Improved Visualization of Oral Microbial Consortia.

Bacteria on the tongue dorsum (TD) form consortia tens to hundreds of microns in diameter organized around a core of epithelial cells. Whole-mount preparations have been instrumental in revealing their organization and specific microbial associations. However, their thickness and intricate 3-dimensional complexity present challenges for a comprehensive spatial analysis. To overcome these challenges, we employed a complementary approach: embedding in hydrophilic plastic followed by sectioning and postsectioning labeling. Samples were labeled by hybridization with multiplexed fluorescent oligonucleotide probes and visualized by spectral imaging and linear unmixing. Application of this strategy to TD biofilms improved the visualization of bacteria that were difficult to resolve in whole-mount imaging. Actinomyces, previously detected as patches, became resolved at the single-cell level. The filamentous taxa Leptotrichia and Lachnospiraceae, located at the core of the consortium, were regularly visualized whereas previously they were rarely detected when using whole mounts. Streptococcus salivarius, heterogeneously detected in whole mounts, were regularly and homogenously observed. Two-dimensional images provide valuable information about the organization of bacterial biofilms. However, they offer only a single plane of view for objects that can extend to hundreds of microns in thickness, and information obtained from such images may not always reflect the complexity of a 3-dimensional object. We combined serial physical sectioning with optical sectioning to facilitate the 3-dimensional reconstruction of consortia, spanning over 100 µm in thickness. Our work showcases the use of hydrophilic plastic embedding and sectioning for examining the structure of TD biofilms through spectral imaging fluorescence in situ hybridization. The result was improved visualization of important members of the human oral microbiome. This technique serves as a complementary method to the previously employed whole-mount analysis, offering its own set of advantages and limitations. Addressing the spatial complexity of bacterial consortia demands a multifaceted approach for a comprehensive and effective analysis.

A 2D ferroelectric vortex pattern in twisted BaTiO3 freestanding layers.

The wealth of complex polar topologies1-10 recently found in nanoscale ferroelectrics results from a delicate balance between the intrinsic tendency of the materials to develop a homogeneous polarization and the electric and mechanical boundary conditions imposed on them. Ferroelectric-dielectric interfaces are model systems in which polarization curling originates from open circuit-like electric boundary conditions, to avoid the build-up of polarization charges through the formation of flux-closure11-14 domains that evolve into vortex-like structures at the nanoscale15-17 level. Although ferroelectricity is known to couple strongly with strain (both homogeneous18 and inhomogeneous19,20), the effect of mechanical constraints21 on thin-film nanoscale ferroelectrics has been comparatively less explored because of the relative paucity of strain patterns that can be implemented experimentally. Here we show that the stacking of freestanding ferroelectric perovskite layers with controlled twist angles provides an opportunity to tailor these topological nanostructures in a way determined by the lateral strain modulation associated with the twisting. Furthermore, we find that a peculiar pattern of polarization vortices and antivortices emerges from the flexoelectric coupling of polarization to strain gradients. This finding provides opportunities to create two-dimensional high-density vortex crystals that would enable us to explore previously unknown physical effects and functionalities.

Valores normativos de pruebas conductuales de procesamiento auditivo para niños chilenos entre 7 y 12 años: una propuesta inicial / Normative values of auditory processing behavioral tests for chilean children between 7 and 12 years old: an initial proposal

Resumen Introducción: El trastorno del procesamiento auditivo en niños está relacionado con trastornos de lenguaje, bajo rendimiento académico, trastornos de aprendizaje y dificultades psicosociales. Existe consenso internacional sobre la utilidad de las pruebas conductuales para su evaluación y diagnóstico. Objetivo: Establecer valores normativos para pruebas conductuales de procesamiento auditivo en población pediátrica chilena con neurodesarrollo y audición normal. Material y Método: Se realizó un estudio observacional de corte transversal. Participaron 153 sujetos entre 7 y 12 años de la Región Metropolitana. Se estudiaron las pruebas de patrones de frecuencia, habla filtrada y dígitos dicóticos. Se construyeron modelos de regresión fraccional para estimar los valores normativos y además se calcularon los puntajes de corte en los percentiles 2.5, 5 y 10. Resultados: No se evidenciaron diferencias significativas entre oídos para las pruebas estudiadas a excepción de la prueba de dígitos dicóticos. Se construyeron modelos únicos para las pruebas patrones de frecuencia y habla filtrada, y modelos independientes para cada oído para la prueba dígitos dicóticos. Todas las estimaciones resultaron significativas y tuvieron niveles aceptables de precisión. Conclusión: Se obtuvieron los valores normativos y puntajes de corte para las tres pruebas estudiadas. Los valores obtenidos fueron similares a los reportados en otras poblaciones considerando los efectos de oído, de edad, sexo y escolaridad.

Abstract Introduction: Auditory processing disorder in children is related to language disorders, poor academic performance, learning disorders, and psychosocial difficulties. There is international consensus on the usefulness of behavioral tests for their evaluation and diagnosis. Aim: To establish normative values for auditory processing behavioral tests in Chilean pediatric population with normal neurodevelopment and hearing thresholds. Material and Method: An observational cross-sectional study was carried out. One hundred fifty-three subjects between 7 and 12 years old from the Metropolitan Region participated. Frequency pattern tests, filtered speech, and dichotic digits were studied. Fractional regression models were built to estimate the normative values, and cut-off scores were also calculated at the 2.5, 5, and 10 percentiles. Results: There were no significant differences between ears for the tests studied except for the dichotic digits test. Single models were built for the frequency and filtered speech patterns tests and independent models for each ear for the dichotic digit test. All estimates were significant and had acceptable levels of precision. Conclusion: The normative values and cut-off scores were obtained for the three tests studied. The values obtained were similar to those reported in other populations considering the effects of ear, age, sex, and education.

Spatial organization of the kelp microbiome at micron scales.

BACKGROUND: Elucidating the spatial structure of host-associated microbial communities is essential for understanding taxon-taxon interactions within the microbiota and between microbiota and host. Macroalgae are colonized by complex microbial communities, suggesting intimate symbioses that likely play key roles in both macroalgal and bacterial biology, yet little is known about the spatial organization of microbes associated with macroalgae. Canopy-forming kelp are ecologically significant, fixing teragrams of carbon per year in coastal kelp forest ecosystems. We characterized the micron-scale spatial organization of bacterial communities on blades of the kelp Nereocystis luetkeana using fluorescence in situ hybridization and spectral imaging with a probe set combining phylum-, class-, and genus-level probes to localize and identify > 90% of the microbial community. RESULTS: We show that kelp blades host a dense microbial biofilm composed of disparate microbial taxa in close contact with one another. The biofilm is spatially differentiated, with clustered cells of the dominant symbiont Granulosicoccus sp. (Gammaproteobacteria) close to the kelp surface and filamentous Bacteroidetes and Alphaproteobacteria relatively more abundant near the biofilm-seawater interface. A community rich in Bacteroidetes colonized the interior of kelp tissues. Microbial cell density increased markedly along the length of the kelp blade, from sparse microbial colonization of newly produced tissues at the meristematic base of the blade to an abundant microbial biofilm on older tissues at the blade tip. Kelp from a declining population hosted fewer microbial cells compared to kelp from a stable population. CONCLUSIONS: Imaging revealed close association, at micrometer scales, of different microbial taxa with one another and with the host. This spatial organization creates the conditions necessary for metabolic exchange among microbes and between host and microbiota, such as provisioning of organic carbon to the microbiota and impacts of microbial nitrogen metabolisms on host kelp. The biofilm coating the surface of the kelp blade is well-positioned to mediate interactions between the host and surrounding organisms and to modulate the chemistry of the surrounding water column. The high density of microbial cells on kelp blades (105-107 cells/cm2), combined with the immense surface area of kelp forests, indicates that biogeochemical functions of the kelp microbiome may play an important role in coastal ecosystems. Video abstract.

Oral Microbiome Geography: Micron-Scale Habitat and Niche.

The mouth presents a multiplicity of local environments in communication with one another via saliva. The spatial organization of microbes within the mouth is shaped by opposing forces in dynamic equilibrium-salivary flow and adhesion, shedding and colonization-and by interactions among and between microbes and the host. Here we review recent evidence confirming that oral microbes are specialized for individual habitats within the mouth and that microbial habitats and niches are defined by micron-scale gradients in combination with short- and long-range interactions. Micron-scale structure illuminates the roles of individual taxa and provides insight into their community ecology and potential pathogenicity.

A Simple Microbiome in the European Common Cuttlefish, Sepia officinalis.

The European common cuttlefish, Sepia officinalis, is used extensively in biological and biomedical research, yet its microbiome remains poorly characterized. We analyzed the microbiota of the digestive tract, gills, and skin in mariculture-raised S. officinalis using a combination of 16S rRNA amplicon sequencing, quantitative PCR (qPCR), and fluorescence spectral imaging. Sequencing revealed a highly simplified microbiota consisting largely of two single bacterial amplicon sequence variants (ASVs) of Vibrionaceae and Piscirickettsiaceae. The esophagus was dominated by a single ASV of the genus Vibrio. Imaging revealed bacteria in the family Vibrionaceae distributed in a discrete layer that lines the esophagus. This Vibrio was also the primary ASV found in the microbiota of the stomach, cecum, and intestine, but occurred at lower abundance, as determined by qPCR, and was found only scattered in the lumen rather than in a discrete layer via imaging analysis. Treatment of animals with the commonly used antibiotic enrofloxacin led to a nearly 80% reduction of the dominant Vibrio ASV in the esophagus but did not significantly alter the relative abundance of bacteria overall between treated versus control animals. Data from the gills were dominated by a single ASV in the family Piscirickettsiaceae, which imaging visualized as small clusters of cells. We conclude that bacteria belonging to the Gammaproteobacteria are the major symbionts of the cuttlefish Sepia officinalis cultured from eggs in captivity and that the esophagus and gills are major colonization sites. IMPORTANCE Microbes can play critical roles in the physiology of their animal hosts, as evidenced in cephalopods by the role of Vibrio (Aliivibrio) fischeri in the light organ of the bobtail squid and the role of Alpha- and Gammaproteobacteria in the reproductive system and egg defense in a variety of cephalopods. We sampled the cuttlefish microbiome throughout the digestive tract, gills, and skin and found dense colonization of an unexpected site, the esophagus, by a microbe of the genus Vibrio, as well as colonization of gills by Piscirickettsiaceae. This finding expands the range of organisms and body sites known to be associated with Vibrio and is of potential significance for understanding host-symbiont associations, as well as for understanding and maintaining the health of cephalopods in mariculture.

La sexualidad en adolescentes de riesgo. Comunidad de Madrid 2013-2014 / Sexuality in adolescents of risk groups autonomous. Community of Madrid 2013-2014

La sexualidad es un componente esencial de la personalidad. Durante la adolescencia -de 10 a 19 años- se producen cambios físicos, psíquicos y hormonales que junto a las transformaciones sociales, favorecen el inicio de conductas de riesgo. A pesar de los esfuerzos realizados para mejorar la salud sexual y reproductiva en los adolescentes y jóvenes españoles por parte de las familias y la sociedad, ellos continúan siendo vulnerables frente a los embarazos y las ITS. Nuestra investigación tiene lugar en el Centro de Primera Acogida de Protección ‘Hortaleza’ que pertenece al Instituto Madrileño de la Familia y el Menor, donde ingresan los/las adolescentes españoles e inmigrantes bajo la condición de guarda o tutela con edades comprendidas desde los 14,9 años hasta los 17,9 años, en la Comunidad de Madrid. Nuestro objetivo es describir las variables cuantitativas de los/las adolescentes ingresados en el centro durante el periodo 2013-2014 y analizar las variables cualitativas de las menores que acudieron a la Consulta de Planificación Familiar (AU)

Sexuality is an essential part of personality. During adolescence -10 to 19 years of age- several physical, psychic, and hormonal changes take place. These changes, together with social transformations, give rise to risk behaviour. In spite of the efforts made by the families and the society to improve the sexual and reproductive health of Spanish youths, they are still vulnerable to unplanned pregnancy and STDs. Our research was carried out in the institution Preliminary Reception Centre for Protection ‘Hortaleza’, which belongs to the Institute for the Child and Family in Madrid, where Spanish and immigrant children and teenagers between the ages of 14.9 and 17.9 are received under custody or tutelage. Our aim is to describe the quantitative variables for adolescents admitted in the Centre between 2013 and 2014 and analyse the qualitative variables for the girls under the age of 18 who turned to our Family Planning Counselling (AU)

Molecular phylogeny of the genus Dactylopius (Hemiptera: Dactylopiidae) and identification of the symbiotic bacteria.

Phylogenetic analyses, from polymerase chain reaction (PCR)-amplified 12S rRNA and 18S rRNA gene sequences from cochineal insects of the genus Dactylopius present in Mexico, showed that D. ceylonicus, D. confusus, and D. opuntiae are closely related. D. coccus constitutes a separate clade, and D. tomentosus is the most distantly related. Bacterial 16S rRNA sequences from all the Dactylopius species sampled showed a common ß-proteobacteria, related to Azoarcus, also found in eggs and in bacteriocytes in D. coccus. We propose the name "Candidatus Dactylopiibacterium carminicum" for this endosymbiont. Other bacterial sequences recovered from the samples were close to those from soil or plant associated bacteria, like Massilia, Herbaspirillum, Acinetobacter, Mesorhizobium, and Sphingomonas, suggesting a possible horizontal transmission from Cactaceae plant sap to Dactylopius spp. during feeding. This is the first molecular analysis of Dactylopius species and of their associated bacteria.