A user-friendly, high-throughput tool for the precise fluorescent quantification of deoxyribonucleoside triphosphates from biological samples

Cells maintain a fine-tuned, dynamic concentration balance in the pool of deoxyribonucleoside 5′-triphosphates (dNTPs). This balance is essential for physiological processes including cell cycle control or antiviral defense. Its perturbation results in increased mutation frequencies, replication arrest and may promote cancer development. An easily accessible and relatively high-throughput method would greatly accelerate the exploration of the diversified consequences of dNTP imbalances. The dNTP incorporation based, fluorescent TaqMan-like assay published by Wilson et al. has the aforementioned advantages over mass spectrometry, radioactive or chromatography based dNTP quantification methods. Nevertheless, the assay failed to produce reliable data in several biological samples. Therefore, we applied enzyme kinetics analysis on the fluorescent dNTP incorporation curves and found that the Taq polymerase exhibits a dNTP independent exonuclease activity that decouples signal generation from dNTP incorporation. Furthermore, we found that both polymerization and exonuclease activities are unpredictably inhibited by the sample matrix. To resolve these issues, we established a kinetics based data analysis method which identifies the signal generated by dNTP incorporation. We automated the analysis process in the nucleoTIDY software which enables even the inexperienced user to calculate the final and accurate dNTP amounts in a 96-well-plate setup within minutes

Early Pliocene Hominid Tooth from Galili, Somali Region, Ethiopia

A fossil hominid tooth was discovered during survey at Galili, Somali region, Ethiopia. The geological and faunal context indicate an Early Pliocene age. The specimen (GLL 33) consists of an almost complete lower right third molar likely representing a male individual of advanced age-at-death. Its comparative metrical, morphological and (micro)structural analysis (supported by a microtomographic record) suggests a tentative taxonomic allocation to Australopithecus cf. A. afarensis

An improved and widely accessible dNTP quantitation tool

Cells maintain a fine-tuned concentration balance in the pool of deoxyribonucleoside 5’- triphosphates (dNTPs). The perturbation of this balance results in increased mutation frequencies suggested to promote cancer development and drug resistance. To study dNTP imbalances and their consequences, an accurate and relatively high-throughput method is necessary. The dNTP quantitation method of our choice is a fluorescence-based, TaqMan-like polymerase assay published by Wilson et al, NAR 2011. This assay has the advantages of being accessible in a standard molecular biology laboratory and having the potential to be automated in contrast to mass spectrometry or radioactive measurements. Although this method works well in diluted samples with high dNTP levels, we observed that the sample matrix largely decreases assay performance. Upon thorough kinetic analysis of the fluorescent dNTP incorporation curves, we found that the Taq polymerase exhibits a dNTP independent, signal generating exonuclease activity and that the polymerization and exonuclease activity are partially inhibited by the sample matrix. Based on our kinetic investigations we suggest several assay modifications and a novel, kineticsbased and automated analysis method. Using these modifications, we measured dNTP pools in widely different organisms including Mycobacterium smegmatis, Staphylococcus aureus and human cancer cells. We found that our improved method is capable of i) determining dNTP concentrations in samples previously proved to be unmeasurable by eliminating the interfering matrix effect, and ii) improving the quantitation limits of the assay. Fundings: NKFIH-PD 124330, NKFIH-K 115993, János Bolyai Research Scholarshi

Generation Times in Wild Chimpanzees and Gorillas Suggest Earlier Divergence Times in Great Ape and Human Evolution

Fossils and molecular data are two independent sources of information that should in principle provide consistent inferences of when evolutionary lineages diverged. Here we use an alternative approach to genetic inference of species split times in recent human and ape evolution that is independent of the fossil record. We first use genetic parentage information on a large number of wild chimpanzees and mountain gorillas to directly infer their average generation times. We then compare these generation time estimates with those of humans and apply recent estimates of the human mutation rate per generation to derive estimates of split times of great apes and humans that are independent of fossil calibration. We date the human–chimpanzee split to at least 7–8 million years and the population split between Neanderthals and modern humans to 400,000–800,000 y ago. This suggests that molecular divergence dates may not be in conflict with the attribution of 6- to 7-million-y-old fossils to the human lineage and 400,000-y-old fossils to the Neanderthal lineage.Human Evolutionary Biolog

The effects of acute and elective cardiac surgery on the anxiety traits of patients with Marfan syndrome

BACKGROUND: Marfan syndrome is a genetic disease, presenting with dysfunction of connective tissues leading to lesions in the cardiovascular and skeletal muscle system. Within these symptoms, the most typical is weakness of the connective tissue in the aorta, manifesting as aortic dilatation (aneurysm). This could, in turn, become annuloaortic ectasia, or life-threatening dissection. As a result, life-saving and preventative cardiac surgical interventions are frequent among Marfan syndrome patients. Aortic aneurysm could turn into annuloaortic ectasia or life-threatening dissection, thus life-saving and preventive cardiac surgical interventions are frequent among patients with Marfan syndrome. We hypothesized that patients with Marfan syndrome have different level of anxiety, depression and satisfaction with life compared to that of the non-clinical patient population. METHODS: Patients diagnosed with Marfan syndrome were divided into 3 groups: those scheduled for prophylactic surgery, those needing acute surgery, and those without need for surgery (n = 9, 19, 17, respectively). To examine the psychological features of the patients, Spielberger's anxiety (STAI) test, Beck's Depression questionnaire (BDI), the Berne Questionnaire of Subjective Well-being, and the Satisfaction with Life scale were applied. RESULTS: A significant difference was found in trait anxiety between healthy individuals and patients with Marfan syndrome after acute life-saving surgery (p 0.1). Finally, a significant, medium size effect was found between patient groups on the Joy in Living scale (F (2.39) = 3.51, p = 0.040, eta2 = 0.15). CONCLUSIONS: Involving psychiatric and mental-health care, in addition to existing surgical treatment interventions, is essential for more successful recovery of patients with Marfan syndrome

The complete genome sequence of a Neandertal from the Altai Mountains

We present a high-quality genome sequence of a Neandertal woman from Siberia. We show that her parents were related at the level of half siblings and that mating among close relatives was common among her recent ancestors. We also sequenced the genome of a Neandertal from the Caucasus to low coverage. An analysis of the relationships and population history of available archaic genomes and 25 present-day human genomes shows that several gene flow events occurred among Neandertals, Denisovans and early modern humans, possibly including gene flow into Denisovans from an unknown archaic group. Thus, interbreeding, albeit of low magnitude, occurred among many hominin groups in the Late Pleistocene. In addition, the high quality Neandertal genome allows us to establish a definitive list of substitutions that became fixed in modern humans after their separation from the ancestors of Neandertals and Denisovans

Reducing the environmental impact of surgery on a global scale: systematic review and co-prioritization with healthcare workers in 132 countries

Abstract Background Healthcare cannot achieve net-zero carbon without addressing operating theatres. The aim of this study was to prioritize feasible interventions to reduce the environmental impact of operating theatres. Methods This study adopted a four-phase Delphi consensus co-prioritization methodology. In phase 1, a systematic review of published interventions and global consultation of perioperative healthcare professionals were used to longlist interventions. In phase 2, iterative thematic analysis consolidated comparable interventions into a shortlist. In phase 3, the shortlist was co-prioritized based on patient and clinician views on acceptability, feasibility, and safety. In phase 4, ranked lists of interventions were presented by their relevance to high-income countries and low–middle-income countries. Results In phase 1, 43 interventions were identified, which had low uptake in practice according to 3042 professionals globally. In phase 2, a shortlist of 15 intervention domains was generated. In phase 3, interventions were deemed acceptable for more than 90 per cent of patients except for reducing general anaesthesia (84 per cent) and re-sterilization of ‘single-use’ consumables (86 per cent). In phase 4, the top three shortlisted interventions for high-income countries were: introducing recycling; reducing use of anaesthetic gases; and appropriate clinical waste processing. In phase 4, the top three shortlisted interventions for low–middle-income countries were: introducing reusable surgical devices; reducing use of consumables; and reducing the use of general anaesthesia. Conclusion This is a step toward environmentally sustainable operating environments with actionable interventions applicable to both high– and low–middle–income countries

Palaeoenvironments during the period of the Neanderthals settlement in Chagyrskaya cave (Altai Mountains, Russia)

The Altai Mountains, situated in the middle of Asia, have been inhabited by human groups since prehistoric times. Many Middle Palaeolithic sites (open-air sites and caves) are located in the northwestern part of the Altai Mountains. The uniqueness of this area is in the simultaneous habitation of different human species, such as Neanderthals, Sapiens and Denisovians. The material culture of the Altai Middle Palaeolithic is mostly homogeneous; however, two caves are distinguished from other sites—Okladnikov Cave and the recently studied Chagyrskaya Cave, located in the Charysh River valley. Palaeolithic assemblages from both caves are comparable with the Mousterian industries recorded in certain regions of Eurasia and represent a special variant of the Altai Middle Palaeolithic industries, known as the Sibiryachikha facies. Anthropological data from these caves suggest that the Sibiryachikha variant was associated with Neanderthals. In this study, we reconstruct the palaeoenvironmental conditions of the period of the Neanderthals settlement in the northwestern part of the Altai Mountains based on bioproxies, such as pollen records and large mammal remains. The time of the Neanderthals settlement of Chagyrskaya Cave is attributed to the termination of MIS4 and is characterised by an arid and continental climate. Dry steppe communities were widespread in the Charysh River valley. It is possible that the Chagyrskaya Cave represents a long-term hunting camp where butchering and processing of game animals were carried out. In the following warmer and more humid period the Neanderthals left the cave. This was around the same time when anatomically modern humans began appearing in Western Siberia