Study of the Relation between the Resonance Behavior of Thickness Shear Mode (TSM) Sensors and the Mechanical Characteristics of Biofilms.

This work analyzes some key aspects of the behavior of sensors based on piezoelectric Thickness Shear Mode (TSM) resonators to study and monitor microbial biofilms. The operation of these sensors is based on the analysis of their resonance properties (both resonance frequency and dissipation factor) that vary in contact with the analyzed sample. This work shows that different variations during the microorganism growth can be detected by the sensors and highlights which of these changes are indicative of biofilm formation. TSM sensors have been used to monitor in real time the development of Staphylococcus epidermidis and Escherichia coli biofilms, formed on the gold electrode of the quartz crystal resonators, without any coating. Strains with different ability to produce biofilm have been tested. It was shown that, once a first homogeneous adhesion of bacteria was produced on the substrate, the biofilm can be considered as a semi-infinite layer and the quartz sensor reflects only the viscoelastic properties of the region immediately adjacent to the resonator, not being sensitive to upper layers of the biofilm. The experiments allow the microrheological evaluation of the complex shear modulus (G* = G' + jG″) of the biofilm at 5 MHz and at 15 MHz, showing that the characteristic parameter that indicates the adhesion of a biofilm for the case of S. epidermidis and E. coli, is an increase in the resonance frequency shift of the quartz crystal sensor, which is connected with an increase of the real shear modulus, related to the elasticity or stiffness of the layer. In addition both the real and the imaginary shear modulus are frequency dependent at these high frequencies in biofilms.

Morphological plasticity of Streptococcus oralis isolates for biofilm production, invasiveness, and architectural patterns.

OBJECTIVE: Streptococcus oralis is an early coloniser of the oral cavity that contributes to dental plaque formation. Many different genotypes can coexist in the same individual and cause opportunistic infections such as bacterial endocarditis. However, little is known about virulence factors involved in those processes. The aim was to analyze the evolving growth of S. oralis colony/biofilm to find out potentially pathogenic features. DESIGN: Thirty-three S. oralis isolates were analyzed for: (1) biofilm production, by spectrophotometric microtiter plate assay; (2) colonial internal architecture, by histological methods and light and electron microscopy; (3) agar invasion, by a new colony-biofilm assay. RESULTS: S. oralis colonies showed two different growth patterns: (1) fast growth rate without invasion or minimally invasive; (2) slow growth rate, but high invasion ability. 12.1% of strains were biofilm non-producers and 24.2% not invasive, compared to 51.5% biofilm high-producers and 39.4% very invasive. Both phenotypic characteristics tended to be mutually exclusive. However, a limited number of strains (15%) co-expressed these features at the highest level. CONCLUSIONS: Morphological plasticity of S. oralis highlighted in this study may have important ecological and clinical implications. Coexistence of strains with different growth patterns could produce a synergic effect in the formation and development of subgingival dental plaque. Moreover, invasiveness might regulate dissemination and colonisation mechanisms. Simultaneous co-expression of high-invasive and high-biofilm phenotypes gives a fitness advantage during colonisation and may confer higher pathogenic potential.

[Biofilm score: is it a differential element within gram negative bacilli?]. / Es la cuantificación del biofilm un elemento diferenciador en la patogenia de bacilos gramnegativos?

The aim of the study was to investigate biofilm formation in Gram negative bacteria and to quantify biofilm production applying a new developed technique that made possible to compare results about biofilm formation within the different Gram negative bacteria species. A total of 153 Gram negative strains corresponding to 12 different bacterium species were studied applying a variation of the optic density measurement technique reported by Stepanovic et al. Data obtained with optic density analysis allow to classify microorganisms in strong biofilm developers, moderate biofilm developers, weak biofilm developers and no biofilm developers. The results were expressed in two ways, using in both cases the same statistical method: without standardization, where controls were different depending on the day optic density measurements were performed, and standardized using a correction factor, using the same control for every strain of all our bacterium species in our study, which allows result homogenization. The obtained results in our study after data analysis and standardization show that over the 153 Gram negative strains in our study, 105 of them were no biofilm developers, representing 63.75% of all the studied bacterium genera. We consider that standardization and quantification of biofilm development in Gram negative bacteria can be useful in clinical practice, because biofilm development ability can lead or focus the gold treatment of pathologies produced by these microorganisms.

¿Es la cuantificación del biofilm un elemento diferenciador en la patogenia de bacilos gramnegativos? / Biofilm score: is it a differential element within gram negative bacilli?

El objetivo del estudio fue investigar la formación de biofilms en bacterias gramnegativas y cuantificar la producción de biofilm mediante la aplicación de una técnica que permitiese una comparación de los resultados de la formación de biofilm entre las diferentes especies de gramnegativos. Se estudiaron un total de 153 cepas de bacilos gramnegativos correspondientes a 12 especies bacterianas por el método de la densidad óptica aplicando una modificación de la técnica descrita por Stepanovic et al. Los valores obtenidos mediante el análisis de la densidad óptica permiten clasificar a los microorganismos en formadores fuertes, moderados, débiles y no formadores. Los resultados obtenidos se han expresado de dos maneras, ambas utilizando el mismo método estadístico: sin estandarizar, donde los controles fueron diferentes dependiendo de los días en los que se realizaron las medidas; y estandarizados mediante un factor de corrección, utilizando el mismo control para todas las cepas de cada especie, lo que permite su homogeneización. Los resultados obtenidos en el estudio tras el análisis y estandarización establecen que de las 153 cepas de gramnegativos estudiados, 105 de ellas fueron no formadoras de biofilms, representando el 63,75% de los géneros estudiados. Consideramos que la estandarización y cuantificación de la producción de biofilm entre las bacterias gramnegativas puede resultar de utilidad en el ámbito clínico, ya que el conocimiento de la capacidad de producción de biofilm puede dirigir o enfocar el tratamiento de elección de las patologías producidas por dichos microorganismos(AU)

The aim of the study was to investigate biofilm formation in Gram negative bacteria and to quantify biofilm production applying a new developed technique that made possible to compare results about biofilm formation within the different Gram negative bacteria species. A total of 153 Gram negative strains corresponding to 12 different bacterium species were studied applying a variation of the optic density measurement technique reported by Stepanovic et al. Data obtained with optic density analysis allow to classify microorganisms in strong biofilm developers, moderate biofilm developers, weak biofilm developers and no biofilm developers. The results were expressed in two ways, using in both cases the same statistical method: without standardization, where controls were different depending on the day optic density measurements were performed, and standardized using a correction factor, using the same control for every strain of all our bacterium species in our study, which allows result homogenization. The obtained results in our study after data analysis and standardization show that over the 153 Gram negative strains in our study, 105 of them were no biofilm developers, representing 63.75% of all the studied bacterium genera. We consider that standardization and quantification of biofilm development in Gram negative bacteria can be useful in clinical practice, because biofilm development ability can lead or focus the gold treatment of pathologies produced by these microorganisms(AU)

Correlation between clinical parameters characterising peri-implant and periodontal health: a practice-based research in Spain in a series of patients with implants installed 4-5 years ago.

OBJECTIVES: To explore peri-implant health (and relation with periodontal status) 4-5 years after implant insertion. STUDY DESIGN: A practice-based dental research network multicentre study was performed in 11 Spanish centres. The first patient/month with implant insertion in 2004 was considered. Per patient four teeth (one per quadrant) showing the highest bone loss in the 2004 panoramic X-ray were selected for periodontal status assessment. Bone losses in implants were calculated as the differences between 2004 and 2009 bone levels in radiographs. RESULTS: A total of 117 patients were included. Of the 408 teeth considered, 73 (17.9%) were lost in 2009 (losing risk: >50% for bone losses ≥7 mm). A total of 295 implants were reviewed. Eight of 117 (6.8%) patients had lost implants (13 of 295 implants installed; 4.4%). Implant loss rate (quadrant status) was 1.4% (edentulous), 3.6% (preserved teeth), and 11.1% (lost teeth) (p=0.037). The percentage of implant loss significantly (p<0.001) increased when the medial/distal bone loss was ≥3 mm. The highest (p≤0.001) pocket depths were found in teeth with ≥5 mm and implants with ≥3 mm bone losses, with similar mean values (≥4 mm), associated with higher rates of plaque index and bleeding by probing. CONCLUSIONS: The significant bi-directional relation between plaque and bone loss, and between each of these two parameters/signs and pocket depths or bleeding (both in teeth and implants, and between them) together with the higher percentage of implants lost when the bone loss of the associated teeth was ≥3 mm suggest that the patient's periodontal status is a critical issue in predicting implant health/lesion.

In vitro interference of tigecycline at subinhibitory concentrations on biofilm development by Enterococcus faecalis.

OBJECTIVES: Since biofilm formation is the hallmark of Enterococcus faecalis isolates, the aim of this study was to quantify biofilm formation in the presence of subinhibitory concentrations of tigecycline. METHODS: Interference of tigecycline on biofilm formation was spectrophotometrically quantified using 20 biofilm-producing E. faecalis isolates with tigecycline MICs of 0.12 (8 strains) or 0.25 mg/L (12 strains). Biofilm production was measured in antibiotic-free tryptic soy broth supplemented with 1% glucose and compared with biofilm production in the same medium with tigecycline at subinhibitory concentrations (0.25× or 0.5× MIC, similar to trough concentrations in serum or concentrations in the colon after a standard dose) by reading the optical density at 450 nm (OD(450)) after staining with Crystal Violet. RESULTS: In the presence of subinhibitory tigecycline concentrations, pooled OD(450) values for the 20 strains [median (IQR)] were significantly lower than those for controls: 0.468 (0.379-0.516) for antibiotic-free controls versus 0.295 (0.200-0.395) for 0.25× MIC tigecycline (P < 0.001) and 0.287 (0.245-0.479) for 0.5× MIC tigecycline (P < 0.001), with significant differences between pooled OD(450) values obtained with each concentration of tigecycline (P = 0.022). In 17 out of 20 (85%) strains the OD(450) obtained with 0.25× MIC tigecycline was significantly (P < 0.05) lower than the basal OD(450), while this occurred in 12 out of 20 (60%) strains with 0.5× MIC. CONCLUSIONS: In vitro tigecycline subinhibitory concentrations were able to interfere with biofilm formation by E. faecalis.

Genotypic versus phenotypic characterization, with respect to susceptibility and identification, of 17 clinical isolates of Staphylococcus lugdunensis.

OBJECTIVES: To compare different methods for the identification and determination of susceptibility to penicillin and methicillin of Staphylococcus lugdunensis. METHODS: Seventeen clinical isolates of S. lugdunensis (identified by PCR amplification and sequencing of the rpoB gene) were studied using the ATB32-Staph, Crystal, Vitek 2 and Wider commercial systems. The clumping factor test and the tube coagulase test were also performed. Beta-lactamase production was studied by chromogenic methods. Methicillin resistance was phenotypically studied by the MRSA slide latex agglutination test, growth in MRSA agar, and the Vitek 2 and Wider systems (based on oxacillin MIC), and genotypically studied by detection of the mecA gene by PCR. RESULTS: The clumping factor test was negative in 35.3% of strains. All isolates were correctly identified to species level by the ATB32-Staph system. Species misidentification rates were 5.9%, 23.5% and 29.4% with the Crystal, the Vitek 2 and the Wider systems, respectively, mostly as Staphylococcus haemolyticus. Beta-lactamase was present in 11.8% of strains. Whereas 76.5% and 47.1% of strains exhibited oxacillin resistance (MIC range 0.5-2 mg/L) by the Vitek 2 system and the Wider system, respectively, none of the strains was positive in the MRSA slide latex agglutination test or grew in MRSA agar. All strains lacked the mecA gene. CONCLUSIONS: The clumping factor test and some commercial systems may misidentify S. lugdunensis. Oxacillin resistance detected by commercial systems is not indicative of the presence of the mecA gene. These facts, together with beta-lactamase production, may preclude adequate treatment of infections by this virulent coagulase-negative Staphylococcus.

Mujer tailandesa con hematuria y eosinofilia / Thai woman with hematuria y eosinophilia

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Infecciones bacterianas mixtas de la cavidad oral / Mixed bacterial infections of the oral cavity

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Cavidad tuberculosa abscesificada de evolución tórpida por sobreinfección con Gemella haemolysans / Torpid course of and abscessified tuberculous cavity due to Gemella haemolysans overgrowth

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Infección por Staphylococcus Lugdunensis: presentación de trece casos / Staphylococcus lugdunensis infection: Report of thirteen cases

MATERIAL Y MÉTODOS. Se estudiaron los aislamientos de S. lugdunensis en nuestro hospital, durante tres años (19972000). Se analizó el factor de afinidad para el fibrinógeno y la coagulasa libre en tubo. La identificación se realizó por los sistemas Crystal GPID (Becton Dickinson) y Wider (Soria Melguizo). El antibiograma se hizo con paneles Wider. Se revisaron las historias clínicas de los pacientes con aislamiento de S. lugdunensis. RESULTADOS. Se obtuvieron trece aislamientos de S. lugdunensis de las siguientes muestras: abscesos (9), heridas quirúrgicas (2), hemocultivo (1) y orina (1). En diez casos el cultivo fue puro, y en tres mixto. Dos casos fueron excluidos por considerar que S. lugdunensis no estaba asociado a la patología. La localización de las lesiones fue: inguinal (4), glúteo (2), mama (2), pared abdominal (1), pie (1), desconocido (1). Diez de los trece aislamientos (76 por ciento) presentaban factor de afinidad para el fibrinógeno. Fueron sensibles a la penicilina 11/13 (84,6 por ciento). En un caso, la cepa era productora de betalactamasa (7,6 por ciento) y en otro resistente a meticilina (7,6 por ciento). CONCLUSIONES. Observamos que S. lugdunensis se asocia a infecciones de piel y tejidos blandos, especialmente abscesos. La infección aparece con mayor frecuencia en pacientes neoplásicos, como sucedió en tres casos de nuestra serie. Insistimos en la necesidad de llevar a cabo una correcta identificación de S. lugdunensis. El aislamiento de un estafilococo coagulasa negativa no debe tranquilizarnos, al menos hasta descartar una especie potencialmente patógena (AU)

Bacteriemia por Campylobacter jejuni en varón de 22 años con anemia hemolítica autoinmune / Bacterimia caused by Campylobacter jejuni in a 22 year old male with autoimmune haemolytic anaemia

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Contribución al diagnóstico precoz de bacteriemia: detección del crecimiento microbiano en medios de cultivo líquidos por ultrasonidos / Contribution to the early diagnosis of bacteremia: microbial growth detection in liquids by ultrasound

La infección nosocomial es un problema importante que afecta a un gran número de pacientes hospitalizados; requiriendo de los laboratorios de cada centro un esfuerzo encaminado a detectar precozmente los microorganismos responsables de bacteriemias y fungemias. Los ultrasonidos son ampliamente utilizados con fines diagnósticos en medicina y es conocida la posibilidad de medir con ellos parámetros físicos y químicos de medios líquidos, usando las variables de la velocidad de propagación y atenuación. Nuestro objetivo fue desarrollar un sistema analítico automatizado, diseñado para detectar, de forma eficaz y no invasora, el crecimiento precoz de microorganismos en medios de cultivo líquidos sembrados con muestras clínicas diversas. Para ello se procedió a la medición de la velocidad de propagación de un pulso ultrasónico a través de frascos de hemocultivo, con muestra de sangre o sin ella, en los que se habían inoculado concentraciones conocidas de microorganismos frecuentemente implicados en cuadros sépticos. El gas carbónico generado por las bacterias en crecimiento modifica la velocidad de propagación del sonido. La monitorización continua del sistema nos permite obtener una representación gráfica del crecimiento microbiano y aplicar una serie de algoritmos diagnósticos relacionados con la pendiente de la velocidad a fin de conocer con la mayor rapidez los cambios físicos que se producen, en el medio líquido, cuando los microorganismos están presentes. El tiempo de detección por nuestro sistema es reducido y se ha mostrado similar o discretamente mejor que otros comercializados. El método es sencillo, económico e innovador y abre la posibilidad de nuevas aplicaciones con fines diagnósticos en microbiología (AU)

Lesión cutánea nodular en ama de casa / Subacute endocarditis due to Actinobacillus actinomycetemcomitans

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