Enterococcus faecalis fiofilm: formation and development in vitro observed by scanning electron microscopy

Se realizó la detección de biofilm de Enterococcus faecalis aislados de conductos radiculares, desarro llándolos en microcubetas realizando tinción con Cristal Violeta 10 por ciento, elución con alcohol y tres procedimientos: sin fijar, fijación con calor y fija ción con formaldehido 10 por ciento. Se evaluó el biofilm formado realizando la lectura con lector de microplaca Versamax-Microplate-Reader (USA). Se incubaron 20 porciones radiculares estériles en el medio TS caldo con E. faecalis (108) durante 48 hs, 4, 7, 14 y 30 días.Transcurrido cada tiempo experimental se procesaron y observaron al microscopio electrónico de barrido (MEB). Se evidenció significativamente mayor formación de biofilm en microplacas cuando se realizó fijación con formaldehido, que al fijar con calor y sin fijar (ANOVA p<0,0001). Al MEB seobservó crecimiento del E. faecalis en todos los tiempos y desarrollo de biofilm a partir de 14 días de incubación. La fijación con formaldehido 10% fue la técnica más apropiada para detectar biofilm de E. faecalis desarrollado enmicroplacas. Mediante el MEB se confirmó la formación de biofilm después de 14 días de incubación...

Detection of viable and viable nonculturable Vibrio cholerae O1 through cultures and immunofluorescence in the Tucumán rivers, Argentina

Vibrio cholerae has been sporadically isolated from rivers in Tucumán, Argentina, since the outbreak in 1991. The aim of this study was to determine the environmental reservoir of the bacterium in these rivers, assessing the presence of Vibrio cholerae non-O1 and O1 (the latter both in its viable culturable and non culturable state) and its relationship to environmental physicochemical variables. 18 water samplings were collected in the Salí River (in Canal Norte and Banda) and the Lules River between 2003 and 2005. Physical-chemical measurements (pH, water temperature, electrical conductivity and dissolved oxygen) were examined. Vibrio cholerae was investigated with conventional culture methods and with Direct Immunofluorescence (DFA-VNC) in order to detect viable non culturable organisms. All isolated microorganisms corresponded to Vibrio cholerae non-O1 and non-O139 (Lules 26 percent, Canal Norte 33 percent and Banda 41 percent). The majority was found during spring and summer and correlated with temperature and pH. Non culturable Vibrio cholerae O1 was detected year round in 38 of the 54 water samples analyzed. Application of the Pearson correlation coefficient revealed that there was no relationship between positive immunofluorescence results and environmental physicochemical parameters. Genes coding for somatic antigen O1 were confirmed in all DFA-VNC-positive samples, whereas the virulence-associated ctxA and tcpA genes were confirmed in 24 samples.

Vibrio cholerae tem sido isolado esporadicamente nos rios da Província de Tucumán, Argentina, desde outubro de 1991. O objetivo deste estudo foi localizar os reservatórios nestes rios, identificar a presença de Vibrio cholerae O1 (em estado cultivável e não cultivável) e relacionar a presença desta bactéria com as variações físico-químicos da água. Foram coletadas dezoito amostras de água do rio Salí (nas localidades de Canal Norte e Banda) e do rio Lules, entre 2003 e 2005. Estas foram submetidas a análises físico-químicos como determinação de pH, temperatura, condutibilidade elétrica e oxigênio dissolvido. A presença de Vibrio cholerae foi verificada por métodos de cultivo convencional e por imunofluorescência direta (DFA-VNC). Todos os microrganismos isolados foram não O1 e não O139 (Lules 26 por cento, Canal Norte 33 por cento e Banda 41 por cento). A maioria foi encontrada na primavera e verão, indicando uma relação com a temperatura e pH. Das 54 amostras analisadas por DFA-VNC, 38 Vibrio cholerae não cultivável, foram detectadas em todas as épocas do ano. As amostras positivas foram confirmadas por PCR para o antígeno somático O1 e para os genes de virulência ctxA e tcpA. Coeficiente de correlação de Pearson revelou que não há relação entre os resultados obtidos por imunofluorescência e a variação dos parâmetros físico-químicos.

In vitro antibacterial effect of different irrigating solutions on Enterococcus faecalis

Se evaluó el efecto antibacteriano (ea) y la concentración inhibitoria mínima (cim) sobre Enterococcus faecalis deNaOCl 2,5 por ciento, Gluconato de Clorhexidina 0,2 por ciento y EDTA17 por ciento. La capacidad antibacteriana fue valorada mediante el test de difusión en agar. El efecto antibacteriano se evaluó empleandotrozos radiculares apicales y medios que fueron esterizados, contaminados con Enterococcus faecalis, sumergidos en lasdiferentes soluciones de irrigación e incubados a 37°c. El recuento de células viables se realizó a 4, 8 y 24 horas.Cim: NaOCl y CHX: 0.2 por ciento, EDTA menor al 5 por ciento. Difusión enagar: NaOCl 2.5 por ciento= 21 mm. CHX 0.2 por ciento= 14mm. EDTA 17 por ciento=20 mm. Efecto sobre la dentina radicular: NaOCl 2.5 por ciento: Enterococcus faecalis fue totalmente inhibido hasta las 24 horas en el áreaapical y hasta las 8 horas en el área media. CHX 0.2 por ciento mostróuna reducción de más de 5 log UFC y EDTA 17 por ciento provocó unareducción de más de 3 log UFC en todos los tiempos testeados en los tercios apical y medio

Was evaluated the minimum inhibitory concentration (MIC) and the antibacterial effect (AE) of 2.5% NaOCl, 0.2% chlorhexidine gluconate (CHX) and 17% EDTA on Enterococcus faecalis. The antibacterial capacity was assessed by diffusion in agar. The AE was evaluated on contaminated root dentin, employing apical and middle portions of human roots, sterilized and contaminated with Enterococcus faecalis, immersed in the irrigation solutions and incubated at 37°C. Viable cells were counted at 0, 4, 8 and 24 hours. MIC: NaOCl and CHX: 0.2%, EDTA below 5%. Diffusion in agar: NaOCl 2.5%= 21 mm. CHX 0.2%= 14mm. EDTA 17%= 20 mm. Effect on root dentin: NaOCl 2.5%: Enterococcus faecalis was totally inhibited for 24 hours in the apical area, and for 8 hours in the middle area. CHX 0.2% elicited a reduction of more than 5 log CFU and EDTA 17% induced a reduction of more than 3 log CFU at all the time points examined in the apical and middle areas.

Ultrastructural studies on the respiratory tract of mice inoculated intranasally with L. fermentum

The ultrastructural modifications of the respiratory tract of mice produced by the intranasally inoculated L. fermentum was evaluated. The genus Lactobacillus has been largely studied from the probiotic effect point of view in different ecological systems. Lactobacilli inoculated in 4 intranasal doses (1 x 10(9) CFU/dose) do not produce fundamental changes at the ultrastructural level in the organs of the respiratory tract studied. The most important finding was the presence of a cellular type with intracellular structures surrounded by membranes exhibiting electron dense paracristalline material parallel arranged.