Miracle-dz/UniFAR-FTUs huggingface dataset:
Direct Cloning from Enrichment Cultures, a Reliable Strategy for Isolation of Complete Operons and Genes from Microbial Consortia

ABSTRACT Enrichment cultures of microbial consortia enable the diverse metabolic and catabolic activities of these populations to be studied on a molecular level and to be explored as potential sources for biotechnology processes. We have used a combined approach of enrichment culture and direct cloning to construct cosmid libraries with large (>30-kb) inserts from microbial consortia. Enrichment cultures were inoculated with samples from five environments, and high amounts of avidin were added to the cultures to favor growth of biotin-producing microbes. DNA was extracted from three of these enrichment cultures and used to construct cosmid libraries; each library consisted of between 6,000 and 35,000 clones, with an average insert size of 30 to 40 kb. The inserts contained a diverse population of genomic DNA fragments isolated from the consortia organisms. These three libraries were used to complement the Escherichia coli biotin auxotrophic strain ATCC 33767 Δ(bio-uvrB). Initial screens resulted in the isolation of seven different complementing cosmid clones, carrying biotin biosynthesis operons. Biotin biosynthesis capabilities and growth under defined conditions of four of these clones were studied. Biotin measured in the different culture supernatants ranged from 42 to 3,800 pg/ml/optical density unit. Sequencing the identified biotin synthesis genes revealed high similarities to biooperons from gram-negative bacteria. In addition, random sequencing identified other interesting open reading frames, as well as two operons, the histidine utilization operon (hut), and the cluster of genes involved in biosynthesis of molybdopterin cofactors in bacteria (moaABCDE).
Reservosome: an endocytic compartment in epimastigote forms of the protozoan Trypanosoma cruzi (Kinetoplastida: Trypanosomatidae). Correlation between endocytosis of nutrients and cell differentiation

Reservosomes are large membrane-bound organelles found at the posterior end of epimastigote forms of Trypanosoma cruzi, but absent in amastigotes and trypomastigotes. We have transferred bloodstream trypomastigotes to LIT medium supplemented with gold-labelled transferrin in order to analyse, at the ultrastructural level, the occurrence of reservosomes and endocytosis during the trypomastigote to epimastigote differentiation. After 24 h, the trypomastigotes differentiated into amastigotes, which adhered to each other forming large clusters. Electron-dense vesicles were detected close to the Golgi complex in cells with intermediary characteristics between amastigotes and epimastigotes, but typical reservosomes at the posterior cell tip were still absent. Transferrin–gold complexes were observed only bound to the surface of clustered cells. After 72 h, epimastigotes were observed being released from the clusters and free-swimming epimastigotes appeared, containing electron-dense vesicles at their posterior region. Typical reservosomes, labelled with transferrin–gold, were observed only in free-swimming epimastigotes. When fully differentiated epimastigotes were incubated with transferrin–gold complexes and then processed for the immunocytochemical detection of cysteine proteinase, all reservosomes were positive for the enzyme, but co-localization of both markers did not occur in all organelles. Our data demonstrate that in T. cruzi epimastigotes endocytosis is strongly related to reservosome biogenesis during the trypomastigote to epimastigote differentiation process.
Study of arterial pattern of 200 renal pedicle through angiotomography.

OBJECTIVE To investigate the prevalence and distribution of renal arteries and their branches in vivo, correlating the particularities found in them with sex and laterality. METHODS Two hundred renal pedicles were studied by CT angiography and its arteries analyzed according to number, position of origin, size, length and trajectory in relation to renal segments. Its frequency and laterality were surveyed regarding gender and age. RESULTS There were multiple arteries in 61.5% of the pedicles (56% in the right and 67% in the left), occurring in 65% of men and 58% of women. The aortic origin to the multiple arteries was more frequent on the right and, more often, the renal arteries originated between vertebrae L1 and L2 as pre-hilar division of the main artery. The average length of the main artery was higher in right kidneys with a single artery. There was no difference between the diameters of the main renal arteries. CONCLUSION There is a higher prevalence of multiple renal arteries than the one described in the literature, with no difference for gender or laterality. The renal arteries originated more frequently between vertebrae L1 and L2, with divisions of the pre-hilar route and main artery to the hilum of the kidney. The average length of the main artery is greater on the right and in kidneys with single artery. There was no difference in diameter between the main renal artery between kidneys with single and multiple arteries.
Forced unfolding of fibronectin type 3 modules: an analysis by biased molecular dynamics simulations.

Titin, an important constituent of vertebrate muscles, is a protein of the order of a micrometer in length in the folded state. Atomic force microscopy and laser tweezer experiments have been used to stretch titin molecules to more than ten times their folded lengths. To explain the observed relation between force and extension, it has been suggested that the immunoglobulin and fibronectin domains unfold one at a time in an all-or-none fashion. We use molecular dynamics simulations to study the forced unfolding of two different fibronectin type 3 domains (the ninth, 9Fn3, and the tenth, 10Fn3, from human fibronectin) and of their heterodimer of known structure. An external biasing potential on the N to C distance is employed and the protein is treated in the polar hydrogen representation with an implicit solvation model. The latter provides an adiabatic solvent response, which is important for the nanosecond unfolding simulation method used here. A series of simulations is performed for each system to obtain meaningful results. The two different fibronectin domains are shown to unfold in the same way along two possible pathways. These involve the partial separation of the "beta-sandwich", an essential structural element, and the unfolding of the individual sheets in a stepwise fashion. The biasing potential results are confirmed by constant force unfolding simulations. For the two connected domains, there is complete unfolding of one domain (9Fn3) before major unfolding of the second domain (10Fn3). Comparison of different models for the potential energy function demonstrates that the dominant cohesive element in both proteins is due to the attractive van der Waals interactions; electrostatic interactions play a structural role but appear to make only a small contribution to the stabilization of the domains, in agreement with other studies of beta-sheet stability. The unfolding forces found in the simulations are of the order of those observed experimentally, even though the speed of the former is more than six orders of magnitude greater than that used in the latter.
Resistance to activated protein C (APCR) in children with acute lymphoblastic leukaemia—the need for a prospective multicentre study

Activated protein C resistance (APCR), usually due to the Arg506→Gln point mutation of the factor V gene, has emerged as the most important hereditary cause of venous thromboembolism. Using an aPTT based method in the presence of APC, together with a DNA technique based on the polymerase chain reaction, we investigated 65 leukaemic children and 65 age-matched healthy controls for the presence of this mutation. In both groups three children showed APCR, All six children showed the common factor V gene mutation, Arg506→Gln. Although no child in the control group presented with thrombosis, all three children with acute lymphoblastic leukaemia had thromboembolic events. Whether the poor anticoagulant response to activated protein C in leukaemic children treated with prednisone, vincristine, daunorubicin and asparaginase affects the risk of thrombotic events requires a more extensive multicentre study.
Intelligent Online Measurement and Management of Energy Meter Data through Advanced Wireless Network

Recently the Electrical supply companies are trying to adopt the electronic measurement of energy consumption data because of reduced manufacturing cost, improved measurement accuracy, increased timely information, miniature size and many other benefits that go well beyond the traditional rotor-plate energy meter type. In this paper with the help of an energy chip, an improved energy metering solution is developed, where automating the progression of measurement through digital wireless communication technique is adopted to get the above benefits alongwith smooth control. The developed energy meter calculates the total average active power mainly for residential consumers. The hardware circuit accepts single phase voltage and currents as its inputs and provides the output in the form of logic data proportional to the average real power. This data is fed to a remote computer server through the wireless ZigBee network that represents the concept of distant wireless metering, practically involving no manpower. This paper also presents a software solution developed for total electrical energy billing and data management system.
Combined subthreshold dose inhibition of myosin light chain phosphorylation and MMP‐2 activity provides cardioprotection from ischaemic/reperfusion injury in isolated rat heart

Phosphorylation and degradation of myosin light chain 1 (MLC1) during myocardial ischaemia/reperfusion (I/R) injury is a well‐established phenomenon. It has been established that MMP‐2 is involved in MLC1 degradation and that this degradation is increased when MLC1 is phosphorylated. We hypothesized that simultaneous inhibition of MLC1 phosphorylation and MMP‐2 activity will protect hearts from I/R injury. As phosphorylation of MLC1 and MMP‐2 activity is important for normal heart function, we used a cocktail consisting combination of low (subthreshold for any protective effect alone) doses of MLC kinase, MMP‐2 inhibitors and subthreshold dose of an MLC phosphatase activator.
Vasorelaxation Induced by a New Naphthoquinone-Oxime is Mediated by NO-sGC-cGMP Pathway

It has been established that oximes cause endothelium-independent relaxation in blood vessels. In the present study, the cardiovascular effects of the new oxime 3-hydroxy-4–(hydroxyimino)-2-(3-methylbut-2-enylnaphtalen-1(4H)-one (OximeS1) derived from lapachol were evaluated. In normotensive rats, administration of Oxime S1 (10, 15, 20 and 30 mg/Kg, i.v.) produced dose-dependent reduction in blood pressure. In isolated aorta and superior mesenteric artery rings, Oxime S1 induced endothelium-independent and concentration-dependent relaxations (10−8 M to 10−4 M). In addition, Oxime S1-induced vasorelaxations were attenuated by hydroxocobalamin or methylene blue in aorta and by PTIO or ODQ in mesenteric artery rings, suggesting a role for the nitric oxide (NO) pathway. Additionally, Oxime S1 (30 and 100 µM) significantly increased NO concentrations (13.9 ± 1.6 nM and 17.9 ± 4.1 nM, respectively) measured by nitric oxide microsensors. Furthermore, pre-contraction with KCl (80 mM) prevented Oxime S1-derived vasorelaxation in endothelium-denuded aortic rings. Of note, combined treatment with potassium channel inhibitors also reduced Oxime S1-mediated vasorelaxation suggesting a role for potassium channels, more precisely Kir, Kv and KATP channels. We observed the involvement of BKCa channels in Oxime S1-induced relaxation in mesenteric artery rings. In conclusion, these data suggest that the Oxime S1 induces hypotension and vasorelaxation via NO pathway by activating soluble guanylate cyclase (sGC) and K+ channels.
SleC Is Essential for Germination of Clostridium difficile Spores in Nutrient-Rich Medium Supplemented with the Bile Salt Taurocholate

ABSTRACT Clostridium difficile is the major cause of infectious diarrhea and a major burden to health care services. The ability of this organism to form endospores plays a pivotal role in infection and disease transmission. Spores are highly resistant to many forms of disinfection and thus are able to persist on hospital surfaces and disseminate infection. In order to cause disease, the spores must germinate and the organism must grow vegetatively. Spore germination in Bacillus is well understood, and genes important for this process have recently been identified in Clostridium perfringens; however, little is known about C. difficile. Apparent homologues of the spore cortex lytic enzyme genes cwlJ and sleB (Bacillus subtilis) and sleC (C. perfringens) are present in the C. difficile genome, and we describe inactivation of these homologues in C. difficile 630Δerm and a B1/NAP1/027 clinical isolate. Spores of a sleC mutant were unable to form colonies when germination was induced with taurocholate, although decoated sleC spores formed the same number of heat-resistant colonies as the parental control, even in the absence of germinants. This suggests that sleC is absolutely required for conversion of spores to vegetative cells, in contrast to CD3563 (a cwlJ/sleB homologue), inactivation of which had no effect on germination and outgrowth of C. difficile spores under the same conditions. The B1/NAP1/027 strain R20291 was found to sporulate more slowly and produce fewer spores than 630Δerm. Furthermore, fewer R20291 spores germinated, indicating that there are differences in both sporulation and germination between these epidemic and nonepidemic C. difficile isolates.
The impact of surgical technique on postoperative hypoparathyroidism in bilateral thyroid surgery: a multivariate analysis of 5846 consecutive patients.

BACKGROUND Limited information exists about risk factors for postoperative hypoparathyroidism after bilateral thyroid surgery. METHODS Between January 1 and December 31, 1998, bilateral thyroid surgery was performed on 5846 patients for benign and malignant thyroid disease. Data were prospectively collected by questionnaires from 45 hospitals. A logistic regression model was used to determine independent risk factors. RESULTS The overall incidence of transient and permanent hypoparathyroidism was 7.3% and 1.5%, respectively. On logistic regression analysis, total thyroidectomy (odds ratio [OR], 4.7), female gender (OR, 1.9), Graves' disease (OR, 1.9), recurrent goiter (OR, 1.7), and bilateral central ligation of the inferior thyroid artery (OR, 1.7) constituted independent risk factors for transient hypoparathyroidism. When the multivariate analysis was confined to permanent hypoparathyroidism, total thyroidectomy (OR, 11.4), bilateral central (OR, 5.0) and peripheral (OR, 2.0) ligation of the inferior thyroid artery, identification and preservation of no or only a single parathyroid gland (OR, 4.1), and Graves' disease (OR, 2.4) emerged as independent risk factors. CONCLUSIONS Extent of resection and surgical technique had a greater impact on the rates of permanent postoperative hypoparathyroidism than thyroid pathologic condition. In bilateral thyroid surgery, peripheral ligation of the inferior thyroid artery at the thyroid capsule should be favored over central ligation, and at least 2 parathyroid glands should be identified and preserved. High-risk procedures, such as total thyroidectomy and Graves' disease, require special surgical training and expertise.
VEGF enhances pulmonary vasculogenesis and disrupts lung morphogenesis in vivo

Vascular endothelial growth factor (VEGF) was expressed in developing respiratory epithelial cells under control of the promoter from the human surfactant protein C (SP‐C) gene. SP‐C‐VEGF transgenic mice did not survive after birth. When obtained by hysterectomy on embryonic day 15 (E15) or 17 (E17), abnormalities in the transgenic mice were confined to the lung and were correlated with the expression of transgene mRNA as revealed by in situ hybridization. On E15 and E17, marked abnormalities in lung morphogenesis were observed in transgenic mice. Lungs consisted of large dilated tubules with increased peritubular vascularity. The mRNA levels of the VEGF receptor, Flk‐1, and the endothelial cell specific receptor tyrosine kinase, Tie‐1, were increased in lung mesenchyme of the transgenic mice. The numbers of acinar tubules and the abundance of mesenchyme were decreased. Endogenous VEGF mRNA was expressed in the respiratory epithelial cells of the developing lungs, and the levels of VEGF mRNA were increased in the SP‐C‐VEGF transgenic mice. Although the normal pattern of immunostaining for SP‐C and Clara cell secretory protein (CCSP) indicated that epithelial cell differentiation was relatively unaltered by the transgene, electron microscopic analysis revealed a lack of alveolar Type I cell differentiation at E18. Expression of VEGF in the developing respiratory epithelium of transgenic mice increased growth of the pulmonary blood vessels, disrupted branching morphogenesis of the lung and inhibited Type I cell differentiation. Dev. Dyn. 1998;211:215–227. © 1998 Wiley‐Liss, Inc.
Effect of repeated caffeine ingestion on repeated exhaustive exercise endurance.

PURPOSE The purpose of this study was to examine the effect of repeated doses of caffeine on repeated exercise endurance. METHODS Nine male caffeine users performed exercise rides (ER) to exhaustion at 80% VO(2max) after ingesting a placebo, 5 mg x kg-1 of caffeine, or 2.5 mg x kg-1 of caffeine 1 h before the ER. Two ER were performed weekly on the same day once in the morning (AM) and 5 h later in the afternoon (PM). There were four treatments containing either caffeine or placebo, i.e., trial A representing 5-mg x kg-1 caffeine in the AM and 2.5-mg x kg-1 caffeine in the PM; trial B, which was placebo in both AM and PM; trial C representing 5-mg x kg-1 caffeine in the AM and placebo in the PM; and trial D representing a placebo in the AM and 5-mg x kg-1 caffeine in the PM. The order of the treatment trials was double blind and randomized. RESULTS Caffeine ingestion significantly increased exercise time to exhaustion in the AM (trial A 24.9 +/- 10.2 min and trial C 21.8 +/- 4.9 vs trial B 18.0 +/- 6.4 min and D 17.7 +/- 4.3 min). This effect was maintained in the PM and greater than placebo (B 18.3 +/- 4.8 min) regardless of whether redosing (trial A 21.5 +/- 8.6 min) or placebo (trial C 21.0 +/- 6.8) followed the initial morning dose. Caffeine dosing in the PM (trial D 22.4 +/- 7.2 min) also increased ER after placebo trial D in the AM. CONCLUSIONS It was concluded that redosing with caffeine after exhaustive exercise in the AM was not necessary to maintain the ergogenic effect of the drug during subsequent exercise 6 h later.
Quantitative evaluation of AgNORs in bone tumours

Aims: Primary tumours of bone present a significant diagnostic and therapeutic challenge at times. Silver stained nucleolar organiser regions (AgNORs) have been widely used in a variety of tissues but with a limited study on bone tumours. Our study was aimed at the evaluation of AgNOR count in various neoplastic lesions of bone. Methods: One hundred biopsies of bone lesions were included in this study. Five samples of foetal lumbar vertebrae obtained from foetal autopsies were taken as control. The study included 58 males and 42 females with age ranging from 5 to 70 years. Fifty‐two cases were malignant while 48 were benign in nature. Silver staining for nuclear organiser regions was performed according to one‐step silver staining technique in these cases. NORs seen as black dots were counted in the nuclei of 100 cells. Results: Our study revealed that the mean count was highest in malignant lesions (4.00±0.69) compared with benign lesions (2.16±0.43) and normal bone (1.32±0.14). Statistically, the AgNOR count showed a significant difference (P<0.001) in all these lesions. Conclusions: The results of the current study revealed that malignant lesions had a greater mean AgNOR count than benign tumours and the normal bone. Thus, quantification of AgNORs strongly correlates with the type as well as aggressiveness of the bone tumour and is diagnostically useful in tumour differentiation.
Volumetric MRI, pathological, and neuropsychological progression in hippocampal sclerosis

Objective: To examine the relationships between age at onset and duration of seizure disorder with severity of hippocampal sclerosis (HS) and cognitive functioning in patients with HS and unilateral temporal lobe epilepsy. Methods: Twenty-six subjects had left temporal lobe seizure onset; 20 had right temporal onset. Measures were age at seizure onset, duration of seizure disorder divided by age (seizure duration), history of febrile convulsion (FC), ratio of the smaller hippocampal volume to the larger (HF) as determined by volumetric MRI, and pathologic HS grade. Results: Results showed that pathologic HS grade and HF were positively related to seizure duration, and negatively related to seizure onset. When subjects were divided into onset prior to age 10 versus later, subjects with earlier onset had higher mean pathologic HS grade and smaller (more asymmetric) mean HF. When subjects were divided into seizure duration <0.5 (i.e., less than half current lifetime) vs greater, subjects with seizure duration ≥0.5 had higher mean pathologic HS grade and lower mean HF. There was also evidence for earlier age at seizure onset and longer seizure duration being associated with worse performance on neuropsychological measures. FC was not related to either seizure duration or age at seizure onset, but patients with a history of FC showed higher pathologic HS grade and lower HF. A history of FC was not related to cognitive functioning. Conclusions: Unilateral HS patients with earlier seizure onset and longer duration of epilepsy have more severe HS and greater hippocampal volume asymmetry. This suggests that HS may be a progressive disorder with risk for cognitive dysfunction.
Differential expression of placental (P)‐cadherin in sertoli cells and peritubular myoid cells during postnatal development of the mouse testis

In previous work, RNA transcripts for placental (P)‐cadherin, a calcium‐dependent cell adhesion molecule, were identified in the rat testis during the first 4 weeks of postnatal development. However, the cells in the testis responsible for P‐cadherin expression have not yet been identified.