EFFECT OF SOME SELECTED FUNGICIDES ON THE GROWTH OF TISSUES CULTURED IN VITRO

Fungi were isolated from contaminated tissue cultures. They were initially characicriscdaccording to the colour and the texture of the colony and an identification number wasgiven to each. Pathogens were inoculated into seven different media in order to determinethe suitable medium for the rapid growth of the fungi and to obtain pure cultures. Theywere incubated at 27±1"C.Five different fungicides (diathanc. systhane, rovral, tumble blight and thiophenatemethyyl)were prepared at IOOppm concentration. Sterile filter paper circles (5mm) werewetted with 101 of each and were placed separately on the agar plates inoculated withfungal spores. They were in duplicates and were incubated for seven days. Using thediameter of the growth free area around the filter paper circles, the resistance or thesusceptibility of the fungi to each fungicide was determined.Most of the fungi were susceptible to dithane and thiophenate-methyl, thus these two werechosen for determination of the growth interference of fungicides on tissue cultures. Thesetwo fungicides were added to the tissue culture media after autoclaving the medium at ()(control), 20ppm and IOOOppmconcentrations. Twenty-five replicates per treatment wereused. After six weeks of incubation, the total numher of newly produced juvenile leaves,percentage or fungal contamination and the percentage number of surviving explants wereobserved. There was a significant difference between the two fungicides used and alsobetween the concentrations used. The interference of the fungicide to the growth of thetissue was minimal when thiophenate-methyl was added at IOppm concentration. Higherconcentrations retard the plant growth significantly.

HABITATS OF Anoectochilus setaceus, Zeuxine flava, Zeuxine regia IN KANNELIYA MAN AND BIOSPHERE RESERVE AND PEAK WILDERNESS SANCTUARY IN SRI LANKA

Anoectochilus setaceus, Zeuxine jlava, and Zeuxine regia are threeendangered medicinal plants belong to family Orchidaceae and sub familyNeottioideae. They are widely used in traditional medicine. Due to itsbeautiful variegated leaves they are also used as ornamental plants andcommonly known as Jewel Orchids.The Methodology used to identify the natural distribution of these specieswas field observation in the sites. In order to identify the places where thesespecies are growing knowledge of the traditional practitioners and villagepeople were used. According to literature, these species are confined totropical wet evergreen, sub montane and mid country wet ever green forests.In this study these three species were observed in particular locations inKanneliya MAB reserve and Peak Wilderness sanctuary.Anoectochilus setaceus is a rather common species found under the shade oftrees among fallen leaves. It was found along the riverbanks in Kanneliyawhile in Peak Wilderness it was found in a valley close to a stream. Theywere confined to small patches with high humidity where it gets very lowintensity of sunlight. Number of plants, which were observed in PeakWilderness sanctuary, was high (150 plants/rrr') while it was lesser (50plants/rrr') in Kanneliya. Distance between two forest patches where A.setaceus was found was about 50m in Peak Wilderness sanctuary while itwas too far (> 100m) in Kanneliya MAB reserve. Some patches had adistance of about 2km.Zeuxine regia was found in disturbed sites in Kanneliya MAB along the trailcloser to village, which situated at the boundary of the forest. In PeakWilderness also this species was found closer to the main trail starts fromSiripagama. These plants were found in places where there is no stream evenwithin 500m. Z. regia was found among rocks where environmentalconditions were very harsh but the soil was rather wet.Zeuxine flava was observed along a trail situated within the village inKanneliya far away from sites where other two species were distributed.Natural abundance is comparatively low for this species. In Peak Wildernessit was found along the same trail with Z. regia and A. setaceus but indifferent pockets. Environmental conditions of Z. flava are totally differentwhen compared with the sites where other two species were found.Anoectochilus setaceus was observed under same environmental conditionsrecorded by previous researchers, while other two species were found fromentirely different environmental conditions from recorded data. Althoughliterature reports that, Z. regia found together with A. setaceus under naturalconditions, such combinations were not observed in both forests. They werefound in entirely different locations

In vitro callus induction of Spilanthes calva DC [Spilanthes acmella auct. non L,.] (Maha Akmella)

Spilanthes calva DC. (Maha Akrnella) is a valuable medicinal plant belongs to Family Asteraceae. Itis widely used in indigenous medicine to treat toothache in most of the Asian countries. Not only it hasanesthetic properties, but also contain secondary metabolites, with the insecticidal properties, whichcould be used as potential bio insecticide. This is an annual plant, which grows to a height about 30ern. After flowering mother plant is dried off. Four to six weeks later seeds are germinated and newseedl ings are produced. Viabil ity of seeds loses with in short period of time. Even though seeds aregerminated percentage of germination is low (about 30%). Rooting of cuttings is also not possible.This is a limitation in using this valuable medicinal plant for commercial production. Therefore it isvery important to develop a protocol for mass propagation through tissue culture and establishing cellcultures will be useful for large-scale chemical extraction in industrial purposes.Leaf discs were used as explant for callus initiation. In order to identify the suitable maturity stage forcallus initiation, leaves were harvested at different maturity stages i.e first, second and third fullyopened leaf.Leaves were washed with Dettol" soap and soaked in a solution of Teepol" for 5 minutes. After thatleaves were washed with running tap water for 45 minutes, In order to surface sterilize. Leaves werewashed with 10% Clorox ™ (5.25% Sodium hypochlorite v/v) for 5 minutes and then with 70% alcoholfor 30 seconds each followed by three successive washings in sterile distilled water. These operationswere carried out inside the laminar airflow cabinet before inoculation. Basal media tested for the study were full strength MS (Murashige and Skoog, 1962) medium and Y2 MS (both macro andmicronutrients) medium. Media were supplemented with different concentrations (1.0 mgl' - 3.0mgl") of BAP and 2,4-0. Cultures were incubated under complete dark at 25± I °C in the growthroom.Study conducted by Haw and Keng (2003) on the same species produced multiple shoots from axillarybud explants without inducing callus in MS medium supplemented with 2.0 mgl.:' BAP. In the presentstudy, callusing was observed within 5 days of incubation in full strength MS medium supplementedwith BAP and 2AO. It took longer period to initiate callus when both macro and micro nutrients in thebasa l rned ium was lowered to ha If and the amount of callus produced was also very low even after 6thweek of incubation. In order to observe the time taken to produce maximum amount callus freshwe iuht was measured after 2".1,4th and 6tltweek of incubation. It was observed that maximum amountofc~llus was produced within 4 weeks in all explant types tested with a maximum of 0.88 g:': 0.23 inleaf discs obtained from first fully opened leaf.In order to determine the best growth regulator combination for callus initiation, calli fresh weightswere measured after fourth week of incubation in different growth regulator combinations tested.Highest amount of calli were in MS medium in the presence of2.25 mgl' BAP and 1.0 mgl' 2.4-0.Fragile calli, which were transulant and mucilaginous in nature were observed within 15 days ofincubation, which could lead to cell suspension cultures. 

IN VITRO SHOOT INITIATION OF Artocarpus heterophyllus Lam. (Jack fruit). Moraceae

Artocarpus heterophyllus is recognized as a versatile multipurpose tree species in alltropical countries. Its popularity is mainly due to its value as timber, food, fodder,fuelwood and a vast array of medicinal and other non-wood products.Seed propagation is the cheapest, easiest and most convenient method but hasdisadvantages in that the progeny may differ from the parent plant due to the high level ofcross-pollination. Vegetative propagation is very useful but has a relatively poor successrate. Developing a tissue culture technique for clonal propagation of the species will bevery useful for qualitative improvement of the plant with in vitro mutagenesis and cellhybridization.Four types of explants (mature embryos, apical meristems of young seedlings, apices frommature plants and nodal segments) were used in order to initiate shoots in vitro. Theembryos from seeds soaked in water for 24 hours produced shoots after 8 weeks ofincubation and the success rate was 50% while embryos from dry seeds only producedroots. There was no significant effect of cold storage (refrigeration) on shoot initiationfrom mature embryos.It has been found 88% of young apical meristems produced shoots in Campbell andDurzan medium compared to 60% in Murashige and Skoog medium. Only 1/ 3 ofproduced multiple shoots. Shoot initiation from nodal segments was a rare event. Callushas been produced from mature apices. Removal of the sheathing cover around the apexenhanced the shoot initiation from mature apices

In vitro propagation of Kaempferia galanga (L)

Kaempferia galanga (L) is an aromatic perennial herb, which is widely used in Ayurvedic medicine.Dry tubers are imported in large scaie to Sri Lanka due to lack of mass production in Sri Lanka.Disease susceptibility and higher cost of production have restricted its cultivation. Propagation ofKaempferia galanga is normally by rh izorne cuttings but disease susceptibi Iity of tender rh izomesrestricts propagation in large scale. Propagation through other vegetative methods is not possible.Rahman et al. (2004) reported the possibility of obtaining plants through somatic embryogenesis butthe survival rate was low. Therefore an attempt was made to develop a protocol for mass propagationof Kaempferia galanga through direct organogenesis.Leaf discs and axi Ilary buds were used as explants. Axi Ilary buds isolated from rh izomes of Kaempferiagalanga mother plants were sprayed with 0.2% Captan ™ 2-3 days before collection. After that, theywere placed on a wet paper Iined tray and covered again with another wet paper. Five to six dayslater young axillary buds were emerged from nodes and they were used as explants. For leaf discexplants leaves were washed with soap and soaked in a solution of Teepol" for 15 minutes, andwashed with running tap water for 45 minutes.Both leaf discs and axillury buds were dipped in 5% Chlorox ™ (5.25% Sodium hyperclorite v/v) for10-15 minutes under sterile conditions. Then they were washed 10% Clorox r for 3 minutes and 70%ethanol for one minute each followed by two successive washings in sterile distilled water. Explantswere cultured on MS basal medium (Murashige and Skoog, 1962) supplemented with differentconcentrations of Benzyl amino purine (BAP) and Indole acetic acid (IAA) (2.00 mgl' - 2.25 rngl'and 0.30 mgl-I- 0.70 mgi' respectively). Sucrose 3% (w/v) and 0.8% agar were added to the media.pH was adjusted to 5.8.Cultures were incubated under 16 hr light 18 hr dark at 26 :.+:: I DCtemperature for 21 days. Callusingwas not observed from both tested explants in any of the media tested. After 15 -18 days of incubationaxillary buds were elongated in all combinations tested. MS supplemented with 2.25 rngl' BAP and0.5 rngl' IAA showed the highest elongation (490 ± 10 mrn).After 25-30 days of incubation in vitro grown shoots were cut and separated from the explant. Thenthey were subcultured on the same medium and incubated in 16 hr light at 26 ± 1 DCtemperature forshoot multiplication. MS medium was used as basal medium with above combinations of growthregulators.The highest multiplication was observed in 2.25 mgl:' BAP and 0.5 mg' IAA (7.0 ±..0.02) shoots perexplant. Further sub culturing on to the same medium induced roots. Seven-weeks old plantlets wereremoved from culture vessels, washed well to remove all agar and transferred to small plastic potscontaining sand, soil and compost in I: I: 1 proportion by volume and kept in shade house covered withpolythene bags, for acclimatization. 100% survival was observed when acclimatized plants weretransferred to the field.

EFFECT OF THE PERIOD OF THE YEAR WHICH EXPLANTS WERE COLLECTED ON SHOOT GROWTH AND THE CALLUS FORMATION OF THE TISSUE CULTURED JACK FRUIT

A method for rapid propagation of mature Jack fruit from apical meristem culture wasdeveloped. Apical meristems were estahlished in Modified Campbell and Durzan mediumsupplemented with NAA and ISA. Cultures were suh cultured in every 4 weeks interval inorder to reduce the accumulation of phenolic compounds. Reducing the accumulatedphenolics at the base of the explant enhanced the growth rate. There was a significantdifference in the growth performance of shoots and callus produced according to the periodof the year in which explants were collected.60% of the apical meristems cultured in modified CD medium supplemented with IBA andNAA produced shoots in November to December period. It was only 30% when the apiceswere cultured in April to May months and decreased to 20% in June - July months. Theshoots produced in November - December period showed a higher vigour (in number ofleaves per shoot. mean leaf width and mean shoot length) than those produced in othermonths. Since jackfruit show seasonal changes in fruit bearing and shedding of leaves. itcan be suggested that the difference in growth performances of tissues cultured in artificialculture media would have been effected by endogenous rhythms. It has been observed thatthe callus production depends on the incubation temperature. Callus was induced at thebase of the shoot hy increasing the incubation temperature from 2S± IIIC to 30± Ifie.Growth of the callus was also retarded hy accumulated phenolic compounds in themedium.

Shrimp waste management Use of dried papaya milk in chitosan manufacture

Chitin is the second most abundant carbon biopolymer on earth, next to cellulose. It is the majorconstituent in the exo-skeleton of crustacean water animals such as shrimps, crabs etc. Shrimp wasteis a major cause for environmental pollution in shrimp cultivating areas such as Puttalam. Currentannual shrimp production in Sri Lanka is about 4000 MT and the shrimp waste produced is about1200 MT. This shrimp waste at present is discharged into environment or buried without any treatment,thereby causing serious environmental pollution problems.Chitosan, which can be obtained from chitin by chemical treatment, is a polysaccharide of very higheconorn ic importance with a wide range of industrial applications. If Sri Lanka can convert its shrimpwaste in to chitosan, it can be a major foreign exchange earner, in addition to solving the problem ofenvironmental pollution caused by shrimp waste.A method for the production of chitosan from shrimp waste using dried papaya milk (OPM) has beendeveloped (Sri Lanka Patent No 13544,2005). It involves the treatment of demineralised (with 4%HCI) shrimp waste with OPM followed by deproteinization with 3% NaOH and deacetylation with50% NaOH. The use ofOPM brings about a 25% reduction in the amount ofNaOH, which is knownto cause environmental pollution problems. Typically, the degree of deacetylation (~O) of resultingchitosan was 66% comparable to DO of conventional methods. Moisture content (11.2%) and ashcontent (0.69%) of resulting chitosan were also comparable to those obtained by 100% chemicalmethods.

Testing the gaugino AMSB model at the Tevatron via slepton pair production

Gaugino AMSB models-- wherein scalar and trilinear soft SUSY breaking terms are suppressed at the GUT scale while gaugino masses adopt the AMSB form-- yield a characteristic SUSY particle mass spectrum with light sleptons along with a nearly degenerate wino-like lightest neutralino and quasi-stable chargino. The left- sleptons and sneutrinos can be pair produced at sufficiently high rates to yield observable signals at the Fermilab Tevatron. We calculate the rate for isolated single and dilepton plus missing energy signals, along with the presence of one or two highly ionizing chargino tracks. We find that Tevatron experiments should be able to probe gravitino masses into the ~55 TeV range for inoAMSB models, which corresponds to a reach in gluino mass of over 1100 GeV.Comment: 14 pages including 6 .eps figure

A comparative study of non-covalent encapsulation methods for organic dyes into silica nanoparticles

Numerous luminophores may be encapsulated into silica nanoparticles (< 100 nm) using the reverse microemulsion process. Nevertheless, the behaviour and effect of such luminescent molecules appear to have been much less studied and may possibly prevent the encapsulation process from occurring. Such nanospheres represent attractive nanoplatforms for the development of biotargeted biocompatible luminescent tracers. Physical and chemical properties of the encapsulated molecules may be affected by the nanomatrix. This study examines the synthesis of different types of dispersed silica nanoparticles, the ability of the selected luminophores towards incorporation into the silica matrix of those nanoobjects as well as the photophysical properties of the produced dye-doped silica nanoparticles. The nanoparticles present mean diameters between 40 and 60 nm as shown by TEM analysis. Mainly, the photophysical characteristics of the dyes are retained upon their encapsulation into the silica matrix, leading to fluorescent silica nanoparticles. This feature article surveys recent research progress on the fabrication strategies of these dye-doped silica nanoparticles

Exclusive Breastfeeding Practices in Relation to Social and Health Determinants: a Comparison of the 2006 and 2011 Nepal Demographic and Health Surveys

Background: Exclusive breastfeeding (EBF) for the first six months can have a significant impact on reducing child morbidity and mortality rates. The objective of this study was to compare the determinants of and trends in EBF in infants =5 months from the 2006 and 2011 Nepal Demographic and Health Surveys. Methods: Data on mother/infant pairs having infants of =5 months from 2006 (n = 482) and 2011 (n = 227) were analysed. The EBF rate, determinants of EBF, and changes in EBF rates between the 2006 and 2011 surveys were examined using Chi-square test and multiple logistic regression. Results: The EBF rate for =5 months in 2006 was 53.2% (95% CI, 47.1%-59.3%) and 66.3% (95% CI, 56.6%-74.8%) in 2011. In 2006, infants =4 months were more likely to be EBF [(aOR) 3.086, 95% CI (1.825-5.206)] after controlling for other factors. A geographic effect was also found in this study, with the odds of EBF higher for infants from the Hills [aOR 3.426, 95% CI (1.568-7.474)] compared to those form the mountains. The odds of EBF were also higher for higher order infants [aOR 1.968, 95% CI (1.020-3.799)]. Infants whose fathers belonged to non-agricultural occupation were less likely to be provided with EBF. Infants who were delivered in the home were more likely to experience EBF [aOR 1.886; 95% CI (1.044-3.407)]. In 2011, infants of age =4 months were more likely [aOR 4.963, 95% CI (2.317-10.629)] to have been breastfed exclusively. While there was an increase in the EBF rate between 2006 and 2011 surveys, the significant increase was noticed only among the infants of four months [32.0%; 95% CI (19.9%-47.0%)] in 2006 to [65.5%; 95% CI (48.1-79.6)] in 2011.Conclusions: The proportion of infants who were EBF was higher in Nepal in 2011survey compared to 2006 survey; however, this is still below the recommended WHO target of 90%. Infant’s age, ecological region, parity and father’s occupation were associated with EBF. Further interventions such as peer counselling, antenatal counselling and involving fathers in the community to promote EBF in Nepal are recommended