An efficient protocol for micropropagation of <Emphasis Type="Italic">Melaleuca alternifolia</Emphasis> Cheel

Melaleuca alternifolia is cultivated for the production of an essential oil useful in the cosmetic and pharmaceutical industries. Despite the economic importance of this species, there is little knowledge about its in vitro propagation. The aim of this study was to establish an efficient protocol for micropropagation of M. alternifolia. With the goal of in vitro multiplication by axillary shoot proliferation, both solid and liquid MS and WPM media were tested with supplementation with BA at 0, 0.55, 1.11, 2.22, 3.33, and 4.44 μM. The best result for shoot multiplication was obtained when either 0.55 μM BA was added into solid MS medium or 1.11 μM BA was added into liquid MS medium, with 5.6 and 11.8 shoots per explant generated, respectively. On solid or liquid WPM medium supplemented with 0.55 μM BA, the proliferation rates were 5.5 and 4.7, respectively. Three auxins (NAA, IAA, and IBA) were tested at 0.53 and 2.64 μM during the rooting stage. Several sucrose concentrations (15, 30, and 45 g L⁻¹) were compared to a sucrose-free medium. Rooting performances on four culture media were then compared: MS, half-strength MS (MS/2), MS + activated charcoal (AC), and MS/2 + AC. The results showed that auxin addition to culture medium is not necessary for in vitro rooting. Rooted microcuttings from different culture media were acclimatized in a greenhouse, and the survival percentage was evaluated. All shoots cultured in an auxin-free MS medium supplemented with sucrose (30 g L⁻¹) produced roots, and all plants survived during acclimatization. Activated charcoal added in rooting medium reduced rooting rates.     

Somatic embryogenesis and plant regeneration in oil palm using the thin cell layer technique

An efficient procedure has been developed for inducing somatic embryogenesis and regeneration of plants from tissue cultures of oil palm (Elaeis guineensis Jacq.). Thin transverse sections (thin cell layer explants) of different position in the shoot apex and leaf sheath of oil palm were cultivated in Murashige and Skoog (MS) (Physiol Plant 15:473–497, 1962) medium supplemented with 0–450 μM picloram and 2,4-D with 3.0% sucrose, 500 mg L⁻¹ glutamine, and 0.3 g L⁻¹ activated charcoal and gelled with 2.5 g L⁻¹ Phytagel. Embryogenic calluses were evaluated 12 wk after inoculation. Picloram (450 μM) was effective in inducing embryogenic calluses in 41.5% of the basal explants. Embryogenic calluses were maintained on a maturation medium composed of basal media, plus 0.6 μM NAA and 12.30 μM 2iP, 0.3 g L⁻¹ activated charcoal, and 500 mg L⁻¹ glutamine, with subcultures at 4-wk intervals. Somatic embryos were converted to plants on MS medium with macro- and micronutrients at half-strength, 2% sucrose, and 1.0 g L⁻¹ activated charcoal and gelled with 2.5 g L⁻¹ Phytagel.     

Partial purification and characterization of trypsin-like proteinases from insecticide-resistant and -susceptible strains of the maize weevil,Sitophilus zeamais

Serine proteinases from three strains of Sitophilus zeamais (Coleoptera: Curculionidae), one susceptible and two resistant to insecticides — one exhibiting fitness cost (resistant cost strain) and the other lacking it (resistant no-cost strain), were partially purified using an aprotinin–agarose affinity column providing purification factors ranging from 36.5 to 51.2%, with yields between 10 and 15% and activity between 529 and 875 µM/min/mg protein with the substrate N-α-benzoyl-l-Arg-p-nitroanilide (L-BApNA). SDS-PAGE of the purified fraction revealed a 56,000 Da molecular mass band in all strains and a 70,000 Da band more visible in the resistant no-cost strain. The purified proteinases from all strains were inhibited by phenylmethyl sulphonyl fluoride (PMSF), N-α-tosyl-l-lysine chloromethyl ketone (TLCK), aprotinin, benzamidine and soybean trypsin inhibitor (SBTI) characterizing them as trypsin-like serine proteinases. Trypsin-like proteinases from the resistant strains exhibited higher affinity for L-BApNA. The resistant no-cost strain exhibited V_max-values 1.5- and 1.7-fold higher than the susceptible and resistance cost strains, respectively. A similar trend was also observed when using N-α-p-tosyl-L-Arg methyl ester (L-TAME) as substrate. These results provide support to the hypothesis that the enhanced serine proteinase activity may be playing a role in mitigating physiological costs associated with the maintenance of insecticide resistance mechanisms in some maize weevil strains.     

Synthesis, characterization, solid-state photo-luminescence and anti-tumor activity of zinc(II) 4′-phenyl-terpyridine compounds

Reactions between 4′-phenyl-terpyridine (L) and several Zn(II) salts (sulfate, nitrate, chloride or acetate) led to the formation of the complexes [Zn₂(μ-O₂SO₂)₂L₂(CH₃OH)₂] (1), [Zn(NO₃)L(H₂O)]NO₃ (2), [Zn(Cl)₂L] (3) and [Zn(CH₃COO)₂L] (4) which were characterized by IR, ¹H NMR and fluorescence spectroscopies, elemental analysis and single crystal X-ray diffraction. In the dinuclear molecule of 1 the Zn atom is hexacoordinated, with a N₃O₃ coordination environment and forms an octagonal ZnOSOZnOSO metallacycle. In the remaining structures, the metal atom is envisaged as possessing highly distorted N₃X₂ (X = O or Cl) square pyramid coordination geometries. The structure of 3 presents two different packing patterns which lead to distinct π-π stackings. In both structures 2 and 4, strong intermolecular hydrogen bonds were identified. The complexes exhibit promising in vitro tumor-inhibiting activities, which are higher than that of cisplatin, against the following human tumor cell lines: promyelocyticfina leukaemia (HL-60), hepatocellular carcinoma (Bel-7402), gastric carcinoma (BGC-823) and nasopharyngeal carcinoma (KB).     

Ground use by northern muriquis (Brachyteles hypoxanthus)

Many arboreal primates descend to the ground, a custom that may occur more frequently in disturbed habitats, and in the presence of researchers to whom the primates are habituated. In this paper, we describe opportunistic observations of ground use in two groups of northern muriquis (Brachyteles hypoxanthus) at the RPPN Feliciano Miguel Abdala, Minas Gerais, Brazil. Members of both groups were observed drinking, resting, feeding, playing, and traveling on the ground to different degrees, and variation in the levels of habituation of the two groups may be responsible for the differences in the distribution of their terrestrial activities. The potential increase in vulnerability to predation or disease owing to ground use has implications for the conservation of this critically endangered species.     

The 1,4-naphthoquinone scaffold in the design of cysteine protease inhibitors

A series of 1,4-naphthoquinone derivatives diversely substituted at C-2, C-3, C-5 and C-8, prepared by reaction of amines, amino acids and alcohols with commercial 1,4-naphthoquinones, has been evaluated against papain and bovine spleen cathepsin B. These 1,4-naphthoquinone derivatives were found to be irreversible inhibitors for both cysteine proteases, with second-order rate constants, k(2), ranging from 0.67 to 35.4M(-1)s(-1) for papain, and from 0.54 to 8.03M(-1)s(-1) for cathepsin B. Some derivatives display a hyperbolic dependence of the first-order inactivation rate constant, k(obs), with the inhibitor concentration, indicative of a specific interaction process between enzyme and inhibitor. The chemical reactivity of the compounds towards cysteine as a model thiol is dependent on the naphthoquinone LUMO energy, whereas papain inactivation is not. The 1,4-naphthoquinone derivatives are inactive against the serine protease, porcine pancreatic elastase.     

Identification of serine proteases from Leishmania braziliensis

Leishmania (V) braziliensis is one of the most important ethiologic agents of the two distinct forms of American tegumentary leishmaniasis (cutaneous and mucosal). The drugs of choice used in leishmaniasis therapy are significantly toxic, expensive and are associated with frequent refractory infections. Among the promising new targets for anti-protozoan chemotherapy are the proteases. In this study, serine proteases were partially purified from aqueous, detergent and extracellular extracts of Leishmania braziliensis promastigotes by aprotinin-agarose affinity chromatography. By zymography, the enzymes purified from the aqueous extract showed apparent activity bands of 60 kDa and 45 kDa; of 130 kDa, 83 kDa, 74 kDa and 30 kDa from the detergent extract; and of 62 kDa, 59 kDa, 57 kDa, 49 kDa and 35 kDa from the extracellular extract. All purified proteases exhibited esterase activity against Nalpha-benzoyl-L-arginine ethyl ester hydrochloride and Nalpha-p-tosyl-L-arginine methyl ester hydrochloride (serine protease substrates) and optimal activity at pH 8. 0. Proteases purified from the aqueous and extracellular extracts were effectively inhibited by benzamidine (trypsin inhibitor) and those from the detergent extract were inhibited by N-tosyl-L-phenyl-alanine chloromethyl ketone (chymotrypsin inhibitor) indicating that all these enzymes are serine proteases. These findings indicate that L. braziliensis serine proteases display some biochemical similarities with L. amazonensis serine proteases, demonstrating a conservation of this enzymatic class in the Leishmania genus. This is the first study to report the purification of a serine protease from Leishmania braziliensis.     

Interaction of the temperature with the parasite mite Acarophenax lacunatus (Cross & Krantz) (Prostigmata: Acarophenacidae) on the development of Rhyzopertha dominica (Fabricius) (Coleoptera: Bostrichidae)

The objective of this study was to evaluate the effect of different temperatures with the association of the mite Acarophenax lacunatus (Cross & Krantz) on the population suppression of Rhyzopertha dominica (Fabricius). The experimental units were petri dishes containing 40 g of whole wheat grains (13% moisture content) infested with 10 adults of R. dominica, under the temperatures of 20, 25, 30 and 35 masculineC, with or without A. lacunatus, in five replicates. Relative humidity of 60 +/- 5% and escotophase of 24h were used for all temperatures. Five days after the infestation, five mites were inoculated in each experimental unit. The evaluations were carried out at 20, 40, 60, 80, 100 and 120 days after R. dominica infestation. The interaction of the temperature with the parasite A. lacunatus was an important tool for the population suppression of R. dominica. In temperatures higher than 25 masculineC, however, despite the reduction of the immature stages of R. dominica, there was a high grain weight loss after 120 days. The maintenance of the temperature of the wheat grains stored at 20 masculineC can be used to complement the biological control of R. dominica with A. lacunatus.     

Biology of Acarophenax lacunatus (Cross & Krantz) (Prostigmata: Acarophenacidae) on Tribolium castaneum (Herbst) (Coleoptera: Tenebrionidae) and Cryptolestes ferrugineus (Stephens) (Coleoptera: Cucujidae)

The aim of the present study was to assess the effect of six different temperatures on the development of Acarophenax lacunatus (Cross & Krantz) using eggs of Tribolium castaneum (Herbst) and Cryptolestes ferrugineus (Stephens) as hosts. The temperature affected the development of A. lacunatus. The largest values for the progeny (19 mites in T. castaneum and 15 mites in C. ferrugineus) were obtained at about 30 degrees C, as also observed for the net reproductive rate (Ro), which revealed that the A. lacunatus population increased 18 times in T. castaneum and 14 times in C. ferrugineus in a generation. The intrinsic rate of increase (r m) gradually increased with temperature, reaching the maximum value at 35 degrees C in T. castaneum (1,608) and C. ferrugineus (1,289). The generation time was negatively correlated with temperature, ranging from 1,60 to 4,85 days in T. castaneum and from 1,96 to 5,34 days in C. ferrugineus. These results suggest that the mite A. lacunatus may be used in programs of biological control of T. castaneum and C. ferrugineus in the tropics.