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Multiple origins of crassulacean acid metabolism and the epiphytic habit in the Neotropical family Bromeliaceae.

The large Neotropical family Bromeliaceae presents an outstanding example of adaptive radiation in plants, containing a wide range of terrestrial and epiphytic life-forms occupying many distinct habitats. Diversification in bromeliads has been linked to several key innovations, including water- and nutrient-impounding phytotelmata, absorptive epidermal trichomes, and the water-conserving mode of photosynthesis known as crassulacean acid metabolism (CAM). To clarify the origins of CAM and the epiphytic habit, we conducted a phylogenetic analysis of nucleotide sequences for 51 bromeliad taxa by using the plastid loci matK and the rps16 intron, combined with a survey of photosynthetic pathway determined by carbon-isotope ratios for 1,873 species representing 65% of the family. Optimization of character-states onto the strict consensus tree indicated that the last common ancestor of Bromeliaceae was a terrestrial C(3) mesophyte, probably adapted to moist, exposed, nutrient-poor habitats. Both CAM photosynthesis and the epiphytic habit evolved a minimum of three times in the family, most likely in response to geological and climatic changes in the late Tertiary. The great majority of epiphytic forms are now found in two lineages: in subfamily Tillandsioideae, in which C(3) photosynthesis was the ancestral state and CAM developed later in the most extreme epiphytes, and in subfamily Bromelioideae, in which CAM photosynthesis predated the appearance of epiphytism. Subsequent radiation of the bromelioid line into less xeric habitats has led to reversion to C(3) photosynthesis in some taxa, showing that both gain and loss of CAM have occurred in the complex evolutionary history of this family.

Bromeliaceae↗

Purification of balansain I, an endopeptidase from unripe fruits of Bromelia balansae Mez (Bromeliaceae).

A new plant endopeptidase was obtained from unripe fruits of Bromelia balansae Mez (Bromeliaceae). Crude extracts were partially purified by ethanol fractionation. This preparation (redissolved ethanol precipitate, REP) showed maximum activity at pH 8.8-9.2, was very stable even at high ionic strength values (no appreciable decrease in proteolytic activity could be detected after 24 h in 1 M sodium chloride solution at 37 degrees C), and exhibited high thermal stability (inactivation required heating for 60 min at 75 degrees C). Anion exchange chromatography allowed the isolation of a fraction purified to mass spectroscopy, SDS-PAGE, and IEF homogeneity, named balansain I, with pI = 5.45 and molecular mass = 23192 (mass spectrometry). The purification factor is low (2.9-fold), but the yield is high (48.3%), a common occurrence in plant organs with high proteolytic activity, where proteases represent the bulk of protein content of crude extracts. Balansain I exhibits a similar but narrower pH profile than that obtained for REP, with a maximum pH value approximately 9.0 and was inhibited by E-64 and other cysteine peptidases inhibitors but not affected by inhibitors of the other catalytic types of peptidases. The alanine and glutamine derivatives of N-alpha-carbobenzoxy-L-amino acid p-nitrophenyl esters was strongly preferred by the enzyme. The N-terminal sequence of balansain I showed a very high homology (85-90%) with other known Bromeliaceae endopeptidases.

Amino Acid Sequence↗

Anti-allergic properties of the bromeliaceae Nidularium procerum: inhibition of eosinophil activation and influx.

New therapeutic approaches for the treatment of allergic diseases can be aided by the development of agents capable of regulating eosinophilic leukocytes. Here, we evaluated the anti-allergic properties of a crude extract of the Brazilian bromeliaceae Nidularium procerum, focusing on its effects on allergic eosinophilia. By studying allergic pleurisy in actively sensitized C57Bl/6 mice, we observed that pretreatment with N. procerum (2 mg/kg; i.p.) reduced pleural eosinophil influx triggered by allergen challenge. N. procerum was also able to reduce lipid body numbers found within infiltrating eosinophils, indicating that N. procerum in vivo is able to affect both migration and activation of eosinophils. Consistently, pretreatment with N. procerum blocked pleural eosinophil influx triggered by PAF or eotaxin, key mediators of the development of allergic pleural eosinophilia. The effect of N. procerum was not restricted to eosinophils, since N. procerum also inhibited pleural neutrophil and mononuclear cell influx. Of note, N. procerum failed to alter the acute allergic reaction, characterized by mast cell degranulation, oedema, and cysteinyl leukotriene release. N. procerum also had direct effects on murine eosinophils, since it inhibited both PAF- and eotaxin-induced eosinophil chemotaxis on an in vitro chemotactic assay. Therefore, N. procerum may be a promising anti-allergic therapy, inasmuch as it presents potent anti-eosinophil activity.

Animals↗

Hieronymain I, a new cysteine peptidase isolated from unripe fruits of Bromelia hieronymi Mez (Bromeliaceae).

A new peptidase, named hieronymain I, was purified to homogeneity from unripe fruits of Bromelia hieronymi Mez (Bromeliaceae) by acetone fractionation followed by cation exchange chromatography (FPLC) on CM-Sepharose FF. Homogeneity of the enzyme was confirmed by mass spectroscopy (MALDI-TOF), isoelectric focusing, and SDS-PAGE. Hieronymain is a basic peptidase (pI > 9.3) and its molecular mass was 24,066 Da. Maximum proteolytic activity on casein (>90% of maximum activity) was achieved at pH 8.5-9.5. The enzyme was completely inhibited by E-64 and iodoacetic acid and activated by the addition of cysteine; these results strongly suggest that the isolated protease should be included within the cysteine group. The N-terminal sequence of hieronymain (ALPESIDWRAKGAVTEVKRQDG) was compared with 25 plant cysteine proteases that showed more than 50% of identity.

Amino Acid Sequence↗

Candida bromeliacearum sp. nov. and Candida ubatubensis sp. nov., two yeast species isolated from the water tanks of Canistropsis seidelii (Bromeliaceae).

Strains belonging to two novel yeast species, Candida bromeliacearum and Candida ubatubensis, were isolated from the bromeliad tank of Canistropsis seidelii (Bromeliaceae) in a sandy coastal plain (restinga) ecosystem site in an Atlantic rainforest of south-eastern Brazil. These species were genetically distinct from all other currently accepted ascomycetous yeasts, based on sequence divergence in the D1/D2 domains of the large-subunit rDNA and in the small-subunit rDNA. The species occupy basal positions in the Metschnikowiaceae clade. The type strains are Candida bromeliacearum UNESP 00-103(T) (=CBS 10002(T)=NRRL Y-27811(T)) and Candida ubatubensis UNESP 01-247R(T) (=CBS 10003(T)=NRRL Y-27812(T)).

Brazil↗

Anti-inflammatory activity in the aqueous crude extract of the leaves of Nidularium procerum: A bromeliaceae from the Brazilian coastal rain forest.

Nidularium procerum LINDMAN, a common bromeliaceae from the Brazilian flora, remains poorly studied regarding its chemical and pharmacological properties. We have recently published that N. procerum has potent analgesic and anti-inflammatory activities. In the present work, we have investigated potential mechanisms involved in the anti-inflammatory effects of N. procerum aqueous extract on lipopolysaccharide (LPS)-, platelet activating factor (PAF)- or formyl-methionyl-leucyl-phenylalanine (fMLP)-induced pleurisy models of inflammation. We found that the aqueous extract of N. procerum leaves (leaf aqueous extract; LAE) inhibits the neutrophil migration, production of inflammatory cytokines interleukin-1 and -6 (IL-1 and IL-6) and the generation of prostaglandin E2 (PGE2) in LPS-induced pleural inflammation in mice. Such inhibitory effect of N. procerum on PGE2 generation was tightly correlated to the inhibition of formation of new cytoplasmic lipid bodies within recruited leukocytes. N. procerum also blocked the in vivo neutrophil influx induced by injection of PAF or fMLP into the mouse pleural cavity and directly inhibited PAF-induced neutrophil chemotaxis in vitro. The data obtained in this study indicate that N. procerum LAE exerts its anti-inflammatory effects by interfering with the capacity of the host to respond to injury at different levels. Among the different functions affected by N. procerum LAE, lipid body formation, PGE2 and cytokine production and neutrophil chemotaxis are readily evidenced in relevant surrogate models. The N. procerum bioactive profile makes it an attractive candidate for future development as a drug or phytomedicine.

Animals↗

Purification and characterization of macrodontain I, a cysteine peptidase from unripe fruits of Pseudananas macrodontes (Morr.) harms (Bromeliaceae).

A new papain-like cysteine peptidase isolated from fruits of Pseudananas macrodontes (Morr.) Harms, a species closely related to pineapple (Ananas comosus L.), has been purified and characterized. The enzyme, named macrodontain I, is the main proteolytic component present in fruit extracts and was purified by acetone fractionation followed by anion-exchange chromatography. Separation was improved by selecting both an adequate pH value and a narrow saline gradient. Optimum pH range (more than 90% of maximum activity with casein) was achieved at pH 6.1-8.5. Homogeneity of the enzyme was confirmed by bidimensional electrophoresis and mass spectroscopy (MS). Molecular mass of the enzyme was 23,459 (MS) and its isoelectric point was 6.1. The alanine, glutamine, and tyrosine derivatives were strongly preferred when the enzyme was assayed on N-alpha-CBZ-l-amino acid p-nitrophenyl esters. The N-terminal sequence of macrodontain (by comparison with the N-terminus of 30 plant proteases with more than 50% homology) showed a great deal of sequence similarity to the other pineapple-stem-derived cysteine endopeptidases, being 85.7, 85. 2, and 77.8% identical to comosain, stem bromelain, and ananain, respectively. It seems clear that the Bromeliaceae endopeptidases are more closely related to each other than to other members of the papain family, suggesting relatively recent divergence.

Amino Acid Sequence↗

Isolation and characterization of hieronymain II, another peptidase isolated from fruits of Bromelia hieronymi Mez (Bromeliaceae).

From unripe fruits of Bromelia hieronymi Mez (Bromeliaceae), a partially purified protease preparation was obtained by acetone fractionation of the crude extract. Purification was achieved by anionic exchange chromatography (FPLC) on Q-Sepharose HP followed by cationic exchange chromatography (SP-Sepharose HP). Homogeneity of the new enzyme, named hieronymain II, was confirmed by SDS-PAGE and mass spectroscopy (MALDI-TOF-TOF). The molecular mass of was 23,411 Da, and maximum proteolytic activity (more than 90% of maximum activity) was achieved at pH 7.5-9.0 on casein and at pH 7.30-8.3 on Z-Phe-Arg-p-nitroanilide. The enzyme was completely inhibited by E-64 and iodoacetic acid and activated by the addition of cysteine. The N-terminal sequence of hieronymain II (AVPQSIDWRVYGAV) was compared with those of 12 plant cysteine proteases which showed more than 70% of identity. Kinetic enzymatic assays were made on Z-Phe-Arg-p-nitroanilide (Km = 0.72mM, kcat = 1.82 seg(-1) , kcat/ Km = 2.54seg(-1) mM(-l)). No detectable activity could be found on PFLNA or Z-Arg-Arg-p-nitroanilide.

Amino Acid Sequence↗

6-Hydroxyluteolin-7-O-(1''-alpha-rhamnoside) from Vriesea sanguinolenta Cogn. and Marchal (Bromeliaceae).

The isolation of 6-hydroxyluteolin-7-O-(1"-alpha-rhamnoside) from the Central American epiphyte Vriesea sanguinolenta Cogn. and Marchal (Bromeliaceae) is described here. Its stereostructure was established by spectroscopic methods and an X-ray structure analysis of its hepta-O-acetyl derivative. This flavonoid glycoside had previously been reported from some Teucrium species (Labiatae), yet without sufficient physical data and spectroscopic evidence.

Animals↗

Betaine distribution in the Bromeliaceae.

Aerial parts of 38 taxa, distributed in 21 genera and all three subfamilies of the Bromeliaceae have been examined for the presence of betaines. Glycinebetaine and trigonelline were isolated from all the plants studied, although the yields of both compounds were low. For glycinebetaine, values (based upon dry weight) varied from 0.0004% for Pitcairnia corallina to 0.087% for Tillandsia usneoides. Trigonelline levels varied from 0.0003 to 0.022%; the highest yields were recorded for Billbergia rubicunda (0.022%) and Vriesia splendens (0.020%).

Journal Article↗

[Presence of Aedes aegypti in Bromeliaceae and plant breeding places in Brazil].

The frequency of Bromeliaceae and other plant breeding places where Aedes aegypti can be found is reported during two consecutive operational cycles (focal treatment) in the city of Rio de Janeiro. These cycles took place from November 12 2000 to March 9 2001 and from March 12 2001 to June 15 2001. This study concentrates on the epidemiological implications resulting from the growing use of these plants as decorative living objects.

Aedes↗

Triatoma ryckmani (Hemiptera: Reduviidae) in the epiphyte Tillandsia xerographica (Bromeliaceae) in the semiarid region of Guatemala.

For the first time, the reduviid bug Triatoma ryckmani Zeledón and Ponce (Hemiptera; Reduviidae) was recorded to inhabit the epiphyte Tillandsia xerographica Rohweder (Bromeliaceae) in the semiarid region of Guatemala. These bromeliads grow mainly in drought-resistant trees with rough bark such as Pereskia lychnidiflora (Cactaceae). In our study site, we investigated 30 T. xerographica, and 53 specimens of T. ryckmani were found. Most T. ryckmani (68.5%) were unfed. Ants (Formicidae) were the predominant (92.2%) insect taxon in T. xerographica. Other insects such as Blattidae (3.0%), Reduviidae (T. ryckmani: 2.5%), Blaberidae (2.2%), Gryllidae (0.1%), and Acrididae (0.1%) were recorded in the bromeliads. T. xerographica is illegally commercialized without previous inspection. This may cause accidental introduction of T. ryckmani to houses and to other countries.

Animals↗

The influence of air pollution on the phyllosphere microflora composition of Tillandsia leaves (Bromeliaceae).

The effect of air pollution on total phyllospheric microflora from two species of the epiphytic neotropical genus Tillandsia (Bromeliaceae) was studied by comparing unpolluted plants living in a forest (Escazú, San José) with polluted ones from an urban site of Costa Rica (San José city). Dilutions of homogenized leaf samples were plated on media suitable for each microbial group. For each microorganism group, total counts were performed and purified strains of randomly chosen colonies were identified. There was a global reduction in the number of living microorganisms due to pollution effects, especially yeasts and bacteria, while nitrogen-fixing microorganisms and fungi were less affected. Our results showed that the phyllosphere microflora of Tillandsia plants living in a tropical urban environment changes in terms of number and species composition of yeasts and bacteria with respect to plants living in unpolluted environment.

Air Pollution↗

[Various aspects of comparative leaf anatomy of 2 Bromeliaceae (Aechema mexicana Baker and Hechtia glomerata Zucc)].

The leaf anatomy of two Bromeliaceae, Hechtia glomerata Zucc., a terrestrial xerophyte of arid zones, and Acehmea mexicana Baker, a tropical moist forest epiphyte, is compared; these species show adaptations to different extremes of adverse environmental conditions as to water uptake. The leaves of A. mexicana have more water storage tissue than those of H. glomerata; on the other hand, H. glomerata is capable of root absorption. In A. mexicana, modified nonfunctional stomata increase transpiration resistance. Also, this species shows more specialized foliar trichomes which permit the uptake of water and possibly of nutrients. The balance between transpiration and absorption rates is regulated in both species by the combined presence of peltate scales and stomata. The degree of adaptation to drought presented by epiphytic species is more complicated than that shown by terrestrial species. Epiphytic plants like A. mexicana may be derived from xerophytic species that have undergone readaptation to a humid climate. The conspicuous negative geotropism in the leaves of A. mexicana causes the formation of basal deposits of water, which play an important functional role in survival.

Plants↗

Antinociceptive effect of Nidularium procerum: a Bromeliaceae from the Brazilian coastal rain forest.

Nidularium procerum, a common plant of the Brazilian flora, has not yet been studied for its pharmacological properties. We report here that extracts of N. procerum show both analgesic and anti-inflammatory properties. Oral (p.o.) or intraperitoneal (i.p.) administration of an aqueous crude extract from leaves of N. procerum (LAE) inhibited the writhing reaction induced by acetic acid (ED50 value = 0.2 mg/kg body weight, i.p.) in a dose-dependent manner. This analgesic property was confirmed in rats using two different models of bradykinin-induced hyperalgesia; there was 75% inhibition of pain in the modified Hargreaves assay, and 100% inhibition in the classical Hargreaves assay. This potent analgesic effect was not blocked by naloxone, nor was it observed in the hot plate model, indicating that the analgesic effect is not associated with the activation of opioid receptors in the central nervous system. By contrast, we found that LAE (0.02 microg/ml) selectively inhibited prostaglandin E2 production by cyclooxygenase (COX)-2, but not COX-1, which is a plausible mechanism for the analgesic effect. A crude methanol extract from the leaves also showed similar analgesic activity. An identical extract from the roots of N. procerum did not, however, block acetic acid-induced writhes, indicating that the analgesic compounds are concentrated in the leaves. Finally, we found that LAE inhibited an inflammatory reaction induced by lipopolysaccharide in the pleural cavity of mice.

Acetic Acid↗

A survey of genetic diversity and reproductive biology of Puya raimondii (Bromeliaceae), the endangered queen of the Andes.

Puya raimondii Harms is an outstanding giant rosette bromeliad found solely around 4000 m above sea level in the Andes. It flowers at the end of an 80 - 100-year or even longer life cycle and yields an enormous (4 - 6 m tall) spike composed of from 15,000 to 20,000 flowers. It is endemic and currently endangered, with populations distributed from Peru to the north of Bolivia. A genomic DNA marker-based analysis of the genetic structure of eight populations representative of the whole distribution of P. raimondii in Peru is reported in this paper. As few as 14 genotypes out of 160 plants were detected. Only 5 and 18 of the 217 AFLP marker loci screened were polymorphic within and among these populations, respectively. Four populations were completely monomorphic, each of the others displayed only one to three polymorphic loci. Less than 4 % of the total genomic variation was within populations and genetic similarity among populations was as high as 98.3 %. Results for seven cpSSR marker loci were in agreement with the existence of a single progenitor. Flow cytometry of seed nuclear DNA content and RAPD marker segregation analysis of progeny plantlets demonstrated that the extremely uniform genome of P. raimondii populations is not compatible with agamospermy (apomixis), but consistent with an inbreeding reproductive strategy. There is an urgent need for a protection programme to save not only this precious, isolated species, but also the unique ecosystem depending on it.

Bromeliaceae↗