Characterization of variation and quantitative trait loci related to terpenoid indole alkaloid yield in a recombinant inbred line mapping population of <Emphasis Type="Italic">Catharanthus roseus</Emphasis>

Improved Catharanthus roseus cultivars are required for high yields of vinblastine, vindoline and catharanthine and/or serpentine and ajmalicine, the pharmaceutical terpenoid indole alkaloids. An approach to derive them is to map QTL for terpenoid indole alkaloids yields, identify DNA markers tightly linked to the QTL and apply marker assisted selection. Towards the end, 197 recombinant inbred lines from a cross were grown over two seasons to characterize variability for seven biomass and 23 terpenoid indole alkaloids content-traits and yield-traits. The recombinant inbred lines were genotyped for 178 DNA markers which formed a framework genetic map of eight linkage groups (LG), spanning 1786.5 cM, with 10.0 cM average intermarker distance. Estimates of correlations between traits allowed selection of seven relatively more important traits for terpenoid indole alkaloids yields. QTL analysis was performed on them using single marker (regression) analysis, simple interval mapping and composite interval mapping procedures. A total of 20 QTL were detected on five of eight LG, 10 for five traits on LG1, five for four traits on LG2, three for one trait on LG3 and one each for different traits on LG three and four. QTL for the same or different traits were found clustered on three LG. Co-location of two QTL for biomass traits was in accord of correlation between them. The QTL were validated for use in marker assisted selection by the recombinant inbred line which transgressively expressed 16 traits contributory to the yield vinblastine, vindoline and catharanthine from leaves and roots that possessed favourable alleles of 13 relevant QTL.     

Prospective role of insecticides of fungal origin: Review

Advances in the application of microbial‐based technology in insect pest management assist us to counter problems created by the application of chemical pesticides. These are mainly strong environmental effects, resistance development and high costs. Among the microbial pesticides, fungal pesticides are now preferred as they are target specific, ecofriendly, lacking in toxic residue and are economical. Being numerous with great diversification, entomopathogenic fungi therefore have great potential to control a large variety of insect pests. Fungi are applied directly in form of spores, mycelia or blastospores or by their metabolites (mycotoxins). Both approaches have very promising roles in insect pest management. However, there are three main obstacles in the development of fungal pesticides: (i) scant production of mycotoxins; (ii) carcinogenic mycotoxicosis in non‐target organisms; and (iii) slow effectiveness. Therefore, to eliminate these problems, attention has recently been paid to a synergistic approach to combating insecticide resistance. Next to synergism, genetic manipulation is also used to enhance the pathogenicity and virulence of fungal insecticides. However, the key risk associated with the release of recombinant microorganisms into the environment is that the novel organism may have unforeseen undesirable characteristics. Therefore, the introduction of synergists in pest control could have great benefit both economically and ecologically. An ideal synergistic approach is the mixing of more than two accelerating components together, i.e. tripartite or multiple synergism to enhance effectiveness. Thus, synergistic approaches have a bright future and require further research and financial support.     

Isolation and Characterization of Sodium Chloride-Resistant Callus Culture of Vigna radiata (L.) Wilczek var. radiata

Callus cultures were initiated from seedling root segments ofmungbean (Vigna radiata (L.) Wilczek var. radiata) cv. K 851on modified PC-L2 basal medium. Growing cells were exposed toincreasing concentrations of NaCl in the medium. A concentrationof 300 mol m^–3 NaCl proved completely inhibitory to growthof the calli. On incubation for 25 d, cells which could toleratethis concentration of NaCl grew to form cell clones. Selectedclones were characterized with regard to their growth behaviour,K⁺, Na⁺ and free proline content when grown under stress aswell as on normal media and were compared with the normal sensitivecallus. The selected callus was capable of growing on mediumcontaining NaCl at the inhibitory concentration. The K⁺ contentof the selected callus was lower in the case of the NaCl mediumthan for the normal medium. However, the selected clones maintainedhigher K⁺ and Na⁺ levels, with increased salinization comparedwith the wild-type cells. Salt-selected cells accumulated higherlevels of free proline under NaCl stress compared to wild-typecells. Under normal conditions, however, the amounts of freeproline in selected and non-selected calli were comparable. Key words: Vigna radiata, callus culture, NaCl stress     

Effects of Phytophthora palmivora on Zygotic Embryos of Papaya In Vitro

Zygotic embryos of Carica papaya were successfully germinatedin vitro on Murashige and Skoog's medium supplemented with 2%activated charcoal. The effects of light, temperature, sucroseand nutrient concentrations in the medium, on growth and developmentof embryos were examined. Strength of the nutrients in the mediumhad no effect on the growth and development of embryos. Thegerminated embryos of different varieties of papaya were inoculatedusing a sporangial suspension of different isolates of Phytophthorapalmivora. In the analysis of variance, varieties, isolatesand variety—isolate interactions differed significantly.The results were compared with the inoculation of glasshouse-grownseedlings; it is suggested that embryo inoculation could bea useful method of detecting resistance at an early stage ofplant development. Papaya, Carica papaya L., embryo culture, Phytophthora palmivora, resistance, in vitro screening     

Interference of Ascorbic Acid with the Starch-lodine Reaction

Starch estimation in various plant tissues is accomplished usinga reaction between starch and iodine. We report that the starch-iodinereaction is strongly inhibited by ascorbic acid, a normal cellularconstituent of higher plants. The presence of 10^–4 M ascorbicacid in the reaction mixture caused an almost complete inhibitionof the starch-iodine reaction. This inhibition may be alleviatedby increasing the amounts of iodine-potassium iodide (IKI) inthe reaction mixture, suggesting the possibility that a complexis formed between ascorbic acid and iodine. Evidence for thiscomes from a shift in absorption maxima of ascorbic acid dueto addition of IKI. Starch-iodine reaction, ascorbic acid, interference     

Cytochemical Observations on Malic Dehydrogenase,Lipase, Nonspecific Esterase and Monoamine Oxidase in Chick Liver Cell Cultures Infected with Trichomonas vaginalis*

SYNOPSIS. In trypsin-dispersed chick liver cell cultures malic dehydrogenase activity is localized in granules distributed thruout the cytoplasm of fibroblasts, epithelial cells, and macrophages. A progressive increase of the enzymic activity in all cell culture elements, except for phagocytes whose external or internal surfaces remain in direct contact with the parasites, accompanies infection of the cultures with a relatively pathogenic strain of Trichomonas vaginalis. In such phagocytes most staining for malic dehydrogenase is lost. Epithelial cells and parasite-free macrophages in experimental cultures also have a diffuse cytoplasmic reaction. No lipase activity is present in fibroblasts, but epithelial cells and macrophages in chick liver cell cultures contain numerous reactive granules. A strong diffuse cytoplasmic reaction is found in the epithelial cells and a weaker one in the control phagocytes. In cultures infected with T. vaginalis the enzyme is lost progressively from the epithelium and from those macrophages which have engulfed the parasites or whose external surfaces are invested with the flagellates; however, no significant changes in lipase activity can be found in parasite-free experimental phagocytes. In all cell types found in chick liver cultures, the reaction for nonspecific esterase is localized in cytoplasmic inclusions of varying size, some of which tend to accumulate along the cell membranes and around the nuclei. In addition, a weak diffuse cytoplasmic reaction is seen in the epithelial cells. Most cells in cultures infected with T. vaginalis have a significant increase in esterase activity, the exception being the macrophages which contain parasites within their cytoplasm or those with flagellates applied closely to their external surfaces. Only a few specifically stained granules are retained by such phagocytes. Monoamine oxidase activity is limited to fine granules dispersed in the cytoplasm of fibroblasts, epithelial cells, and macrophages of control cultures. Infection of chick liver cultures with T. vaginalis results in lowered enzyme activity in non-phagocytic cells as well as in macrophages with engulfed flagellates and in those whose external surface are invested with the parasites. The number of reactive inclusions appears to increase in trichomonad-free phagocytes of experimental cultures.     

Biology of the Collembola Xenylla grisea Axelson and Lepidocyrtus cyaneus f. cinereus Folsom

• 1 Xenylla grisea Axelson and Lepidocyrtus cyaneus f. cinereus Folsom were reared on brown whole wheat flour in culture jars containing a base of plaster of paris and charcoal.
• 2 Sexes in X.grisea may be differentiated by the combination of three characters: length, colour and body shape. No parthenogenesis was observed in either species.
• 3 At room temperature (25–27°C), females of X.grisea began laying eggs after the fourth moult and required 10–15 days (mean of 12.6) to reach sexual maturity from the day of hatching. With L.cyaneus f. cinereus, average periods from hatching to maturity were 16.8, 27.4 and 43.2 days at temperatures of 26.5, 22 and 15°C, respectively.

     

The Melanotropic Peptide, [Nle4, d-Phe7]α-MSH,Stimulates Human Melanoma Tyrosinase Activity and Inhibits Cell Proliferation

Seventeen human melanoma cell (HMC) lines, both melanotic and amelanotic, were incubated in the continuous presence of a potent melanotropic peptide hormone analog, [Nle⁴,d -Phe⁷]α-MSH, for 72 hr with daily changes of medium. Only one cell line (HD, melanotic) consistently responded to the hormone analog by increased tyrosinase activity. Three (one melanotic, two amelanotic) of the HMC lines also failed to respond to the peptide by either increased or decreased enzyme activity when incubated continuously in the presence of the peptide for longer periods of time (6,15,27,43 days). The HD cell line, however, again responded with increasingly enhanced basal enzyme activity the longer the cells were incubated in the presence of the melanotropin. One amelanotic cell line (C8161) responded with enhanced enzyme activity when grown to confluency in the continuous presence of the peptide. Basal tyrosinase activity of the C8161 cell line may have increased as cell density in the flasks increased. These results suggest that under conditions of increased cell number, phenotypic expression of tyrosinase activity in so called “amelanotic” (tyrosinase-negative) cells is increased and can be enhanced further by stimulation with a melanotropic peptide. Under conditions of increased cell number, the presence of [Nle⁴,d -Phe⁷]α-MSH caused morphological differentiation (shape change); the cells became enlarged and very dendritic. The number of cells in monolayer (surface of the flask) and in the medium were drastically reduced in both melanotic and “amelanotic” cell lines incubated with [Nle⁴,d -Phe⁷]α-MSH. The data support other published reports that melanotropic peptides inhibit human melanoma cell growth (proliferation) in vitro, most likely through a cytostatic mechanism. [Nle⁴,d -Phe⁷]α-MSH also exhibited a prolonged (residual) inhibitory action on HD cell proliferation. In other words, inhibition of cell growth (proliferation) of the HMCs was evident even several days after removal of the melanotropic peptide from the incubation medium.     

Resistance to cyst nematode (Heterodera cajani) in pigeonpea cultivars and in wild relatives of Cajanus*

Heterodera cajani is an important nematode pest of pigeonpea in India. Evaluation of 58 pigeonpea cultivars and 61 accessions of Cajanus acutifolius, C. cajanifolius, C. grandifolius, C. lanceolatus, C. lineatus, C. mollis, C. pla-tycarpus, C. reticulatus, C. scarabaeoides, C. sericeus, C. volubilis, Flemingia macrophylla, F. stricta, F. strobilifera, Rhynchosia aurea, R. bracteata, R. cana, R. densiflora, R. minima, R. rothii, R. suaveolens and R. sublobata revealed that the tested pigeonpea cultivars lacked resistance to H. cajani. Eight accessions of wild relatives were resistant and 20 accessions were moderately resistant. Based on the white cyst number on roots and low plant-to-plant variation, two accessions of C. scarabaeoides (ICPWs 111 and 128), three accessions of Flemingia spp. (ICPWs 194, 202 and 203), and one accession each of R. rothii (ICPW 257), R. densiflora (ICPW 224), and R. aurea (ICPW 210) were identified as resistant and promising for use in intergeneric hybridisation programmes.