Loopholes in the regulation of invasive species: genetic identifications identify mislabeling of prohibited aquarium plants

Numerous invasive aquatic species introductions can be traced to the aquarium trade. Many potentially harmful aquarium species may be difficult to identify based on morphology alone. As such, some prohibited or invasive species may be available for purchase if they are mislabeled as species without restrictions. Here we compare molecular identifications to internet vendors’ identifications for accessions of a popular genus of aquarium plants that are difficult to distinguish morphologically (Myriophyllum; watermilfoils). Specifically, we identified the extensive mislabeling of M. heterophyllum—an invasive species in the northeastern and western US. Furthermore, genotypes of M. heterophyllum found in our aquarium survey have also been found in invasive populations, suggesting their potential introduction through escape from aquaria, water gardens, or nurseries. Two additional taxa were sold under incorrect names. Finally, our survey revealed that Myriophyllum taxa present in the aquarium trade generally have poorly known distributions and ecologies, and therefore their invasive potential is unknown. Our study confirms that molecular identification methods can provide a valuable tool to survey commercial pathways for potentially harmful species that are otherwise difficult to identify.     

An evolutionary framework for association testing in resequencing studies

Sequencing technologies are becoming cheap enough to apply to large numbers of study participants and promise to provide new insights into human phenotypes by bringing to light rare and previously unknown genetic variants. We develop a new framework for the analysis of sequence data that incorporates all of the major features of previously proposed approaches, including those focused on allele counts and allele burden, but is both more general and more powerful. We harness population genetic theory to provide prior information on effect sizes and to create a pooling strategy for information from rare variants. Our method, EMMPAT (Evolutionary Mixed Model for Pooled Association Testing), generates a single test per gene (substantially reducing multiple testing concerns), facilitates graphical summaries, and improves the interpretation of results by allowing calculation of attributable variance. Simulations show that, relative to previously used approaches, our method increases the power to detect genes that affect phenotype when natural selection has kept alleles with large effect sizes rare. We demonstrate our approach on a population-based re-sequencing study of association between serum triglycerides and variation in ANGPTL4.     

Enhanced prostacyclin formation and Wnt signaling in sclerostin deficient osteocytes and bone

We show that prostacyclin production is increased in bone and osteocytes from sclerostin (Sost) knockout mice which have greatly increased bone mass. The addition of prostacyclin or a prostacyclin analog to bone forming osteoblasts enhances differentiation and matrix mineralization of osteoblasts. The increase in prostacyclin synthesis is linked to increases in β-catenin concentrations and activity as shown by enhanced binding of lymphoid enhancer factor, Lef1, to promoter elements within the prostacyclin synthase promoter. Blockade of Wnt signaling reduces prostacyclin production in osteocytes. Increased prostacyclin production by osteocytes from sclerostin deficient mice could potentially contribute to the increased bone formation seen in this condition.     

Immobilization of Escherichia coli JM103[pUC8] in kappa-carrageenan coupled with recombinant protein release by in situ cell membrane permeabilization.

Immobilization of Escherichia coli JM103[pUC8] was carried out with kappa-carrageenan as the support matrix. Substantial natural excretion of beta-lactamase, attributable to the less intact membrane of plasmid-harboring cells, was observed in immobilized cell cultures. Nevertheless, a significant portion of the beta-lactamase produced was retained in the cells. As compared to suspension cultures, much higher beta-lactamase activities, especially in the extracellular liquid, and much longer retention of plasmid-bearing cells (improved plasmid stability) were observed in immobilized cell cultures. Further enhancement in excretion of the recombinant protein (beta-lactamase) was achieved by permeabilization of cell membrane by periodic exposure of the immobilized cell cultures to ethylenediaminetetraacetic acid (EDTA). While the presence of EDTA led to some suppression of cell growth in suspension cultures, cell growth in gel beads was not affected by EDTA to the same extent, possibly due to lesser exposure of immobilized cells to EDTA. Exposure of immobilized cell cultures to EDTA presumably inhibited plasmid replication and led in turn to diversion of cellular resources for the support of expression of plasmid genes. Indeed, treatment of the immobilized cell cultures with EDTA resulted in increased production of beta-lactamase when compared to the enzyme production in EDTA-free cultures. More frequent addition of EDTA increased the period of retention of plasmid-bearing cells in these cultures but did not have any noticeable adverse effect on synthesis of beta-lactamase. Improvement in plasmid stability in EDTA-treated immobilized cell cultures was ascribed to the reduction in the growth rate differential between plasmid-free and plasmid-bearing cells, since plasmid-free cells were subject to more reduction in specific growth rate than were plasmid-bearing cells.     

Woody plant encroachment in granite barrens on the Frontenac Arch,eastern Ontario,Canada

Aims

Woody plant encroachment is a widespread phenomenon affecting treeless or sparsely treed habitats. We aimed to determine the extent and timing of tree and shrub encroachment into rock barrens of eastern Ontario over the last century, and to assess implications for their ongoing management.

Location

Queen's University Biological Station in the Frontenac Arch ecoregion.

Methods

We quantified the extent of change in woody vegetation in 290 rock barrens using aerial photography from 1925, 1965, and 2008. Composition and structure of woody plant communities in 10 barrens was subsequently quantified in the field using plot-based sampling. Cores or cross-sections were obtained from individuals >1.5 m height and dendrochronological techniques were used to determine their age and identify temporal patterns of any woody encroachment.

Results

Aerial photography indicated that the mean proportion of woody plant cover in barrens increased 22.5% from 1925 to 2008. Dendroecological analysis supported this. Few trees were present prior to 1900 and most established since 1960. Fraxinus americana, Juniperus virginiana, and Juniperus communis were the most common woody species colonizing the barrens. Remnants of large Pinus strobus stumps with extensive charring were found in 90% of the sampled barrens at a mean density of 22.6 stumps ha⁻¹.

Conclusions

Rock barrens on the Frontenac Arch have changed substantially over the past century; gradually being colonized by trees and shrubs and losing their distinctly open character. Active management — including prescribed fire and mechanical thinning — may be necessary if there is a desire to maintain these barrens and the rare species they support as components of the region's biodiversity. However, identification of a reference state for restoration is complicated by the fact that the structure and composition of these habitats were undoubtedly altered by European land clearance in the 19th century, and that some of these areas likely existed as pine woodlands before that.     

Uncovering the evolutionary origin of blue anthocyanins in cereal grains

Functional divergence after gene duplication plays a central role in plant evolution. Among cereals, only Hordeum vulgare (barley), Triticum aestivum (wheat) and Secale cereale (rye) accumulate delphinidin‐derived (blue) anthocyanins in the aleurone layer of grains, whereas Oryza sativa (rice), Zea mays (maize) and Sorghum bicolor (sorghum) do not. The underlying genetic basis for this natural occurrence remains elusive. Here, we mapped the barley Blx1 locus involved in blue aleurone to an approximately 1.13 Mb genetic interval on chromosome 4HL, thus identifying a trigenic cluster named MbHF35 (containing HvMYB4H, HvMYC4H and HvF35H). Sequence and expression data supported the role of these genes in conferring blue‐coloured (blue aleurone) grains. Synteny analyses across monocot species showed that MbHF35 has only evolved within distinct Triticeae lineages, as a result of dispersed gene duplication. Phylogeny analyses revealed a shared evolution pattern for MbHF35 in Triticeae, suggesting that these genes have co‐evolved together. We also identified a Pooideae‐specific flavonoid 3′,5′‐hydroxylase (F3′5′H) lineage, termed here Mo_F35H2, which has a higher amino acid similarity with eudicot F3′5′Hs, demonstrating a scenario of convergent evolution. Indeed, selection tests identified 13 amino acid residues in Mo_F35H2 that underwent positive selection, possibly driven by protein thermostablility selection. Furthermore, through the interrogation of barley germplasm there is evidence that HvMYB4H and HvMYC4H have undergone human selection. Collectively, our study favours blue aleurone as a recently evolved trait resulting from environmental adaptation. Our findings provide an evolutionary explanation for the absence of blue anthocyanins in other cereals and highlight the importance of gene functional divergence for plant diversity and environmental adaptation.     

Continued optimization of the MLPCN probe ML071 into highly potent agonists of the hM1 muscarinic acetylcholine receptor

This Letter describes the continued optimization of the MLPCN probe molecule ML071. After introducing numerous cyclic constraints and novel substitutions throughout the parent structure, we produced a number of more highly potent agonists of the M(1) mACh receptor. While many novel agonists demonstrated a promising ability to increase soluble APPα release, further characterization indicated they may be functioning as bitopic agonists. These results and the implications of a bitopic mode of action are presented.     

Dipsticks for rapid detection of Plasmodium in vectoring Anopheles mosquitoes

Malaria remains the most serious vector-borne disease, affecting some 300-500 million people annually, transmitted by many species of Anopheles mosquitoes (Diptera: Culicidae). Monoclonal antibodies developed against specific circumsporozoite (CS) proteins of the main malaria parasites Plasmodium falciparum and P. vivax have been used previously for enzyme-linked immunosorbent assays (ELISA), widely employed for detection of malaria sporozoites in vector Anopheles for local risk assessment, epidemiological studies and targeting vector control. However, ELISA procedures are relatively slow and impractical for field use. To circumvent this, we developed rapid wicking assays that identify the presence or absence of specific peptide epitopes of CS protein of the most important P. falciparum and two strains (variants 210 and 247) of the more widespread P. vivax. The resulting assay is a rapid, one-step procedure using a 'dipstick' wicking test strip. In laboratory assessment, dipsticks identified 1 ng/ mL of any of these three CS protein antigens, with sensitivity nearly equal to the CS standard ELISA. We have developed and are evaluating a combined panel assay that will be both qualitative and quantitative. This quick and easy dipstick test (VecTest Malaria) offers practical advantages for field workers needing to make rapid surveys of malaria vectors.     

Polymorphism analysis of the prion gene in BSE-affected and unaffected cattle

Polymerase chain reaction (PCR) primers designed to amplify the octapeptide repeat region of the bovine prion gene were used to test the association of genotypes with bovine spongiform encephalitis (BSE) in 56 BSE-affected and 177 unaffected animals. Three alleles (A, B, C) were detected as single-strand conformation polymorphisms (SSCPs) and two alleles (1,2 representing six or five copies of the octapeptide repeat respectively) were detected as amplified double-strand fragment length polymorphisms (AMFLPs). Observed genotypes of SSCPs and AMFLPs were analysed by x-square. The SSCP genotypes of nuclear family members of animals with BSE and BSE-affected animals were different (P < 0.001, P < 0.01) from unrelated animals of the same breed without BSE. No genotypic differences were found between the BSE-affected animals and their relatives (P > 0.469). No AMFLP genotypic differences were detected between BSE-affected animals, their relatives, unrelated animals of the same breed or animals of different breeds (P > 0.05). These data suggest that BSE-affected animals and their relatives are more likely to have the AA SSCP genotype than unrelated animals of the same breed or animals of different breeds.