Identification of functionally active aerobic methanotrophs in sediments from an arctic lake using stable isotope probing

Arctic lakes are a significant source of the greenhouse gas methane (CH₄), but the role that methane oxidizing bacteria (methanotrophs) play in limiting the overall CH₄ flux is poorly understood. Here, we used stable isotope probing (SIP) techniques to identify the metabolically active aerobic methanotrophs in upper sediments (0–1 cm) from an arctic lake in northern Alaska sampled during ice‐free summer conditions. The highest CH₄ oxidation potential was observed in the upper sediment (0–1 cm depth) with 1.59 µmol g wet weight^?1 day^?1 compared with the deeper sediment samples (1–3 cm, 3–5 cm and 5–10 cm), which exhibited CH₄ oxidation potentials below 0.4 µmol g wet weight^?1 day^?1. Both type I and type II methanotrophs were directly detected in the upper sediment total communities using targeted primer sets based on 16S rRNA genes. Sequencing of 16S rRNA genes and functional genes (pmoA and mxaF) in the ¹³C‐DNA from the upper sediment indicated that type I methanotrophs, mainly Methylobacter, Methylosoma, Methylomonas and Methylovulum miyakonense, dominated the assimilation of CH₄. Methylotrophs, including the genera Methylophilus and/or Methylotenera, were also abundant in the ¹³C‐DNA. Our results show that a diverse microbial consortium acquired carbon from CH₄ in the sediments of this arctic lake.     

Comparison of sex pheromone traps for monitoring pink hibiscus mealybug (Hemiptera: Pseudococcidae)

The pink hibiscus mealybug, Maconellicoccus hirsutus (Green) (Hemiptera: Pseudococcidae), is a highly polyphagous pest that invaded Florida in 2002 and has recently been reported from several locations in Louisiana. Although identification of its sex pheromone in 2004 improved monitoring capabilities tremendously, the effectiveness and efficiency of different pheromone trap designs for capturing males has not been evaluated. We deployed green Delta, Pherocon IlB, Pherocon V, Jackson, and Storgard Thinline traps in Homestead, FL, and compared the number of male M. hirsutus captured per trap, the number captured per unit of trapping surface area, the amount of extraneous material captured, and the time taken to count trapped mealybugs. Pheromone-baited traps with larger trapping surfaces (green Delta, Pherocon IIB, and Pherocon V) captured more males per trap than those with smaller surfaces (Jackson and Storgard Thinline), and fewest males were captured by Storgard Thinline traps. However, Jackson traps captured as many or more males per square centimeter of trapping surface as those with larger surfaces, and the time required to count males in Jackson traps was significantly less than in green Delta, Pherocon IIB, and Pherocon V traps. Although all trap designs accumulated some debris and nontarget insects, it was rated as light to moderate for all designs. Based on our measures of effectiveness and efficiency, the Jackson trap is most suitable for monitoring M. hirsutus populations. Additionally, unlike the other traps evaluated, which must be replaced entirely or inspected in the field and then redeployed, only the sticky liners of Jackson traps require replacement, enhancing the efficiency of trap servicing.     

Compartmental modeling of whole-body vitamin A kinetics in unsupplemented and vitamin A-retinoic acid-supplemented neonatal rats

Little is known about the contribution of different tissues to whole-body vitamin A (VA) kinetics in neonates. Here, we have used model-based compartmental analysis of tissue tracer kinetic data from unsupplemented (control) and VA-retinoic acid (VARA)-supplemented neonatal rats to determine VA kinetics in specific tissues under control and supplemented conditions. First, compartmental models for retinol kinetics were developed for individual tissues, and then an integrated compartmental model incorporating all tissues was developed for both groups. The models predicted that 52% of chylomicron (CM) retinyl ester was cleared by liver in control pups versus 22% in VARA-treated pups, whereas about 51% of VA was predicted to be extrahepatic in 4- to 6-day-old unsupplemented neonatal rats. VARA increased CM retinyl ester uptake by lung, carcass, and intestine; decreased the release into plasma of retinol that had been cleared by liver and lung as CM retinyl esters; stimulated the uptake of retinol from plasma holo-retinol binding protein into carcass; and decreased the retinol turnover out of the liver. Overall, neonatal VA trafficking differed from that previously described for adult animals, with a larger contribution of extrahepatic tissues to CM clearance, especially after VA supplementation, and a significant amount of VA distributed in extrahepatic tissues.     

Carbon Accumulation Patterns During Post-Fire Succession in Cajander Larch (Larix cajanderi) Forests of Siberia

Increased fire activity within boreal forests could affect global terrestrial carbon (C) stocks by decreasing stand age or altering tree recruitment, leading to patterns of forest regrowth that differ from those of pre-fire stands. To improve our understanding of post-fire C accumulation patterns within boreal forests, we evaluated above- and belowground C pools within 17 Cajander larch (Larix cajanderi) stands of northeastern Siberia that varied in both years since fire and stand density. Early-successional stands (<20-year old) exhibited low larch recruitment, and consequently, low density, aboveground larch biomass, and aboveground net primary productivity (ANPP_tree). Mid-successional stands (21- to 70-year old) were even-aged with considerable variability in stand density. High-density mid-successional stands had 21 times faster rates of ANPP_tree than low-density stands (252 vs. 12?g?C?m^?2?y^?1) and 26 times more C in aboveground larch biomass (2,186 vs. 85?g?C?m^?2). Density had little effect on total soil C pools. During late-succession (>70-year old), aboveground larch biomass, ANPP_tree, and soil organic layer C pools increased with stand age. These stands were low density and multi-aged, containing both mature trees and new recruits. The rapid accumulation of aboveground larch biomass in high-density, mid-successional stands allowed them to obtain C stocks similar to those in much older low-density stands (~8,000?g?C?m^?2). If fire frequency increases without altering stand density, landscape-level C storage could decline, but if larch density also increases, large aboveground C pools within high-density stands could compensate for a shorter successional cycle.     

A nesting of vipers: Phylogeny and historical biogeography of the Viperidae (Squamata: Serpentes)

Despite their medical interest, the phylogeny of the snake family Viperidae remains inadequately understood. Previous studies have generally focused either on the pitvipers (Crotalinae) or on the Old World vipers (Viperinae), but there has been no comprehensive molecular study of the Viperidae as a whole, leaving the affinities of key taxa unresolved. Here, we infer the phylogenetic relationships among the extant genera of the Viperidae from the sequences of four mitochondrial genes (cytochrome b, NADH subunit 4, 16S and 12S rRNA). The results confirm Azemiops as the sister group of the Crotalinae, whereas Causus is nested within the Viperinae, and thus not a basal viperid or viperine. Relationships among the major clades of Viperinae remain poorly resolved despite increased sequence information compared to previous studies. Bayesian molecular dating in conjunction with dispersal-vicariance analysis suggests an early Tertiary origin in Asia for the crown group Viperidae, and rejects suggestions of a relatively recent, early to mid-Tertiary origin of the Caenophidia.     

Defective formation of PKA/CnA-dependent annexin 2-S100A10/CFTR complex in DeltaF508 cystic fibrosis cells

Cystic fibrosis (CF) is characterised by impaired epithelial ion transport and is caused by mutations in the cystic fibrosis conductance regulator protein (CFTR), a cAMP/PKA and ATP-regulated chloride channel. We recently demonstrated a cAMP/PKA/calcineurin (CnA)-driven association between annexin 2 (anx 2), its cognate partner –S100A10 and cell surface CFTR. The complex is required for CFTR and outwardly rectifying chloride channel function in epithelia. Since the cAMP/PKA-induced Cl⁻ current is absent in CF epithelia, we hypothesized that the anx 2–S100A10/CFTR complex may be defective in CFBE41o cells expressing the commonest F508del-CFTR (ΔF-CFTR) mutation. Here, we demonstrate that, despite the presence of cell surface ΔF-CFTR, cAMP/PKA fails to induce anx 2–S100A10/CFTR complex formation in CFBE41o− cells homozygous for F508del-CFTR. Mechanistically, PKA-dependent serine phosphorylation of CnA, CnA–anx 2 complex formation and CnA-dependent dephosphorylation of anx 2 are all defective in CFBE41o− cells. Immunohistochemical analysis confirms an abnormal cellular distribution of anx 2 in human and CF mouse epithelia.

Thus, we demonstrate that cAMP/PKA/CnA signaling pathway is defective in CF cells and suggest that loss of anx 2–S100A10/CFTR complex formation may contribute to defective cAMP/PKA-dependent CFTR channel function.     

Quantitative profiling of nucleotides and related phosphate-containing metabolites in cultured mammalian cells by liquid chromatography tandem electrospray mass spectrometry

A method has been developed for the quantitative profiling of over twenty nucleotides and related phosphorylated species using ion-pair reversed-phase liquid chromatography hyphenated to negative ion tandem electrospray mass spectrometry. The influence of mobile phase pH and ion-pairing agent concentration were assessed to optimise separation and peak shapes. Full quantitative analysis was obtained for the nucleotides by reference to structurally related calibration standards. The developed method was applied to profile changes in nucleotides and related compounds in monolayer cultured Chinese hamster ovary (CHO) cells expressing the beta(2) adrenoceptor when exposed to pharmacological stimuli. These experiments demonstrate the potential of the LC-MS/MS method to detect changes in nucleotide drug targets as well as the simultaneous monitoring of levels of other nucleotides.