Quantitative assessment of the tetracycline resistance gene pool in cheese samples by real-time TaqMan PCR

A TaqMan real-time PCR assay was developed to quantify the tetS gene pool present in retail cheeses. This protocol offers a rapid, specific, sensitive, and culture-independent method for assessing antibiotic resistance genes in food samples rich in fats and proteins.     

The PM2 virion has a novel organization with an internal membrane and pentameric receptor binding spikes

Biological membranes are notoriously resistant to structural analysis. Excellent candidates to tackle this problem in situ are membrane-containing viruses where the membrane is constrained by an icosahedral capsid. Cryo-EM and image reconstruction of bacteriophage PM2 revealed a membrane bilayer following the internal surface of the capsid. The viral genome closely interacts with the inner leaflet. The capsid, at a resolution of 8.4 A, reveals 200 trimeric capsomers with a pseudo T = 21 dextro organization. Pentameric receptor-binding spikes protrude from the surface. It is evident from the structure that the PM2 membrane has at least two important roles in the life cycle. First, it acts as a scaffold to nucleate capsid assembly. Second, after host recognition, it fuses with the host outer membrane to promote genome entry. The structure also sheds light on how the viral supercoiled circular double-stranded DNA genome might be packaged and released.     

The Allee Effect in Mechanistic Models Based on Inter-individual Interaction Processes

Recently, Eskola and Geritz (Bull. Math. Biol. 69:329–346, 2007) showed that several discrete-time population models can be derived mechanistically within a single ecological framework by varying the within-season patterns of reproduction and inter-individual aggression. However, these models do not have the Allee effect. In this paper, we modify the original modelling framework by adding different mate finding processes, and thus derive mechanistically several population models with the Allee effect.     

Transfer of germ plasm from the secondary to the primary gene pool in pennisetum

Summary Germ plasm from the A-genome of Pennisetum purpureum Schum. (AABB) of the secondary gene pool was transferred to cultivated pearl millet (AA) [P. glaucum (L.) R. Br.] by pollinating cytoplasmicnuclear male-sterile (cms) pearl millet with fertile allohexaploid pearl millet x P. purpureum hybrids (AAAABB). Certain allohexaploids used as pollinators on cms pearl millet resulted in 14-chromosome diploid pearl millet progenies. Three types of diploid pearl millet plants were produced in addition to the expected 28-chromosome AAAB-genome plants: (1) cms plants with only the A-genome, (2) cms plants with the A- and A-genomes, and (3) fertile plants with the A- and A-genomes. The latter group has allowed the utilization of genes for fertility restoration, stiff stalk, maturity, height, and morphological characteristics from the A-genome of P. purpureum in the pearl millet breeding program. Production of monoploid gametes by the allohexaploids appeared to be genetically controlled.     

Function of herpes simplex virus type 1 gD mutants with different receptor-binding affinities in virus entry and fusion

We have studied the receptor-specific function of four linker-insertion mutants of herpes simplex virus type 1 glycoprotein D (gD) representing each of the functional regions of gD. We used biosensor analysis to measure binding of the gD mutants to the receptors HVEM (HveA) and nectin-1 (HveC). One of the mutants, gD(inverted Delta 34t), failed to bind HVEMt but showed essentially wild-type (WT) affinity for nectin-1t. The receptor-binding kinetics and affinities of the other three gD mutants varied over a 1,000-fold range, but each mutant had the same affinity for both receptors. All of the mutants were functionally impaired in virus entry and cell fusion, and the levels of activity were strikingly similar in these two assays. gD(inverted Delta 34)-containing virus was defective on HVEM-expressing cells but did enter nectin-1-expressing cells to about 60% of WT levels. This showed that the defect of this form of gD on HVEM-expressing cells was primarily one of binding and that this was separable from its later function in virus entry. gD(inverted Delta 243t) showed WT binding affinity for both receptors, but virus containing this form of gD had a markedly reduced rate of entry, suggesting that gD(inverted Delta 243) is impaired in a postbinding step in the entry process. There was no correlation between gD mutant activity in fusion or virus entry and receptor-binding affinity. We conclude that gD functions in virus entry and cell fusion regardless of its receptor-binding kinetics and that as long as binding to a functional receptor occurs, entry will progress.     

Parental investment, sexual selection and sex ratios

Conventional sex roles imply caring females and competitive males. The evolution of sex role divergence is widely attributed to anisogamy initiating a self‐reinforcing process. The initial asymmetry in pre‐mating parental investment (eggs vs. sperm) is assumed to promote even greater divergence in post‐mating parental investment (parental care). But do we really understand the process? Trivers [Sexual Selection and the Descent of Man 1871–1971 (1972), Aldine Press, Chicago] introduced two arguments with a female and male perspective on whether to care for offspring that try to link pre‐mating and post‐mating investment. Here we review their merits and subsequent theoretical developments. The first argument is that females are more committed than males to providing care because they stand to lose a greater initial investment. This, however, commits the ‘Concorde Fallacy’ as optimal decisions should depend on future pay‐offs not past costs. Although the argument can be rephrased in terms of residual reproductive value when past investment affects future pay‐offs, it remains weak. The factors likely to change future pay‐offs seem to work against females providing more care than males. The second argument takes the reasonable premise that anisogamy produces a male‐biased operational sex ratio (OSR) leading to males competing for mates. Male care is then predicted to be less likely to evolve as it consumes resources that could otherwise be used to increase competitiveness. However, given each offspring has precisely two genetic parents (the Fisher condition), a biased OSR generates frequency‐dependent selection, analogous to Fisherian sex ratio selection, that favours increased parental investment by whichever sex faces more intense competition. Sex role divergence is therefore still an evolutionary conundrum. Here we review some possible solutions. Factors that promote conventional sex roles are sexual selection on males (but non‐random variance in male mating success must be high to override the Fisher condition), loss of paternity because of female multiple mating or group spawning and patterns of mortality that generate female‐biased adult sex ratios (ASR). We present an integrative model that shows how these factors interact to generate sex roles. We emphasize the need to distinguish between the ASR and the operational sex ratio (OSR). If mortality is higher when caring than competing this diminishes the likelihood of sex role divergence because this strongly limits the mating success of the earlier deserting sex. We illustrate this in a model where a change in relative mortality rates while caring and competing generates a shift from a mammalian type breeding system (female‐only care, male‐biased OSR and female‐biased ASR) to an avian type system (biparental care and a male‐biased OSR and ASR).     

Expansion of Parasite-Specific CD4+ and CD8+ T Cells Expressing IL-10 Superfamily Cytokine Members and Their Regulation in Human Lymphatic Filariasis

Background

Lymphatic filariasis (LF) is known to be associated with an increased production of IL-10. The role of the other IL-10 family members in the pathogenesis of infection and/or disease is not known.

Methodology/Principal Findings

We examined the expression patterns of IL-10 family members – IL-19, IL-24 and IL-26 in LF. We demonstrate that both CD4⁺ and CD8⁺ T cells express IL-19, IL-24 and IL-26 and that the frequency of CD4⁺ T cells expressing IL-19 and IL-24 (as well as IL-10) is significantly increased at baseline and following filarial antigen stimulation in patients with LF in comparison to individuals with filarial lymphedema and uninfected individuals. This CD4⁺ T cell expression pattern was associated with increased production of IL-19 and IL-24 by filarial – antigen stimulated PBMC. Moreover, the frequency of CD4⁺ and CD8⁺ T cells expressing IL-26 was significantly increased following filarial antigen stimulation in filarial lymphedema individuals. Interestingly, IL-10 blockade resulted in diminished frequencies of IL-19⁺ and IL-24⁺ T cells, whereas the addition of recombinant IL-10 resulted in significantly increased frequency of IL-19⁺ and IL-24⁺ T cells as well as significantly up regulated IL-19 and IL-24 gene expression, suggesting that IL-10 regulates IL-19 and IL-24 expression in T cells. In addition, IL-1β and IL-23 blockade also induced a diminution in the frequency of IL-19⁺ and IL-24⁺ T cells, indicating a novel role for these cytokines in the induction of IL-19 and IL-24 expressing T cells. Finally, elimination of infection resulted in significantly decreased frequencies of antigen – specific CD4⁺ T cells expressing IL-10, IL-19 and IL-24.

Conclusions

Our findings, therefore, suggest that IL-19 and IL-24 are associated with the regulation of immune responses in active filarial infection and potentially with protection against development of pathology, while IL-26 is predominantly associated with pathology in LF.     

Autophagy and KRT8/keratin 8 protect degeneration of retinal pigment epithelium under oxidative stress

Contribution of autophagy and regulation of related proteins to the degeneration of retinal pigment epithelium (RPE) in age-related macular degeneration (AMD) remain unknown. We report that upregulation of KRT8 (keratin 8) as well as its phosphorylation are accompanied with autophagy and attenuated with the inhibition of autophagy in RPE cells under oxidative stress. KRT8 appears to have a dual role in RPE pathophysiology. While increased expression of KRT8 following autophagy provides a cytoprotective role in RPE, phosphorylation of KRT8 induces pathologic epithelial-mesenchymal transition (EMT) of RPE cells under oxidative stress, which is mediated by MAPK1/ERK2 (mitogen-activated protein kinase 1) and MAPK3/ERK1. Inhibition of autophagy further promotes EMT, which can be reversed by inhibition of MAPK. Thus, regulated enhancement of autophagy with concurrent increased expression of KRT8 and the inhibition of KRT8 phosphorylation serve to inhibit oxidative stress-induced EMT of RPE cells as well as to prevent cell death, suggesting that pharmacological manipulation of KRT8 upregulation through autophagy with combined inhibition of the MAPK1/3 pathway may be attractive therapeutic strategies for the treatment of AMD.