Integrative and Conjugative Element ICETh1 Functions as a Pangenomic DNA Capture Module in Thermus thermophilus.

Transjugation is an unconventional conjugation mechanism in Thermus thermophilus (Tth) that involves the active participation of both mating partners, encompassing a DNA secretion system (DSS) in the donor and an active natural competence apparatus (NCA) in the recipient cells. DSS is encoded within an integrative and conjugative element (ICETh1) in the strain Tth HB27, whereas the NCA is constitutively expressed in both mates. Previous experiments suggested the presence of multiple origins of transfer along the genome, which could generate genomic mosaicity among the progeny. Here, we designed transjugation experiments between two closely related strains of Tth with highly syntenic genomes, containing enough single nucleotide polymorphisms to allow precise parenthood analysis. Individual clones from the progeny were sequenced, revealing their origin as derivatives of our ICETh1-containing intended “donor” strain (HB27), which had acquired separate fragments from the genome of the ICETh1-free HB8 cells, which are our intended recipient. Due to the bidirectional nature of transjugation, only assays employing competence-defective HB27 derivatives as donors allowed the recovery of HB8-derived progeny. These results show a preference for a retrotransfer mechanism in transjugation in ICETh1-bearing strains, supporting an inter-strain gene-capture function for ICETh1. This function could benefit the donor-capable host by facilitating the acquisition of adaptive traits from external sources, ultimately increasing the open pangenome of Thermus, maximizing the potential repertoire of physiological and phenotypical traits related to adaptation and speciation.post-print11136 K

Leishmania mitochondrial genomes: Maxicircle structure and heterogeneity of minicircles

The mitochondrial DNA (mtDNA), which is present in almost all eukaryotic organisms, is a useful marker for phylogenetic studies due to its relative high conservation and its inheritance manner. In Leishmania and other trypanosomatids, the mtDNA (also referred to as kinetoplast DNA or kDNA) is composed of thousands of minicircles and a few maxicircles, catenated together into a complex network. Maxicircles are functionally similar to other eukaryotic mtDNAs, whereas minicircles are involved in RNA editing of some maxicircle-encoded transcripts. Next-generation sequencing (NGS) is increasingly used for assembling nuclear genomes and, currently, a large number of genomic sequences are available. However, most of the time, the mitochondrial genome is ignored in the genome assembly processes. The aim of this study was to develop a pipeline to assemble Leishmania minicircles and maxicircle DNA molecules, exploiting the raw data generated in the NGS projects. As a result, the maxicircle molecules and the plethora of minicircle classes for Leishmania major, Leishmania infantum and Leishmania braziliensis have been characterized. We have observed that whereas the heterogeneity of minicircle sequences existing in a single cell hampers their use for Leishmania typing and classification, maxicircles emerge as an extremely robust genetic marker for taxonomic studies within the clade of kinetoplastidsThis work was supported by grants (to B.A. and J.M.R.) from Proyecto del Ministerio de Economía, Industria y Competitividad SAF2017-86965-R, and by the Network of Tropical Diseases Research RICET (RD16/0027/0008); both grants are co-funded with FEDER funds. The CBMSO receives institutional grants from the Fundación Ramón Areces and from the Fundación Banco de Santande

Into the Thermus Mobilome: Presence, Diversity and Recent Activities of Insertion Sequences Across Thermus spp.

: A high level of transposon-mediated genome rearrangement is a common trait among microorganisms isolated from thermal environments, probably contributing to the extraordinary genomic plasticity and horizontal gene transfer (HGT) observed in these habitats. In this work, active and inactive insertion sequences (ISs) spanning the sequenced members of the genus Thermus were characterized, with special emphasis on three T. thermophilus strains: HB27, HB8, and NAR1. A large number of full ISs and fragments derived from different IS families were found, concentrating within megaplasmids present in most isolates. Potentially active ISs were identified through analysis of transposase integrity, and domestication-related transposition events of ISTth7 were identified in laboratory-adapted HB27 derivatives. Many partial copies of ISs appeared throughout the genome, which may serve as specific targets for homologous recombination contributing to genome rearrangement. Moreover, recruitment of IS1000 32 bp segments as spacers for CRISPR sequence was identified, pointing to the adaptability of these elements in the biology of these thermophiles. Further knowledge about the activity and functional diversity of ISs in this genus may contribute to the generation of engineered transposons as new genetic tools, and enrich our understanding of the outstanding plasticity shown by these thermophiles.post-print2.276 K

Gene annotation and transcriptome delineation on a de novo genome assembly for the reference Leishmania major friedlin strain

Leishmania major is the main causative agent of cutaneous leishmaniasis in humans. The Friedlin strain of this species (LmjF) was chosen when a multi-laboratory consortium undertook the objective of deciphering the first genome sequence for a parasite of the genus Leishmania. The objective was successfully attained in 2005, and this represented a milestone for Leishmania molecular biology studies around the world. Although the LmjF genome sequence was done following a shotgun strategy and using classical Sanger sequencing, the results were excellent, and this genome assembly served as the reference for subsequent genome assemblies in other Leishmania species. Here, we present a new assembly for the genome of this strain (named LMJFC for clarity), generated by the combination of two high throughput sequencing platforms, Illumina short-read sequencing and PacBio Single Molecular Real-Time (SMRT) sequencing, which provides long-read sequences. Apart from resolving uncertain nucleotide positions, several genomic regions were reorganized and a more precise composition of tandemly repeated gene loci was attained. Additionally, the genome annotation was improved by adding 542 genes and more accurate coding-sequences defined for around two hundred genes, based on the transcriptome delimitation also carried out in this work. As a result, we are providing gene models (including untranslated regions and introns) for 11,238 genes. Genomic information ultimately determines the biology of every organism; therefore, our understanding of molecular mechanisms will depend on the availability of precise genome sequences and accurate gene annotations. In this regard, this work is providing an improved genome sequence and updated transcriptome annotations for the reference L. major Friedlin strai

Change in Nasal Airflow, Associated with the Treatment of Rapid Maxillary Expansion in Children with Maxillary Compression

Purpose: To determine the change of nasal airflow (ANF) in children with maxillary compression after performing rapid expansion treatment. Materials and methods: The sample consisted of 30 patients who attended the Orthodontics Postgraduate Program of the Autonomous University of Nuevo Leon (UANL). They underwent clinical history, study models, lateral cephalogram and orthopantomography. Those who presented maxillary compression were referred to the Regional Center of Allergy and Clinical Immunology (CRAIC) of the University Hospital, UANL., Where a rhinomanometry (T1) was performed to know the values of nasal airflow (NAD) and nasal resistance (RN). before starting the maxillary expansion treatment (ERM). ERM was performed with a Hyrax type expansion screw. Three subsequent registrations were performed with rhinomanometry, the first was one month after starting the ERM (T2), the second to the third month (T3) and the third to the ninth month (T4). Changes in the nasal airflow were compared after of the maxillary rapid expansion, with the student’s t test, with p≤0.05, for related samples. Results: The values of the FAN increased as the measurements were made, on the other hand, the RN decreased progressively. The results were not statistically significant from one time to another. We did not find a statistically significant difference in relation to gender, nor to the present symptomatology. The group of 9 to 10 years was especially favored in relation to the two groups of greater age. Conclusion: The treatment of MRS is an effective treatment to relieve maxillary compression, helps improve nasal capacity, increasing FAN and decreasing the NR; however, it is not recommended in order to improve nasal permeability exclusively

Leishmania infantum (JPCM5) transcriptome, gene models and resources for an active curation of gene annotations

Leishmania infantum is one of the causative agents of visceral leishmaniases, the most severe form of leishmaniasis. An improved assembly for the L. infantum genome was published five years ago, yet delineation of its transcriptome remained to be accomplished. In this work, the transcriptome annotation was attained by a combination of both short and long RNA-seq reads. The good agreement between the results derived from both methodologies confirmed that transcript assembly based on Illumina RNA-seq and further delimitation according to the positions of spliced leader (SAS) and poly-A (PAS) addition sites is an adequate strategy to annotate the transcriptomes of Leishmania, a procedure previously used for transcriptome annotation in other Leishmania species and related trypanosomatids. These analyses also confirmed that the Leishmania transcripts boundaries are relatively slippery, showing extensive heterogeneity at the 5′- and 3′-ends. However, the use of RNA-seq reads derived from the PacBio technology (referred to as Iso-Seq) allowed the authors to uncover some complex transcription patterns occurring at particular loci that would be unnoticed by the use of short RNA-seq reads alone. Thus, Iso-Seq analysis provided evidence that transcript processing at particular loci would be more dynamic than expected. Another noticeable finding was the observation of a case of allelic heterozygosity based on the existence of chimeric Iso-Seq reads that might be generated by an event of intrachromosomal recombination. In addition, we are providing the L. infantum gene models, including both UTRs and CDS regions, that would be helpful for undertaking whole-genome expression studies. Moreover, we have built the foundations of a communal database for the active curation of both gene/transcript models and functional annotations for genes and proteinsThis research was supported by the Spanish Ministerio de Ciencia, Innovación (MICINN), Agencia Estatal deInvestigación(AEI), grant number PID2020-117916RB-I00, and Instituto de Salud Carlos III, grant CB21/13/00018 (CIBERINFEC). An institutional grant from Fundacion RamonAreces is also acknowledge

Estado gingival y presencia de bacterias del complejo rojo en escolares de 12 años

The objective of this study was to determine the gingival state and presence of red complex bacteria in saliva samples of 12-year-old schoolchildren. A calibrated periodontist evaluated biofilm index (BI) (Silness and Löe, 1964), presence of calculus, and gingival index (GI) (Silness and Löe, 1967) in sixty two 12-year-old students of Carmen Lyra School. Saliva samples were collected from each student. The DNA of each sample was extracted and amplified by the polymerase chain reaction (PCR) technique, using specific primers. The BI was 1.18. Calculus was present in 40.40% of the schoolchildren examined; 19.4% was supragingival calculus and 21% both supragingival and subgingival calculus. The GI was 0.97, which according to Silness and Löe is mild gingivitis. Gingivitis was present in 96.8% of the children examined. Regarding the PCR tests: 18 of the samples (31.58%) did not present any of the bacteria analyzed and the remaining 39 samples (68.42%) were positive for at least the presence of red complex bacteria. Within the limitations of this study, it is concluded that the prevalence of gingivitis and calculus is high in the sample examined, and the gingival state observed in the study population, may be related to the presence of red complex bacteria.El objetivo de este estudio era determinar el estado gingival y la presencia de bacterias del complejo rojo en muestras de saliva de niños de 12 años de la Escuela Carmen Lyra. Una periodoncista calibrada evaluó en 62 estudiantes de 12 años de la Escuela Carmen Lyra, el índice de biofilme (IB) (Silness y Löe, 1964), la presencia de cálculo y el índice gingival (IG) (Silness y Löe, 1967). Se recolectaron muestras de saliva de cada estudiante. El ADN de cada muestra fue extraído y amplificado por medio de la prueba PCR, empleando primers específicos, para determinar la presencia de bacterias del complejo rojo. El IB fue de 1.18. El cálculo estuvo presente en el 40.40% de la muestra, se encontró 19.4% de cálculo en supragingival y 21% tanto en supragingival como en subgingival. El IG fue de 0.97, que de acuerdo con Silness y Löe es una gingivitis leve. La gingivitis estuvo presente en el 96.8 % de los niños examinados. Con respecto a las pruebas PCR: 18 de las muestras (31.58 %) no presentaron ninguna de las bacterias analizadas y las 39 muestras restantes (68.42 %) fueron positivas por lo menos a la presencia de las bacterias del complejo rojo. Dentro de las limitaciones de este estudio, se concluye que la prevalencia de gingivitis y cálculo es alta en la muestra examinada y el estado gingival observado puede estar relacionado con la presencia de bacterias del complejo rojo.UCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias Básicas::Centro de Investigación en Biología Celular y Molecular (CIBCM)UCR::Vicerrectoría de Docencia::Salud::Facultad de Odontologí

ARAMIS: From systematic errors of NGS long reads to accurate assemblies

NGS long-reads sequencing technologies (or third generation) such as Pacific BioSciences (PacBio) have revolutionized the sequencing field over the last decade improving multiple genomic applications like de novo genome assemblies. However, their error rate, mostly involving insertions and deletions (indels), is currently an important concern that requires special attention to be solved. Multiple algorithms are available to fix these sequencing errors using short reads (such as Illumina), although they require long processing times and some errors may persist. Here, we present Accurate long-Reads Assembly correction Method for Indel errorS (ARAMIS), the first NGS long-reads indels correction pipeline that combines several correction software in just one step using accurate short reads. As a proof OF concept, six organisms were selected based on their different GC content, size and genome complexity, and their PacBio-assembled genomes were corrected thoroughly by this pipeline. We found that the presence of systematic sequencing errors in long-reads PacBio sequences affecting homopolymeric regions, and that the type of indel error introduced during PacBio sequencing are related to the GC content of the organism. The lack of knowledge of this fact leads to the existence of numerous published studies where such errors have been found and should be resolved since they may contain incorrect biological information. ARAMIS yields better results with less computational resources needed than other correction tools and gives the possibility of detecting the nature of the found indel errors found and its distribution along the genome. The source code of ARAMIS is available at https://github.com/genomics-ngsCBMSO/ARAMIS.gi

Formation of a magnetite/hematite epitaxial bilayer generated with low energy ion bombardment

We have used a low-energy ion bombardment to fabricate an epitaxial single-crystalline magnetite/hematite bilayer grown on Au(111). This non-conventional fabrication method involves the transformation of the upper layers of a single-crystalline hematite thin film to single-crystalline magnetite, a process driven by the preferential sputtering of oxygen atoms and favoured by the good structural matching of both phases. We show the reversibility of the transformation between hematite and magnetite, always keeping the epitaxial and single- crystalline character of the films. The magnetic characterization of the bilayer grown using this method shows that the magnetic response is mainly determined by the magnetite thin film, exhibiting a high coercivity. Published by AIP Publishing

New Structured Materials in the Study of the Mechanobiological Processes Related to the Heart Failure

Cardiovascular diseases are the number one of death globally. According to the World Health organization 17.7 million people died from cardiovascular diseases in 2015, representing 31% of all global deaths.  In these diseases the cardiac homeostasis is disrupted by a non-appropriate myocardium remodelling. The cardiac extracellular matrix (ECM) provides not only the biochemical environment but also a natural scaffold surrounding and connecting cardiac cells and distributing mechanical forces throughout the organ. Thus, the properties of the ECM are essential for the maintenance of the functional myocardium. Alterations in cardiac ECM structure associated with heart failure influence cell-matrix and cell-cell adhesions modifying cell shape and mechanotransduction.The need to understand the cardiac ECM remodelling mechanisms that allow us to identify new therapeutic targets lead us to create biomimetic scaffolds which emulate the structure, topography, mechanics and chemical composition of ECM.Here, we present the development of modulable materials for the manufacturing, by using photopolymerizable materials, of structured hydrogels with myocardium properties of stiffness and elastic modulus in physiological and pathological conditions