Suitability of machine learning models for prediction of clinically defined Stage III/IV periodontitis from questionnaires and demographic data in Danish cohorts.

AIM: To evaluate if, and to what extent, machine learning models can capture clinically defined Stage III/IV periodontitis from self-report questionnaires and demographic data. MATERIALS AND METHODS: Self-reported measures of periodontitis, demographic data and clinically established Stage III/IV periodontitis status were extracted from two Danish population-based cohorts (The Copenhagen Aging and Midlife Biobank [CAMB] and The Danish Health Examination Survey [DANHES]) and used to develop cross-validated machine learning models for the prediction of clinically established Stage III/IV periodontitis. Models were trained using 10-fold cross-validations repeated three times on the CAMB dataset (n = 1476), and the resulting models were validated in the DANHES dataset (n = 3585). RESULTS: The prevalence of Stage III/IV periodontitis was 23.2% (n = 342) in the CAMB dataset and 9.3% (n = 335) in the DANHES dataset. For the prediction of clinically established Stage III/IV periodontitis in the CAMB cohort, models reached area under the receiver operating characteristics (AUROCs) of 0.67-0.69, sensitivities of 0.58-0.64 and specificities of 0.71-0.80. In the DANHES cohort, models derived from the CAMB cohort achieved AUROCs of 0.64-0.70, sensitivities of 0.44-0.63 and specificities of 0.75-0.84. CONCLUSIONS: Applying cross-validated machine learning algorithms to demographic data and self-reported measures of periodontitis resulted in models with modest capabilities for the prediction of Stage III/IV periodontitis in two Danish cohorts.

Effects of glyphosate spray-drift on plant flowering.

Recent studies have shown that sub-lethal doses of herbicides may affect plant flowering, however, no study has established a direct relationship between the concentrations of deposited herbicide and plant flowering. Here the aim was to investigate the relationship between herbicide spray drift deposited on non-target plants and plant flowering in a realistic agro-ecosystem setting. The concentrations of the herbicide glyphosate deposited on plants were estimated by measuring the concentration of a dye tracer applied together with the herbicide. The estimated maximal and average deposition of glyphosate within the experimental area corresponded to 30 g glyphosate/ha (2.08% of the label rate of 1440 g a.i./ha) and 2.4 g glyphosate/ha (0.15% label rate), respectively, and the concentrations decreased rapidly with increasing distance from the spraying track. However, there were not a unique relation between distance and deposition, which indicate that heterogeneities of turbulence, wind speed and/or direction can strongly influence the deposition from 1 min to another during spraying. The effects of glyphosate on cumulative flower numbers and flowering time were modelled using Gompertz growth models on four non-target species. Glyphosate had a significantly negative effect on the cumulative number of flowers on Trifolium pratense and Lotus corniculatus, whereas there were no significant effects on Trifolium repens, and a positive, but non-significant, effect on number of flowers on Cichorium intybus. Glyphosate did not affect the flowering time of any of the four species significantly. Lack of floral resources is known to be of major importance for pollinator declines. The implications of the presented results for pesticide risk assessment are discussed.

Porphyromonas gingivalis-induced production of reactive oxygen species, tumor necrosis factor-α, interleukin-6, CXCL8 and CCL2 by neutrophils from localized aggressive periodontitis and healthy donors: modulating actions of red blood cells and resolvin E1.

BACKGROUND AND OBJECTIVES: Porphyromonas gingivalis is regarded as a significant contributor in the pathogenesis of periodontitis and certain systemic diseases, including atherosclerosis. P. gingivalis occasionally translocates from periodontal pockets into the circulation, where it adheres to red blood cells (RBCs). This may protect the bacterium from contact with circulating phagocytes without affecting its viability. MATERIAL AND METHODS: In this in vitro study, we investigated whether human peripheral blood neutrophils from 10 subjects with localized aggressive periodontitis (LAgP) and 10 healthy controls release the proinflammatory cytokines interleukin (IL)-6, tumor necrosis factor α (TNF-α), the chemokine (C-X-C motif) ligand 8 (CXCL8; also known as IL-8) and chemokine (C-C motif) ligand 2 (CCL2; also known as monocyte chemotactic protein-1) and intracellular reactive oxygen species (ROS) in response to challenge with P. gingivalis. In addition, the impact of RBC interaction with P. gingivalis was investigated. The actions of resolvin E1 (RvE1), a known regulator of P. gingivalis induced neutrophil responses, on the cytokine and ROS responses elicited by P. gingivalis in cultures of neutrophils were investigated. RESULTS: Upon stimulation with P. gingivalis, neutrophils from subjects with LAgP and healthy controls released similar quantities of IL-6, TNF-α, CXCL8, CCL2 and intracellular ROS. The presence of RBCs amplified the release of IL-6, TNF-α and CCL2 statistically significant in both groups, but reduced the generation of ROS in the group of healthy controls, and showed a similar tendency in the group of subjects with LAgP. RvE1 had no impact on the production of intracellular ROS, TNF-α, IL-6, CXCL8 and CCL2 by neutrophils from either group, but tended to reduce the generation of ROS in subjects with LAgP in the absence of RBCs. CONCLUSIONS: Our data support that binding to RBCs protects P. gingivalis from ROS and concomitantly enhances neutrophil release of proinflammatory cytokines providing a selective advantage for P. gingivalis growth.

In vitro complement activation, adherence to red blood cells and induction of mononuclear cell cytokine production by four strains of Aggregatibacter actinomycetemcomitans with different fimbriation and expression of leukotoxin.

BACKGROUND AND OBJECTIVE: The periodontal pathogen Aggregatibacter actinomycetemcomitans has been proposed as pro-atherogenic, and complement-mediated adherence to red blood cells (RBCs) may facilitate its systemic spread. We investigated the ability of four strains of A. actinomycetemcomitans with differential expression of leukotoxin A (LtxA) and fimbriae to activate complement, adhere to RBCs and elicit cytokine responses by mononuclear cells (MNCs). MATERIAL AND METHODS: Aggregatibacter actinomycetemcomitans serotype b strains HK 921, HK 1651, HK 2092 and HK 2108 were fluorescence-labeled, incubated with human whole blood cells in the presence of autologous serum, and assessed for RBC adherence by flow cytometry and for capacity to induce cytokine production by cytometric bead array analysis. The levels of IgG to A. actinomycetemcomitans serotype b were quantified by ELISA, as was consumption of complement. RESULTS: The JP2 clone variants HK 1651 and, to a lesser extent, HK 2092, consumed complement efficiently, while HK 2108 (= strain Y4) consumed complement poorly. Nonetheless, the four tested strains adhered equally well to RBCs in the presence of autologous serum, without causing RBC lysis. The JP2 clone variant HK 2092, selectively lacking LtxA production, induced higher production of tumor necrosis factor (TNF)-α, interleukin (IL)-1ß, IL-6 and IL-10 by MNCs than did the other three strains, while the four strains induced similar production of IL-12p70. RBCs facilitated the HK 2092-induced production of TNF-α and IL-1ß, and IL-6 was enhanced by RBCs, and this facilitation could be counteracted by blockade of complement receptor 3 (CD11b/CD18). CONCLUSION: Our data suggest that the JP2 clone of A. actinomycetemcomitans, most closely resembled by the variant HK 1651, activates complement well, while strain Y4, represented by HK 2108, activates complement poorly. However, all strains of A. actinomycetemcomitans adhere to RBCs and, when capable of producing LtxA, prevent production of inflammatory cytokines by MNCs. This "immunologically silent" immune adherence may facilitate systemic spread and atherogenesis.

Influence of complement on neutrophil extracellular trap release induced by bacteria.

BACKGROUND AND OBJECTIVES: Neutrophil extracellular trap (NET) release has generally been studied in the absence of serum, or at low concentrations of untreated or heat-inactivated serum. The influence of serum complement on NET release therefore remains unclear. We examined the DNA release induced by Staphylococcus aureus and three oral bacteria: Actinomyces viscosus, Aggregatibacter actinomycetemcomitans and Fusobacterium nucleatum subsp. vincettii. MATERIAL AND METHODS: Bacteria-stimulated NET release from the neutrophils of healthy donors was measured fluorometrically. Various complement containing and complement blocking conditions were used, including heat inactivation of the serum and antibody blockade of complement receptors 1 (CR1, CD35) and 3 (CR3, CD11b/CD18). RESULTS: While the presence of serum markedly enhanced NET release induced by S. aureus, A. actinomycetemcomitans, and to a lesser extent by A. viscosus, there was no enhancement of NET release induced by F. nucleatum. The serum-mediated enhancement of NET release by A. actinomycetemcomitans was neutralized by heat inactivation of serum complement, while this was not the case for S. aureus. Blockade of CR1, significantly reduced NET release induced by S. aureus, A. actinomycetemcomitans and A. viscosus, while blockade of CR3, had no effect. However, opsonization of S. aureus with antibodies may also have contributed to the enhancing effect of serum, independently of complement, in that purified IgG promoted NET release. CONCLUSIONS: In conclusion, complement opsonization promotes NET release induced by a variety of bacteria, including A. actinomycetemcomitans, and CR1 plays a dominant role in the process. Complement consumption or deficiency may compromise NETosis induced by some bacterial species, including A. actinomycetemcomitans. Within biofilms, the complement-inactivating abilities of some bacteria may protect other species against NETosis, while these are more vulnerable when adopting a planktonic lifestyle.

The complement system and its role in the pathogenesis of periodontitis: current concepts.

Periodontitis is a highly prevalent inflammatory disease in tooth supporting tissues, induced by bacteria growing in a biofilm on tooth surfaces. Components of the complement system are present in the periodontal tissue and the system is activated in periodontitis. Continuous complement activation and modulation by bacteria within the biofilm in periodontal pockets, however, may enhance local tissue destruction, providing the biofilm with both essential nutrients and space to grow. A more profound understanding of the mechanisms involved in complement-derived tissue degradation may facilitate the development of new treatment concepts for periodontitis. Further studies on the role of complement in periodontitis pathogenesis may also contribute to the understanding of why some individuals fail to resolve periodontitis. Here, we review evidence that links complement to the pathogenesis of periodontitis with an emphasis on interaction of complement with bacteria from periodontitis-associated biofilm.

Gene flow and population structure of a common agricultural wild species (Microtus agrestis) under different land management regimes.

The impact of landscape structure and land management on dispersal of populations of wild species inhabiting the agricultural landscape was investigated focusing on the field vole (Microtus agrestis) in three different areas in Denmark using molecular genetic markers. The main hypotheses were the following: (i) organic farms act as genetic sources and diversity reservoirs for species living in agricultural areas and (ii) gene flow and genetic structure in the agricultural landscape are influenced by the degree of landscape complexity and connectivity. A total of 443 individual voles were sampled within 2 consecutive years from two agricultural areas and one relatively undisturbed grassland area. As genetic markers, 15 polymorphic microsatellite loci (nuclear markers) and the central part of the cytochrome-b (mitochondrial sequence) were analysed for all samples. The results indicate that management (that is, organic or conventional management) was important for genetic population structure across the landscape, but that landscape structure was the main factor shaping gene flow and genetic diversity. More importantly, the presence of organically managed areas did not act as a genetic reservoir for conventional areas, instead the most important predictor of effective population size was the amount of unmanaged available habitat (core area). The relatively undisturbed natural area showed a lower level of genetic structuring and genetic diversity compared with the two agricultural areas. These findings altogether suggest that political decisions for supporting wildlife friendly land management should take into account both management and landscape structure factors.

Interaction between density and Cu toxicity for Enchytraeus crypticus--comparing first and second generation effects.

Density of organisms varies considerably in nature depending e.g. on seasonality or food availability. A recent investigation on interaction between Cu and density using Enchytraeus crypticus showed that density itself (5-50 per 20 gr dry soil) had an impact on population and individual growth [up to 3000 individuals per test vessel], but the interaction between density and Cu toxicity was not significant. Here, a follow-up study was performed, in which the interactions between density and Cu-exposure were investigated along a two-generation exposure using E. crypticus (three factorial: 1. density (5-50), 2. Cu (0-300 mg/kg) and generation (G1-G2)). After G1, the juveniles were retrieved and further exposed under the same conditions along a G2 (using a refined density set - 10 and 50). Results showed an interaction between density and Cu in the reproduction of E. crypticus, this being significant in G2, showing lower toxicity for higher density of organisms whereas in the 1st generation the opposite occurred. Hence, there was an interaction seen along G1 to G2, i.e. animals from density 50 in G1 when further exposed at density 50 in G2 had lower Cu toxicity compared to when further exposed at density 10. Possible explanations include the hydra effect (overcompensation in G2 due to stress in G1) or that for density 50 the organisms are exposed to less Cu than at lower densities, this by e.g. (1) organisms avoid exposure by lumping/clustering which would limit exposure to Cu and (2) there would be less available Cu contaminated soil per individual at high density hence less exposure.

Evolutionary rate variation and RNA secondary structure prediction.

Predicting RNA secondary structure using evolutionary history can be carried out by using an alignment of related RNA sequences with conserved structure. Accurately determining evolutionary substitution rates for base pairs and single stranded nucleotides is a concern for methods based on this type of approach. Determining these rates can be hard to do reliably without a large and accurate initial alignment, which ideally also has structural annotation. Hence, one must often apply rates extracted from other RNA families with trusted alignments and structures. Here, we investigate this problem by applying rates derived from tRNA and rRNA to the prediction of the much more rapidly evolving 5'-region of HIV-1. We find that the HIV-1 prediction is in agreement with experimental data, even though the relative evolutionary rate between A and G is significantly increased, both in stem and loop regions. In addition we obtained an alignment of the 5' HIV-1 region that is more consistent with the structure than that currently in the database. We added randomized noise to the original values of the rates to investigate the stability of predictions to rate matrix deviations. We find that changes within a fairly large range still produce reliable predictions and conclude that using rates from a limited set of RNA sequences is valid over a broader range of sequences.

Sublethal soil copper concentrations increase mortality in the earthworm Aporrectodea caliginosa during drought.

Simultaneous effects of drought and a sublethal concentration of copper on the earthworm Aporrectodea caliginosa were investigated in the laboratory. Experiments were performed with exposure to 150 mg Cu kg(-1) dry soil and different levels of drought during a 3-wk period. Soil water potentials from pF 1.5 (wet) to pF 5 (very dry) were obtained. The soil water potential resulting in 50% mortality was significantly reduced from pF 4.48 in worms not exposed to copper to pF 4.09 in copper-exposed worms, thus demonstrating synergy-like effects of drought and copper. Development of estivation cells was significantly depressed in copper-exposed worms compared to worms not exposed to copper. For all drought levels (except the highest, where 100% mortality occurred), copper-exposed earthworms with no estivation cells had more than twice as high mortality and, at severe drought exposure, also lower water content than those which had developed the estivation cells. With increasing drought level, the whole-body burden of copper increased from about 40 microg Cu g(-1) dry weight to about 90 microg Cu g(-1). When the worms were exposed to drought, the osmolality in their body fluids increased. This was due to a greater concentration of already existing solutes resulting from extensive loss of water, and not to mobilization or synthesis of additional solutes.

Male fitness of oilseed rape (Brassica napus), weedy B. rapa and their F(1) hybrids when pollinating B. rapa seeds.

The likelihood that two species hybridise and backcross may depend strongly on environmental conditions, and possibly on competitive interactions between parents and hybrids. We studied the paternity of seeds produced by weedy Brassica rapa growing in mixtures with oilseed rape (B. napus) and their F(1) hybrids at different frequencies and densities. Paternity was determined by the presence of a transgene, morphology, and AFLP markers. In addition, observations of flower and pollen production, and published data on pollen fertilisation success, zygote survival, and seed germination, allowed us to estimate an expected paternity. The frequency and density of B. napus, B. rapa, and F(1) plants had a strong influence on flower, pollen, and seed production, and on the paternity of B. rapa seeds. Hybridisation and backcrossing mostly occurred at low densities and at high frequencies of B. napus and F(1), respectively. F(1) and backcross offspring were produced mainly by a few B. rapa mother plants. The observed hybridisation and backcrossing frequencies were much lower than expected from our compilation of fitness components. Our results show that the male fitness of B. rapa, B. napus, and F(1) hybrids is strongly influenced by their local frequencies, and that male fitness of F(1)hybrids, when pollinating B. rapa seeds, is low even when their female fitness (seed set) is high.

The hnRNP A1 protein regulates HIV-1 tat splicing via a novel intron silencer element.

The generation of >30 different HIV-1 mRNAs is achieved by alternative splicing of one primary transcript. The removal of the second tat intron is regulated by a combination of a suboptimal 3' splice site and cis-acting splicing enhancers and silencers. Here we show that hnRNP A1 inhibits splicing of this intron via a novel heterogeneous nuclear ribonucleoprotein (hnRNP) A1-responsive intron splicing silencer (ISS) that can function independently of the previously characterized exon splicing silencer (ESS3). Surprisingly, depletion of hnRNP A1 from the nuclear extract (NE) enables splicing to proceed in NE that contains 100-fold reduced concentrations of U2AF and normal levels of SR proteins, conditions that do not support processing of other efficiently spliced pre-mRNAs. Reconstituting the extract with recombinant hnRNP A1 protein restores splicing inhibition at a step subsequent to U2AF binding, mainly at the time of U2 snRNP association. hnRNP A1 interacts specifically with the ISS sequence, which overlaps with one of three alternative branch point sequences, pointing to a model where the entry of U2 snRNP is physically blocked by hnRNP A1 binding.

CRM1 mediates the export of ADAR1 through a nuclear export signal within the Z-DNA binding domain.

RNA editing of specific residues by adenosine deamination is a nuclear process catalyzed by adenosine deaminases acting on RNA (ADAR). Different promoters in the ADAR1 gene give rise to two forms of the protein: a constitutive promoter expresses a transcript encoding (c)ADAR1, and an interferon-induced promoter expresses a transcript encoding an N-terminally extended form, (i)ADAR1. Here we show that (c)ADAR1 is primarily nuclear whereas (i)ADAR1 encompasses a functional nuclear export signal in the N-terminal part and is a nucleocytoplasmic shuttle protein. Mutation of the nuclear export signal or treatment with the CRM1-specific drug leptomycin B induces nuclear accumulation of (i)ADAR1 fused to the green fluorescent protein and increases the nuclear editing activity. In concurrence, CRM1 and RanGTP interact specifically with the (i)ADAR1 nuclear export signal to form a tripartite export complex in vitro. Furthermore, our data imply that nuclear import of (i)ADAR1 is mediated by at least two nuclear localization sequences. These results suggest that the nuclear editing activity of (i)ADAR1 is modulated by nuclear export.

Interaction between the transcription factor SPBP and the positive cofactor RNF4. An interplay between protein binding zinc fingers.

The activator of stromelysin 1 gene transcription, SPBP, interacts with the RING finger protein RNF4. Both proteins are ubiquitously expressed and localized in the nucleus. RNF4 facilitates accumulation of specific SPBP-DNA complexes in vitro and acts as a positive cofactor in SPBP-mediated transactivation. SPBP harbors an internal zinc finger of the PHD/LAP type. This domain can form intra-chain protein-protein contacts in SPBP resulting in negative modulation of SPBP-RNF4 interaction.

Fixation of advantageous alleles in partially self-fertilizing populations. The effect of different selection modes.

The expected fixation probability of an advantageous allele was examined in a partially self-fertilizing hermaphroditic plant species using the diffusion approximation. The selective advantage of the advantageous allele was assumed to be increased viability, increased fecundity, or an increase in male fitness. The mode of selection, as well as the selfing rate, the population size, and the dominance of the advantageous allele, affect the fixation probability of the allele. In general it was found that increases in selfing rate decrease the fixation probability under male sexual selection, increase fixation probability under fecundity selection, and increase when recessive and decrease when dominant under viability selection. In some cases the highest fixation probability of advantageous alleles under fecundity or under male sexual selection occurred at an intermediary selfing rate. The expected mean fixation times of the advantageous allele were also examined using the diffusion approximation.

Coevolution of a plant host-pathogen gene-for-gene system in a metapopulation model without cost of resistance or cost of virulence

A metapopulation model of a one-locus gene-for-gene system in a plant host and a biotrophic pathogen is described. The model allows subpopulations to go extinct, and, due to characteristic differences in life-history strategies, the plant host is assumed to be recolonized from a seed bank, whereas the pathogen is recolonized by migration. It is shown that variation in the gene-for-gene system can be maintained at a noticeable level without assuming cost of resistance or cost of virulence, if the probability of extinction depends on the host mean fitness in the subpopulation. The level of variation in the pathogen population increases with increasing extinction rate, genetic drift and fitness of the infected host, but decreases with increasing migration rate. Generally, these effects are magnified for life cycles in which selection occur before genetic drift and after migration. The metapopulation model generates positive associations between the virulence allele and the resistance allele without assuming cost of resistance or cost of virulence. Copyright 1999 Academic Press.

Mapping the RNA binding sites for human immunodeficiency virus type-1 gag and NC proteins within the complete HIV-1 and -2 untranslated leader regions.

Encapsidation of HIV-1 genomic RNA is mediated by specific interactions between the RNA packaging signal and the Gag protein. During maturation of the virion, the Gag protein is processed into smaller fragments, including the nucleocapsid (NC) domain which remains associated with the viral genomic RNA. We have investigated the binding of glutathione- S -transferase (GST) Gag and NC fusion proteins from HIV-1, to the entire HIV-1 and -2 leader RNAencompassing the packaging signal. We have mapped the binding sites at conditions where only about two complexes are formed and find that GST-Gag and GST-NC fusion proteins bind specifically to discrete sites within the leader. Analysis of the HIV-1 leader indicated that GST-Gag strongly associates with the PSI stem-loop and to a lesser extent with regions near the primer binding site. GST-NC binds the same regions but with reversed preferences. The HIV-1 proteins also interact specifically with the 5'-leader of HIV-2 and the major site of interaction mapped to a stem-loop, with homology to the HIV-1 PSI stem-loop structure. The different specificities of Gag and NC may reflect functionally distinct roles in the viral replication, and suggest that the RNA binding specificity of NC is modulated by its structural context.

Inbreeding depression and dominance-suppression competition after inbreeding in rapeseed (Brassica napus).

Rapeseed plants, of the summer annual variety Topas, that had been selfed twice consecutively were compared to outcrossed half-sibs for inbreeding depression in a rapeseed population at mating equilibrium. The effect of dominance-suppression competition was included in the effect of inbreeding. Both female-and male-fitness characters showed significant inbreeding depression. Biomass decreased 17% with inbreeding and was highly correlated with seed weight. The total number of flowers decreased 15% with inbreeding. There was a significant effect of lines. The possible importance of experimental design in studies that estimate inbreeding depression is discussed.

Environmental variation for outcrossing rate in rapeseed (Brassica napus).

Rapeseed (Brassica napus) is a predominantly selfpollinated crop with about one-third outcrossing. The outcrossing rate may be influenced by environmental factors, and hence changes in the heterozygosity level of a variety may occur during multiplication. In an investigation on environmental variation in outcrossing, we estimated the outcrossing rate in the Swedish spring rapeseed cv 'Topas' by isozyme analysis and found that outcrossing varied from 12% to 47% over five locations in Sweden, Denmark and Germany. Among flowers at different positions on the same plant, average outcrossing varied from 11% at the top to 39% at the bottom of the plant. In conclusion, environmental factors can greatly influence the outcrossing rate in rapeseed, and an investigation therefore merit further studies.