Gene Cloning and Biochemical Characterization of an Alcohol Dehydrogenase from Euglena gracilis1

ABSTRACT. Euglena gracilis is a freshwater free‐living organism able to grow with ethanol as carbon source; to facilitate this metabolism several alcohol dehydrogenase (ADH) activities have been detected. We report the gene cloning, over‐expression, and biochemical characterization of a medium‐chain NAD⁺‐dependent ADH from E. gracilis (EgADH). The enzyme's amino acid sequence displayed the highest percentages of similarity and identity with ADHs of bacteria and fungi. In the predicted three‐dimensional model, all the residues involved in Zn²⁺, cofactor, and substrate binding were conserved. A conventional signal peptide for import into mitochondria could not be clearly identified. The protein of 37 kDa was over‐expressed, purified to homogeneity, and kinetically characterized. The enzyme's optimal pH was 7.0 for ethanol oxidation displaying a V_m of 11.7±3.6 U/mg protein and a K_m of 3.2±0.7 mM for this substrate. Isopropanol and isopentanol were also utilized, although with less efficiency. It showed specificity for NAD⁺ with a K_m value of 0.39±0.1 mM and Mg²⁺ or Zn²⁺ were essential for activity. The recombinant EgADH reported here may help to elucidate the roles that different ADHs have on the metabolism of short‐ and long‐chain alcohols in this microorganism.     

Fatty acids of leaf wax esters in Coincya Rouy (Brassicaceae)

A total of 46 populations belonging to the genus Coincya Ruoy (Brassicaceae) from the Iberian Peninsula was examined for their wax ester fatty acids composition. Size of ester fatty acids in the genus Coincya varies between 14 and 28 carbon atoms. This series is dominated by acids with an even number of carbon atoms. The most abundant acids are Ojg, C^y, C^^o and C₂₆ among evens and C₁₇ and C₂₁ among odds. A positive correlation was observed between chemical composition and environmental adaptation, and also with certain morphological characters. Changes relate to low temperatures or to high relative humidity, and there is a positive correlation between chemical composition and leaf glaucousness.     

Primary growth and morphological markers of interannual growth limits in Cupressaceae from Patagonia

The present study shows experimentally that primary growth of the native Patagonian Cupressaceae Austrocedrus chilensis (D. Don) Pic.-Serm et Bizz., Fitzroya cupressoides (Molina) I. M. Johnst. and Pilgerodendron uviferum (D. Don) Florin is potentially continuous. External morphological markers of interannual growth limits have been identified for the main axes of these species after several years of observation. Such limits correspond with axis portions in which internodes are relatively short and leaves have a small distal free end forming an acute angle with the axis. The causality of these results and their utility for studying the productivity, adaptation to different environments, population management and conservation of these threatened species are discussed.  © 2004 The Linnean Society of London, Botanical Journal of the Linnean Society, 2004, 146 , 285–293.     

Sex ratio, reproductive mode and genetic diversity in Triops cancriformis

1. Aquatic invertebrates display a wide array of alternative reproductive modes from apomixis to hermaphroditism and cyclical parthenogenesis. These have important effects on genetic diversity and population structure. Populations of the 'living fossil' Triops cancriformis display a range of sex ratios, and various reproductive modes are thought to underlie this variation. Using sex ratio information and histological analyses European populations have been inferred to be gonochoric (with separate males and females), selfing hermaphroditic and androdioecious, a rare reproductive mode in which selfing hermaphrodites coexist with variable proportions of males. In addition, some populations have been described as meiotic parthenogens.
2. Here we use population genetic analysis using microsatellite loci in populations with a range of sex ratios including a gonochoric population, and marker segregation patterns in heterozygote individuals reared in isolation, to clarify the reproductive mode in this species.
3. Our data show that populations in general have very low levels of genetic diversity. Non-gonochoric populations show lower genetic diversity, more heterozygote deficiencies, higher inbreeding coefficients and stronger linkage disequilibria than the gonochoric population. The maintenance of some heterozygosity in populations is consistent with some male influence in T. cancriformis populations, as would be expected from an androdioecious reproductive system. Results of marker segregation in eggs produced in isolation from non-gonochoric populations indicate that meiosis occurs and are consistent with two reproductive modes: selfing hermaphroditism and a type of ameiotic parthenogenesis.
4. Overall, our data indicate that androdioecy and selfing hermaphroditism are the most likely reproductive modes of non-gonochoric European Triops populations. Triops populations are strongly structured, suggesting high genetic drift and low levels of gene flow.     

Phylogeny of western Mediterranean Leptodirini, with an emphasis on genital characters (Coleoptera: Leiodidae: Cholevinae)

Abstract The tribe Leptodirini (Leiodidae: Cholevinae) is one of the largest radiations of Coleoptera in the subterranean environment. Although subjected to systematic and evolutionary studies, the phylogeny remains poorly understood. We assessed the phylogeny of the western Mediterranean lineages (Iberian Peninsula, Pyrenees and Sardinia) based on a cladistic analysis of fourteen characters of external morphology and twenty characters of the male and female genitalia, studied in 182 species belonging to thirty‐nine genera. We tested the monophyly of the traditional two main divisions of the group (infraflagellates and supraflagellates), as well as that of some ‘phyletic series’. The final matrix contained fifty‐eight terminal taxa, twenty‐four of which had different character state combinations. The strict consensus of the sixty most parsimonious trees recovered a monophyletic Leptodirini, but not their separation into infraflagellates and supraflagellates. The supraflagellates formed a paraphyletic group with respect to the infraflagellates (corresponding to our sampled ‘Speonomus’ series), with Notidocharis sister to all other included Leptodirini, and Speonomidius sister to Leptodirini excluding Notidocharis. The series ‘Spelaeochlamys’, including the Sardinian genera but excluding Pseudochlamys, was recovered as monophyletic with weak support. The ‘Quaestus’ series formed a polytomy with Pseudochlamys plus the ‘Speonomus’ series (including Bathysciola), which was recovered as monophyletic with strong support. Speonomus, Bathysciola, Quaestus and Troglophyes were para‐ or polyphyletic. Our results suggested the respective monophyletic origin of the Leptodirini from the Pyrenees (Pseudochlamys plus the ‘Speonomus’ series) and the Mediterranean coast plus Sardinia (series ‘Spelaeochlamys’). On the contrary, the Leptodirini of the Atlantic north coast of the Iberian Peninsula (series ‘Quaestus’ and ‘Speonomidius’) were not monophyletic.     

Population structure,diversity, and phylogeography in the near‐threatened Eurasian black vultures Aegypius monachus (Falconiformes; Accipitridae) in Europe: insights from microsatellite and mitochondrial DNA variation

The Eurasian black vulture (Aegypius monachus) has experienced a severe decline during the last two centuries and is globally classified as near‐threatened. This has led to the extinction of many traditional breeding areas in Europe and resulted in the present patchy distribution (Iberian and Balkan peninsulas) in the Western Palearctic. In the present study, we describe the current genetic status of the European populations using both mitochondrial cytochrome b sequences and nuclear microsatellite markers, comparing with those found in Asia (Mongolia and Caucasus region). Although, mitochondrial (mt)DNA revealed a relatively low genetic variability (haplotype diversity), no evidence of genome‐wide genetic erosion exists because nuclear diversity exhibits normal levels and strong differentiation. A highly philopatric dispersal behaviour must be invoked to explain the existence of a clear pattern that revealed by the phylogeographic analysis, which indicates a sharp East–West clinal distribution and an allopatric differentiation. The distribution of mtDNA haplotypes one in the Iberian population and two in Balkan population and the significance divergence at nuclear loci fulfill the definitions of those populations as evolutionary significant units. We discuss how management strategies should aim at the maintenance (or increase) of current genetic variability levels, suggesting that independent conservation plans are urgently required to protect these two breeding European populations from extinction. © 2008 The Linnean Society of London, Biological Journal of the Linnean Society, 2008, 95 , 859–872.     

Development and variation of the anuran webbed feet (Amphibia, Anura)

Webbed feet evolved convergently in most groups of aquatic tetrapods. However, extensive webbing is not always limited to an aquatic life style. In Anurans, hind limbs display great variation, including absence, of interdigital membranes, which is explained by differential growth rates of digital and interdigital tissues during early limb development. In order to explore web diversification in anurans, this paper presents analyses of: (1) hind limb early development and its relationship to the expression of interdigital membranes; (2) intraordinal variation of interdigital membranes in adult feet; and (3) intraordinal variation of metatarsal and digit lengths, including comments on metatarsal development. Study of limb development is carried out in larval series of 12 anuran species. Analysis of intraordinal variation comprises a sample of adults of 111 species. We recognize two configurations in the autopodium bud: (1) paddle-like shape with digits differentiated within the confines of interdigital tissues, and (2) pointed autopodium with digits differentiated beyond interdigital tissues. These early differences are conserved in adult morphology, in which allometry and isometry of digit IV (and metatarsal IV) with respect to other digits (and metatarsals) result in asymmetrical and paddle-like autopodium, respectively. The paddle-like autopodium is restricted to fossil and extant pipids and the hylids Pseudis and Lysapsus , whereas the asymmetrical one is present in most anurans. Both configurations seem to represent an early divergence of the autopodium shape. The paddle-like configuration observed in hylids appears as a reversion to an ancient condition that results from a conserved program of limb development.  © 2008 The Linnean Society of London, Zoological Journal of the Linnean Society , 2008, 152 , 39–58.     

A new species of Armeria (Plumbaginaceae) from southern Spain with molecular and morphometric evidence on its origin

The taxonomic status of the populations of Armeria from the Horconera massif (Cordoba province), previously misidentified or referred to without an explicit identification, is re-examined on the basis of morphological and molecular data. A multivariate analysis indicates that they can be discriminated from morphologically and geographically close taxa using morphometric characters. According to these results, and taking into account the previous evidence that supports extensive reticulation in the genus, a new species, A. trianoi , is formally described. Molecular evidence from our previous works, in particular the nuclear ribosomal ITS sequence data, provides the frame for a discussion on the origin of the new entity. It is suggested that A. trianoi may be the result of introgression of sympatric populations of A. villosa subsp. longiaristata into a pre-existing taxon, possibly A. filicaulis var. minor presently occurring in Sierra Tejeda some 50 km apart.     

A new aglaspidid arthropod from the Upper Cambrian of Tasmania

Abstract: The aglaspidid arthropod, Australaglaspis stonyensis gen. et sp. nov., is described from the Idamean (Upper Cambrian) of Stony Point, Montagu, north‐western Tasmania. The dorsal exoskeleton comprises a semicircular cephalon with a well‐defined marginal rim, acute genal angles, oval eyes, subtriangular glabella area and a subtrapezoidal hypostome. The trunk bears eleven somites with well‐developed pleural spines that progressively curve backwards. Paired postventral plates cover the last trunk tergites and the base of the tailspine, which is long and characterized by a medial cleft. Bilobed ventral impressions indicate the presence of homopodous appendages, forming a food groove along the trunk. The fossil assemblage and its mode of preservation suggest that Australaglaspis originally possessed a phosphatic cuticle, but chemical traces of it have been lost because of regional metamorphism and weathering. Australaglaspis appears to be closely related to the North American aglaspidid Chraspedops modesta Raasch, 1939, based on its medially cleft tailspine.     

Habitat effects on resource tracking ability: do wintering Blackcaps Sylvia atricapilla track fruit availability?

If resource availability shapes population distribution, changes in resource abundance should cause parallel changes in population numbers. However, tracking ability may be disrupted by different environmental and behavioural factors that act at different spatial and temporal scales. Here we analyse the ability of wintering Blackcap Sylvia atricapilla populations to track spatio-temporal variation in fruit availability in southern Spain in two habitats (forests and shrublands) with different population structure. Former studies had shown that forests are equally used by both adult migrant and local Blackcaps, whereas shrublands are nearly monopolized by juvenile migrants. These differences might affect resource tracking: it should be disrupted in forests, as local birds remain over winter in their breeding territories, but not in shrublands where similarly competitive juvenile migrants can freely track the spatial distribution of fruits. We analysed the fruit-tracking ability of Blackcap populations among sites and years in both habitat types using a habitat-matching model, which predicts spatio-temporal changes in population abundance proportional to changes in resource availability. We counted Blackcaps and fruiting shrubs (dominated by Lentiscs Pistacia lentiscus and Wild Olives Olea europaea sylvestris ) during four winters in forest and shrubland patches. The abundance of fruits was always higher in shrublands than in forests. In shrublands, Blackcaps seemed to move freely across fruit-rich habitat patches, tracking changes in fruiting-shrub abundance among sites and years. However, such tracking was not observed in forests. This supports the view that fruit-tracking ability may be constrained by local factors, such as the social structure of populations occurring in different habitat types, which introduces spatio-temporal variation in the way fruit availability shapes the abundance distribution of these birds in their Mediterranean wintering grounds.