End Of Fragment Research Articles

Proximal-end bias from in-vitro reconstituted nucleosomes and the result on downstream data analysis.

The most basic level of eukaryotic gene regulation is the presence or absence of nucleosomes on DNA regulatory elements. In an effort to elucidate in vivo nucleosome patterns, in vitro studies are frequently used. In vitro, short DNA fragments are more favorable for nucleosome formation, increasing the likelihood of nucleosome occupancy. This may in part result from the fact that nucleosomes prefer to form on the terminal ends of linear DNA. This phenomenon has the potential to bias in vitro reconstituted nucleosomes and skew results. If the ends of DNA fragments are known, the reads falling close to the ends are typically discarded. In this study we confirm the phenomenon of end bias of in vitro nucleosomes. We describe a method in which nearly identical libraries, with different known ends, are used to recover nucleosomes which form towards the terminal ends of fragmented DNA. Finally, we illustrate that although nucleosomes prefer to form on DNA ends, it does not appear to skew results or the interpretation thereof.

Polymer Chain Reaction (PCR): Principle and Applications

The new, standard molecular biologic system for duplicating DNA enzymatically devoid of employing a living organism, like E. coli or yeast, represents polymerases chain reaction (PCR). This technology allows an exponential intensification of a minor quantity of DNA molecule several times. Analysis can be straightforward with more DNA available.
 A thermal heat cycler performs a polymerization chain reaction that involves repeated cycles of heating and cooling the reactant tubes at the desired temperature for each reaction step. A heated deck is positioned on the upper reaction tube to avoid evaporating the reaction mixture (normally volumes range from 15 to 100 l per tube), or an oil layer can be placed on a reaction mixture surface.
 The amplified DNA fragment is determined based on selecting primers in addition to the starting and end of the DNA fragment. The primers stand for short, artificial DNA stripes, no higher than fifty (typically 18-25bp) nucleotides have been based on a starting and ending of DNA fragment to be amplified. DNA-polymerase connects and starts a new DNA strand synthesis
 The PCR products can be visualized by dual foremost methods: (1) staining of the product of DNA amplified by a chemical dye like bromide ethidium, or (2) marking of fluorescent dyes (fluorophores) PCR primers or nucleotides before amplification of PCRs.
 PCR offers some benefits. First, it is a simple method of understanding and using and quick results. It has an extremely sensitive technology with the potential for sequencing, cloning, and analyzing millions or milliards of copies of a particular product.

Polygenic basis and biomedical consequences of telomere length variation

Telomeres, the end fragments of chromosomes, play key roles in cellular proliferation and senescence. Here we characterize the genetic architecture of naturally occurring variation in leukocyte telomere length (LTL) and identify causal links between LTL and biomedical phenotypes in 472,174 well-characterized UK Biobank participants. We identified 197 independent sentinel variants associated with LTL at 138 genomic loci (108 new). Genetically determined differences in LTL were associated with multiple biological traits, ranging from height to bone marrow function, as well as several diseases spanning neoplastic, vascular and inflammatory pathologies. Finally, we estimated that, at the age of 40 years, people with an LTL >1 s.d. shorter than the population mean had a 2.5-year-lower life expectancy compared with the group with ≥1 s.d. longer LDL. Overall, we furnish new insights into the genetic regulation of LTL, reveal wide-ranging influences of LTL on physiological traits, diseases and longevity, and provide a powerful resource available to the global research community.

DONETSK MINING AND METALLURGICAL CENTER AND PRODUCTION HUBS OF THE EASTERN EUROPEAN STEPPE OF THE BRONZE AGE: A COMPARATIVE ANALYSIS OF PRODUCTION ACTIVITIES

Large-scale works have been carried out at the beginning of this century on the background of the set of Bronze Age monuments in the zone of Kartamysh copper ore occurrence in the Bakhmut basin of Donbass. Taking into account some previously researched monuments of mining and metallurgical activities within other ore occurrences of the Donetsk ridge, those works enable to compare the monuments of the Donetsk Mining and Metallurgical Center (DMMC) with other specialized monuments of Srubnaya cultural community in the copper ore territories of the Eastern European steppe.
 The monuments of Kartamysh archaeological microdistrict, as well as the other monuments of the Donetsk mining and metallurgical center, located in the zone of copper ore occurrences in the Bakhmut basin of Donbass, give evidences of all the cycles of ancient metal production. The majority of them are the evidence of the mining and ore-dressing cycle. Thus, the considerable volumes of mined and dressed ore, found in Kartamysh, as well as in other ore occurrences of the Bakhmut basin, currently suggest that the monuments in the ore territory of Donbass mainly operate within mining system. Similar specialization is observed in other mining and metallurgical areas in Eastern Europe, such as Mikhailo-Ovsyanka (Povolzhye) and Kargaly (South Ural). A feature of the DMMC is far lesser intensity of metallurgy and metalworking in its cultural frames. Limited range of DMMC monuments indicate that the production of metal products was focused only on domestic consumption.
 Fracturing of the Donbass bedrock, in contrast to the monolithic bedrock of Povolzhye, and especially to the Southern Urals, facilitated the effective use of stone tools in the process of mining. In the MMC, operating within the eastern production zone of the Srubnaya cultural community (Mikhailo-Ovsyanka, Kargaly), miners and metallurgists had to develop metallurgy more actively, since the features of geology in these regions required the use of metal tools to extract copper ores. Obviously, this circumstance explains a large number of end fragments of metal pickaxes found at Kargaly, as well as casting molds for casting these tools.
 The functioning of the full-scale cycle of ancient metal production and even visually recorded scale of ancient mining activity in the zone of copper ore occurrences of the Bakhmut basin gives reasons to assert the existence of mining and metallurgical center focused on large-scale production in the Donetsk Ridge in the Bronze Age. The main products of the DMMC were not metal items or even ingots, as it had been considered previously, but enriched copper ore (concentrate) as raw material for metallurgical production.
 Enriched ore was the commodity to be exchanged by the miners and metallurgists of the Donetsk center. It could be possibly done via professional traders with neighboring and distant tribes, and was exchanged for livestock and agriculture products, as well as for various household items.
 The analysis, carried out in the paper, proves that other mining and metallurgical complexes of the Eastern European steppe (Kargaly, Mikhailo-Ovsyanka), which had been previously explored, also operated at the same time and in similar to the DMMC production system.

A Fragment of Robert Burns: from his Letter to Robert Ainslie, 16 October 1788

This paper describes the background research on a fragment of a letter from Robert Burns to his great friend Robert Ainslie purchased at auction in Edinburgh in 2020. The author wished to determine the history of the fragment and whether it was possible to reunite the fragment with the rest of the letter. Though the details of the fragment's history over the last two hundred and forty years could not be fully determined the paper describes a fascinating story involving multiple auctions, a Victorian autograph album, one of Queen Victoria's ladies of the bedchamber and even, possibly, Sir Walter Scott. It establishes beyond reasonable doubt that this fragment is the signed end of another letter fragment to be found in the Edinburgh edition belonging to Robert Ainslie which is now in the G. Ross Roy Collection of Robert Burns in the University of South Carolina.

Nuclease deficiencies alter plasma cell-free DNA methylation profiles

The effects of DNASE1L3 or DNASE1 deficiency on cell-free DNA (cfDNA) methylation were explored in plasma of mice deficient in these nucleases and in DNASE1L3-deficient humans. Compared to wild-type cfDNA, cfDNA in DNASE1L3-deficient mice was significantly hypomethylated, while cfDNA in DNASE1-deficient mice was hypermethylated. The cfDNA hypomethylation in DNASE1L3-deficient mice was due to increased fragmentation and representation from open chromatin regions (OCRs) and CpG islands (CGIs). These findings were absent in DNASE1-deficient mice, demonstrating the preference of DNASE1 to cleave in hypomethylated OCRs and CGIs. We also observed a substantial decrease of fragment ends at methylated CpGs in the absence of DNASE1L3, thereby demonstrating that DNASE1L3 prefers to cleave at methylated CpGs. Furthermore, we found that methylation levels of cfDNA varied by fragment size in a periodic pattern, with cfDNA of specific sizes being more hypomethylated and enriched for OCRs and CGIs. These findings were confirmed in DNASE1L3-deficient human cfDNA. Thus, we have found that nuclease-mediated cfDNA fragmentation markedly affects cfDNA methylation level on a genome-wide scale. This work provides a foundational understanding of the relationship between methylation, nuclease biology, and cfDNA fragmentation.

Shorting Resection and Correction of the Leg Length in the Treatment of Posttramatic Tibial Defects Complicated by Osteomyelitis

Background.Severe fractures of the shin bones are often accompanied by the formation of defects in the tibia, suppuration and soft tissue necrosis. In the case of surgical treatment of fractures, infectious complications reach 3.6-9.1%. One of the methods of treatment of infected defects is resection of the ends of the tibia with shortening. This operation has proven to be effective in the treatment of fresh fractures. The relevance of the work is due to the prospects of using this technique in the treatment of the consequences of fractures with the formation of infected defects of the tibia.Aim of the study. Improving the results of treatment of patients with post-traumatic defects of the tibia complicated by osteomyelitis by performing a shortening resection with simultaneous or sequential correction of the segment length.Material and methods.The results of treatment of 65 patients with diaphyseal post-traumatic tibial defects complicated by osteomyelitis were analysed. They were divided into 2 groups. Group 1 was formed by 31 (47.7%) patients, they underwent shortening resection of the ends of bone fragments in the defect zone with simultaneous lengthening at another level. Group 2 included 34 (52.3%) patients who underwent a shortening resection of the tibia without lengthening. In all cases, the Ilizarov apparatus was used as a fixator.Results.The technique for assessing the size of the true defect of the tibia was optimized taking into account the initial shortening of the segment and the distance between the proximal and distal fragments after resection of their ends. A treatment regimen was developed depending on the level of localization of the tibial defect, and the results of treatment of patients in the compared groups were assessed.Conclusion.Shortening resection is an effective treatment for patients with post-traumatic tibial defects complicated by osteomyelitis. Depending on the level of localization of the defect, it is advisable to carry out treatment according to one of two possible options. When the defect is localized in the upper and middle third of the tibia, shortening resection in an isolated form is shown. If the defect is localized in the lower third of the tibia, it is possible to supplement the shortening resection with an osteotomy in the upper third with Ilizarov lengthening.

Studying SNPs in SUN and OVATE Genes Responsible for Fruit Shape in Tomato

Introduction: Commercial tomato (Solanum lycopersicum L.), one of the most widely grown vegetable crops worldwide, belongs to the Solanaceae family. The marketability of the commercial tomato mostly depends on the fruit quality. Tomato fruit quality is determined mainly by color, texture, shape and flavor. Fruit shape, one of the important traits affecting the quality of tomato fruit, is controlled by multiple minor genes and quantitatively inherited. Two important genes, involved in fruit shape, are SUN and OVATE genes. The SUN gene, which is a member of the IQD (IQ-domain) gene family and the Calmodulin binding protein, controls fruit length. The more expression of both SUN and OVATE genes leads to increased fruit length. Moreover, the increased expression of OVATE gene reduces the size of flower and leaf components. Due to the important role of these genes in tomato fruit shape, identification of single nucleotide polymorphisms (SNPs) as a new generation of robust, frequent and reliable bi-allelic markers, in the coding regions of these genes might be necessary for generating functional markers associated with fruit shape. Materials and Methods: Seeds of 96 tomato genotypes from 12 populations were grown in the research greenhouse of Faculty of Agriculture and Natural Resources of Urmia University. The genotypes had been collected from different regions of West Azerbaijan of Iran and Turkey (Igdir). The young and green plant leaves were used for genomic DNA extraction. The quality and quantity of the extracted DNA was assessed using spectrophotometry and agarose gel electrophoresis. To identify SNPs in SUN and OVATE genes, specific primers were designed by using FastPCR and Gene Runner software for amplifying fragments from coding regions of these genes in 96 tomato genotypes. Then, the amplified fragments of both genes were digested by using restriction enzymes TruI and PstI. Due to the lack of polymorphism in the digested patterns obtained by the used enzymes, four individuals from populations with close geographical distance were selected and amplified. The amplified bands were then purified by a purification kit (Kiagen, USA) and sequenced (Bioneer, South Korea). Sequencing was performed from both ends of the PCR fragments using both the forward and reverse primers used in the PCR reactions. The exon and intron regions of the sequenced fragments were identified by Softberry software. Following the retrieval of the sequenced fragments of each gene using FastPCR and Softberry software, multiple sequence alignment using Clustal Omega was used to identify SNPs in the exon and intron of the genes. Results and Discussion: Digestion of the amplified fragments of the genes using TruI and PstI restriction enzymes produced no polymorphism in the studied genotypes. Thus, four individuals were selected from geographically different populations and gene fragments were amplified, purified and sequenced in these genotypes. Sequencing of the amplified fragment of SUN gene revealed an intron region with a size of 369 bp. Out of the 10 SNPs detected in the SUN gene, four was found in the exon region, while the number of SNPs in intron was six. Of the total SNPs found in the SUN gene, the percentage of transition and transversion substitutions was 80 (50% T/C and 30% A/G) and 20 (T/G), respectively. In the OVATE gene, five SNPs were identified. The percentage of transition (40% G/A and 40% C/T) and transversion (20% G/T) substitutions in this genes were the same as SUN. The ratio of transition to transversion substitutions was 1:4 for both genes. The average number of SNPs in a 100 bp fragment in exonic and itronic region of SUN was 0.9 and 1.62, respectively, while it was 0.5 for exonic region of OVATE gene. Conclusion: The results of the current study revealed low polymorphisms and point mutations in the exon regions of SUN and OVATE genes, suggesting that the coding regions of these genes were conserved during the tomato evolution. Also, the number of SNPs in intron was more than those of exon. Considering the important role of fruit quality, especially fruit shape, in tomato market, the SNPs found in the current study may be used in genetic diversity studies, genetic map preparation, and saturation and identification of the functional markers associated with tomato fruit shape. These markers could accelerate tomato breeding programs aimed fruit shape improvement.

Characterization of DNA lesions associated with cell-free DNA by targeted deep sequencing

BackgroundRecently, a next-generation sequencing (NGS)-based method has been used for the successful detection of circulating tumor DNA (ctDNA) in various cancer types. Thus, the use of NGS on liquid biopsies will improve cancer diagnosis and prognosis. However, the low-allelic fraction of ctDNA poses a challenge for the sensitive and specific detection of tumor variants in cell-free DNA (cfDNA). To distinguish true variants from false positives, the characteristics of errors that occur during sample preparation and sequencing need to be elucidated.MethodsWe generated capture-based targeted deep sequencing data from plasma cfDNA and peripheral blood leucocyte (PBL) gDNA to profile background errors. To reveal cfDNA-associated DNA lesions, background error profiles from two sample types were compared in each nucleotide substitution class.ResultsIn this study, we determined the prevalence of single nucleotide substitutions in cfDNA sequencing data to identify DNA damage preferentially associated with cfDNA. On comparing sequencing errors between cfDNA and cellular genomic DNA (gDNA), we observed that the total substitution error rates in cfDNA were significantly higher than those in gDNA. When the substitution errors were divided into 12 substitution error classes, C:G>T:A substitution errors constituted the largest difference between cfDNA and gDNA samples. When the substitution error rates were estimated based on the location of DNA-fragment substitutions, the differences in error rates of most substitution classes between cfDNA and gDNA samples were observed only at the ends of the DNA fragments. In contrast, C:G>T:A substitution errors in the cfDNA samples were not particularly associated with DNA-fragment ends. All observations were verified in an independent dataset.ConclusionsOur data suggested that cytosine deamination increased in cfDNA compared to that in cellular gDNA. Such an observation might be due to the attenuation of DNA damage repair before the release of cfDNA and/or the accumulation of cytosine deamination after it. These findings can contribute to a better understanding of cfDNA-associated DNA damage, which will enable the accurate analysis of somatic variants present in cfDNA at an extremely low frequency.

Abstract 2286: Use of a synthetic spike-in ladder to measure NGS library complexity

Abstract Background: Detection of rare variants in traditional and liquid biopsies is becoming increasingly important for cancer diagnosis, monitoring and treatment. Next Generation Sequencing (NGS) has the capacity to detect known and novel variants in small biological samples with a high level of sensitivity. However, the limit of detection is affected by sample DNA concentration and integrity, technical error and number of unique molecules captured and sequenced (library complexity). Unique molecular identifiers (UMl) and random fragment end analysis commonly are used to measure complexity yet both are reported to have systematic biases leading to a non-random read distribution when measured, altering interpretation of results and potentially skewing data. Here we describe the use of a spike in complexity calibration ladder comprising synthetic DNA internal standard competitors (IS) as an orthogonal measure of library complexity. Methods: An Endogenous Complexity Calibration Ladder (ECCL) comprising multiple unique synthetic IS at different concentrations was created. All ECCL IS share homology with each other and endogenous human SCGB1A1 sequence but each contain nucleotide changes at different positions along the sequence string so that they can be distinguished. The ECCL was mixed with several IS targeting different TP53 exons in a fixed proportion. The ECCL/TP53 IS mixture is designed to be used in either amplicon or hybrid-capture library preparation. Here, amplicon libraries were generated. Molecule numbers and ratios between IS were determined using deep sequencing of multiple replicates. The ECCL/TP53 IS mix was spiked into a commercial human gDNA prior to NGS library preparation. Serial dilutions (1.5-, 3-, 6-, 12- and 24-fold) and intentional addition of inhibitors were conducted to stress and test the system. The ECCL/TP53 IS mixture also was spiked into gDNA from 60 primary airway epithelial cell (AEC) specimens prior to library preparation and NGS to measure TP53 variant fraction while controlling for complexity. Results: Observed complexity measured using the ECCL was close to expected for each serially diluted mixture of gDNA and ECCL/TP53 IS. The precision of complexity measurement for less dilute and less inhibited samples was limited by absence of finer titration of the least concentrated IS in ECCL. Use of ECCL enabled measurement of inter-sample variation in complexity among the 60 AEC specimens. Conclusions: The ECCL controls for both sample and library specific variation in complexity, and enables better estimation of the lower limit of detection for variant allele fraction (VAF). Additionally, it is compatible with the use of target specific IS which allow for improved measurement of technical sequencing error. The variation in complexity observed in primary AEC gDNA samples demonstrates the need to account for this when assessing rare variant allele fraction. Citation Format: Erin L. Crawford, Tian Chen, Daniel J. Craig, James C. Willey. Use of a synthetic spike-in ladder to measure NGS library complexity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2286.

Breather arrest in a chain of damped oscillators with Hertzian contact

Breather propagation in a damped oscillatory chain with Hertzian nearest-neighbour coupling is investigated. The breather propagation exhibits an unusual two-stage pattern. The first stage is characterized by power-law decay of the breather amplitude. This stage extends over finite number of the chain sites. Drastic drop of the breather amplitude towards the end of this finite fragment is referred to as breather arrest. At the second stage, the breather exhibits very small amplitudes with hyper-exponential decay. Numeric results are rationalized by considering a simplified model of two damped linear oscillators coupled by Hertzian contact forces. Initial excitation of one of these oscillators results in a finite number of beating cycles in the system. This simplified model reliably predicts main features of the breather arrest. More general coupling potentials and effect of pre-compression on the breather propagation are also discussed.

3D X-ray Microscopy as a Tool for in Depth Analysis of the Interfacial Interaction between a Single Carbon Fiber and an Epoxy Matrix after Mechanical Loading

The benefit of fiber-reinforced composites originates from the interaction between the fiber reinforcement and the matrix. This interplay controls many of its mechanical properties and is of utmost importance to enable its unique performance as a lightweight material. However, measuring the fiber−matrix interphase strength with micromechanical tests, like the Broutman test, is challenging, due to the many, often unknown boundary conditions. Therefore, this study uses state-of-the-art, high-resolution X-ray computed microtomography (XRM) as a tool to investigate post mortem the failure mechanisms of single carbon fibers within an epoxy matrix. This was conducted at the example of single carbon fiber Broutman test specimens. The capabilities of today’s XRM analysis were shown in comparison to classically obtained light microscopy. A simple finite element model was used to enhance the understanding of the observed fracture patterns. In total, this research reveals the possibilities and limitations of XRM to visualize and assess compression-induced single fiber fracture patterns. Furthermore, comparing two different matrix systems with each other illustrates that the failure mechanisms originate from differences in the fiber−matrix interphases. The carbon fiber seems to fail due to brittleness under compression stress. Observation of the fiber slippage and deformed small fracture pieces between the fragments suggests a nonzero stress state at the fragment ends after fiber failure. Even more, these results demonstrate the usefulness of XRM as an additional tool for the characterization of the fiber−matrix interphase.

A Study on Cooling System of HTS Quadrupole Magnets for Heavy Ion Accelerator

Quadrupole magnets located at the front end of an in-flight fragment (IF) separator in a heavy ion accelerator have large heat loads due to neutron radiation. Thus, high-temperature superconducting (HTS) magnets are used instead of low-temperature superconducting (LTS) magnets because of their high cooling efficiency and thermal stability. The cooling system of the HTS magnet uses circulation of cold helium gas. The circulating helium gas is provided from the distribution box, where helium gas lines are divided into 7 pairs of small helium lines corresponding to 7 HTS magnets. To cool down the 7 HTS magnets with different heat loads, the mass flow rate of each magnet should be individually controlled by monitoring the temperature of the magnets. This paper describes the cooling system for the 7 HTS magnets considering the heat load of each magnet and the thermohydraulic characteristics of the cooling system, including a heat exchanger, a cryogenic blower, control valves, etc. The helium mass flow rates for each magnet and the temperature at the outlet of the magnet are determined so that the magnets operate below 45 K. The optimal helium mass flow rates for each magnet were determined by analyzing the correlation between the magnet outlet temperature and flow rate.

Comprehensive profiling of mRNA polyadenylation in specific cell types in vivo by cTag-PAPERCLIP.

The ability to generate cell-type specific mRNA polyadenylation (pA) maps from complex tissues is crucial for understanding how alternative polyadenylation (APA) is regulated in individual cell types in their physiological environment under different conditions. In this chapter, we discuss cTag-PAPERCLIP, a recently developed method combining the well-established CLIP (crosslinking immunoprecipitation) technique and the Cre-lox system to achieve customized cell-type specific APA profiling from mouse tissue without cell purification or enrichment. In cTag-PAPERCLIP, selective expression of GFP-tagged poly(A) binding protein (PABP-GFP) in the desired cell type is achieved through Cre-mediated activation of a latent knock-in allele of PABP-GFP. Immunoprecipitation of PABP-GFP then allows mRNA 3' end fragments in the desired cell type to be specifically retrieved from ultraviolet (UV)-irradiated whole tissue lysate. The mRNA fragments are subsequently turned into a cDNA library to provide a comprehensive APA map and an mRNA expression profile of the chosen cell type through deep sequencing.

Characterization of DNA-PK-bound end fragments using GLASS-ChIP.

Endonucleolytic cleavage of DNA ends by the human Mre11-Rad50-Nbs1 (MRN) complex occurs in a manner that is promoted by DNA-dependent Protein Kinase (DNA-PK). A method is described to isolate DNA-PK-bound fragments released from chromatin in human cells using a modified Gentle Lysis and Size Selection chromatin immunoprecipitation (GLASS-ChIP) protocol. This method, combined with real-time PCR or next-generation sequencing, can identify sites of MRN endonucleolytic cutting adjacent to DNA-PK binding sites in human cells.

Construction of a Highly Diverse mRNA Library for in vitro Selection of Monobodies.

Recently, we developed transcription/translation coupled with the association of puromycin linker (TRAP) display as a quick in vitro selection method to obtain antibody-like proteins. For the in vitro selection, it is important to prepare mRNA libraries among which the diversity is high. Here, we describe a method for the preparation of monobody mRNA libraries with greater than 1013 theoretical diversity. First, we synthesized two long single-stranded DNAs that corresponded to fragments of monobody DNA, with random codons in the BC and FG loops. These oligonucleotides were ligated by T4 DNA ligase with the support of guide oligonucleotides containing 3' ends that were protected by a modification. After amplifying the product DNAs by PCR, one end of each DNA fragment was digested with the type II restriction enzyme BsaI, and the resulting DNA fragments were ligated using T4 DNA ligase. After amplification of the DNA product, mRNAs were synthesized by T7 RNA polymerase. This method is simple and could be used for the preparation of mRNA libraries for various antibody-like proteins. Graphic abstract: Construction of a highly diverse mRNA library.

A three-dimensional finite element analysis of stress distribution in maxillary central incisor with a horizontal mid root fracture after various management protocols.

Context:The incidence of root fractures is about 1.2%–7% in permanent teeth and horizontal mid root fractures are most common in maxillary anteriors.Aims:This study aims to evaluate the stress distribution in a maxillary central incisor with mid root fracture after various management protocols using three-dimensional finite element (FE) analysis.Materials and Methods:Four three dimensional FE models were constructed: Model 1-Normal tooth, Model 2-tooth with horizontal mid root fracture, Model 3-tooth with horizontal mid root fracture, coronal fragment filled with Biodentine, and access cavity sealed with composite resin cement and apical fragment left untreated and Model 4-tooth with horizontal mid root fracture restored with intraradicular fiber post as a splint and access cavity sealed with composite resin cement. The properties of materials were assigned and a force of 100 N was applied at 45° angle to the longitudinal axis of the tooth onto the palatal surface incisal to the cingulum.Statistical Analysis:Von-Mises stress along the length of the root analyzed and compared.Results:The maximum stress distribution was observed at the palatal end of the coronal radicular fragment in both without treatment and with biodentine treatment. The stress distribution was concentrated more at the junction of the splinted area in the intraradicular fiber post splinted model.Conclusions:The model with intraradicular fiber post as a splint exhibited more stress concentration at the splinted area due to bending forces. These stresses may adversely affect the healing around the fracture site and affect the longevity.

Experimental analysis of high velocity impacts of composite fragments on aluminium plates

The improvement of engines is one of the ways to diminish the fuel consumption in civil aircrafts, and Open Rotors engines are one of the best promises in order to achieve a sensible efficiency increment. These engines have large composite blades that could, in the event of failure, impact against the fuselage, totally or partially. In this case, composite fragments could behave as impactors. In order to design fuselages for this event and adopt these new engines in the future, it is necessary to understand the impact behaviour of a composite fragment against a deformable structure. To this end, unidirectional and woven composites fragments were impacted at high velocity (up to 150 m/s) against aluminium panels at different impact velocities. The composite fragments were made using AS4/8552 (UD) and AGP-193PW (woven) prepregs manufactured by Hexcel Composites, both using AS4 fibres and 8552 epoxy matrix. High speed video cameras were used to record the impact process and to measure both the impact and the residual velocity and hence the energy absorbed.

Features of titanium and aluminum particles combustion

The results of the study of metal particles combustion process in free fall in air are presented. Titanium particles with a size of 120-540 microns and particles-aluminum agglomerates with a size of 540-660 microns were used. The characteristic times of the beginning and the end of fragmentation, the end of combustion were found with help of video filming. The laws of particles motion, in particular, their velocity and coordinate at the moment of the beginning of fragmentation, were determined. The size of titanium particles at which the fragmentation mode changes (“star” and “spruce branch”) were estimated. Condensed combustion products of particles were selected and investigated.

Phylogenetic Analysis of Candida auris Isolates From Clinical Samples of Surgical Intensive Care Units

Background: Between January and September of 2019, 15 patients acquired Candida auris infection in our surgical intensive care unit (SICU). Although the outbreak was controlled by enhancing horizontal measures and a change of the environmental disinfectant from a quaternary ammonium compound to a peroxide-based product; we wanted to understand whether patient-to-patient spread was occurring or the environment was the major contributor. Methods: We submitted all the 15 isolates from clinical samples for WGS and phylogenetic analysis; along with operation theater (OT) and ICU environment random swabs for metagenomic analysis. DNA sample QC DNA extraction was done using a Qiagen QiAmp DNA mini kit (cat. no. 51306). The DNA samples were subjected to QIAXPERT and Qubit for quantifying the amount of DNA in the extracted sample. Also, the 260/280-nm ratio was examined for the purity of the same. They were also subjected for agarose gel electrophoresis. For the DNA library prep protocol, whole-genome libraries were prepared from 21 samples using NEBNext Ultra IITM DNA Library Prep Kit (Cat. No: E7645L). The adapter sequences were added to the ends of DNA fragments to generate paired-end libraries. The resulting adaptor-ligated libraries were purified and index tags were added by amplification, followed by purification. Libraries were assessed to check the quality and quantity using Agilent 2200 Tape station (cat. no. 3-PM 863NA). For the sequencing protocol, prepared libraries were quantified using Qubit High Sensitivity reagent. The obtained libraries were diluted to final concentration of 2 nm in 10 µL and were subjected to cluster amplification. Once the cluster generation was completed, the flow cells were loaded on to the sequencer. Sequencing was carried out in Hi Seq X10 to generate 2X150-bp sequence reads at >100X sequencing depth (~1.5 Gb). A minimum of 75% of the sequenced bases were of Q30 value. Sequenced data were processed to generate FASTQ files and were uploaded on the FTP server for download and secondary data analysis. Results: Overall, 2% of the DNA from the OT and SICU environment showed Candida auris. The phylogenetic analysis confirmed 2 different clusters. Furthermore, 13 of the clinical isolates belonged to the same cluster, confirming that patient-to-patient transmission had occurred, which was subsequently confirmed by line listing the patients. Conclusions:Candida auris can efficiently spread from patient to patient, resulting in outbreaks. It can also persist in the healthcare environment causing ongoing propagation of these outbreaks.Funding: NoneDisclosures: None