Projection pursuit nonparametric regression applied to field counts of the aphid Macrosiphum euphorbiae (Homoptera: Aphididae) on tomato crops in greenhouses

The number of plants that sampling requires in aphid population studies often exceeds one hundred. Thus, only quick and nondestructive methods can be used to sample this pest in a reasonable time interval. We propose a visual method for estimating the density of the aphid Macrosiphum euphorbiae (Thomas) on tomato plants reared in greenhouses. After approximately 1min of visual observation plants can be assigned to abundance classes, the boundaries of which are roughly the powers of square root 10. Precise counts were collected simultaneously on sets of reference plants from the same greenhouse. Projection pursuit nonparametric regression was then used to provide unbiased estimates of aphid densities from the abundance classes and several easily gathered explanatory variables. The robustness of the method was evaluated by testing the models on the complementary data sets from plants in which the aphid densities were precisely counted. In both single and twin-row cultural conditions, for the reference and complementary data sets, the order of magnitude of the error was less than one class rank per plant. The investigation time was reduced by approximately 10-fold compared with the exact counting method. This easy-to-teach field method could be useful in large-scale population surveys and for optimizing integrated pest management strategies.     

Substitution bias, rapid saturation, and the use of mtDNA for nematode systematics

Only relatively recently have researchers turned to molecular methods for nematode phylogeny reconstruction. Thus, we lack the extensive literature on evolutionary patterns and phylogenetic usefulness of different DNA regions for nematodes that exists for other taxa. Here, we examine the usefulness of mtDNA for nematode phylogeny reconstruction and provide data that can be used for a priori character weighting or for parameter specification in models of sequence evolution. We estimated the substitution pattern for the mitochondrial ND4 gene from intraspecific comparisons in four species of parasitic nematodes from the family Trichostrongylidae (38-50 sequences per species). The resulting pattern suggests a strong mutational bias toward A and T, and a lower transition/transversion ratio than is typically observed in other taxa. We also present information on the relative rates of substitution at first, second, and third codon positions and on relative rates of saturation of different types of substitutions in comparisons ranging from intraspecific to interordinal. Silent sites saturate extremely quickly, presumably owing to the substitution bias and, perhaps, to an accelerated mutation rate. Results emphasize the importance of using only the most closely related sequences in order to infer patterns of substitution accurately for nematodes or for other taxa having strongly composition-biased DNA. ND4 also shows high amino acid polymorphism at both the intra- and interspecific levels, and in higher level comparisons, there is evidence of saturation at variable amino acid sites. In general, we recommend using mtDNA coding genes only for phylogenetics of relatively closely related nematode species and, even then, using only nonsynonymous substitutions and the more conserved mitochondrial genes (e.g., cytochrome oxidases). On the other hand, the high substitution rate in genes such as ND4 should make them excellent for population genetics studies, identifying cryptic species, and resolving relationships among closely related congeners when other markers show insufficient variation.      

Phylogeny of the Drosophila saltans species group based on combined analysis of nuclear and mitochondrial DNA sequences

Nucleotide sequences from two nuclear loci, alcohol dehydrogenase and internal transcribed spacer-1 of the nuclear ribosomal DNA repeats, and two mitochondrial genes, cytochrome oxidase I and cytochrome oxidase II, were determined from nine species in the Drosophila saltans species group. The partition homogeneity test and partitioned Bremer support were used to measure incongruence between phylogenetic hypotheses generated from individual partitions. Individual loci were generally congruent with each other and consistent with the previously proposed morphological hypothesis, although they differed in level of resolution. Since extreme conflict between partitions did not exist, the data were combined and analyzed simultaneously. The total evidence method gave a more resolved and highly supported phylogeny, as indicated by bootstrap proportions and decay indices, than did any of the individual analyses. The cordata and elliptica subgroups, considered to have diverged early in the history of the D. saltans group, were sister taxa to the remainder of the saltans group. The sturtevanti subgroup, represented by D. milleri and D. sturtevanti, occupies an intermediate position in this phylogeny. The saltans and parasaltans subgroups are sister clades and occupy the most recently derived portion of the phylogeny. As with previous morphological studies, phylogenetic relationships within the saltans subgroup were not satisfactorily resolved by the molecular data.      

Phylogenetic analysis supports horizontal transfer of P transposable elements

Nucleotide sequence comparisons were used to investigate the evolution of P transposable elements and the possibility that horizontal transfer has played a role in their occurrence in natural populations of Drosophila and other Diptera. The phylogeny of P elements was examined using published sequences from eight dipteran taxa and a new, partial sequence from Scaptomyza elmoi. The results from a number of different analyses are highly consistent and reveal a P-element phylogeny that contradicts the phylogeny of the species. At least three instances of horizontal transfer are necessary to explain this incongruence, but other explanations cannot be ruled out at this time.      

Novel bacterial molybdenum and tungsten enzymes: three-dimensional structure, spectroscopy, and reaction mechanism

The molybdenum enzymes 4-hydroxybenzoyl-CoA reductase and pyrogallol-phloroglucinol transhydroxylase and the tungsten enzyme acetylene hydratase catalyze reductive dehydroxylation reactions, i.e., transhydroxylation between phenolic residues and the addition of water to a triple bond. Such activities are unusual for this class of enzymes, which carry either a mononuclear Mo or W center. Crystallization and subsequent structural analysis by high-resolution X-ray crystallography has helped to resolve the reaction centers of these enzymes to a degree that allows us to understand the interaction of the enzyme and the respective substrate(s) in detail, and to develop a concept for the respective reaction mechanism, at least in two cases.     

Hybridization dependent cleavage of internally modified disulfide-peptide nucleic acids

Selectivity of the cleavage of single stranded over hybridized forms of internally modified disulfide-peptide nucleic acids (PNA) has been optimized using a series of phosphines and thiols, which have different sizes and charges. For the most selective cleaver found (tris-(carboxyethyl)-phosphine), reactivity of single stranded PNA is 33 times higher than that of the PNA-DNA duplex. Selectivity of single stranded disulfide-PNA cleavage has been explained in terms of electrostatic interaction between the substrate and the cleaver.     

Dearomatizing benzene ring reductases

The high resonance energy of the benzene ring is responsible for the relative resistance of aromatic compounds to biodegradation. Nevertheless, bacteria from nearly all physiological groups have been isolated which utilize aromatic growth substrates as the sole source of cell carbon and energy. The enzymatic dearomatization of the benzene nucleus by microorganisms is accomplished in two different manners. In aerobic bacteria the aromatic ring is dearomatized by oxidation, catalyzed by oxygenases. In contrast, anaerobic bacteria attack the aromatic ring by reductive steps. Key intermediates in the anaerobic aromatic metabolism are benzoyl-CoA and compounds with at least two meta-positioned hydroxyl groups (resorcinol, phloroglucinol and hydroxyhydroquinone). In facultative anaerobes, the reductive dearomatization of the key intermediate benzoyl-CoA requires a stoichiometric coupling to ATP hydrolysis, whereas reduction of the other intermediates is readily achieved with suitable electron donors. Obligately anaerobic bacteria appear to use a totally different enzymology for the reductive dearomatization of benzoyl-CoA including selenocysteine- and molybdenum- containing enzymes.     

Mechanism of carbon tetrachloride-induced hepatotoxicity. Hepatocellular damage by reactive carbon tetrachloride metabolites

CCl4-induced liver damage was modeled in monolayer cultures of rat primary hepatocytes with a focus on involvement of covalent binding of CCl4 metabolites to cell components and/or peroxidative damage as the cause of injury. (1) Covalent binding of 14C-labeled metabolites was detected in hepatocytes immediately after exposure to CCl4. (2) Low oxygen partial pressure increased the reductive metabolism of CCl4 and thus covalent binding. (3) [14C]-CCl4 was bound to lipids and to proteins throughout subcellular fractions. Binding occurred preferentially to triacylglycerols and phospholipids, with phosphatidylcholine containing the highest amount of label. (4) The lipid peroxidation potency of CCl4 revealed subtle differences compared to other peroxidative substances, viz., ADP-Fe3+ and cumol hydroperoxide, respectively. (5) CCl4, but not the other peroxidative substances, decreased the rate of triacylglycerol secretion as very low density lipoproteins. (6) The anti-oxidant vitamin E (alpha-tocopherol) blocked lipid peroxidation, but not covalent binding, and secretion of lipoproteins remained inhibited. (7) The radical scavenger piperonyl butoxide prevented CCl4-induced lipid peroxidation as well as covalent binding of CCl4 metabolites to cell components, and also restored lipoprotein metabolism. The results confirm that covalent binding of the CCl3* radical to cell components initiates the inhibition of lipoprotein secretion and thus steatosis, whereas reaction with oxygen, to form CCl3-OO*, initiates lipid peroxidation. The two processes are independent of each other, and the extent to which either process occurs depends on partial oxygen pressure. The former process may result in adduct formation and, ultimately, cancer initiation, whereas the latter results in loss of calcium homeostasis and, ultimately, apoptosis and cell death.     

Differential purification of methionine-tRNA synthetase and lysine-tRNA synthetase from rabbit liver

Purification procedures for methionine-tRNA synthetase and lysine-tRNA synthetase from rabbit liver cytosol are presented. The purification factor for both enzymes is higher than that reported previously. The data suggest that methionine-tRNA synthetase and lysine-tRNA synthetase occur as a mixture in rabbit liver cytosol rather than as a complex.