Bile-induced biofilm formation in Bacteroides thetaiotaomicron requires magnesium efflux by an RND pump.

Bacteroides thetaiotaomicron is a prominent member of the human gut microbiota contributing to nutrient exchange, gut function, and maturation of the host's immune system. This obligate anaerobe symbiont can adopt a biofilm lifestyle, and it was recently shown that B. thetaiotaomicron biofilm formation is promoted by the presence of bile. This process also requires a B. thetaiotaomicron extracellular DNase, which is not, however, regulated by bile. Here, we showed that bile induces the expression of several Resistance-Nodulation-Division (RND) efflux pumps and that inhibiting their activity with a global competitive efflux inhibitor impaired bile-dependent biofilm formation. We then showed that, among the bile-induced RND-efflux pumps, only the tripartite BT3337-BT3338-BT3339 pump, re-named BipABC [for Bile Induced Pump A (BT3337), B (BT3338), and C (BT3339)], is required for biofilm formation. We demonstrated that BipABC is involved in the efflux of magnesium to the biofilm extracellular matrix, which leads to a decrease of extracellular DNA concentration. The release of magnesium in the biofilm matrix also impacts biofilm structure, potentially by modifying the electrostatic repulsion forces within the matrix, reducing interbacterial distance and allowing bacteria to interact more closely and form denser biofilms. Our study therefore, identified a new molecular determinant of B. thetaiotaomicron biofilm formation in response to bile salts and provides a better understanding on how an intestinal chemical cue regulates biofilm formation in a major gut symbiont.IMPORTANCEBacteroides thetaiotaomicron is a prominent member of the human gut microbiota able to degrade dietary and host polysaccharides, altogether contributing to nutrient exchange, gut function, and maturation of the host's immune system. This obligate anaerobe symbiont can adopt a biofilm community lifestyle, providing protection against environmental factors that might, in turn, protect the host from dysbiosis and dysbiosis-related diseases. It was recently shown that B. thetaiotaomicron exposure to intestinal bile promotes biofilm formation. Here, we reveal that a specific B. thetaiotaomicron membrane efflux pump is induced in response to bile, leading to the release of magnesium ions, potentially reducing electrostatic repulsion forces between components of the biofilm matrix. This leads to a reduction of interbacterial distance and strengthens the biofilm structure. Our study, therefore, provides a better understanding of how bile promotes biofilm formation in a major gut symbiont, potentially promoting microbiota resilience to stress and dysbiosis events.

Ordered Mesoporous Electrodes for Sensing Applications.

Electrochemical sensors have become increasingly relevant in fields such as medicine, environmental monitoring, and industrial process control. Selectivity, specificity, sensitivity, signal reproducibility, and robustness are among the most important challenges for their development, especially when the target compound is present in low concentrations or in complex analytical matrices. In this context, electrode modification with Mesoporous Thin Films (MTFs) has aroused great interest in the past years. MTFs present high surface area, uniform pore distribution, and tunable pore size. Furthermore, they offer a wide variety of electrochemical signal modulation possibilities through molecular sieving, electrostatic or steric exclusion, and preconcentration effects which are due to mesopore confinement and surface functionalization. In order to fully exploit these advantages, it is central to develop reproducible routes for sensitive, selective, and robust MTF-modified electrodes. In addition, it is necessary to understand the complex mass and charge transport processes that take place through the film (particularly in the mesopores, pore surfaces, and interfaces) and on the electrode in order to design future intelligent and adaptive sensors. We present here an overview of MTFs applied to electrochemical sensing, in which we address their fabrication methods and the transport processes that are critical to the electrode response. We also summarize the current applications in biosensing and electroanalysis, as well as the challenges and opportunities brought by integrating MTF synthesis with electrode microfabrication, which is critical when moving from laboratory work to in situ sensing in the field of interest.

Bacteroides thetaiotaomicron uses a widespread extracellular DNase to promote bile-dependent biofilm formation.

Bacteroides thetaiotaomicron is a gut symbiont that inhabits the mucus layer and adheres to and metabolizes food particles, contributing to gut physiology and maturation. Although adhesion and biofilm formation could be key features for B. thetaiotaomicron stress resistance and gut colonization, little is known about the determinants of B. thetaiotaomicron biofilm formation. We previously showed that the B. thetaiotaomicron reference strain VPI-5482 is a poor in vitro biofilm former. Here, we demonstrated that bile, a gut-relevant environmental cue, triggers the formation of biofilm in many B. thetaiotaomicron isolates and common gut Bacteroidales species. We determined that bile-dependent biofilm formation involves the production of the DNase BT3563 or its homologs, degrading extracellular DNA (eDNA) in several B. thetaiotaomicron strains. Our study therefore shows that, although biofilm matrix eDNA provides a biofilm-promoting scaffold in many studied Firmicutes and Proteobacteria, BT3563-mediated eDNA degradation is required to form B. thetaiotaomicron biofilm in the presence of bile.

Conversion of the OmpF Porin into a Device to Gather Amyloids on the E. coli Outer Membrane.

Protein amyloids are ubiquitous in natural environments. They typically originate from microbial secretions or spillages from mammals infected by prions, currently raising concerns about their infectivity and toxicity in contexts such as gut microbiota or soils. Exploiting the self-assembly potential of amyloids for their scavenging, here, we report the insertion of an amyloidogenic sequence stretch from a bacterial prion-like protein (RepA-WH1) in one of the extracellular loops (L5) of the abundant Escherichia coli outer membrane porin OmpF. The expression of this grafted porin enables bacterial cells to trap on their envelopes the same amyloidogenic sequence when provided as an extracellular free peptide. Conversely, when immobilized on a surface as bait, the full-length prion-like protein including the amyloidogenic peptide can catch bacteria displaying the L5-grafted OmpF. Polyphenolic molecules known to inhibit amyloid assembly interfere with peptide recognition by the engineered OmpF, indicating that this is compatible with the kind of homotypic interactions expected for amyloid assembly. Our study suggests that synthetic porins may provide suitable scaffolds for engineering biosensor and clearance devices to tackle the threat posed by pathogenic amyloids.

Gnotobiotic rainbow trout (Oncorhynchus mykiss) model reveals endogenous bacteria that protect against Flavobacterium columnare infection.

The health and environmental risks associated with antibiotic use in aquaculture have promoted bacterial probiotics as an alternative approach to control fish infections in vulnerable larval and juvenile stages. However, evidence-based identification of probiotics is often hindered by the complexity of bacteria-host interactions and host variability in microbiologically uncontrolled conditions. While these difficulties can be partially resolved using gnotobiotic models harboring no or reduced microbiota, most host-microbe interaction studies are carried out in animal models with little relevance for fish farming. Here we studied host-microbiota-pathogen interactions in a germ-free and gnotobiotic model of rainbow trout (Oncorhynchus mykiss), one of the most widely cultured salmonids. We demonstrated that germ-free larvae raised in sterile conditions displayed no significant difference in growth after 35 days compared to conventionally-raised larvae, but were extremely sensitive to infection by Flavobacterium columnare, a common freshwater fish pathogen causing major economic losses worldwide. Furthermore, re-conventionalization with 11 culturable species from the conventional trout microbiota conferred resistance to F. columnare infection. Using mono-re-conventionalized germ-free trout, we identified that this protection is determined by a commensal Flavobacterium strain displaying antibacterial activity against F. columnare. Finally, we demonstrated that use of gnotobiotic trout is a suitable approach for the identification of both endogenous and exogenous probiotic bacterial strains protecting teleostean hosts against F. columnare. This study therefore establishes an ecologically-relevant gnotobiotic model for the study of host-pathogen interactions and colonization resistance in farmed fish.

Capsular Polysaccharide Cross-Regulation Modulates Bacteroides thetaiotaomicron Biofilm Formation.

Bacteroides thetaiotaomicron is one of the most abundant gut symbiont species, whose contribution to host health through its ability to degrade dietary polysaccharides and mature the immune system is under intense scrutiny. In contrast, adhesion and biofilm formation, which are potentially involved in gut colonization and microbiota structure and stability, have hardly been investigated in this intestinal bacterium. To uncover B. thetaiotaomicron biofilm-related functions, we performed a transposon mutagenesis in the poorly biofilm-forming reference strain VPI-5482 and showed that capsule 4, one of the eight B. thetaiotaomicron capsules, hinders biofilm formation. We then showed that the production of capsules 1, 2, 3, 5, and 6 also inhibits biofilm formation and that decreased capsulation of the population correlated with increased biofilm formation, suggesting that capsules could be masking adhesive surface structures. In contrast, we showed that capsule 8 displayed intrinsic adhesive properties. Finally, we demonstrated that BT2934, the wzx homolog of the B. thetaiotaomicron glycosylation locus, competes with capsule production and impacts its adhesion capacity. This study therefore establishes B. thetaiotaomicron capsule regulation as a major determinant of B. thetaiotaomicron biofilm formation, providing new insights into how modulation of different B. thetaiotaomicron surface structures affects in vitro biofilm formation.IMPORTANCE The human gut harbors a complex bacterial community that plays important roles in host health and disease, including nutrient acquisition, maturation of the immune system, and resistance to infections. The capacity to adhere to surfaces and form communities called biofilms is believed to be important for niche colonization and maintenance of gut bacteria. However, little is known about the adhesion capacity of most gut bacteria. In this study, we investigated biofilm formation in Bacteroides thetaiotaomicron, one of the most abundant bacteria of the normal mammalian intestine. We identified that B. thetaiotaomicron capsules, a group of eight surface-exposed polysaccharidic layers mediating important interactions with the gut environment, are also major determinants of biofilm formation that mask or unmask adhesion factors. Studying how B. thetaiotaomicron regulates its adhesion properties will allow us to better understand the physiology and specific properties of this important gut symbiont within anaerobic biofilms.

Uso de timolol en hemangiomas de la infancia / Using timolol in hemangiomas of infancy

Los hemangiomas de la infancia (HI) son los tumores vasculares benignos más frecuentes de la niñez. Ocasionalmente requieren tratamiento y, en aquellos en que esté indicada la intervención, se utilizan como terapias sistémicas los corticoides y, más recientemente, el propranolol. En el año 2010, se publicó por primera vez un trabajo sobre el uso de timolol tópico para HI no complicados, superficiales y de tamaño pequeño. Se presentan tres pacientes con HI que fueron tratados con timolol tópico, en los cuales se obtuvo un resultado satisfactorio

Infantile hemangiomas (IH) are the most common vascular benign tumors of childhood. Occasionally, treatment is required and in those in which intervention is indicated, the use of oral steroids and -most recently- propranolol, are established systemic therapies. In 2010 the use of topical timolol for the treatment of uncomplicated, superficial and small sized IH, was first published. We report 3 patients with IH treated with topical timolol showing satisfactory results

[Acute infectious gastroenteritis in Rio Cuarto: clinic, diagnosis and epidemiology with special reference to viral infections]. / Gastroenteritis aguda infecciosa en Rio Cuarto: clínica, diagnóstico y epidemiología con especial referencia a infecciones virales.

The objectives of the present study are to describe the clinics view of acute infectious gastroenteritis (GE) at the community, in primary and secondary attention health centers, with special references to viral aetiology (VIGE); to correlate with drinkable water and excrete treatment; to develop for the first time a fast diagnostic using electron microscopy in Río Cuarto, Córdoba, Argentina, considering the university and community collaboration in the viral diagnostic. It has been during one year, 122 cases of acute GE, over its clinics epidemiology and diagnostic aspects (1999-2000). With conventional laboratory methods, it has been determined the bacteria and micotic aetiology (NOVIGE); the virology diagnostic with electron microscopy; and the use of the statistics for the data analysis. The microbial findings has been: pathogenics bacteria (31.7%), fungus (17%), parasites (1.2%), rotavirus (16.4%), calicivirus (1.6%), adenovirus and coronavirus (1.6%). The clinics findings are presents like digestive and extra-digestive signs, separated in NOVIGE (and no-diagnosticated) and VIGE groups. The seasonal viral and no-viral distributions are present in fig. 5. There are statistics signification: the NOVIGE in summer period (49%) against the VIGE (12.5%) (p < 0.0003); the VIGE in winter period (87.5%) against the VIGE in summer period (12.5%) (p < 0.0003); the GE aetiology (VIGE and NOVIGE) in associated with age groups (p < 0.0003); the vomits preceding diarrhoea in VIGE was 58% and in NOVIGE (19.5%) (P < 0.001). It has been presented without significative differences all the clinics signs and laboratory examinations; the breeding signs for the younger 2 years group compared with the 2 years older; the age correlation (< and > 2 years) and season (winter-summer) with the 16% for the first in VIGE (62.5% rotavirus); the absence of health systems at the NOVIGE (70%) and with both (29%); and others epidemiology considerations of the sequence. With came to conclusion that in our city: 1. there are VIGE with signficative participation of rotavirus; 2. its distribution are winter and age group, also considering the other age groups and virosis; 3. the NOVIGE may difference at the clinic sign like vomit, between all the possible sintomatology; 4. the principal cause of diarrhoea and the no seasonal distribution are the NOVIGE; 5. there are not a strong relationship of diarrhoea by shortage environmental health in this study; 6. it's possible in Río Cuarto to made a fast and direct virology diagnostic using an electron microscopy.

Gastroenteritis aguda infecciosa en Rio Cuarto: clínica, diagnóstico y epidemiología con especial referencia a infecciones viralesa / Acute infectious gastroenteritis at Rio Cuarto: clinic, diagnosis, and epidemiology with special reference to viral infectious

El presente estudio tiene como objetivo determinar las etiologías y describir los cuadros clínicos de gastroenteritis aguda infecciosa (GE) en la comunidad de Río Cuarto, Argentina, en especial las de orígen viral (GEVI) y su relación con el abastecimiento de agua potable y el tratamiento de exccretas; y desarrollar las técnicas de diagnóstico viral rápido para microscopía electrónica (ME) convencional para apoyar tecnológicamente desde la universidad la resolución de los problemas de salud pública. Se analizaron 122 casos de GE aguda durante um año en sus aspectos clíncos, epidemiológicos y diagnósticos, de un total de 185 pacientes consultantes de 0 a 80 años. Se determinó la etiología bacteriana, micótica y parasitaria (GENOVI) con métodos convencionales de laboratorio; el diagnóstico virológico se realizó con ME, y el análisis estadístico de los datos con prueba de Chi cuadrado y test de Fisher. Los hallazgos microbiológicos fueron: de etiología bacteriana (31,7); fúngicas (17,1); parasitarias (1m2); rotavirus (16,4); calicivirus (1,6); adenovirus y coronavirus (1,6). Lo hallazgos clínicos se separaron en signos digestivos y extradigestivos, tanto para las GENOVI como las GEVI. Las GENOVI no manifiestan distribución estacional )n.s.) y conservan su importancia etiológica con Escherichia coli como agente principal (31,7). La diferencias en la distribución estaciona GEVI vs. GENOVI fue significativa, demostrándose que en el verano las GENOVI representaron el 49 vs. GEVI con el 12, (p<0.003). En período invernal las GEVI tuvieron prevalencia del 87,5 vs.GEVI en el período de verano con un 12,5 (p<0.0003). Las GE de etiología VI y NOVI estuvieron asociadas en ambos grupos etarios, menores de 2 años vs. mayores de 2 años (p<0.003). El vómito precedido de la diarrea en las GEVI fue del 58 vs. las GENOVI en el 19,52 (p<0.001). Estuvieron presente, sin cambios significativos (n.s.) todos los signos clínicos y de laboratorio examinados. El desarrollo de los signos clíncios, comparando el grupo stario < de 2 años vs. > de 2 años tuvo una correlación etaria, y la variación estacional (invierno vs. verano) con el 16 para GEVI (representada por rotavirus en el 62,5 de los casos) (p<0.005). En < de 2 años y en invierno, el rotavirus fue responsable del 100 de las GEVI. Se concluye que en nuestra ciudad existe una significativa relevancia de rotavirus en las GEVI...

Gastroenteritis aguda infecciosa en Rio Cuarto: clínica, diagnóstico y epidemiología con especial referencia a infecciones virales. / [Acute infectious gastroenteritis in Rio Cuarto: clinic, diagnosis and epidemiology with special reference to viral infections]

The objectives of the present study are to describe the clinics view of acute infectious gastroenteritis (GE) at the community, in primary and secondary attention health centers, with special references to viral aetiology (VIGE); to correlate with drinkable water and excrete treatment; to develop for the first time a fast diagnostic using electron microscopy in Río Cuarto, Córdoba, Argentina, considering the university and community collaboration in the viral diagnostic. It has been during one year, 122 cases of acute GE, over its clinics epidemiology and diagnostic aspects (1999-2000). With conventional laboratory methods, it has been determined the bacteria and micotic aetiology (NOVIGE); the virology diagnostic with electron microscopy; and the use of the statistics for the data analysis. The microbial findings has been: pathogenics bacteria (31.7

), fungus (17

), parasites (1.2

), rotavirus (16.4

), calicivirus (1.6

), adenovirus and coronavirus (1.6

). The clinics findings are presents like digestive and extra-digestive signs, separated in NOVIGE (and no-diagnosticated) and VIGE groups. The seasonal viral and no-viral distributions are present in fig. 5. There are statistics signification: the NOVIGE in summer period (49

) against the VIGE (12.5

) (p < 0.0003); the VIGE in winter period (87.5

) against the VIGE in summer period (12.5

) (p < 0.0003); the GE aetiology (VIGE and NOVIGE) in associated with age groups (p < 0.0003); the vomits preceding diarrhoea in VIGE was 58

and in NOVIGE (19.5

) (P < 0.001). It has been presented without significative differences all the clinics signs and laboratory examinations; the breeding signs for the younger 2 years group compared with the 2 years older; the age correlation (< and > 2 years) and season (winter-summer) with the 16

for the first in VIGE (62.5

rotavirus); the absence of health systems at the NOVIGE (70

) and with both (29

); and others epidemiology considerations of the sequence. With came to conclusion that in our city: 1. there are VIGE with signficative participation of rotavirus; 2. its distribution are winter and age group, also considering the other age groups and virosis; 3. the NOVIGE may difference at the clinic sign like vomit, between all the possible sintomatology; 4. the principal cause of diarrhoea and the no seasonal distribution are the NOVIGE; 5. there are not a strong relationship of diarrhoea by shortage environmental health in this study; 6. its possible in Río Cuarto to made a fast and direct virology diagnostic using an electron microscopy.

Gastroenteritis aguda infecciosa en Rio Cuarto: clínica, diagnóstico y epidemiología con especial referencia a infecciones viralesa / Acute infectious gastroenteritis at Rio Cuarto: clinic, diagnosis, and epidemiology with special reference to viral infectious

El presente estudio tiene como objetivo determinar las etiologías y describir los cuadros clínicos de gastroenteritis aguda infecciosa (GE) en la comunidad de Río Cuarto, Argentina, en especial las de orígen viral (GEVI) y su relación con el abastecimiento de agua potable y el tratamiento de exccretas; y desarrollar las técnicas de diagnóstico viral rápido para microscopía electrónica (ME) convencional para apoyar tecnológicamente desde la universidad la resolución de los problemas de salud pública. Se analizaron 122 casos de GE aguda durante um año en sus aspectos clíncos, epidemiológicos y diagnósticos, de un total de 185 pacientes consultantes de 0 a 80 años. Se determinó la etiología bacteriana, micótica y parasitaria (GENOVI) con métodos convencionales de laboratorio; el diagnóstico virológico se realizó con ME, y el análisis estadístico de los datos con prueba de Chi cuadrado y test de Fisher. Los hallazgos microbiológicos fueron: de etiología bacteriana (31,7); fúngicas (17,1); parasitarias (1m2); rotavirus (16,4); calicivirus (1,6); adenovirus y coronavirus (1,6). Lo hallazgos clínicos se separaron en signos digestivos y extradigestivos, tanto para las GENOVI como las GEVI. Las GENOVI no manifiestan distribución estacional )n.s.) y conservan su importancia etiológica con Escherichia coli como agente principal (31,7). La diferencias en la distribución estaciona GEVI vs. GENOVI fue significativa, demostrándose que en el verano las GENOVI representaron el 49 vs. GEVI con el 12, (p<0.003). En período invernal las GEVI tuvieron prevalencia del 87,5 vs.GEVI en el período de verano con un 12,5 (p<0.0003). Las GE de etiología VI y NOVI estuvieron asociadas en ambos grupos etarios, menores de 2 años vs. mayores de 2 años (p<0.003). El vómito precedido de la diarrea en las GEVI fue del 58 vs. las GENOVI en el 19,52 (p<0.001). Estuvieron presente, sin cambios significativos (n.s.) todos los signos clínicos y de laboratorio examinados. El desarrollo de los signos clíncios, comparando el grupo stario < de 2 años vs. > de 2 años tuvo una correlación etaria, y la variación estacional (invierno vs. verano) con el 16 para GEVI (representada por rotavirus en el 62,5 de los casos) (p<0.005). En < de 2 años y en invierno, el rotavirus fue responsable del 100 de las GEVI. Se concluye que en nuestra ciudad existe una significativa relevancia de rotavirus en las GEVI... (AU)