Limnology of Crater Lakes in Los Tuxtlas,Mexico

Investigations on the abundance, biomass and position of heterotrophic flagellates (HF) in the benthic microbial food web of a melt water stream on King George Island, Antarctic Peninsula, were undertaken during the Antarctic summer from 23rd December 1997 until 13th March 1998. Abundance and biomass of potential HF resources (picophotoautotrophic and non-photoautotrophic bacteria) as well as potential predators on HF (ciliates and meiofauna) were also investigated. HF abundance ranged from approximately 9 × 10³ to 81 × 10³ cells cm^–3, values which fall into the same range as those found in lower latitudes. Numerically important benthic HF were euglenids, kinetoplastids, thaumatomastigids and especially chrysomonads. Most species identified have been shown to have a worldwide distribution. Abundance of the benthic ciliates ranged from 27 to 950 cells cm^–3. Mean bacterial abundance was 1.9 × 10⁷ and 5.2 × 10⁸ cells cm^–3 for picophotoautotrophic and non-photoautotrophic benthos, respectively. The well-developed microbial community was able to support the large number of nematods, gastotrichs, tardigrads and rotifers with abundances reaching more than 1000 individuals cm^–3. The largest portion of heterotrophic biomass was formed by the meiofauna with a mean of 63 g C cm^–3, followed by that of the heterotrophic bacteria with 4.80 g C cm^–3. Picophotoautotrophic bacteria contributed a mean of 1.37 g C cm^–3. HF and ciliates mean biomass was 0.61 and 1.99 g C cm^–3, respectively, with the HF biomass comprising between <10 and 70% of the total protozoan biomass. The data obtained in this study identify the melt water stream as a hot-spot of heterotrophic microbial and meiofaunal activity during the austral summer. The HF in the melt water stream formed a diverse group in terms of taxa and potential feeding types. Chrysomonads, kinetoplastids, euglenids and thaumatomastigida were the most abundant taxa. A classification into feeding types identified an average of 34% of the total HF as bacterivorous while all others were able to utilise other, larger organisms as resources. Potential trophic interactions between HF and bacteria and higher trophic levels are discussed.     

Tryptophan spectroscopy studies and black lipid bilayer analysis indicate that the oligomeric structure of Cry1Ab toxin from Bacillus thuringiensis is the membrane-insertion intermediate

During intoxication, the Cry protoxins must change from insoluble crystals into membrane-inserted toxins, which form ionic pores. Binding of Cry1A toxins to the cadherin receptor promotes the formation of a 250 kDa oligomer. In this work, we analyzed for the first time the structural changes presented by Cry1Ab toxin upon membrane insertion. Trp fluorescence of pure monomeric and oligomeric structures in solution and in a membrane-bound state was analyzed. Cry1Ab has nine Trp residues, seven of them in pore-forming domain I. Trp quenching analysis with iodide indicated that oligomerization caused a 27% reduction in the level of Trp exposed to the solvent. Most of the oligomeric structure (96%) inserts into the membrane as a function of the lipid:protein ratio, in contrast to the monomer (10%). Additionally, the membrane-associated oligomer presented a blue shift of 5 nm in lambda(max) of the emission spectrum, indicating a more hydrophobic environment for some Trp residues. In agreement with this, iodide was unable to quench the Trp of the membrane-bound oligomer, suggesting that a significant part of the protein may be buried in the membrane. Quenching analysis using brominated and spin-labeled phospholipids in the vesicles indicates that most of the Trp residues are located close to the membrane-water interface. Finally, ionic currents in black lipid bilayers revealed that the oligomeric structure has kinetics different from those of the monomer, producing stable channels with a high probability of being open in contrast to the monomer that exhibited unstable opening patterns. These data show that the oligomer, in contrast to the monomer, is able to interact efficiently with phospholipid membranes forming stable pores.     

Transmembrane reorientation of the substrate-binding site in vesicular acetylcholine transporter

Active transport of acetylcholine (ACh) by vesicular ACh transporter (VAChT) is driven by a proton-motive force established by V-ATPase. A published microscopic kinetics model predicts the ACh-binding site is primarily oriented toward the outside for nontransporting VAChT and toward the inside for transporting VAChT. The allosteric ligand [(3)H]vesamicol cannot bind when the ACh-binding site is outwardly oriented and occupied by ACh, but it can bind when the ACh site is inwardly oriented. The kinetics model was tested in the paper reported here using rat VAChT expressed in PC12(A1237) cells. Equilibrium titrations of [(3)H]vesamicol binding and ACh competition show that ATP blocks competition between vesamicol and ACh in over one-half of the VAChT. NaCl did not mimic ACh chloride, and bafilomycin A(1) and FCCP completely blocked the ATP effect, which shows that it is mediated by a proton-motive force. The data are consistent with reorientation of over one-half of the ACh-binding sites from the outside to the inside of vesicles upon activation of transport. The observations support the proposed microscopic kinetics model, and they should be useful in characterizing effects of mutations on the VAChT transport cycle.     

Low temperature regulates sucrose-phosphate synthase activity in Colobanthus quitensis (Kunth) Bartl. by decreasing its sensitivity to Pi and increased activation by glucose-6-phosphate

Colobanthus quitensis (Kunth) Bartl. is widely distributed from Mexico to the Antarctic. C. quitensis is a freezing resistant species that accumulates sucrose in response to cold. We tested the hypothesis that low temperature modifies the kinetic properties of C. quitensis sucrose phosphate synthase (SPS) to increase its activity and ability to synthesize sucrose during cold acclimation. Cold acclimation caused a fourfold increment in sucrose concentration and a 100% increase in SPS activity, without changes in the level of SPS protein. Cold acclimation did not affect the optimal temperature and pH for SPS activity. However, it caused a tenfold increase in the inhibition constant (K _i) for inorganic phosphate (Pi) calculated as a function of fructose-6-phosphate (Fruc-6-P). SPS from cold acclimated plants also exhibited a higher reduction of its Michaelis constant (K _m) for glucose-6-phosphate (Gluc-6-P) with respect to non-acclimated plants. We suggest that the increase in C. quitensis SPS K _i for Pi and the increase in activation by Gluc-6-P in response to cold keep SPS activated, leading to high sucrose accumulation. This may be an important adaptation that allows efficient accumulation of sucrose during the harsh Antarctic summer.     

Relationship between grain yield, osmotic adjustment and benzoxazinone content in Triticum aestivum L. cultivars

Fifteen wheat genotypes were grown under water deficit to ascertain the role of osmotic adjustment (OA) and the concentration of benzoxazinones in sustaining grain yield. A positive correlation between osmotic adjustment capacity and yield was observed in wheat genotypes cultivated under field conditions. The weight gain of plants exposed to drought was in agreement with the OA values (R(2) = 0.93). However, when wheat plants were infested by cereal aphids, this correlation was not found. The benzoxazinones 2,4-dihydroxy-1,4-benzoxa-zin-3-one (DIBOA) and 2,4-dihydroxy-7-methoxy-1,4 benzoxazin-3-one (DIMBOA) are defensive secondary metabolites present in wheat and others cereals. The content of these compounds varied in wheat genotypes and increased with drought and aphid infestation. A positive correlation between weight gain of irrigated-infested plants and drought-infested plants and the contents of benzoxazinones was observed. These results suggest that plants with better OA capacity and high benzoxazinone content should have better field yields.     

Hydroxamic acids in Secale cereale L. and the relationship with their antifeedant and allelopathic properties

Contents of the hydroxamic acids 2,4-dihydroxy-1,4-benzoxazin-3-one (DIBOA), and 2,4-dihydroxy-7-methoxy-1,4-benzoxazin-3-one (DIMBOA) in leaves and roots of 14 cultivars of rye, Secale cereale L., were determined. Dynamics of accumulation in three cultivars were evaluated. DIBOA was the main cyclic hydroxamic acid in leaves but the contents differed significantly between the cultivars. Both DIBOA and DIMBOA were present in the roots. Maximum concentration of DIBOA in leaves and DIMBOA in roots was reached between 48-54 h and 54-72 h after germination, respectively. Antifeedant activity of DIBOA towards the aphid Rhopalosiphum padi and the feeding behavior were studied by electronic recording in barley leaves treated with different contents of DIBOA. The deleterious activity of DIBOA could arise by starvation and/or a toxic effect. Additionally, allelopathic potential of pure DIBOA and aqueous extracts of leaves and roots of rye (Tetra-Baer) on the germination of lettuce (Lactuca sativa) and rye (Tetra-Baer) seeds was evaluated. A high percentage of germination inhibition of pure DIBOA and the extracts of leaves and roots was observed. The activity is in agreement with the contents of hydroxamic acids in the plants. The substrates had no allelopathic effect on rye seeds.     

Genetic variability of Spodoptera frugiperda Smith (Lepidoptera: Noctuidae) populations from Latin America is associated with variations in susceptibility to Bacillus thuringiensis cry toxins

Bacillus thuringiensis strains isolated from Latin American soil samples that showed toxicity against three Spodoptera frugiperda populations from different geographical areas (Mexico, Colombia, and Brazil) were characterized on the basis of their insecticidal activity, crystal morphology, sodium dodecyl sulfate-polyacrylamide gel electrophoresis of parasporal crystals, plasmid profiles, and cry gene content. We found that the different S. frugiperda populations display different susceptibilities to the selected B. thuringiensis strains and also to pure preparations of Cry1B, Cry1C, and Cry1D toxins. Binding assays performed with pure toxin demonstrated that the differences in the toxin binding capacities of these insect populations correlated with the observed differences in susceptibility to the three Cry toxins analyzed. Finally, the genetic variability of the three insect populations was analyzed by random amplification of polymorphic DNA-PCR, which showed significant genetic diversity among the three S. frugiperda populations analyzed. The data presented here show that the genetic variability of S. frugiperda populations should be carefully considered in the development of insect pest control strategies, including the deployment of genetically modified maize in different geographical regions.