Genotyping of <Emphasis Type="Italic">Mycobacterium tuberculosis</Emphasis> clinical isolates in two cities of Turkey: Description of a new family of genotypes that is phylogeographically specific for Asia Minor

Background  

Population-based bacterial genetics using repeated DNA loci is an efficient approach to study the biodiversity and phylogeographical structure of human pathogens, such as Mycobacterium tuberculosis, the agent of tuberculosis. Indeed large genetic diversity databases are available for this pathogen and are regularly updated. No population-based polymorphism data were yet available for M. tuberculosis in Turkey, at the crossroads of Eurasia.     

The budding yeast GSK-3 homologue Mck1 is an essential component of the spindle position checkpoint

The spindle position checkpoint (SPOC) is a mitotic surveillance mechanism in Saccharomyces cerevisiae that prevents cells from completing mitosis in response to spindle misalignment, thereby contributing to genomic integrity. The kinase Kin4, one of the most downstream SPOC components, is essential to stop the mitotic exit network (MEN), a signalling pathway that promotes the exit from mitosis and cell division. Previous work, however, suggested that a Kin4-independent pathway contributes to SPOC, yet the underlying mechanisms remain elusive. Here, we established the glycogen-synthase-kinase-3 (GSK-3) homologue Mck1, as a novel component that works independently of Kin4 to engage SPOC. Our data indicate that both Kin4 and Mck1 work in parallel to counteract MEN activation by the Cdc14 early anaphase release (FEAR) network. We show that Mck1''s function in SPOC is mediated by the pre-replication complex protein and mitotic cyclin-dependent kinase (M-Cdk) inhibitor, Cdc6, which is degraded in a Mck1-dependent manner prior to mitosis. Moderate overproduction of Cdc6 phenocopies MCK1 deletion and causes SPOC deficiency via its N-terminal, M-Cdk inhibitory domain. Our data uncover an unprecedented role of GSK-3 kinases in coordinating spindle orientation with cell cycle progression.     

Characterization and 454 pyrosequencing of Major Histocompatibility Complex class I genes in the great tit reveal complexity in a passerine system

ABSTRACT: BACKGROUND: The critical role of Major Histocompatibility Complex (Mhc) genes in disease resistance and their highly polymorphic nature make them exceptional candidates for studies investigating genetic effects on survival, mate choice and conservation. Species that harbor many Mhc loci and high allelic diversity are particularly intriguing as they are potentially under strong selection and studies of such species provide valuable information as to the mechanisms maintaining Mhc diversity. However comprehensive genotyping of complex multilocus systems has been a major challenge to date with the result that little is known about the consequences of this complexity in terms of fitness effects and disease resistance. RESULTS: In this study, we genotyped the Mhc class I exon 3 of the great tit (Parus major) from two nest-box breeding populations near Oxford, UK that have been monitored for decades. Characterization of Mhc class I exon 3 was adopted and bidirectional sequencing was carried using the 454 sequencing platform. Full analysis of sequences through a stepwise variant validation procedure allowed reliable typing of more than 800 great tits based on 214,357 reads; from duplicates we estimated the repeatability of typing as 0.94. A total of 862 alleles were detected, and the presence of at least 16 functional loci was shown - the highest number characterized in a wild bird species. Finally, the functional alleles were grouped into 17 supertypes based on their antigen binding affinities. CONCLUSIONS: We found extreme complexity at the Mhc class I of the great tit both in terms of allelic diversity and gene number. The presence of many functional loci was shown, together with a pseudogene family and putatively non-functional alleles; there was clear evidence that functional alleles were under strong balancing selection. This study is the first step towards an in-depth analysis of this gene complex in this species, which will help understanding how parasite-mediated and sexual selection shape and maintain host genetic variation in nature. We believe that study systems like ours can make important contributions to the field of evolutionary biology and emphasize the necessity of integrating long-term field-based studies with detailed genetic analysis to unravel complex evolutionary processes.     

Genome-wide identification of genes that play a role in boron stress response in yeast

Boron is an essential micronutrient for plants and it is either necessary or beneficial for animals. Studies identified only few genes related to boron metabolism thus far and details of how boron is imported into cells and used in cell metabolism are largely unknown. In order to identify genes that play roles in boron metabolism, we screened the entire set of yeast haploid deletion mutants and identified 6 mutants that were resistant to toxic levels of boron, and 21 mutants that were highly sensitive to boron treatment. Furthermore, we performed a proteomic approach to identify additional proteins that are significantly up-regulated by boron treatment. Our results revealed many genes and pathways related to boron stress response and suggest a possible link between boron toxicity and translational control.     

Investigation of PLLA/PCL Blends and Paclitaxel Release Profiles

Blends of poly (l-lactide) (PLLA) and poly (ε-caprolactone) (PCL) with and without paclitaxel were prepared via solution casting. DSC analysis as well as SEM analysis of the PLLA/PCL blend solution cast films showed that these blends are all phase separated.%PLLA crystallinity was found to increase with increasing PCL content (up till 15 wt.%). The PCL phase is found to homogeneously disperse in the PLLA matrix as spherical domains where the pore diameters of the PCL domains significantly increased with increasing PCL content. The degradation profiles matched with the slower degrading component PCL rather than PLLA and also increasing PCL content of the blends increased the degradation rate relatively. The increased crystallinity of the PLLA phase with increasing PCL contents confirmed that the degradation occurred through PCL phase. Cell proliferation on PLLA/PCL blends showed that all these blends were suitable for the support of cellular growth. Apoptosis assay with the paclitaxel-loaded PLLA/PCL blends showed an increase in cell death throughout 7 days of incubation where the cell death was increased with increasing PCL contents. This was attributed to the faster release of paclitaxel which was at least partially affected by the faster degradation rate at increasing PCL contents. The paclitaxel release was shown to be degradation controlled in the initial stages followed by a faster diffusion-controlled release in the later stages. These polymer blends were found to be very suitable paclitaxel release agents for which the paclitaxel release times can be altered with the composition of the blend and the film thickness.     

Mhc supertypes confer both qualitative and quantitative resistance to avian malaria infections in a wild bird population

Major histocompatibility complex (Mhc) genes are believed to play a key role in the genetic basis of disease control. Although numerous studies have sought links between Mhc and disease prevalence, many have ignored the ecological and epidemiological aspects of the host–parasite interaction. Consequently, interpreting associations between prevalence and Mhc has been difficult, whereas discriminating alleles for qualitative resistance, quantitative resistance and susceptibility remains challenging. Moreover, most studies to date have quantified associations between genotypes and disease status, overlooking the complex relationship between genotype and the properties of the Mhc molecule that interacts with parasites. Here, we address these problems and demonstrate avian malaria (Plasmodium) parasite species-specific associations with functional properties of Mhc molecules (Mhc supertypes) in a wild great tit (Parus major) population. We further show that correctly interpreting these associations depends crucially on understanding the spatial variation in risk of infection and the fitness effects of infection. We report that a single Mhc supertype confers qualitative resistance to Plasmodium relictum, whereas a different Mhc supertype confers quantitative resistance to Plasmodium circumflexum infections. Furthermore, we demonstrate common functional properties of Plasmodium-resistance alleles in passerine birds, suggesting this is a model system for parasite–Mhc associations in the wild.     

Repetitive Long-Term Hyperbaric Oxygen Treatment (HBOT) Administered after Experimental Traumatic Brain Injury in Rats Induces Significant Remyelination and a Recovery of Sensorimotor Function

Cells in the central nervous system rely almost exclusively on aerobic metabolism. Oxygen deprivation, such as injury-associated ischemia, results in detrimental apoptotic and necrotic cell loss. There is evidence that repetitive hyperbaric oxygen therapy (HBOT) improves outcomes in traumatic brain-injured patients. However, there are no experimental studies investigating the mechanism of repetitive long-term HBOT treatment-associated protective effects. We have therefore analysed the effect of long-term repetitive HBOT treatment on brain trauma-associated cerebral modulations using the lateral fluid percussion model for rats. Trauma-associated neurological impairment regressed significantly in the group of HBO-treated animals within three weeks post trauma. Evaluation of somatosensory-evoked potentials indicated a possible remyelination of neurons in the injured hemisphere following HBOT. This presumption was confirmed by a pronounced increase in myelin basic protein isoforms, PLP expression as well as an increase in myelin following three weeks of repetitive HBO treatment. Our results indicate that protective long-term HBOT effects following brain injury is mediated by a pronounced remyelination in the ipsilateral injured cortex as substantiated by the associated recovery of sensorimotor function.     

Screening for free radical scavenging and cell aggregation inhibitory activities by secondary metabolites from Turkish Verbascum species

Free radical scavenging and cell aggregation inhibitory activities of 36 secondary metabolites isolated from the methanolic extracts of Verbascum cilicicum Boiss., V. lasianthum Boiss. ex Bentham, V pterocalycinum var. mutense Hub.-Mor., and V. salviifolium Boiss. (Scrophulariaceae) were investigated. The isolated compounds, 6-O-vaniloyl ajugol (1), ilwensisaponin A (2), ilwensisaponin C (3), verbascoside (4), beta-hydroxyacteoside (5), martynoside (6), poliumoside (7), forsythoside B (8), angoroside A (9), dehydrodiconiferyl alcohol-9-O-beta-D-glucopyranoside (10), dehydrodiconiferyl alcohol-9'-O-beta-D-glucopyranoside (11), apigenin 7-O-beta-glucopyranoside (12), luteolin 7-O-beta-glucopyranoside (13), luteolin 3'-O-beta-glucopyranoside (14) and chrysoeriol 7-O-beta-glucopyranoside (15), exhibited a dose-dependent inhibition of bioautographic and spectrophotometric DPPH activities. Verbascoside (4) was the most active (IC50 4.0 microg/ml) comparing it to vitamin C (IC50 4.4 microg/ml) to inhibit phorbol 12-myristate 13-acetate (PMA)-induced peroxide-catalyzed oxidation of 2',7'-dichlorofluorescein (DCFH) by reactive oxygen species (ROS) within human promyelocytic HL-60 cells. Ilwensisaponin A (2) (MIC 6.9 microg/ml) showed moderate in vitro activity on lymphocyte-associated antigen-1 (LFA-1)/intercellular adhesion molecule-1 (ICAM-1)-mediated aggregation using the HL-60 cell line [positive control was cytochalasin B (MIC 2.3 microg/ml)]. None of the other compounds showed free radical scavenging and cell aggregation inhibitory activities.     

Spatial determinants of infection risk in a multi‐species avian malaria system

Spatially‐variable processes can be an important element of host–parasite interactions, but their longer term demographic and evolutionary effects depend on the magnitude of variation in space, the scale at which variation occurs and the degree to which such processes are temporally stable. Here, we use multiple years of data from a study of two closely related tit species (Paridae), infected with two congeneric species of avian malaria (Plasmodium), to evaluate the roles of extrinsic and intrinsic factors in driving spatial heterogeneity in infection risk, and to address questions of scale and temporal stability in these vector‐driven host–parasite interactions. We show that the two malaria parasite species exhibit markedly different spatial epidemiology: P. relictum infections are effectively randomly distributed in space, with no temporal consistency, whereas P. circumflexum infections exhibit pronounced spatial structuring that is stable over the six years of this study and similar in both host species. We show that both conspecific and heterospecific host density contribute to elevated infection risk, but that the main determinants of elevated risk of P. circumflexum infection risk are habitat features probably associated with vector distribution and abundance. We discuss the implications of these findings, both for our understanding of the epidemiology of malaria in the wild, but also in terms of the longer‐term evolutionary and demographic consequences that spatially variable parasite‐mediated selection may have on host populations.