Genetic diversity and population structure of endangered rosewood from the Peruvian Amazon using ISSR markers

Rosewood, Aniba rosaeodora is an endangered species in Amazon forests and its natural stands have been heavily depleted due to over-exploitation for the cosmetic industry. This study aimed to investigate the genetic diversity and population structure of 90 rosewood accessions from eight localities in the Peruvian Amazon through 11 Inter Simple Sequence Repeats (ISSR) primers. The ISSR primers produced a sum of 378 bands, of which 375 (99.2%) were polymorphic, with an average polymorphism information content (PIC) value of 0.774. The mean effective number of alleles (Ne), Shannon informative index (I), gene diversity (He) and total gene diversity (Ht) were 1.485, 0.294, 0.453 and 0.252, respectively. Analysis of molecular variance (AMOVA) showed the presence of maximum variability within populations (88%). The Structure algorithm, neighbor joining and principal coordinate analysis (PCoA) grouped the 90 rosewood accessions into three main populations (A, B and C). Diversity indices at the inter-population level revealed a greater genetic diversity in population A, due to higher gene flow. The neighbor-joining analysis grouped populations A and B, while population C was found to be divergent at the inter population level. We concluded that population A reflects higher genetic diversity and should be prioritized for future management and conservation plans. (AU)

Minerals and phytochemical analysis of bitter melon fruits and its components in some indigenous and exotic cultivars / Análise fitoquímica e de minerais de frutos de melão amargo e seus componentes em alguns cultivares indígenas e exóticos

The present study was designed to compare the mineral contents and phytochemicals in fruits of some indigenous and exoticbitter melon cultivars. Skin, flesh, seeds and whole fruit of each cultivar were dried at room temperature and ground to fine powder which was packed separately in air-tight plastic jars for analysis. The data revealed that all the cultivars possessed appreciable amount of minerals. Indigenous cultivars possessed higher mineral contents than the exotic cultivars. Black King was found to be desirable cultivar with high amount of potassium, phosphorous, magnesium, calcium, iron and zinc with values of 258.83, 92.92, 46.74, 48.42, 3.90 and 1.78 mg/100g, respectively. Among fruit parts, flesh portion was rich in potassium, phosphorous, magnesium, sodium and calcium followed by skin and seed. The seeds possessed relatively high amount of iron (4.44 mg/100g) and zinc (2.80 mg/100g). On the whole, flesh part of Black King is predominated in mineral profile while least quantity was observed in the seeds of Noor. All the cultivars possessed same types of phytochemicals although may vary in their abundance. Phenols, flavanoids, saponins and ascorbic acid were present in abundance in skin and flesh while seeds were found rich in alkaloids and glycoalkaloid. The variations in the chemical composition in fruit and components due to changes in agro-climatic conditions suggest that there is scope for developing cultivars to be grown in wide range of agronomic, climatic and geographic conditions with no effect on its nutrient and phytochemical profile.

O presente estudo foi projetado para comparar os teores de minerais e fitoquímicos em frutos de alguns cultivares indígenas e exóticos de melão amargo. Casca, polpa, sementes e frutos inteiros de cada cultivar foram secos à temperatura ambiente e moídos até obter-se um pó fino que foi embalado separadamente em frascos de plástico hermeticamente fechados para análise. Os dados revelaram que todos os cultivares possuíam quantidade apreciável de minerais. Cultivares indígenas possuíam maior conteúdo mineral que os cultivares exóticos. Descobriu-se que o Black King foi o cultivar desejável com alta quantidade de potássio, fósforo, magnésio, cálcio, ferro e zinco com valores de 258.83, 92.92, 46.74, 48.42, 3.90 e 1.78 mg/100g, respectivamente. Entre as partes das frutas, a polpa era mais rica em potássio, fósforo, magnésio, sódio e cálcio, seguida pela casca e pela semente. As sementes possuíam quantidades relativamente elevadas de ferro (4.44 mg/100g) e zinco (2.80 mg/100g). No geral, na polpa do Black King predomina o perfil mineral, enquanto uma menor quantidade foi observada nas sementes de Noor. Todos os cultivares possuíam os mesmos tipos de fitoquímicos, embora possam variar em sua abundância. Fenóis, flavonóides, saponinas e ácido ascórbico estavam presentes em abundância na casca e na polpa enquanto as sementes eram ricas em alcalóides e glicoalcalóides. As variações na composição química de frutos e componentes devido a mudanças nas condições agroclimáticas sugerem que há espaço para o desenvolvimento de cultivares a serem cultivadas em ampla escala de condições agronômicas, climáticas e geográficas, sem efeito no seu perfil nutricional e fitoquímico.

Recombinant production and biochemical and in silico characterization of lactate dehydrogenase from Geobacillus thermodenitrificans DSM-465

Background: Lactate dehydrogenase (LDH) is an enzyme of glycolytic pathway, ubiquitously found in living organisms. Increased glycolysis and LDH activity are associated with many pathologic conditions including inflammation and cancer, thereby making the enzyme a suitable drug target. Studies on conserved structural and functional domains of LDH from various species reveal novel inhibitory molecules. Our study describes Escherichia coli production and characterization of a moderately thermostable LDH (LDH-GT) from Geobacillus thermodenitrificans DSM-465. An in silico 3D model of recombinant enzyme and molecular docking with a set of potential inhibitors are also described. Results: The recombinant enzyme was overexpressed in E. coli and purified to electrophoretic homogeneity. The molecular weight of the enzyme determined by MALDI-TOF was 34,798.96 Da. It exhibited maximum activity at 65°C and pH 7.5 with a KM value for pyruvate as 45 µM. LDH-GT and human LDH-A have only 35.6% identity in the amino acid sequence. On the contrary, comparison by in silico structural alignment reveals that LDH-GT monomer has approximately 80% identity to that of truncated LDH-A. The amino acids "GEHGD" as well as His179 and His193 in the active site are conserved. Docking studies have shown the binding free energy changes of potential inhibitors with LDH-A and LDH-GT ranging from −407.11 to −127.31 kJ mol−1 . Conclusions: By highlighting the conserved structural and functional domains of LDH from two entirely different species, this study has graded potential inhibitory molecules on the basis of their binding affinities so that they can be applied for in vivo anticancer studies

Eosinophilia with PDGFRB gene rearrangement and isolated del 5q; an extremely rare presentation of myeloid neoplasm

Introduction: Myeloproliferative neoplasms with PDGFRA, PDGFRB and FGFR1 rearrangements are reported to be very rare entities. Myeloid neoplasms with PDGFRB rearrangement have prominent eosinophilia, neutrophilia or monocytosis and presence of t (5;12)(q31~q33;p12-13) or variant translocation. Case Report: We report a 55 years old female patient who presented with eosinophilia along with predominant eosinophil precursors on bone marrow and concomitant isolated del (5q) and PDGFRB gene rearrangement which is an infrequent and rare finding. Conclusion: isolated acquired deletion of the long arm of chromosome 5 (del 5q) also known as 5q- syndrome, is a distinct hematologic disorder reported to be primary myelodys plastic syndrome which is found in females of middle age and usually presents with macrocytic anemia, oval macrocytes, with white blood cell counts normal or reduced, platelet counts normal or elevated and bone marrow exhibitery throid hypoplasia with megakaryocytes showing hypolobated nuclei

A 3-way Rare Variant Ph Translocation t(7;9;22) in a patient with Chronic Myeloid Leukemia.

Chronic Myeloid Leukemia (CML) is associated with translocation between chromosome 9 & chromosome 22, t(9;22)(q34;q11.2) and with the formation of BCR-ABL fusion gene. In few newly diagnosed CML cases, complex cytogenetics variants of the Philadelphia (Ph) chromosome can be observed with the involvement of a third chromosome other than chromosome 9 & chromosome 22. Here in, we describe a 45 yrs old male, diagnosed as CML in Chronic phase with a complex translocation involving chromosomes 7, 9 & 22. Cytogenetics investigations for confirmation of CML revealed 46,XY, t(7;9;22)(q11.2;q34;q11.2) in 100% metaphases counted. Fluorescence in-situ hybridization (FISH) showed BCR-ABL fusion signals pattern in 100% nucleated cells analyzed. A review of the literature revealed that CML patients with this translocation tend to have an aggressive course and poor outcome. Additional 3-way chromosome translocations associated with CML are also reviewed.

Kinetics Study of Extracellular Detergent Stable Alkaline Protease from Rhizopus oryzae

In this study, extracellular alkaline protease was produced from Rhizopus oryzae in submerged fermentation using dairy waste (whey) as a substrate. Fermentation kinetics was studied and various parameters were optimized. The strain produced maximum protease at initial medium pH of 6.0 medium depth of 26 mm, inoculum size of 2% at incubation temperature of 35ºC for 168 h of fermentation. Alkaline protease was purified to homogeneity by ammonium sulphate fractionation followed by sephadex G-100 chromatography. The molecular mass of alkaline protease was 69 kDa determined by 10% SDS-PAGE. The optimum pH and temperature of alkaline protease was 9.0 and 40ºC, respectively. Metal profile of the enzyme showed that the enzyme was non-metallic in nature. The Km , Kcat , Vmax and Kcat/Km values of purified protease were 7.0 mg/mL, 3.8 x102 S-1, 54.30 µmol/min and 54.28 s-1 mg -1.mL respectively, using casein as substrate. The purified alkaline protease had stability with commercial detergents. .

Investigation of Genetic Diversity among Medicago species Using RAPD Markers.

Aims: Medicago is known as the Queen of forage with potential economic importance to our society. The present study aimed at the use of RAPD-PCR DNA marker to identify the genetic fingerprints affinities of six species of Alfalfa. Place and Duration of Study: The study was conducted at the Department of Genetics, Garden Campus, Hazara University, Mansehra Pakistan during February, 2011 to August, 2013. Methodology: In this study, six species of Medicago namely TWAL (Tetraploid Wisconsin Alfalfa Line), Medicago arborea, Medicago falcata, Medicago sativa, Medicago lupulina and Medicago polymorpha were used to explore the diversity of alfalfa. Seven out of 120 decamers produced 34 polymorphic loci with 100% polymorphism to identify the different species of Medicago crop. The range of polymorphic loci was observed from 300 to 700 bp. Eleven species specific loci were generated by seven decamers. Primer B-18 generated single specific locus 700 bp against genomic DNA of M. lupulina and it is important to identify particular species of Alfalfa. The bivariate data were recorded as the presence of locus 1 and absence 0 and then this data was transferred into A and C respectively to make it suitable for DNAMAN software (version 5.2.2.0; Applied Biostatistics Inc). Moreover, cluster analysis was performed using sequence alignment and divergence function of the DNAMAN against the bivariate data collected from the products of decamers. All members clustered in a unique pattern except M. falcata and M. lupulina those shared 86% homology. Three distinct groups were observed during UPGMA (Unweighted pair Group Method with Arithmetic Mean). During the phylogenetic study, TWAL was observed to have genetic diversity from other five species of Alfalfa. Conclusion: So, the present study is enabling us to discriminate different species of Alfalfa and it could be useful to identify and authenticate different species of the same genus of medicinal important plant from the Flora of Pakistan.

Ethanol production from agricultural wastes using Sacchromyces cervisae

The main objective of this study was production of ethanol from three lignocellulosic biomasses like sugarcane bagasse, rice straw and wheat straw by Sacchromyces cervisae. All the three substrates were ground to powder form (2 mm) and pretreated with 3%H2O2 + 2% NaOH followed by steaming at 130 °C for 60 min. These substrates were hydrolyzed by commercial cellulase enzyme. The whole fermentation process was carried out in 500 mL Erlenmeyer flask under anaerobic conditions in submerged fermentation at 30 °C for three days of incubation period. FTIR analysis of the substrates indicated significant changes in the alteration of the structure occurred after pretreatment which leads to efficient saccharification. After pretreatment the substrates were hydrolyzed by commercial cellulase enzyme and maximum hydrolysis was observed in sugarcane bagasse (64%) followed by rice straw (40%) and wheat straw (34%). Among all these tested substrates, sugarcane bagasse (77 g/L) produced more ethanol as compared to rice straw (62 g/L) and wheat straw (44 g/L) using medium composition of (%) 0.25 (NH4)2SO4, 0.1 KH2PO4, 0.05 MgSO4, 0.25 Yeast extract by S. cervisae.

Physico-chemical analysis and antimicrobial potential of Apis dorsata, Apis mellifera and Ziziphus jujube honey samples from Pakistan

Objective: To evaluate physico-chemical properties and antimicrobial potential of indigenous honey samples against different reference strains including Escherichia coli ATCC 8739, Enterobacter aerogenes ATCC 13048, Pseudomonas aeroginosa ATCC 9027, Bacillus subtilis ATCC 6633, Staphylococcus aureus ATCC 25923, Salmonella typhi ATCC 14028, Klebsiella pneumonia ATCC 13883, Aspergillus niger ATCC 16404, Rhizopus oligosporus PCSIR1, Candida albicans ATCC 14053 and Candida utilis ATCC 9950. Methods: By using standard methods samples were evaluated for their antimicrobial properties including additive effect of starch and non-peroxidase activity, antioxidative properties (phenol contents, flavonoid contents, 1,1-diphenyl-2-picrylhydrazyl radical scavenging activity). Prior to this evaluation, complete physico-chemical properties including pH, color, ash contents, protein contents, moisture contents, hydroxymethyl furfural contents, total sugar contents, reducing sugar and non-reducing sugar contents were analyzed. Results: Relatively higher ash contents were found in the Siddar honey i.e. (0.590 0±0.033 6)%and small honey showed relatively higher protein contents i.e. (777.598±9.880) mg/kg. The moisture contents of tested honey samples ranged between 13.8%-16.6%, total sugar contents from 61.672%-72.420%and non-reducing sugar contents from 1.95%-3.93%. Presences of phenolic contents indicate higher antioxidant potential of these honey samples. All bacteria showed clear inhibition zones in response to tested honey samples whereas fungi and yeast showed inhibition at higher concentrations of these honey samples. For Escherichia coli, Bacillus subtilis, Salmonella typhi, Pseudomonas aeroginosa and Aspergillus niger, overall the small honey showed the higher activity than other honey samples. Conclusion: Physico-chemical analysis of honey samples confirmed good quality of honey according to the standards set by European Union Commission and Codex Alimentarius Commission. Evaluation of these honey samples confirms antimicrobial potential of particular types of honeys indigenous to Pakistan.

Statistical Prediction Model for Relapse Rate in Chronic Hepatitis C Patients Treated with Conventional Interferon and Ribavirin Therapy.

Objective: To determine the significantly associated factors with the relapse rate in chronic hepatitis C patients treated with conventional interferon and their predictive strength through the regression model. Material & Methods: In a retrospective analysis of 244 patients, result of PCR, gender, fatty liver, diabetes, abnormal ALT at start and end of treatment were the qualitative variables. Age, weight, ALT at start and end of treatment, hemoglobin, platelets and WBC at start of treatment were quantitative variables. Bivariate, multivariate analysis and odds ratio were computed to verify statistically significant association with relapse rate by running binary logistic regression model. Results: Out of total 244 patients there were 54.1% male and 45.9% female. Eighty two (33.6%) patients had weight > 70 Kg, 30 (12.3%) had fatty liver, 18 were (7.4%) diabetic, 12 (4.9%) had normal ALT at start of therapy and 140 (57.4%) had abnormal ALT at the end. Eighty four (34.4%) patients relapsed while 160 (65.6%) maintained SVR after 6 month to 2 years of completion. In bivariate analysis, age, weight, fatty liver, high fever, decrease and increase in Hb were found significant. The binary logistic regression revealed the significant association of weight (OR=84.813; p=0.000), high fever (OR=4.478; p= 0.038) and Hb increase at 1st month (OR=0.037; p=0.013) with relapse rate. Nagelkerke R Square and Cox & Snell R Square statistics explained 71.1% and 51.1% variation in the model respectively and 93.1% area under the curve gave it very good prediction strength.

Enhanced production of alkaline protease by a mutant of Bacillus licheniformis N-2 for dehairing

The purpose of the present investigations was to improve the yield of alkaline protease for leather dehairing by subjecting the indigenous proteolytic strain Bacillus licheniformis N-2 to various mutagenic treatments viz. UV irradiations, NTG (N-methyl-N-nitro-N-nitrosoguinidine) and MMS (methyl methane sulfonate). After screening on skim milk agar plates, a total of nine positive mutants were selected for shake flask experiments. Among these, the best proteolytic mutant designated as UV-9 showed 1.4 fold higher alkaline protease activity in preoptimized growth medium than the parent strain. The fermentation profile and kinetic parameters such u(h-1), Yp/s, Yp/x, Yx/s, q s, Qs, q p and Qp also indicated the superiority of the selected mutant UV-9 for alkaline protease production over the parent strain and rest of the mutants. The dehairing capability of mutant UV-9 alkaline protease was analyzed by soaking goat skin pieces for different time intervals (3-15 h) at 40 º C. A complete dehairing without degradation of collagen was achieved after 12 h, indicating its commercial exploitation in leather industry.

Germination behaviour of seeds from herbicide treated plants of Chenopodium album L

The carry-over effect of sub-lethal herbicides was investigated on the germination of seeds collected from surviving Chenopodium album plants, which had received 1/8, 1/8 twice, 1/8 three times, 1/4, 1/2, 1/1 doses of either pre-emergence ioxynil or post-emergence bentazone in a previous onion (Allium cepa) crop. Seeds were also collected from surviving C. album plants, which had received 1/4, 1/2, 1/1 of either pre-emergence pendimethalin, propachlor and linuron, or 1/8, 1/8 twice, 1/8 three times, 1/4, 1/2, 1/1 of post-emergence ioxynil or linuron in a previous leek (Allium porrum) crop. Seeds of surviving plants were collected and tested for germination at temperature of 5, 10, 15, 20 and 25°C. The effect of different temperatures on the total number of germinated seeds was significant. Germination was minimum at low temperatures (5°C or 10°C). Herbicides did not show any effect on germination of C. album and resulted in the same final germination percentage as seeds collected from the unsprayed control plots.

Avaliou-se o efeito residual de doses sub-letais de herbicidas sobre a germinação de sementes de plantas de Chenopodium album sobreviventes em uma cultura prévia de cebola (Allium cepa), que recebeu 2x, 3x, 1/4, 1/2 ou a dose recomendada de ioxynil em pré-emergência ou bentazone em pós-emergência. As sementes foram também coletadas de plantas de C. album sobreviventes de um campo de alho-porró (Allium porrum) que havia sido tratado com 1/4, 1/2 ou a dose recomendada de pendimethalin, propachlor e linuron em pré-emergência, ou ainda 2x, 3x, 1/4, 1/2 ou a dose recomendada de ioxynil ou linuron em pós-emergência. As sementes destas plantas sobreviventes foram coletadas e testadas quanto à germinação a temperaturas de 5°C, 10°C, 15°C, 20°C e 25°C. Verificou-se que o efeito das temperaturas na germinação destas sementes foi significativa. A germinação foi mínima a baixas temperaturas (5°C e 10°C). Os herbicidas não causaram nenhum efeito na germinação das sementes de C. album tendo resultado no mesmo nível daquelas das plantas controles de culturas não pulverizadas.