture media for their growth kinetics. Isolate BGLR6 followed by BGLR5 showed the highest growth and biomass production in Algae culture medium and BG-11 medium respectively. These isolates upon molecular identification were found to be Asterarcys quadricellulare BGLR5 (MF661929) and Spirulina subsalsa BGLR6 (MF191711). The cultivation conditions for the enhanced production of biomass and other functional components (chlorophyll, carbohydrate, lipid and protein) of BGLR5 and BGLR6 were first screened by Plackett–Burman design and then the significant factors were optimized by Central Composite design (CCD). The optimal cultural conditions for BGLR5 and BGLR6 as per the model were pH of 9.92 & 11.5; temperature of 21.84 & 20°C; light intensity of 80.99 & 81 μmol m-2 s-1; growth period of 25 days (both); 15.00 mM NH4Cl/ CaCl2; 12.28 & 5.00 mM NaNO3 and 7.09 & 2.00 mM K2HPO4 respectively. Under these conditions, the response variables generated a desirability index of 84.10 and 94.91% for BGLR5 and BGLR6 respectively. A 0.42 and 1.60-fold increase in dry cell biomass yield was achieved in BGLR5 and BGLR6, compared to the basal condition (0.8858 and 1.0890 g biomass L-1) respectively. The biogas potential of the microalgal biomass under controlled temperature (35 ± 2°C) conditions revealed that in case of BGLR5, the highest biogas yield (51.712 Lkg-1algal biomass) was obtained in the hydrothermally (100°C 30min) pretreated biomass whilst in BGLR6, the highest biogas yield (42.73 Lkg-1algal biomass) was obtained in enzymatically (10% 24hr) pretreated biomass. Further, co-digestion of 50% paddy straw and 50% BGLR5 biomass (replacement of paddy straw by equal amount of microalgal biomass) produced 168.32 L biogas kg-1 feedstock with the ultimate biogas yield potential of 361.81 mLg-1 VS, reduced lag phase (λ:2.81 d) and increased rate of biogas production (Rm: 8.19 mLg-1d -1) which was higher not only in comparison to the anaerobic digestion of paddy straw and algal biomass individually but also to that of co-digestion of BGLR6 biomass with paddy straw. Likewise, 50 and 100% supplementation of BGLR5 biomass led to enhanced biogas production. Utilization of algal biomass cultivated in the open pond for co-digestion study with paddy straw in field scale digesters resulted in an increase of 17.27% biogas compared to control. The enhancement of methane content from 46.4% (control) to 66.5% (digester containing algal biomass) was also achieved. Hence, the present study signifies that the Asterarcys quadricellulare BGLR5 and Spirulina subsalsa BGLR6 biomass could be utilized as a cofeedstock with paddy straw for biogas production.
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Development of diagnostic protocol for assessment of microbiological quality of fresh vegetables
(Punjab Agricultural University, Ludhiana, 2017) Sood, Bhavish; Param Pal
The epidemiological surveillance for microbiological quality of seven fresh vegetables (N=725) sourced from Village fields along Buddha Nallah, Supermarkets and Street vendors was conducted and concluded the high incidence of 11 important food borne pathogens with highest positive percentage of K. pneumonia (69%) in village fields, Y. enterocolitica in supermarkets (63.54%) and vendor samples (78.98%). The highest mean total plate count was observed in Spinach (>6 log cfu g-1) collected from village fields and vendors. Most probable number (MPN) analysis reflected the deteriorated quality of irrigation water with index range upto >1100/100ml (above desirable limit 1000/100ml). Pathogenic bacterial load was dominated in internal part of tomato (3.98 log cfu g-1) and on the surface region of cucumber (4.12 log cfu g-1). Phenotying of isolates of 11 pathogens was carried out to study the diversification of pathogens on basis of biochemical, virulence and antibiogram profiling. Clustering analysis of isolates (Unweighted Pair Group Method with Arithmetic Mean) revealed their significant diversity among themselves which provides insight of the indigenous pathogens inhabiting the vegetable commodity. Antibiotic profiling using 25 antibiotics belonging to 10 classes was established for all the isolates and complete tolerance to five antibiotics [Clindamycin (2mcg), Tetracycline (30mcg), Cloxacillin (5mcg), Metronidazole (5mcg) and Imipenem (10mcg)] was observed in Salmonella enterica. The bacterial isolates showed high MAR index range of 0.08-0.84. The species-specific virulence markers (enterotoxin, effector proteins encoding genes) used in PCR assay to detect the gene distribution pattern among the pathogens and their presence prove the potentiality of causing illness. A Bacteriological Food testing Kit (BFTK) (12h detection time) was devised based on the biochemical characteristics of indicator (Escherichia coli, Staphylococcus aureus, Shigella spp., Salmonella spp., Listeria monocytogenes) and emerging (Yersinia enterocolitica, Aeromonas hydrophila, Campylobacter jejuni, Bacillus cereus and Klebsiella pneumoniae) and validated with food samples (N=1000) against BIS Standard Methods at 5% level of significance. Another rapid yet inexpensive detection technique based on the Multiplex PCR was devised in which hblD gene (430bp fragment) of Bacillus cereus; ystA gene (79bp fragment) of Yersinia enterocolitica, invA gene (280bp fragment) of Salmonella enterica and iap gene in Listeria monocytogenes (225bp fragment) were targeted. Furthermore, a combined culturing (BFTK) and molecular tool (multiplex PCR) was found promising to detect pathogens in time slot of 15 h using specific markers with detection limit of 106 cells per target bacteria. A simulation study on concentration method stepped with vacuum filtration, low speed centrifugation and high speed centrifugation has shown high rate of recovery of all the 11 pathogens with sensitivity limit not less than 104 cfu/10g by reducing the sample to final volume of 2ml. A useful intervention was made for decontamination of vegetables by effectively using 1000ppm of citric acid in wash water.
Open Access
STUDY ON ERGOSTEROL CONTENT AND PROTEIN PROFILE OF MEDICINAL MUSHROOM, Ganoderma lucidum
(PAU Ludhiana, 2013) Anna Goyal; H. S., SODHI
Ganoderma lucidum, a specie belonging to the class basidiomycetes, family polyporaceae of the order aphyllophorales has been widely used as a source of potent nutracuetical products. Present study was planned to identify and characterize the role of proteins and ergosterol in the developmental process of Ganoderma lucidum. Four strains of Ganoderma lucidum (GL I - IV) showed a gradual increase in biomass to give 25.52g to 31.72g of biomass after three weeks of growth in mushroom complete medium broth with maximum in strain GL-III. Ganoderma lucidum strains were grown on wheat straw supplemented with 5% wheat bran with maximum biological efficiency for GL-I strain (31.27%) followed by GL-II (26.76%) and number of fruit bodies were 927 and 693 each weighing 33.7g and 38.6g, respectively. Ganoderma lucidum strain GL-I showed maximum ergosterol content (4601μg/g) whereas strain GL-III showed only 32μg/g. From spawn run, maximum ergosterol was obtained from GL-IV strain followed by strain GL-II while at pin head formation and fruit body formation ergosterol content was better for GL-II. Ergosterol content of fruit body of GL-1 was observed maximum (7009μg/g). Overall observation indicated that the ergosterol content increased with each stage of cultivation process i.e. from spawn run to pinhead and finally to fruit body formation. The intracellular and extracellular enzymatic studies have indicated enhanced activity during spawn run on solid substrate in comparison to that grown in the broth. The esterase and peroxidase activity significantly increased during the pinning of the cultures thus, indicating a positive role of these enzymes in fructification process. The FTIR analysis of proteins made during different stages of cultivation namely spawn run, pin head formation and fruiting indicated that the fruiting strains (GL-I and GL-II) have an ordered protein structure with hydrophobic amino acids. In case of GL-IV, unordered structure was obtained that could be related to the role of hydrophobin proteins in mushroom fructification process. Another observation on GL-IV indicated the presence of basic amino acids and aromatic amino acids with very low amount of acidic amino acids like aspartic acid and glutamic acid. The observation recorded during present study indicated a positive role of hydrophobic amino acids and hydrophobin proteins in mushroom fructification process. Ganoderma lucidum strain GL-II was also grown on selenium fortified mushroom minimal medium at different concentrations of 5ppm to 25ppm of sodium selenate. Scanning electron micrographs exhibited gradual decrease in hyphal diameter, spore number and spore diameter with increase in selenium concentration and the spore structures were altered. A significant decrease in spore diameter is observed in concentration of 20ppm and 25ppm (5.60 and 1.26 μm, respectively) as compared to control (10.04 μm). The SEM-EDS studies showed no selenium traces on the hyphal surface, however, on the contrary, SEM-EDS studies of crushate samples revealed selenium traces indicating selenium absorption as the cytosolic moieties as selenoproteins. Atomic absorption spectroscopy indicated an increasing trend in the uptake by the hyphal biomass as the concentration of sodium selenate increased with maximum absorption at concentrations of 15 ppm and 25 ppm (9.9%). It was concluded that fortification till 15 ppm can be used as stress was not that prominent and culture could grow rapidly without significant alteration in structure and morphology to enhance its biomedicinal properties. Present study has indicated that during the mushroom development process, ergosterol content increases with a positive role of proteins like peroxidases and hydrophobins at each stage of morphogenesis.
Open Access
Elucidation of phorate metabolism by bacterial isolates from agricultural soils for bioremediation
(PAU, 2013) Jariyal, Monu; Gupta, V. K.
Fifteen phorate metabolizing bacterial species isolated from sugarcane field soils were identified using 16SrDNA sequence homology. Based upon relative phorate degradation, Brevibacterium frigoritolerans, Bacillus aerophilus and Pseudomonas fulva were found to cause more than 98 per cent phorate reduction. These bacterial species could grow over a wide range of pH (4.0-11.0) and temperature (25-37°C), but optimally at pH of 6.0-6.5 and 37°C, in shaking cultures. Whereas, B. frigoritolerans was salt sensitive, P. fulva & B. aerophilus grew optimally in 3.0 and 4.0 per cent NaCl, respectively. All the three bacterial species grew optimally in the presence of glucose and peptone (1.0 % each). Only B. aerophilus carried a plasmid of around 4 kbp, but curing of this plasmid did not affect the phorate degradation establishing that phorate degradation genes are borne on chromosomal DNA. Induction by phorate resulted in only nondifferentiating protein profiles in all the three bacterial species establishing that phorate degradation is a constitutive character. In soils amended with upto 300 mg kg -1 phorate, these bacterial species within 42 days actively metabolized phorate by between 89.81 and 95.62 per cent with maximum capacity shown by P. fulva. This phorate degradation was further improved to 98.31 per cent, by mixed cultures of all the three bacterial species which constituted most effective bioremediation consortia for significantly relieving soils from phorate residues. The investigations in this study has isolated three bacterial species and established their potential for active bioremediation of phorate both in liquid cultures and agricultural soils.
Open Access
Characterization of cellulase producing thermophilic fungi for saccharification of sweet sorghum (Sorghum bicolor L. Moench) bagasse in order to produce ethanol
(PAU, 2014) Reetika; Kochar, G.S.
Diminishing fossil fuel reserves and increase in greenhouse gas emissions resulting in environment pollution problems have led to a search for alternate sources of fuel. The availability of agricultural lignocellulosic biomass such as bagasse, straw, dry leaves etc. in huge quantities render such a biomass an ideal substrate for bioethanol production. Sweet sorghum, because of its inherent characteristics, such as salinity and lodging tolerance, ability to grow under water and nutrient deficient soils, rapid growth, high sugar accumulation and high biomass production makes it a potential candidate for bioethanol production. The present study was envisaged with the major objective to isolate, thermophilic cellulolytic fungi capable of producing cellulases with improved functional characteristics which could be eventually used for effective hydrolysis of sweet sorghum bagasse (SSB) for bioconversion of sugars produced during hydrolysis to bioethanol. Out of the ten isolates screened for cellulase production through solid state fermentation, two isolates WAT-1 and RWY showed relatively higher filter paper cellulase and β-glucosidase activity at temperatures ranging from 45-55 °C. Therefore, WAT-1 and RWY were identified on the basis of microscopic, macroscopic and molecular characteristics as strains of Aspergillus fumigatus and A. terreus, respectively. Enzyme characterization studies for endoglucanase produced by A. fumigatus and A. terreus revealed that the optimum temperature and pH were 60 ºC and 4.0 and 50 ºC and 5.0, respectively. The kinetic studies of the cellulase produced by A. fumigatus revealed Km value of 19.46 mg/mL and vmax with 74.86 mg/mL/min. Whereas, for cellulase produced by A. terreus Km value was 16.09 mg/mL and vmax was 64.60 mg/mL/min. Alkali pretreatment was found to be more effective than the dilute acid pretreatment or biological treatment. Thus, enzymatic hydrolysis of the alkali pretreated SSB on optimized parameters using response surface methodology (4% sodium hydroxide, 140 ºC, 15 min and 800 micron particle size) led to production of 61.29, 16.28 and 3.33 g/L glucose, xylose and arabinose, respectively. Hydrolysis of alkali-treated SSB using optimized parameters for cellulase obtained from A. terreus (30 FPU/gds crude enzyme, 20 % substrate loading, 50 ºC for 33 h) resulted in 75 g/L glucose, 10 g/L xylose and 3 g/L arabinose. Hydrolysis of alkali-treated SSB with crude concentrated cellulase from A. fumigatus using optimized parameters (30 FPU/gds crude enzyme, 20% substrate loading, 55 ºC for 24 h) yielded 85 g/L glucose, 13 g/L xylose and 4 g/L arabinose. Saccharification using cellulases in presence of 0.2% Tween 20 increased glucose production by 15-20%. Simultaneous saccharification and fermentation at 35 ºC, 20% substrate loading, 10% inoculum concentration for 24 h resulted in 19 g/L, 15 g/L and 23 g/L ethanol, respectively using cellulases from A. fumigatus and A. terreus and commercial enzymes and Pichia kudriavzevii for fermentation. Separate hydrolysis and fermentation (SHF) using statistically optimized parameters (35 ºC, 10% inoculum concentration for 16 h) resulted in 40 g/L , 49 g/Land 51 g/L ethanol while using sugars produced by crude cellulases obtained from A. terreus, A. fumigatus and commercial enzymes. The present study thus revealed that sweet sorghum bagasse (SSB) could be ideally used as a substrate for bioethanol production using the optimized pretreatment, hydrolysis and fermentation parameters.

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