Sterile Tissue Preparation and Callus Induction of Curcuma longa Linn.

Curcuma longa Linn. (family Zingiberaceae), commonly known as ‘turmeric’, is native to Southeast Asia. Turmeric has been used for color, flavor as a spice in cuisine and employed for treatment of various diseases. The major component in yellow-pigmented fraction of turmeric is curcuminoids. Curcuminoid production in callus of C. longa Linn. is our focus of study. Sterile techniques to obtain germ-free of C. longa Linn. explants were investigated and the results showed that immersing rhizome buds in 70% ethanol for 5 min, followed by 0.10% HgCl2 for 10 min offered approximately 66% survival rate. Multiple shoots were generated from the aseptic rhizome explants cultured on Murashige and Skoog (MS) agar medium fortified with 3.00 µM of 6-Benzylaminopurine (BA) and 0.50 µM of 1-Naphthaleneacetic acid (NAA) at 25 ± 2°C under a photoperiod of 16 h light and 8 h dark. The sterile leaf sheath and root were subsequently used for callus induction which produced various responses when cultured on MS agar medium fortified with different concentrations of 2,4-dichlorophenoxy acetic acid (2, 4-D), Thidiazuron (TDZ) and BA. The highest induction yields of friable callus were obtained from leaf sheath segments cultured on MS agar medium fortified with 0.50 mg/l 2, 4-D which are the conditions proposed for successful production of callus culture of C. longa Linn. Keywords: Callus induction, Curcuma longa Linn., Turmeric, Plant tissue culture

PHYTOCHEMICAL ANALYSIS OF CALLUS TWO VARIETIES ORTHOSIPHON ARISTATUS (BLUME) MIQ ON MURASHIGE AND SKOOG MEDIA: A STRATEGIC STEP OF SECONDARY METABOLITE PRODUCTION

Objective: The research aimed to provide new information regarding the secondary metabolites content of purple and white-purple Orthosiphon aristatus (Blume) Miq. callus, which can then be used as a basis for developing towards cell suspension and ultimately producing secondary metabolites using bioreactors. Methods: Callus induction of two varieties of O. aristatus were performed by inoculating sterile leaf explants grown on Murashige and Skoog basal media supplemented with 2,4-dichlorophenoxyacetis acid 0.4 ppm. The secondary metabolites were analysed and quantified using high-performance liquid chromatography with gradient elution. Results: The results showed the growth of callus two varieties of O. aristatus in growth media MS with 2,4-D 0.4 ppm. Rosmarinic acid content in the acetone extract of the purple variety callus was 1.28% w/w, and the white-purple variety was 2.22% w/w. Conclusion: This study could form the basis for the development of rosmarinic acid production by In vitro culture modification.

Cyanogenesis prospection in galled and non-galled tissues of Microgramma squamulosa (Polypodiaceae)

Cyanogenic glycosides are defense substances that can produce hydrocyanic acid when they undergo hydrolysis as a result of herbivory, a process called cyanogenesis. Galls are neoformed structures of plant tissues induced by species-specific interactions between an inducer organism and a host plant. Earlier studies in Microgramma species have demonstrated that has a variation in cyanogenesis within and between populations, as well as in different plant organs. Microgramma squamulosa is an epiphytic fern that may contain stem galls induced by Tortrimosaica polypodivora (Lepidoptera: Tortricidae). Thus, the aim of the present study was to assess cyanogenesis seasonally and in different tissues (galled and non-galled) of M. squamulosa. The study was conducted in populations located in the Rio de Janeiro state, Brazil. Cyanogenesis was assessed using the Feigl-Anger paper test. A total of 260 galled and non-galled tissues were analyzed, 45 gall samples, 67 sterile leaves, 103 stems and 2 croziers. Cyanogenesis was detected in only three sterile leaf samples. In none of the samples were the stems or galls cyanogenic. The results corroborate the hypothesis that the stems of Microgramma squamulosa galled by Tortrimosaica polypodivora are not cyanogenic.

Identifikasi Bakteri Endofit Daun Ficus Minahassae (Teijsm. & De Vriese) Miq. Berdasarkan Gen 16s rRNA

Bakteri endofit telah ditemukan hampir di semua tumbuhan yang telah diteliti. Bakteri-bakteri ini mengkolonisasi jaringan internal tumbuhan inang. Di Sulawesi Utara terdapat salah satu tumbuhan ara, yaitu Ficus minahassae, yang hanya tumbuh lokal di daerah ini dan Filipina. Tujuan dari penelitian ini yaitu untuk mengidentifikasi dari bakteri endofit yang mendiami daun F. minahassae. Isolasi bakteri endofit dilakukan dengan menebarkan ekstrak daun yang steril di atas permukaan media Nutrient Agar. Isolat murni yang diperoleh diidentifikasi menggunakan penanda gen 16S rRNA. Dari hasil penanaman ekstrak daun F. minahassae diperoleh dua isolat, yaitu YL1 yang koloninya berwarna kuning, dan YL2 yang koloningnya berwarna krem. Dengan melakukan proses BLAST di GenBank, YL1 memiliki kemiripan 100 % dengan Brachybacterium muris, sedangkan YL2 memiliki kemiripan 99 % dengan Pseudacidovorax intermedius. Analisis filogenetik yang dilakukan menggunakan metode UPGMA yang terintegrasi pada piranti lunak Geneious memperlihatkan perbedaan taksa dari bakteri endofit yang ditemukan pada daun F. minahassae.Endophytic bacteria have been found in virtually every plant studied. These bacteria colonize the internal tissue of the host plant. In North Sulawesi there is a fig plant named Ficus minahassae which is endemic to the area and the Philippines. This research was aimed to identify of the endophytic bacteria inhabit the endosphere of the leaf of F. minahassae. Isolation of endophytic bacteria was performed by spreading the sterile leaf extract onto NA media. The pure isolates were identified using 16S rRNA gene marker. There were two isolates, designated as YL1 and YL2, were isolate from the leaf of F. minahassae. The color of isolate YL1 was yellowish and YL2 was creme. Using BLAST nucleotide, YL1 showed 100 % similarity with Brachybacterium muris, and YL2 showed 99 % similarity with Pseudacidovorax intermedius. Phylogenetic analysis performed using UPGMA method integrated on Geneious software showed different taxa from endophytic bacteria found on F. minahassae leaf.

Pteris langsonensis (Pteridaceae), a new brake fern species from Lang Son Province, northern Vietnam

A new fern species, Pteris langsonensis belonging to the P. cadieri complex (Pteridaceae), is described and illustrated from Lang Son Province, northern Vietnam. P. langsonensis is most similar to P. plumbea, but the former has typical dimorphic leaves and fertile pinnae much contracted and with a width of no more than half of sterile pinnae, while the latter has slightly dimorphic leaves. The new species is also similar to P. cretica, but the former has smaller habit and slightly toothed sterile leaf margins.

Clarification of Zamia acuminata and a new Zamia species from Coclé Province, Panama

Zamia acuminata has remained an obscure, poorly understood species for over a century due to possibly misinterpreted or erroneous locality data on the unicate sterile type specimen, a very brief protologue description, the misidentification of the plants from El Valle de Antón in Panama as Z. acuminata, and the erroneous determinations of plants of Z. acuminata from Costa Rica as Z. fairchildiana. Recently collected material from San José Province in Costa Rica is here determined to be identical to the single sterile leaf material of the holotype of Zamia acuminata. We consider Z. acuminata to be a Costa Rican endemic species restricted to the western Talamanca mountain range in San José Province, and that the Zamia from El Valle de Antón in Panama, which has previously been referred to as Zamia acuminata, to be a new species, here described as Zamia nana.

Effect of Timing of Application of the Biological Control Agent Microsphaeropsis ochracea on the Production and Ejection Pattern of Ascospores by Venturia inaequalis

Field and in vitro trials were conducted to establish the influence of the biological control agent Microsphaeropsis ochracea on the ejection pattern of ascospores by Venturia inaequalis and on apple scab development, and to establish the best timing of application. The ejection pattern of ascospores was similar on leaves sprayed with M. ochracea and on untreated leaves. Fall application of M. ochracea combined with a delayed-fungicide program was evaluated in orchards with intermediate and high scab risk. For both orchards, it was possible to delay the first three and two infection periods in 1998 and 1999, respectively, without causing significant increase or unacceptable leaf and fruit scab incidence. To evaluate the best timing of application, sterile leaf disks were inoculated with V. inaequalis and then with M. ochracea 0, 2, 4, 6, 8, 10, 12, 14, and 16 weeks later. After incubation under optimal conditions for pseudothecia development, the number of ascospores was counted. Similarly, M. ochracea was sprayed on scabbed leaves on seven occasions from August to November 1999 and 2000. Leaves were overwintered on the orchard floor and ascospore production was evaluated the following spring. Ascospore production was reduced by 97 to 100% on leaf disks inoculated with M. ochracea less than 6 weeks after inoculation with V. inaequalis, but ascospore production increased with increasing period of time when M. ochracea was applied 8 to 16 weeks after the inoculation with V. inaequalis. In the orchard, the greatest reduction in production of ascospores (94 to 96% in 2000 and 99% in 2001) occurred on leaves sprayed with M. ochracea in August. The production of ascospores was reduced by 61 to 84% in 2000 and 93% in 2001 on leaves sprayed with M. ochracea in September, reduced by 64 to 86% in 2000 and 74 to 89% in 2001 on leaves sprayed in October, and reduced by 54 and 67% in 2000 and 2001, respectively, on leaves sprayed in November. It was concluded that M. ochracea should be applied in August or September and that ascospore maturation models and delayed-fungicide program could be used in orchards treated with this biological control agent.

Hormonal control of ear abortion in a stress-sensitive maize (Zea mays) inbred

A cold-sensitive maize (Zea mays L.) inbred was used as a model for investigating the interactions between growth regulators, reproductive development, and environmental stress. In this genotype, a chilling treatment given just before floral transition caused the topmost ear to abort and be replaced at maturity by a sterile, leaf-like, structure. Exogenous applications of the synthetic auxin 2,4-dichlorophenoxyacetic acid or of the cytokinin benzyladenine respectively mimicked or prevented the abortive response caused by chilling. Chilling also induced a moderate decrease in the content of endogenous indoleacetic acid (IAA) in the apical shoot tissues. By contrast, zeatin-type cytokinins decreased dramatically (5–8 fold), both in the apical shoot tissues and in the xylem exudate of chilled plants. Overall, the ratio of free-IAA to zeatin-cytokinins was increased in the apical shoot of chilled plants. Our results suggest that: (1) ear abortion induced by chilling might be related to an altered cytokinin content; (2) the number of developing ears may be limited by the endogenous levels of cytokinins just before floral transition; and (3) cytokinins may have a potential for increasing yield in maize.