DNA polymerase kappa from Trypanosoma cruzi localizes to the mitochondria, bypasses 8-oxoguanine lesions and performs DNA synthesis in a recombination intermediate

DNA polymerase kappa (Polκ) is a low-fidelity polymerase that has the ability to bypass several types of lesions. The biological role of this enzyme, a member of the DinB subfamily of Y-family DNA polymerases, has remained elusive. In this report, we studied one of the two copies of Polκ from the protozoan Trypanosoma cruzi (TcPolκ). The role of this TcPolκ copy was investigated by analysing its subcellular localization, its activities in vitro , and performing experiments with parasites that overexpress this polymerase. The TcPOLK sequence has the N-terminal extension which is present only in eukaryotic DinB members, but its C-terminal region is more similar to prokaryotic and archaeal counterparts since it lacks C₂HC motifs and PCNA interaction domain. Our results indicate that in contrast to its previously described orthologues, this polymerase is localized to mitochondria. The overexpression of TcPOLK increases T. cruzi resistance to hydrogen peroxide, and in vitro polymerization assays revealed that TcPolκ efficiently bypasses 8-oxoguanine lesions. Remarkably, our results also demonstrate that the DinB subfamily of polymerases can participate in homologous recombination, based on our findings that TcPolκ increases T. cruzi resistance to high doses of gamma irradiation and zeocin and can catalyse DNA synthesis within recombination intermediates.     

Ion Channel Blockers as Antimicrobial Agents,Efflux Inhibitors,and Enhancers of Macrophage Killing Activity against Drug Resistant Mycobacterium tuberculosis

Given the ability of M. tuberculosis to survive as an intracellular pathogen and its propensity to develop resistance to the existing antituberculosis drugs, its treatment requires new approaches. Here the antimycobacterial properties of verapamil, thioridazine, chlorpromazine, flupenthixol and haloperidol were investigated against a panel of drug resistant M. tuberculosis strains, both in vitro and on human-infected macrophages. These compounds are efflux inhibitors that share among them the characteristic of being ion channel blockers. In vitro, all compounds exhibited synergistic inhibitory activities when combined with isoniazid and rifampicin, and were able to inhibit active efflux, demonstrating their role as efflux inhibitors. Gene expression analysis showed that M. tuberculosis efflux genes were overexpressed in response to antibiotic exposure, in vitro and within macrophages, irrespective of their resistance pattern. These compounds displayed a rapid and high killing activity against M. tuberculosis, associated with a decrease in intracellular ATP levels demonstrating that the bactericidal action of the ion channel blockers against M. tuberculosis clinical strains is associated with their interference with energy metabolism. The compounds led to a decrease in the intracellular mycobacterial load by increasing phagosome acidification and activating lysosomal hydrolases. The results presented in this study enable us to propose the following mechanism of action for these compounds: a) in the bacteria, the compounds generate a cascade of events involving the inhibition of the respiratory chain complexes and energy production for efflux activity. Indirectly, this reduce the resistance level to antituberculosis drugs potentiating their activity; b) on the host cell, the treatment with the ion channel blockers increases phagosome acidification and induces the expression of phagosomal hydrolases, leading to bacterial growth restriction irrespective of their resistance pattern. This work highlights the potential value ion channel blockers as adjuvants of tuberculosis chemotherapy, in particular for the development of new therapeutic strategies, with strong potential for treatment shortening against drug susceptible and resistant forms of tuberculosis. Medicinal chemistry studies are now needed to improve the properties of these compounds, increasing their M. tuberculosis efflux-inhibition and killing-enhancement activity and reduce their toxicity for humans, therefore optimizing their potential for clinical usage.     

Application of a roding survey method to the sedentary Eurasian Woodcock <Emphasis Type="Italic">Scolopax rusticola</Emphasis> population in Pico Island,Azores

The Eurasian Woodcock Scolopax rusticola is a resident species in Azores, Madeira and Canaries (Macaronesian archipelagos) in contrast to its mainly migrant continental populations. The biology and ecology of these insular populations are still poorly known; however, woodcocks are hunted in Azores and Madeira. This work aims to continuing bridging the gap between scientific knowledge and game management of woodcocks in Azores. Working at Pico Island, the main objectives were: (a) to characterize the roding activity of these insular populations, (b) to select the best period of the breeding season to perform a roding survey and (c) to evaluate how sensitive is this method to variations in abundance because of hunting. From January to July 2002, the number of contacts with roding birds (males) was recorded, at dusk, at three observation points. Roding started in the beginning of February and lasted until late June/early July. The weekly variation in the number of contacts was similar in all points, strongly suggesting that the roding survey should be performed between the beginning of March and middle April. Two of the three observation points, located in an area where woodcock hunting is allowed in alternate years, were surveyed between March 1 and April 15, 2001–2005. The two points presented a similar annual variation pattern in the number of contacts, also suggesting stability in abundance. Sex ratio among birds bagged during three consecutive hunting seasons (October–November, 2002–2004) or among birds collected periodically during two winters (2000–2002) was equal to one, suggesting that annual fluctuations by the roding survey can be extended to the female fraction.     

Albinism in plants – far beyond the loss of chlorophyll: Structural and physiological aspects of wild‐type and albino royal poinciana (Delonix regia) seedlings

• The partial or complete loss of chlorophylls, or albinism, is a rare phenomenon in plants. In the present study, we provide the first report of the occurrence in albino Delonix regia seedlings and describe the morpho‐physiological changes associated with albinism.
• Wild‐type (WT) and albino seedlings were characterized. Leaflets samples were processed following common procedures for analysis with light, scanning and transmission electron microscopy. The chlorophyll a fluorescence parameters and the carbohydrate, lipid and soluble protein content were also determined in leaf and cotyledon samples of both albino and WT seedlings.
• Albino seedlings showed reduced growth. They also had lower chlorophyll and protein content in foliar tissues than WT seedlings, in addition to lower concentrations of lipids and carbohydrates stored in cotyledons. The chloroplasts of albino seedlings were poorly developed, with an undefined internal membrane system and the presence of plastoglobules. Wild‐type seedlings had a uniseriate and hypoestomatic epidermis. The mesophyll was dorsiventral, consisting of a layer of palisade parenchyma and two to four layers of spongy parenchyma. In albino seedlings, the spongy parenchyma was compact, with few intercellular spaces, and the thickness of the mesophyll was larger, resulting in increased thickness of the leaf blade. Albino seedlings had higher stomatal density and number of pavement cells, although the stomata had smaller dimensions.
• In addition to the partial loss of chlorophylls, albino D. regia showed changes at physiological and structural levels, demonstrating the crucial nature of photosynthetic pigments during the development and differentiation of plant leaf tissues/cells.

     

Correlation between circulating biomarkers of oxidative stress of maternal and umbilical cord blood at birth

The objective of the work was to study the relationship between the oxidative state of the mother and the newborn at the moment of birth. We measured oxidative stress markers (carbonyl groups, lipid peroxides and total antioxidant capacity (TAC)) and found a good correlation between the oxidative state of the normal mother and the neonate, since a high mother oxidative stress corresponds to an even higher oxidative stress of the newborn in umbilical cord blood. We also found that smoking mothers and their newborns had a higher concentration of the carbonyl group, lipid peroxides and less TAC. Newborns from these mothers weighed significantly less than others at birth. These data suggest a need for interest in monitoring the oxidative state of mothers during the pregnancy period, especially taking into account that the oxidative level could be involved in later risks of metabolic diseases for both mother and newborn.     

Frequency of the musculus palmaris longus studied in vivo in some Amazon indians

The frequency of the Musclus palmaris longus was studied by observing its tendon in vivo in 379 Amazon Indians belonging to the following tribes: Tucano, Tariana, Tiriyo, Desana, Piratapuya, Macu and Arapaso. The muscle was not observed in 14 individuals (3.7% ± 1.0) either unilaterally or bilaterally and in 24 of 758 limbs (3.1% ± 0.6). No statistically significant difference was observed in the frequency of the muscle among the different tribes. Bilateral absence was significantly more frequent than the unilateral one; absence in females was more frequent than in males. The frequency of palmaris longus agenesis in Amazon Indians agrees with reported values for Negroids and Mongoloids and is considerably lower than that generally reported for the Caucasoids.     

Dynamic Analysis and Pattern Visualization of Forest Fires

This paper analyses forest fires in the perspective of dynamical systems. Forest fires exhibit complex correlations in size, space and time, revealing features often present in complex systems, such as the absence of a characteristic length-scale, or the emergence of long range correlations and persistent memory. This study addresses a public domain forest fires catalogue, containing information of events for Portugal, during the period from 1980 up to 2012. The data is analysed in an annual basis, modelling the occurrences as sequences of Dirac impulses with amplitude proportional to the burnt area. First, we consider mutual information to correlate annual patterns. We use visualization trees, generated by hierarchical clustering algorithms, in order to compare and to extract relationships among the data. Second, we adopt the Multidimensional Scaling (MDS) visualization tool. MDS generates maps where each object corresponds to a point. Objects that are perceived to be similar to each other are placed on the map forming clusters. The results are analysed in order to extract relationships among the data and to identify forest fire patterns.     

Distinct Phenotypes Caused by Mutation of MSH2 in Trypanosome Insect and Mammalian Life Cycle Forms Are Associated with Parasite Adaptation to Oxidative Stress

BackgroundDNA repair mechanisms are crucial for maintenance of the genome in all organisms, including parasites where successful infection is dependent both on genomic stability and sequence variation. MSH2 is an early acting, central component of the Mismatch Repair (MMR) pathway, which is responsible for the recognition and correction of base mismatches that occur during DNA replication and recombination. In addition, recent evidence suggests that MSH2 might also play an important, but poorly understood, role in responding to oxidative damage in both African and American trypanosomes.Conclusions/SignificanceTaken together, these results indicate MSH2 displays conserved, dual roles in MMR and in the response to oxidative stress. Loss of the latter function results in life cycle dependent differences in phenotypic outcomes in T. brucei MSH2 mutants, most likely because of the greater burden of oxidative stress in the insect stage of the parasite.