Determination of the Structure of the Catabolic N-Succinylornithine Transaminase (AstC) from Escherichia coli

Escherichia coli possesses two acyl ornithine aminotransferases, one catabolic (AstC) and the other anabolic (ArgD), that participate in L-arginine metabolism. Although only 58% identical, the enzymes have been shown to be functionally interchangeable. Here we have purified AstC and have obtained X-ray crystal structures of apo and holo-AstC and of the enzyme complexed with its physiological substrate, succinylornithine. We compare the structures obtained in this study with those of ArgD from Salmonella typhimurium obtained elsewhere, finding several notable differences. Docking studies were used to explore the docking modes of several substrates (ornithine, succinylornithine and acetylornithine) and the co-substrate glutamate/α-ketogluterate. The docking studies support our observations that AstC has a strong preference for acylated ornithine species over ornithine itself, and suggest that the increase in specificity associated with acylation is caused by steric and desolvation effects rather than specific interactions between the substrate and enzyme.     

Measurement of the Infection and Dissemination of Bluetongue Virus in Culicoides Biting Midges Using a Semi-Quantitative RT-PCR Assay and Isolation of Infectious Virus

Background

Culicoides biting midges (Diptera: Ceratopogonidae) are the biological vectors of globally significant arboviruses of livestock including bluetongue virus (BTV), African horse sickness virus (AHSV) and the recently emerging Schmallenberg virus (SBV). From 2006–2009 outbreaks of BTV in northern Europe inflicted major disruption and economic losses to farmers and several attempts were made to implicate Palaearctic Culicoides species as vectors. Results from these studies were difficult to interpret as they used semi-quantitative RT-PCR (sqPCR) assays as the major diagnostic tool, a technique that had not been validated for use in this role. In this study we validate the use of these assays by carrying out time-series detection of BTV RNA in two colony species of Culicoides and compare the results with the more traditional isolation of infectious BTV on cell culture.

Methodology/Principal Findings

A BTV serotype 1 strain mixed with horse blood was fed to several hundred individuals of Culicoides sonorensis (Wirth & Jones) and C. nubeculosus (Mg.) using a membrane-based assay and replete individuals were then incubated at 25°C. At daily intervals 25 Culicoides of each species were removed from incubation, homogenised and BTV quantified in each individual using sqPCR (C_q values) and virus isolation on a KC-C. sonorensis embryonic cell line, followed by antigen enzyme-linked immunosorbent assay (ELISA). In addition, comparisons were also drawn between the results obtained with whole C. sonorensis and with individually dissected individuals to determine the level of BTV dissemination.

Conclusions/Significance

C_q values generated from time-series infection experiments in both C. sonorensis and C. nubeculosus confirmed previous studies that relied upon the isolation and detection of infectious BTV. Implications on the testing of field-collected Culicoides as potential virus vectors by PCR assays and the use of such assays as front-line tools for use in diagnostic laboratories in this role are discussed.     

Development of a multi-assay approach for monitoring coral diversity using eDNA metabarcoding

Coral Reefs - Cumulative anthropogenic pressures have triggered a global decline in the health of marine ecosystems, and coral reefs, in particular, are in crisis. With climate and...     

Character displacement drives floral variation in Pelargonium (Geraniaceae) communities

Interactions between plant community members are an underexplored driver of angiosperm floral variation. We investigate character displacement as a potential contributor to floral variation in Pelargonium communities. Pelargoniums all place pollen on the ventral sides of their pollinators, potentially leading to interspecific pollen transfer (IPT) in sympatry. We show that the positions of pollen placement and receipt are determined by anther and style exsertion lengths. Using field experiments, we demonstrate that heterospecific species experience higher IPT if they have similar style lengths than when they have greater style length differences. Using crosses, we show that IPT has negative consequences on seed set. In combination, these results suggest that character displacement in style length is likely to reduce IPT and increase female fitness in sympatry. Patterns of style length variation across 29 different Pelargonium communities suggest that character displacement has occurred in multiple communities. Furthermore, analyses using a wide-ranging species pair show that style lengths are more different between sympatric populations than they are between allopatric populations. In addition to pollinators as agents of floral divergence, this study suggests that variation in Pelargonium community structure has driven style length variation through character displacement.     

Variation in abundance and harvest of sooty shearwaters (Puffinus griseus) by Rakiura Maori on Putauhinu Island,New Zealand

Abstract

Sooty shearwater (Puffinus griesus, titi) abundance, harvest levels and chick mass were monitored repeatedly on Putauhinu Island, south‐west of Rakiura (Stewart Island) between 1997 and 2005. Putauhinu is the second largest of the Titi Islands and has a relatively high density of chicks distributed over most of the island, so it supports what is likely the second‐largest population of sooty shearwaters in the Rakiura region (after Taukihepa, Big South Cape Island). Rakiura Maori harvested chicks from five “manu” (family birding areas) that covered 56% of the 128.4 ha of breeding colony of the island. Chick density was lower on the unharvested area in the interior of the island than on harvested areas. Burrow entrance density was higher where there was more ground cover (mainly fern) vegetation, but these areas had lower burrow occupancy, so overall chick density was similar at different levels of ground cover. Twenty‐six harvesters present on Putauhinu in 2005 took 31 280 chicks in total, equivalent to 8.4% (95% CI = 6.6–12%) of the available chicks on the entire island. Seasonal variation in total chicks harvested (CV 15–22%) was not related to chick abundance or mass. Refuges, including impenetrable patches of vegetated ground within manu, the unharvested centre of the island, and even nearby unharvested islands, will ameliorate localised impacts of harvest if density‐dependent immigration is operating.     

Conservation status of New Zealand frogs, 2009

Abstract

A reappraisal of the conservation status of the New Zealand frog fauna is presented using the 2008 version of the New Zealand Threat Classification System. Of New Zealand's four extant endemic species, three are judged to be ‘Threatened’ (Leiopelma hamiltoni being ‘Nationally Critical’, and L. pakeka and L. archeyi being ‘Nationally Vulnerable’) and one ‘At Risk’ (L. hochstetteri ‘Declining’). Three Leiopelma species are listed as extinct—they are known from bone deposits in caves throughout the country until some time in the last 1000 years. Three introduced and naturalised Litoria species are abundant in New Zealand although two (L. aurea and L. raniformis) are threatened in their country of origin (Australia). An additional unidentified frog taxon from northern Great Barrier Island is listed as ‘Data Deficient’.     

Byssal proteins of the freshwater zebra mussel,Dreissena polymorpha

The freshwater zebra mussel (Dreissena polymorpha) is a notorious biofouling organism. It adheres to a variety of substrata underwater by means of a proteinaceous structure called the byssus, which consists of a number of threads with adhesive plaques at the tips. The byssal proteins are difficult to characterize due to extensive cross-linking of 3,4-dihydroxyphenylalanine (DOPA), which renders the mature structure largely resistant to protein extraction and immunolocalization. By inducing secretion of fresh threads and plaques in which cross-linking is minimized, three novel zebra mussel byssal proteins were identified following extraction and separation by gel electrophoresis. Peptide fragment fingerprinting was used to match tryptic digests of several gel bands against a cDNA library of genes expressed uniquely in the mussel foot, the organ which secretes the byssus. This allowed identification of a more complete sequence of Dpfp2 (D. polymorpha foot protein 2), a known DOPA-containing byssal protein, and a partial sequence of Dpfp5, a novel protein with several typical characteristics of mussel adhesive proteins.