A novel serine protease cryptolepain from Cryptolepis buchanani: purification and biochemical characterization

A novel protease is purified to homogeneity from the latex of a medicinally important plant Cryptolepis buchanani of family Apocynaceae (formerly Asclepiadaceae). The enzyme named cryptolepain has a molecular mass of 50.5 kDa. The isoelectric point and extinction coefficient (epsilon280nm1%) are 6.0 and 26.4, respectively. Cryptolepain contains 15 tryptophans, 41 tyrosines, and eight cysteine residues forming four disulfide bridges. The detectable carbohydrate moiety in the enzyme was found to be 6-7%. Cryptolepain hydrolyzes denatured natural substrates like casein, azocasein, and azoalbumin with high specific activity. The protease is exclusively inhibited by serine protease inhibitors phenylmethansulfonyl fluoride and diisopropyl fluorophosphate. Hydrolysis of azoalbumin by the cryptolepain is optimal in the pH range of 8-10 and temperatures of 65-75 degrees C. The enzyme shows high stability against pH (2.5-11.5), temperature (up to 80 degrees C), and chemical denaturants. The Km value of the enzyme was found to be 10 microM with azocasein as the substrate. The N-terminal sequence of cryptolepain is unique and shows only little homology to other known serine proteases, which makes this enzyme an ideal candidate for our ongoing biochemical and structure-function investigations of proteases. Easy availability of the latex and simple purification procedures make the enzyme a good system for exploring the biophysical chemistry of serine proteases as well as applications in the food industry.     

Deep Tillage,Soil Moisture Regime,and Optimizing N Nutrition for Sustaining Soil Health and Sugarcane Yield in Subtropical India

A field experiment was conducted at ICAR-Indian Institute of Sugarcane Research, Lucknow, with three tillage practices (T₁: Control- two times ploughing with harrow and cultivator, each followed by planking before sugarcane planting; T₂: Deep tillage with disc plough (depth 25–30 cm) before planting followed by harrowing, cultivator, and planking; and T₃: Subsoiling at 45–50 cm and deep tillage with disc plough/moldboard plough (depth 25–30 cm) followed by harrowing, cultivator, and planking before planting, two soil moisture regimes (M₁: 0.5 irrigation water (IW)/cumulative pan evaporation (?CPE) ratio and M₂: 0.75 IW/CPE ratio) at 7.5 cm depth of IW, and four N levels (N₁- 0, N₂- 75, N₃- 150, and N₄-225 kg N ha^?1) in sugarcane plant crop. Deep tillage and subsoiling increased porosity and reduced bulk density in surface/subsurface soil. Further, these physical changes also improved soil biological and chemical properties responsible for higher crop growth and yield. Deep tillage and subsoiling reduced the compaction by 6.12% in 0–15 cm depth in sugarcane plant crop at maximum tillering stage. The highest N uptake (158.5 kg ha^?1) was analyzed with deep tillage and subsoiling compared to all other tillage practices. Maintaining suboptimal moisture regime with deep tillage and subsoiling showed the highest IW use efficiency (157.16 kg cane kg^?1 N applied). Mean soil microbial biomass carbon (SMBC) in ratoon crop was higher compared to plant crop. During initial tillering stage, ratoon crop showed higher SMBC with application of deep tillage and subsoiling (1209 mg CO₂-C g^?1 soil day^?1) at 0–15 cm depth and 1082.9 mg CO₂-C g^?1 soil day^?1 at 15–30 cm depth. Thus, it could be concluded that besides improving sugarcane yield, soil health could be sustained by adopting subsoiling (45–50 cm depth) and deep tillage (20–25 cm depth), with soil moisture regime of 0.75 IW/CPE and application of 150 kg N ha^?1 in sugarcane (plant crop).     

Estimation of endogenous levels of osteopontin,total antioxidant capacity and malondialdehyde in seminal plasma: Application for fertility assessment in buffalo (Bubalus bubalis) bulls

This study was attempted to identify subfertile bulls by quantifying the endogenous levels of osteopontin (OPN), total antioxidant capacity (TAC) and malondialdehyde (MDA) in seminal plasma of buffalo bulls. On the basis of conception rate, buffalo bulls were classified into two groups: high‐fertile (conception rate >50%) and subfertile bulls (conception rate <40%). A total of 100 ejaculates (10 ejaculates from each bull) were collected through artificial vagina method. The concentration of OPN, TAC and catalase (CAT) of high‐fertile bulls was found to be higher (p < .05) than that of subfertile bulls. Further, MDA level in seminal plasma was found to be lower (p < .05) in high‐fertile bulls compared with subfertile bulls. The fertility status had no effect on the superoxide dismutase (SOD) concentration in seminal plasma of both the groups. The levels of OPN (r = .678, p = 0.013) and TAC (r = .648, p = .042) were found to be positively correlated with bull fertility and the level of MDA (r = ?.718, p = .019) was found to be negatively correlated with bull fertility. However, the fertility of bulls was not found to be significantly correlated with SOD, CAT and sperm motility. In conclusion, seminal OPN, TAC and MDA tended to be more realistic in identification of subfertile bulls from breeding herds.     

Interspecific compatibility barriers,development of interspecific hybrids through embryo rescue and lineage of Trifolium alexandrinum (Egyptian clover)—important tropical forage legume

Genetic improvement of Trifolium alexandrinum L. is hampered because of narrow genetic base and lack of interspecies compatibility information. The study was envisaged to understand the interspecies incompatibility and the affinity between T. alexandrinum and 22 species of the genus representing different sections and to develop interspecific crosses. Development of interspecific hybrids with T. resupinatum, T. lappaceum, T. subterraneum, T. vesiculosum and T. echinatum, through embryo rescue, revealed their affinity with T. alexandrinum. Failure of pollen germination or restricted pollen tube growth showed its distant relatedness with T. clypeatum, T. balansae, T. purpureum, T. leucanthum, T. hirtum, T. pilulare and T. hybridum. Pollen tubes reached up to ovule among the crosses with T. alpestre, T. repens and T. nigrescens, whereas it travelled up to ovary among the crosses with T. angustifolium, T. grandiflorum and T. dasyurum indicating need of manipulation to overcome barrier. Thus, incompatibility of T. alexandrinum with many Trifolium species could be overcome through embryo rescue with intensive crossing. Egyptian clover ecotype ‘Fahli’ was more compatible with other species than ‘Mescavi’.     

Effect of soil application of nickel on growth,micronutrient concentration and uptake in barley (Hordeum vulgare L.) grown in Inceptisols of Varanasi

A pot experiment was conducted in a glass house on low nickel containing alluvial soil in the Department of Soil Science and Agricultural Chemistry, Institute of Agricultural Sciences, Banaras Hindu University, Varanasi, during 2012–13 and 2013–14, to study the response of barley to soil application of nickel (Ni). There were ten treatments of Ni (0, 2.5, 5, 10, 15, 20, 30, 40, 50 and 60 mg kg^?1) studied with recommended dose of fertilizers nitrogen, phosphorus, potassium and sulfur (N:P:K:S :: 40:30:30:20 mg kg^?1).The results showed a significant increase in plant height, number of tillers, chlorophyll content, straw and grain yield, and 1,000 grains weight with application of 10 mg Ni kg^?1 soil during both years of study. The micronutrient concentration and uptake in straw and grain increased with application of <15 mg Ni kg^?1 soil and beyond that declined significantly. Diethylenetriaminepentaacetic acid-extractable micronutrient iron, manganese, copper, zinc and nickel (Fe, Mn, Cu, Zn and Ni) content in soil increased with increasing level of Ni. The maximum urease activity in post-harvest soil was noticed with application of 40 mg Ni kg^?1 soil. The microbial population viz. bacteria, fungi and actinomycetes were higher with 5, 30 and 10 mg Ni kg^?1 soil, respectively.     

Impact of low potassium fertilization on potassium transformation under different crop management systems in western plain of arid India

Acquaintance of potassium (K) transformation in soil is crucial for K management in management system of arid India. Present study revealed that the extractable K like ammonium acetate–potassium (NH₄OAc–K), nitric acid (HNO₃)–K, and hydrochloric acid (HCl)–K under different systems were followed the order: irrigated low input > rainfed low input > irrigated high input > no input. The pearl millet systems in irrigated low input practice maintained a higher NH₄OAc–K than irrigated high input practice. The exchangeable-K (Exch-K), non-exchangeable-K (Non-exch K) and lattice-K were observed highest in irrigated low input system and lowest in no input system. It is noticed that Exch-K contributed 2.3–2.6% over total-K, which readily available to plants. The Exch-K was highly significant correlated with K pools and soil properties, indicating that rapid establishment of equilibrium between the pools. Thus, knowing K reserve and distribution of K forms can help in understanding K replenishment capacity of arid soil under long-term intensive cultivation.     

Pathological studies on the southern blight of China aster (<Emphasis Type="Italic">Callistephus chinensis</Emphasis>) caused by <Emphasis Type="Italic">Sclerotium rolfsii</Emphasis>

A severe outbreak of southern blight disease of China aster was observed during the post rainy season (September–November 2015) in the Mysore and Mandya Districts of Karnataka, Southern India. The disease incidence ranged between 12 and 47%. The typical disease symptoms include water-soaked lesions on leaves, stems and on the lower stem surfaces followed by quick wilting of the whole plant with abundant production of sclerotia near the stem-soil interface. The associated fungal pathogen was isolated on potato dextrose agar (PDA) medium, on which numerous reddish-brown sclerotia were seen. A total of 26 fungal isolates were isolated and studied for the mycelial compatibility. Isolate SrCCM 1 was used for pathogenicity analysis. The results of the study showed that, there was no variation among the isolates tested. Molecular identification of the pathogen by ITS-rDNA sequences of S. rolfsii showed 100% similarity with reference sequences. Based on the cultural, morphological and molecular characteristics, the fungal pathogen was identified as Sclerotium rolfsii Sacc. (Sexual morph: Athelia rolfsii (Curzi) C.C. Tu & Kimbr). Pathogenicity tests were performed on healthy leaves, roots and stems. Typical disease symptoms on leaves, stems and roots were evident after 5, 8 and 10 days of post-inoculation. Sclerotium rolfsii is known to cause diseases in economically important crop plants. However, no reports are available on the occurrence of S. rolfsii on China aster in India.     

Rules of benefit distribution: A dynamic impact on Nepalese community forestry organizations

Participatory approach has been an official part of Community Forestry (CF) since 1989 when Master Plan for the Forestry Sector (1989) was introduced in Nepal. However, many problems related to benefit distribution from CF have emerged because of the way decision-making is influenced by social and institutional structures present at community level, particularly in terms of dominance by wealthy and caste elite to formulate rules. The study used a conceptual approach using elite theory with models that looked at Executive Committee (EC) that used to formulate distribution rules. The study uses Community Forestry User Groups (CFUGs) level data from 31 CFUGs and household data from 310 households in Baglung district, Nepal. The study examined the factors linked with rules of distribution that determined the relative distribution of forest products. The higher the representation of the poor and disadvantaged households on the EC, the greater the benefits to them in terms of greater quantities distributed and longer collection periods. The policy implication of this study is that the forestry organizations help the poor and underprivileged households to build up capacity to undertake leadership roles that influence the formulation of rules through which organizational elite models in favor to them become part of the elite decision-making.