An Efficient Method of Pennisetum x Advena ‘Rubrum’ Seedlings Production Using Temporary Immersion Bioreactor and Agar Cultures

The aim of this study was to propose an efficient method of Pennisetum x advena ‘Rubrum’ micropropagation. Agar cultures with MS medium supplemented with BAP in various concentrations (0.5 mg/L-2 mg/L) and a temporary immersion bioreactor system (TIS) with liquid medium MS with an addition of 1 mg/L BAP were used. For rooting ½ MS medium with different auxin combinations (IBA, NAA) and activated charcoal was utilized. The most efficient method turned out to be TIS which produced 36.9 new plants in four weeks. The seedlings were slender in shape, bright green in colour with no signs of hyperhydricity. The most suitable agar medium produced 19.5 new plants in an eight week period. Rooting should be carried on ½ MS supplemented with 0.5 mg/L IBA and 0.5 mg/L NAA with an 84% rooting rate. The addition of activated charcoal inhibited rooting.

Characterization and pathogenicity of a Pythium aphanidermatum isolate causing ‘damping off’ in pepper seedlings

<p>The production of the pepper seedling (<em>Capsicum annuum</em>) is affected by the fungal complex that causes the ‘damping-off’, in which some species of the oomycete <em>Pythium</em> spp., stand out. The objective of the present study was to identify the causal agent of the death of pepper plants and evaluate its pathogenicity in pepper seeds and seedlings. A fast and aggressive growing oomycete was isolate from pepper plants, morphologically identified as P. aphanidermatum based on its sexual and asexual reproduction structures and, by molecular techniques. This isolate had a high degree of<em> in vitro</em> pathogenicity in pre-emergence and post-emergence in chile, showing 100% mortality. In addition, it presented a high rate of mycelial growth in different culture media (V8-Agar, Corn Agar, Corn Potato Agar, Potato Dextrose Agar, Czapek &amp; Oat Agar), being in V8-Agar medium the only medium where it developed reproduction structures sexual and asexual. The isolation presented a mycelial growth rate of 58.3 ± 0.3 mm / day at 26 ± 2 °C in PDA medium. Due to its rapid growth and its high degree of pathogenicity <em>in vitro</em>, it is an unusual and aggressive isolate for pepper seedlings.</p>

Screening, Isolation and Biochemical Characterization of Laccase Producing Bacteria from Contaminated Sites

Screening and isolation of Laccase producing bacteria from Guntur District soil was carried out to assess the diversity of Lignocellulose degrading bacteria. Isolation and identification of environmental friendly bacteria for lignin degradation becomes an essential one, because all the researchers are mainly concentrating on fungal strains. However, bacteria seem to play a leading role in decomposing lignin. For isolation of Laccase producing bacteria nutrient agar medium containing guaiacol was used. Total nine bacterial strains were isolated from soil samples collected from different regions of Guntur district. Preliminary screening of bacterial strains was carried out on guaiacol containing nutrient agar medium for laccase production. Formation of green colour using ABTS (2,2'- azinobis(3-ethylbenzthiazoline-6-sulfonate) confirms the capability of laccase production by the bacterial strains. Nine bacterial strains showed positive results. High laccase producing bacterial isolates were examined for morphological and biochemical characteristics according to Bergey’s Manual of Systematic Bacteriology. The predominant isolates were identified as Bacillus and Enterobacter species.

Evaluation of Lactobacillus spp. Based on Phenotypical Profile as Direct-Fed Microbial Candidate for Poultry Nutrition

The present study was conducted to isolate, identify and characterize a lactic acid bacteria strain from turkey ileum content (46-day-old). The new strain was phenotypical confirmed as Lactobacillus acidophilus (L. acidophilus) and conserved under the code IBNA 09. Bacterial profile of L. acidophilus was compared with other strains known as L. paracasei CCM 1837 and L. plantarum ATCC 8014, based on cultural, morphological, biochemical and enzymatic activity (amylase and cellulase). The strains appear as Gram positive bacilli, thin, non-spore-forming, isolated, diplo form, in short chains or in small irregular piles on Man Rogosa and Sharp (MRS) broth and agar medium. The identification and biochemical traits were performed by catalase assay, API 50 CHL V 5.1 soft (L. acidophilus biotype 2, 99.9% ID; good identification to the genus L. paracasei spp. paracasei 1 or 3, 48-51% ID; L. plantarum 1, 99.9% ID) and ABIS online (L. acidophilus ~ 88%; L. paracasei spp. paracasei, ~ 90%; L. plantarum, ~91%). The highest total score of extracellular amylase activity was recorded by L. acidophilus IBNA 09 at 24-48 h (5.10 ± 0.176 U/mL, 4.99 ± 0.409 U/mL), follow by L. paracasei CCM 1837(0.12 ± 0.002 U/mL, 0.15 ± 0.001 U/mL). During entire period, cellulase production was observed only for L. acidophilus (0.28 ± 0.019 U/mL), comparative with L. paracasei where the activity was observed in the first 24 h, respectively at 72 h for L. plantarum. These results suggest that L. acidophilus IBNA 09 possesses potential probiotic traits as a suitable candidate for amylase and cellulase production, and starter culture can improve cereal fermentation and the process of digestion in poultry nutrition.

Transformation and characterization of human insulin precursor gene in Pichiapastoris X-33

The case of diabetes increases significantly and has been projected to reach 592 million people in 2035. Consequently, the necessity of insulin will rise manifold and an efficient production system for insulin production is required to meet the market demands. The human insulin precursors that enzymatically converted to human insulin can be produced using Escherichia coli, Saccharomyces cerevisiae, or Pichia pastoris. In this study, Pichia pastoris is used for production human insulin precursor because the resulting recombinant protein can be folded accordingly and secreted to the external environment of the cell that simplifies the purification process. The study was initiated with the insertion of a synthetic gene of human insulin precursor into the pPICZaA to create recombinant pPICZaA-IP plasmid. The recombinant plasmid was transformed into Escherichia coli Top10 which then isolated and digested by the SacI enzyme. The linearize pPICZaA-IP plasmid was transfected into Pichia pastoris X-33 by electroporator. The result of transformation process, a total of 20 colonies of P pastoris X-33 were selected and inoculated in YPD agar medium containing Zeocin. The two colonies of P pastoris were characterized by PCR and sequencing showed that the recombinant pPICZaA-IP plasmid was successfully integrated into selected colonies of P pastoris.

Isolation and Identification Cellulolytic Bacteria from Termite Gut Obtained from Indralaya Peatland area

The production of second-generation bioethanol as renewable energy has developed very rapidly and has become a promising alternative energy source. Bioethanol production using biomass can be obtained alternatively from cellulose in wood, sawdust, organic waste, and agricultural waste. This research used termites obtained from Indralaya peatland area as organisms that can decompose cellulose into glucose with the cellulase enzymes produced by bacteria in their digestive tract. Cellulases are enzymes capable of hydrolyzing lignocellulose into glucose. The study aimed to isolate and identify of cellulolytic bacteria from termite gut obtained from Indralaya peatland area. The bacterial isolates were classified by using morphological and biochemical standard methods, and identification based on Bergey’s Manual of Determinative Bacteriology. Cellulolytic bacteria of termite gut were isolated and cultured on CMC (Carboxymethyl cellulose) agar medium. The activity of cellulolytic bacterial was conducted based on halo area and cellulolytic index on CMC agar medium. Among 64 isolates of bacteria, 24 isolates were identified as cellulolytic bacteria. Futhermore, our isolates with higher cellulolytic index were identified as the Staphylococcus, Microbacterium, Bacillus, and Brevibacterium genus.

Cultural, Morphological and Biochemical Variability Studies among the Isolates of Alternaria solani, the Causal Agent of Early Blight Disease of Tomato

Present experiment was conducted at College of Horticulture, Bengaluru (KA) during year 2017–18 to study the cultural, morphological and biochemical variations among the isolates of the pathogen Alternaria solani, the causal agent of early blight disease in tomato. The results revealed variation among the isolates collected from different regions of Karnataka state, India with regard to the colony characteristics viz., colony colour, mycelial growth pattern, margin of the colony and zonations whereas the maximum mycelial growth in terms of diameter (90 mm) was observed in the isolates Bagalkot (BaBG) and Chikkamagaluru (CMH) on Czapek’s (Dox) agar medium while the least growth (36.33) was noticed in Bidar (BiHH) isolate. The isolate could grow better on Czapek’s (Dox) agar medium as among the 3 media tested Czapek’s (Dox) agar medium produced maximum growth of 80.70 mm and the least growth (63.70 mm) was noticed in V-8 juice agar. The morphological studies revealed that all the conidia of various isolates varied in length (25.07–42.90 µm), breadth (10.53–21.52 µm) and number of horizontal septa (2–7), longitudinal septa (0–4). Biochemical studies among the isolates revealed significant variation in their enzyme activities. The peroxidase activity was more in Chikamagaluru (CMH) isolate (81.80 Unit g-1 FW) least activity was found in Bidar (BiHH) isolate 11.78 Unit g-1 FW whereas the esterase activity was more Bengaluru (BYC) isolate (69.01 Unit g-1 FW) least activity was found in Bagalkot (BaBG) isolate 11.78 Unit g-1 FW. Existence of variation among the isolates of Alternaria solani evident from the results obtained.