Gene Loss and Lineage-Specific Restriction-Modification Systems Associated with Niche Differentiation in the Campylobacter jejuni Sequence Type 403 Clonal Complex

Campylobacter jejuni is a highly diverse species of bacteria commonly associated with infectious intestinal disease of humans and zoonotic carriage in poultry, cattle, pigs, and other animals. The species contains a large number of distinct clonal complexes that vary from host generalist lineages commonly found in poultry, livestock, and human disease cases to host-adapted specialized lineages primarily associated with livestock or poultry. Here, we present novel data on the ST403 clonal complex of C. jejuni, a lineage that has not been reported in avian hosts. Our data show that the lineage exhibits a distinctive pattern of intralineage recombination that is accompanied by the presence of lineage-specific restriction-modification systems. Furthermore, we show that the ST403 complex has undergone gene decay at a number of loci. Our data provide a putative link between the lack of association with avian hosts of C. jejuni ST403 and both gene gain and gene loss through nonsense mutations in coding sequences of genes, resulting in pseudogene formation.     

Efficient measurement error correction with spatially misaligned data

Association studies in environmental statistics often involve exposure and outcome data that are misaligned in space. A common strategy is to employ a spatial model such as universal kriging to predict exposures at locations with outcome data and then estimate a regression parameter of interest using the predicted exposures. This results in measurement error because the predicted exposures do not correspond exactly to the true values. We characterize the measurement error by decomposing it into Berkson-like and classical-like components. One correction approach is the parametric bootstrap, which is effective but computationally intensive since it requires solving a nonlinear optimization problem for the exposure model parameters in each bootstrap sample. We propose a less computationally intensive alternative termed the "parameter bootstrap" that only requires solving one nonlinear optimization problem, and we also compare bootstrap methods to other recently proposed methods. We illustrate our methodology in simulations and with publicly available data from the Environmental Protection Agency.     

Sub-lethal effects of pesticide residues in brood comb on worker honey bee (Apis mellifera) development and longevity

Background

Numerous surveys reveal high levels of pesticide residue contamination in honey bee comb. We conducted studies to examine possible direct and indirect effects of pesticide exposure from contaminated brood comb on developing worker bees and adult worker lifespan.

Methodology/Principal Findings

Worker bees were reared in brood comb containing high levels of known pesticide residues (treatment) or in relatively uncontaminated brood comb (control). Delayed development was observed in bees reared in treatment combs containing high levels of pesticides particularly in the early stages (day 4 and 8) of worker bee development. Adult longevity was reduced by 4 days in bees exposed to pesticide residues in contaminated brood comb during development. Pesticide residue migration from comb containing high pesticide residues caused contamination of control comb after multiple brood cycles and provided insight on how quickly residues move through wax. Higher brood mortality and delayed adult emergence occurred after multiple brood cycles in contaminated control combs. In contrast, survivability increased in bees reared in treatment comb after multiple brood cycles when pesticide residues had been reduced in treatment combs due to residue migration into uncontaminated control combs, supporting comb replacement efforts. Chemical analysis after the experiment confirmed the migration of pesticide residues from treatment combs into previously uncontaminated control comb.

Conclusions/Significance

This study is the first to demonstrate sub-lethal effects on worker honey bees from pesticide residue exposure from contaminated brood comb. Sub-lethal effects, including delayed larval development and adult emergence or shortened adult longevity, can have indirect effects on the colony such as premature shifts in hive roles and foraging activity. In addition, longer development time for bees may provide a reproductive advantage for parasitic Varroa destructor mites. The impact of delayed development in bees on Varroa mite fecundity should be examined further.     

Large-scale cortical functional organization and speech perception across the lifespan

Aging is accompanied by substantial changes in brain function, including functional reorganization of large-scale brain networks. Such differences in network architecture have been reported both at rest and during cognitive task performance, but an open question is whether these age-related differences show task-dependent effects or represent only task-independent changes attributable to a common factor (i.e., underlying physiological decline). To address this question, we used graph theoretic analysis to construct weighted cortical functional networks from hemodynamic (functional MRI) responses in 12 younger and 12 older adults during a speech perception task performed in both quiet and noisy listening conditions. Functional networks were constructed for each subject and listening condition based on inter-regional correlations of the fMRI signal among 66 cortical regions, and network measures of global and local efficiency were computed. Across listening conditions, older adult networks showed significantly decreased global (but not local) efficiency relative to younger adults after normalizing measures to surrogate random networks. Although listening condition produced no main effects on whole-cortex network organization, a significant age group x listening condition interaction was observed. Additionally, an exploratory analysis of regional effects uncovered age-related declines in both global and local efficiency concentrated exclusively in auditory areas (bilateral superior and middle temporal cortex), further suggestive of specificity to the speech perception tasks. Global efficiency also correlated positively with mean cortical thickness across all subjects, establishing gross cortical atrophy as a task-independent contributor to age-related differences in functional organization. Together, our findings provide evidence of age-related disruptions in cortical functional network organization during speech perception tasks, and suggest that although task-independent effects such as cortical atrophy clearly underlie age-related changes in cortical functional organization, age-related differences also demonstrate sensitivity to task domains.     

Phylogenetic relationships of honey bees (Hymenoptera:Apinae:Apini) inferred from nuclear and mitochondrial DNA sequence data

Two different genomic regions (ND2 mitochondrial gene and EF1-alpha intron) were PCR amplified, cloned and sequenced for the ten known honey bee species collected within their natural range distribution. DNA sequences were analyzed using parsimony, distance and maximum likelihood methods to investigate phylogenetic relationships within Apis. The phylogenetic analyses strongly supported the basic topology recoverable from morphometric analysis, grouping the honey bees into three major clusters: giant bees (A. dorsata, A. binghami, and A. laboriosa), dwarf bees (A. andreniformis and A. florea), and cavity-nesting bees (A. mellifera, A. cerana, A. koschevnikovi, A. nuluensis, and A. nigrocincta). However, the clade of Asian cavity-nesting bees included paraphyletic taxa. Exemplars of Apis cerana collected from divergent portions of its range were less related to each other than were sympatric A. cerana, A. nuluensis, and A. nigrocincta taxa. Nucleotide sequence divergence between allopatrically distributed western (A. mellifera) and eastern (A. cerana, A. koschevnikovi, A. nigrocincta, and A. nuluensis) cavity-nesting species, around 18% for the mitochondrial gene and 10-15% for the nuclear intron, suggested an earlier divergence for these groups than previously estimated from morphometric and behavioral studies. This latter finding neccessitates reevaluation of the hypothesized origin of extant European, African, and west Asian Apis mellifera. Sequence divergence between A. laboriosa and A. dorsata was consistent with behavioral data and supports the species status of A. laboriosa.     

Chain termination and inhibition of Saccharomyces cerevisiae poly(A) polymerase by C-8-modified ATP analogs

The nucleotide substrate specificity of yeast poly(A) polymerase (yPAP) toward various C-2- and C-8-modified ATP analogs was examined. 32P-Radiolabeled RNA oligonucleotide primers were incubated with yPAP in the absence of ATP to assay polyadenylation using unnatural ATP substrates. The C-2-modified ATP analogs 2-amino-ATP and 2-chloro (Cl)-ATP were excellent substrates for yPAP. 8-Amino-ATP, 8-azido-ATP, and 8-aza-ATP all produced chain termination of polyadenylation, and no primer extension was observed with the C-8-halogenated derivatives 8-Br-ATP and 8-Cl-ATP. The effects of modified ATP analogs on ATP-dependent poly(A) tail synthesis by yPAP were also examined. Whereas C-2 substitution (2-amino-ATP and 2-Cl-ATP) had little effect on poly(A) tail length, C-8 substitution produced moderate (8-amino-ATP, 8-azido-ATP, and 8-aza-ATP) to substantial (8-Br-ATP and 8-Cl-ATP) reduction in poly(A) tail length. To model the biochemical consequences of 8-Cl-Ado incorporation into RNA primers, a synthetic RNA primer containing a 3'-terminal 8-Cl-AMP residue was prepared. Polyadenylation of this modified RNA primer by yPAP in the presence of ATP was blocked completely. To probe potential mechanisms of inhibition, two-dimensional NMR spectroscopy experiments were used to examine the conformation of two C-8-modified AMP nucleotides, 8-Cl-AMP and 8-amino-AMP. C-8 substitution in adenosine analogs shifted the ribose sugar pucker equilibrium to favor the DNA-like C-2'-endo form over the C-3'-endo (RNA-like) conformation, which suggests a potential mechanism for polyadenylation inhibition and chain termination. Base-modified ATP analogs may exert their biological effects through polyadenylation inhibition and thus may provide useful tools for investigating polyadenylation biochemistry within cells.     

A glycoprotein hormone expressed in corticotrophs exhibits unique binding properties on thyroid-stimulating hormone receptor

Corticotroph-derived glycoprotein hormone (CGH), also referred to as thyrostimulin, is a noncovalent heterodimer of glycoprotein hormone alpha 2 (GPHA2) and glycoprotein hormone beta 5 (GPHB5). Here, we demonstrate that both subunits of CGH are expressed in the corticotroph cells of the human anterior pituitary, as well as in skin, retina, and testis. CGH activates the TSH receptor (TSHR); (125)I-CGH binding to cells expressing TSHR is saturable, specific, and of high affinity. In competition studies, unlabeled CGH is a potent competitor for (125)I-TSH binding, whereas unlabeled TSH does not compete for (125)I-CGH binding. Binding and competition analyses are consistent with the presence of two binding sites on the TSHR transfected baby hamster kidney cells, one that can interact with either TSH or CGH, and another that binds CGH alone. Transgenic overexpression of GPHB5 in mice produces elevations in serum T(4) levels, reductions in body weight, and proptosis. However, neither transgenic overexpression of GPHA2 nor deletion of GPHB5 produces an overt phenotype in mice. In vivo administration of CGH to mice produces a dose-dependent hyperthyroid phenotype including elevation of T(4) and hypertrophy of cells within the inner adrenal cortex. However, the distinctive expression patterns and binding characteristics of CGH suggest that it has endogenous biological roles that are discrete from those of TSH.     

A modelling approach to estimate the effect of exotic pollinators on exotic weed population dynamics: bumblebees and broom in Australia

The role of mutualisms in contributing to species invasions is rarely considered, inhibiting effective risk analysis and management options. Potential ecological consequences of invasion of non‐native pollinators include increased pollination and seed set of invasive plants, with subsequent impacts on population growth rates and rates of spread. We outline a quantitative approach for evaluating the impact of a proposed introduction of an invasive pollinator on existing weed population dynamics and demonstrate the use of this approach on a relatively data‐rich case study: the impacts on Cytisus scoparius (Scotch broom) from proposed introduction of Bombus terrestris. Three models have been used to assess population growth (matrix model), spread speed (integrodifference equation), and equilibrium occupancy (lattice model) for C. scoparius. We use available demographic data for an Australian population to parameterize two of these models. Increased seed set due to more efficient pollination resulted in a higher population growth rate in the density‐independent matrix model, whereas simulations of enhanced pollination scenarios had a negligible effect on equilibrium weed occupancy in the lattice model. This is attributed to strong microsite limitation of recruitment in invasive C. scoparius populations observed in Australia and incorporated in the lattice model. A lack of information regarding secondary ant dispersal of C. scoparius prevents us from parameterizing the integrodifference equation model for Australia, but studies of invasive populations in California suggest that spread speed will also increase with higher seed set. For microsite‐limited C. scoparius populations, increased seed set has minimal effects on equilibrium site occupancy. However, for density‐independent rapidly invading populations, increased seed set is likely to lead to higher growth rates and spread speeds. The impacts of introduced pollinators on native flora and fauna and the potential for promoting range expansion in pollinator‐limited ‘sleeper weeds’ also remain substantial risks.     

Evaluation of temperature gradient gel electrophoresis for the analysis of prey DNA within the guts of invertebrate predators

The utility of temperature gradient gel electrophoresis (TGGE) as a means of analysing the gut contents of predators was evaluated. Generalist predators consume multiple prey species and a species-specific primer approach may not always be a practical means of analysing predator responses to prey diversity in complex and biodiverse ecosystems. General invertebrate primers were used to amplify the gut contents of predators, generating banding patterns that identified component prey remains. There was no evidence of dominance of the polymerase chain reaction (PCR) by predator DNA. When applied to field samples of the carabid predator Pterostichus melanarius (Illiger) nine banding patterns were detected, including one for aphids. To further distinguish between species, group-specific primers were designed to separate species of earthworm and aphid. TGGE of the earthworm PCR products generated banding patterns that varied with haplotype in some species. Aphid and earthworm DNA could be detected in the guts of carabids for up to 24 h using TGGE. In P. melanarius, with low numbers of prey per insect gut (mean<3), interpretation of banding patterns proved to be tractable. Potential problems of interpretation of TGGE gels caused by multiple prey bands, cryptic bands, haplotype variation, taxonomic uncertainties (especially with regard to earthworms), secondary predation, scavenging and presence of parasites and parasitoids in the prey or the predators, are discussed. The results suggest that PCR, using combinations of general invertebrate and group-specific primers followed by TGGE, provides a potentially useful approach to the analysis of multiple uncharacterized prey in predators.