Assessment of prey overlap between a native (Polistes humilis) and an introduced (Vespula germanica) social wasp using morphology and phylogenetic analyses of 16S rDNA

Abstract In newly invaded communities, interspecific competition is thought to play an important role in determining the success of the invader and its impact on the native community. In southern Australia, the native Polistes humilis was the predominant social wasp prior to the arrival of the exotic Vespula germanica (Hymenoptera: Vespidae). Both species forage for similar resources (water, pulp, carbohydrate and protein prey), and concerns have arisen about potential competition between them. The aim of this study was to identify the protein foods that these wasps feed on. As many prey items are masticated by these wasps to the degree that they cannot be identified using conventional means, morphological identification was complemented by sequencing fragments of the mitochondrial 16S rRNA gene. GenBank searches using blast and phylogenetic analyses were used to identify prey items to at least order level. The results were used to construct complete prey inventories for the two species. These indicate that while P. humilis is restricted to feeding on lepidopteran larvae, V. germanica collects a variety of prey of invertebrate and vertebrate origin. Calculated values of prey overlap between the two species are used to discuss the implications of V. germanica impacting on P. humilis. Results obtained are compared to those gained by solely 'conventional' methods, and the advantages of using DNA-based taxonomy in ecological studies are emphasized.     

Exposure and Health Risk from Swimming in Outdoor Pools Contaminated by Trichloroethylene

A method was developed that evaluated the exposure and health risk to children from swimming in outdoor pools filled with contaminated water. It was found that dermal absorption of trichloroethylene (TCE) was significantly larger than the inhalation component of the exposures. It was estimated that the inhalation route accounted for only 1% of the total exposure, whereas accidental ingestion was 7% of the child's total intake and the dermal absorbed dose was 92% of the total exposure. The relative percentage of the total exposure estimated for each exposure route indicated that the dermal exposure route and accidental ingestion of pool water should not be ignored for volatile compounds when evaluating exposure.

The method utilized was simple enough to use computer spreadsheets for the calculations and can be easily adapted to various swimming scenarios and age groups. This method also included an assessment of the uncertainty in the exposure and risk estimates. The range of estimated exposures was 40 μ g to 442 μ g of TCE per swimming season. All values in this range were below the health benchmark for both non-carcinogenic and carcinogenic endpoints.     

Characterization of thymic progenitors in adult mouse bone marrow

Thymic cellularity is maintained throughout life by progenitor cells originating in the bone marrow. In this study, we describe adult mouse bone cells that exhibit several features characteristic of prothymocytes. These include 1) rapid thymic engraftment kinetics following i.v. transplantation, 2) dramatic expansion of thymic progeny, and 3) limited production of hemopoietic progeny other than thymocytes. The adult mouse bone marrow population that is depleted of cells expressing any of a panel of lineage-specific Ags, stem cell Ag-1 positive, and not expressing the Thy1.1 Ag (Thy1.1(-)) (Thy1.1(-) progenitors) can repopulate the thymus 9 days more rapidly than can hemopoietic stem cells, a rate of thymic repopulation approaching that observed with transplanted thymocytes. Additionally, Thy1.1(-) progenitors expand prolifically to generate thymocyte progeny comparable in absolute numbers to those observed from parallel hemopoietic stem cell transplants, and provide a source of progenitors that spans multiple waves of thymic seeding. Nevertheless, the Thy1.1(-) population yields relatively few B cells and rare myeloid progeny posttransplant. These observations describe the phenotype of an adult mouse bone marrow population highly enriched for rapidly engrafting, long-term thymocyte progenitors. Furthermore, they note disparity in B and T cell expansion from this lymphoid progenitor population and suggest that it contains the progenitor primarily responsible for seeding the thymus throughout life.     

Distribution,levels, and activation of MEK1 in Alzheimer's disease

Extracellular-signal-regulated kinase (ERK) has been implicated in the pathogenesis of Alzheimer's disease (AD), but the upstream cascade leading to ERK activation has not been elucidated. In this study, we focused on one of the physiological activators of ERK, mitogen-activated protein kinase (MAPK)/ERK kinase 1 (MEK1). Although there was no significant difference in the level and distribution of total MEK1 between AD and age-matched control cases, increased levels of activated phospho-MEK1 were specifically localized to neuronal intracytoplasmic granular structures in severe AD (Braak stage V-VI). The considerable overlap between MEK1 and its downstream effector, phospho-ERK, suggests both a functional and mechanistic link. Nuclear localization of phospho-MEK1 was a prominent feature in both mild AD cases (Braak stage III-IV) and control cases with limited pathology (Braak stage I-II). Since MEK1 is normally cytoplasmic due to the active export from nucleus because of the presence of nuclear export signal in its amino-terminus, we suspect that the apparent nuclear accumulation of phospho-MEK1 in AD patients at early stages suggests that abnormal nuclear trafficking may contribute to the pathogenesis of AD. By immunoblot analyses, phospho-MEK1 was significantly increased in AD over control cases. Together, these findings lend further credence to the notion that the ERK pathway is dysregulated in AD and also indicate an active role for this pathway in disease pathogenesis.     

Accuracy and consistency in application of a probabilistic approach to reporting breast fine needle aspiration

OBJECTIVE: To determine the application of a probabilistic/categorical approach for reporting breast fine needle aspiration (FNA) and its dependence on the cytopathologist's level of experience. STUDY DESIGN: All breast surgical specimens that had preoperative breast FNA at our institution during a 3-year period were identified. The cytologic results were reported as 1 of 6 categories: positive, suspicious, atypical, epithelial proliferative, unremarkable and nondiagnostic, according to well-defined criteria. Five cytopathologists were responsible for all cytology sing-out during the study period. The histologic and cytologic diagnoses were correlated. RESULTS: A total of 297 cases were identified. Overall, there were no false positive cases (positive predictive value [PPV] = 100%). Two false negative cases (negative predictive value [NPV] = 96%) were due to sampling error. This indicates that the PPV and NPV for each of the 5 pathologists were also all 100% except for the 1 pathologist who had two false negative cases due to sampling errors. The probability of finding carcinoma on histology for suspicious and atypical cytologic categories ranged from 67% to 100% and 8% to 31%, respectively, for the individual pathologists. Fifteen cases were signed out by > or = 2 pathologists. The involvement of consultants was significantly associated with diagnosis (P = .02). Ten of the 15 cases were in the suspicious (5) or atypical (5) category. CONCLUSION: The probabilistic approach with defined diagnostic criteria is an accurate method and can be consistently applied in reporting breast FNA. Although use of the indeterminate (suspicious and atypical) categories is variable, a definite and considerable difference in the probability of carcinoma between these 2 categories was observed for all pathologists. The involvement of consultants did not move the cases out of these indeterminate categories.     

Epimerization at carbon-5' of (5'R)-[5'-2H]adenosylcobalamin by ribonucleoside triphosphate reductase: cysteine 408-independent cleavage of the Co-C5' bond

The adenosylcobalamin-dependent ribonucleoside triphosphate reductase (RTPR) from Lactobacillus leichmannii catalyzes the reduction of ribonucleoside triphosphates to deoxyribonucleoside triphosphates. RTPR also catalyzes the exchange of the C5'-hydrogens of adenosylcobalalamin with solvent hydrogen. A thiyl radical located on Cys 408 is generated by reaction of adenosylcobalamin at the active site and is proposed to be the intermediate for both the nucleotide reduction and the 5'-hydrogen exchange reactions. In the present research, a stereochemical approach is used to study the mechanism of the Co-C5' bond cleavage of adenosylcobalamin in the reaction of RTPR. When stereoselectively deuterated coenzyme, (5'R)-[5'-(2)H(1)] adenosylcobalamin (5'R/S = 3:1), was incubated with RTPR or the Cys 408 viariants, C408A-RTPR and C408S-RTPR in the presence of dGTP, the deuterium at the 5'-carbon was stereochemically scrambled, leading to epimerization of the (5'S)-[5'-(2)H(1)]- and (5'R)-[5'-(2)H(1)]-isotopomers. Observation of epimerization with mutated RTPR proves that transient cleavage of the Co-C5' bond occurs in the absence of the thiol group on Cys 408. The rate constants for epimerization by RTPR, C408A-RTPR, and C408S-RTPRs in the presence of dGTP are 5.1, 0.28, and 0.42 s(-1), respectively. Only the wild-type RTPR catalyzes the 5'-hydrogen exchange reaction. Both epimerization and 5'-hydrogen exchange reactions are stimulated by the allosteric effector dGTP, and epimerization is not detected in the absence of the effector. Mechanistic implications with respect to wt-RTPR-mediated carbon cobalt bond homolysis and the intermediacy of the 5'-deoxyadenosyl radical will be presented.     

Asymmetric organization of the pore region of the epithelial sodium channel

Epithelial sodium channels (ENaCs) are composed of three homologous subunits that have regions preceding the second transmembrane domain (also referred as pre-M2) that form part of the channel pore. To identify residues within this region of the beta-subunit that line the pore, we systematically mutated residues Gln(523)-Ile(536) to cysteine. Wild type and mutant mouse ENaCs were expressed in Xenopus oocytes, and a two-electrode voltage clamp was used to examine the properties of mutant channels. Cysteine substitutions of 9 of 13 residues significantly altered Li(+) to Na(+) current ratios, whereas only cysteine replacement of beta Gly(529) resulted in K(+)-permeable channels. Besides beta G525C, large increases in the inhibitory constant of amiloride were observed with mutations at beta Gly(529) and beta Ser(531) within the previously identified 3-residue tract that restricts K(+) permeation. Cysteine substitution preceding (beta Phe(524) and beta Gly(525)), within (beta Gly(530)) or following (beta Leu(533)) this 3-residue tract, resulted in enhanced current inhibition by external MTSEA. External MTSET partially blocked channels with cysteine substitutions at beta Gln(523), beta Phe(524), and beta Trp(527). MTSET did not inhibit alpha beta G525C gamma, although previous studies showed that channels with cysteine substitutions at the corresponding sites within the alpha- and gamma-subunits were blocked by MTSET. Our results, placed in context with previous observations, suggest that pore regions from the three ENaC subunits have an asymmetric organization.     

Creation of genome-wide protein expression libraries using random activation of gene expression

Here we report the use of random activation of gene expression (RAGE) to create genome-wide protein expression libraries. RAGE libraries containing only 5 x 10(6) individual clones were found to express every gene tested, including genes that are normally silent in the parent cell line. Furthermore, endogenous genes were activated at similar frequencies and expressed at similar levels within RAGE libraries created from multiple human cell lines, demonstrating that RAGE libraries are inherently normalized. Pools of RAGE clones were used to isolate 19,547 human gene clusters, approximately 53% of which were novel when tested against public databases of expressed sequence tag (EST) and complementary DNA (cDNA). Isolation of individual clones confirmed that the activated endogenous genes can be expressed at high levels to produce biologically active proteins. The properties of RAGE libraries and RAGE expression clones are well suited for a number of biotechnological applications including gene discovery, protein characterization, drug development, and protein manufacturing.     

Differential ANG II generation in plasma and tissue of mice with decreased expression of the ACE gene

We utilized mice with homozygous disruption of angiotensin-converting enzyme (ACE) (-/-), mice with heterozygous deletion of ACE (+/-), and wild-type mice (+/+) to test the hypothesis that genetic variation in ACE modulates tissue and plasma angiotensin (ANG) II concentrations. With the use of ANG I as substrate, kidney, heart, and lung ACE activity was reduced 80% in -/- mice compared with +/+ mice. However, ANG II concentrations and ANG II-to-ANG I ratios in the kidney, heart, and lung did not differ among genotypes. In contrast, plasma ANG II concentrations in -/- mice were <2 fmol/ml, whereas plasma ANG I concentrations were extremely high (765 fmol/ml). Chymase activity was increased 14-fold in the kidney (P < 0.05) and 1.5-fold in the heart (P < 0.05) of -/- versus +/+ mice but did not differ among genotypes in the lung. ANG II formation from enzymes other than ACE and chymase contributed <2% of total ANG II formation in all genotypes. These data suggest that ACE is essential to ANG II formation in the vascular space, whereas chymase may provide an important mechanism in maintaining steady-state ANG II levels in tissue.