RESUMEN
With the recent insights into the Streptococcus milleri group (SMG) as pulmonary pathogens in patients with cystic fibrosis (CF), we sought to characterize 128 isolates from the sputum of adults with CF, along with 45 isolates from patients with invasive diseases for comparison. The tests performed included Lancefield grouping; tests for hemolysis; tests for the production of hyaluronidase, chondroitin sulfatase, DNase, proteases, and hydrogen peroxide; and PCR for the detection of the intermedilysin gene (ily). We also generated biochemical profiles with the Rapid ID Strep 32 API system and tested cell-free supernatants for the presence of the signal molecule autoinducer-2 (AI-2) using a Vibrio harveyi bioassay with a subset of CF strains. The S. intermedius isolates from both strain collections were similar, while the S. constellatus and S. anginosus isolates yielded several biotypes that differed in prevalence between the two strain collections. Beta-hemolytic, Lancefield group C S. constellatus comprised 74.4% of the S. constellatus isolates from patients with CF but only 13.3% of the corresponding isolates from patients with invasive infections. This was the only S. constellatus biotype associated with pulmonary exacerbations. Hyaluronidase-positive S. anginosus was detected only among the isolates from patients with CF. Strain-to-strain variability in AI-2 expression was evident, with the mean values being the highest for S. anginosus, followed by S. constellatus and then S. intermedius. Cluster analysis and 16S rRNA sequencing revealed that the species of SMG could be accurately determined with a minimum of three phenotypic tests: tests for the Lancefield group, hyaluronidase production, and chondroitin sulfatase production. Furthermore, isolates from patients with invasive infections clustered with isolates from the sputum of patients with CF, suggesting that the respiratory tract isolates were equally pathogenic.
Asunto(s)
Técnicas de Tipificación Bacteriana , Fibrosis Quística/complicaciones , Esputo/microbiología , Infecciones Estreptocócicas/microbiología , Streptococcus milleri (Grupo)/clasificación , Streptococcus milleri (Grupo)/aislamiento & purificación , Adulto , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Bacteriocinas/genética , ADN Bacteriano/química , ADN Bacteriano/genética , ADN Ribosómico/química , ADN Ribosómico/genética , Enzimas/metabolismo , Genotipo , Humanos , Fenotipo , Reacción en Cadena de la Polimerasa/métodos , ARN Ribosómico 16S/genética , Análisis de Secuencia de ADN , Serotipificación , Streptococcus milleri (Grupo)/genética , Streptococcus milleri (Grupo)/metabolismo , Adulto JovenRESUMEN
The aim of this study was to determine the in vitro antibacterial effect of two experimental glass ionomer cements (GICs) on Streptococcus milleri, Staphylococcus aureus, and Enterococcus faecalis after 24 and 48 h incubation by using the agar diffusion inhibitory test. Silver zeolite (SZ) was added at 0.2 and 2% mass fraction concentration to GIC (Endion). The control group was Endion with no SZ. Each of them were prepared to uniform size using a custom-made Teflon mold, and the GIC materials were prepared to form disks (n = 5 per group). The effect of these materials on the growth of three bacteria associated with endodontic infections was determined using the agar diffusion inhibitory test. The amounts of silver ion release from these materials were measured with atomic absorption spectrophotometry at 10 min, 24- and 48-h periods. The pH of samples was measured with a pH-meter at 10 min, 24- and 48-h periods. After the incubation period, the agar plates were evaluated and the degrees of bacterial inhibition were measured in millimeters. A comparison of the mean of the test materials was statistically different in each group of specimens (p < 0.05). Between the two tested materials 2% SZ containing GIC showed the largest zone of inhibition on the agar plates of all the tested strains (p < 0.05). The most inhibition in bacterial growth occurred in E. faecalis. Adding 2% SZ to GIC resulted in a significant increase in the silver release into deionized water. This study demonstrated that GIC had an inhibitory affect on Streptococcus milleri, Staphylococcus aureus, and Enterococcus faecalis and that adding SZ increases that affect proportional to its concentration.
Asunto(s)
Cementos de Ionómero Vítreo/química , Materiales de Obturación del Conducto Radicular/química , Plata/química , Zeolitas/química , Antiinfecciosos/farmacología , Cavidad Pulpar , Difusión , Enterococcus faecalis/metabolismo , Concentración de Iones de Hidrógeno , Ensayo de Materiales , Pruebas de Sensibilidad Microbiana , Espectrofotometría Atómica/métodos , Staphylococcus aureus/metabolismo , Streptococcus milleri (Grupo)/metabolismo , Factores de TiempoRESUMEN
OBJECTIVE: To determine which cell fraction(s) of Streptococcus mitis biovar 1 serve as the best source of antigens recognized by salivary SIgA antibodies in infants. DESIGN: Whole cells of 38 reference and wild-type isolates of S. mitis, Streptococcus oralis, Streptococcus gordonii, Enterococcus casseliflavus, and Enterococcus faecalis were fractionated into cell walls (CW), protease-treated cell walls (PTCW), cell membranes (CM) and cell protein (CP). Whole cells and these fractions were tested for binding by rabbit anti-S. mitis SK145 and anti-S. oralis SK100 sera, and also by salivary SIgA antibodies from infants and adults. RESULTS: Anti-SK145 and anti-SK100 sera bound whole cells and fractions of all strains of S. mitis and S. oralis variably. Cluster analysis of antibody binding data placed the strains into S. mitis, S. oralis and 'non-S. mitis/non-S. oralis' clusters. Antigens from CW and CM best discriminated S. mitis from S. oralis. CM bound the most infant salivary SIgA antibody and PTCW bound the least. In contrast, adult salivary SIgA antibody bound all of the cell fractions and at higher levels. CONCLUSIONS: Presumably the relatively short period of immune stimulation and immunological immaturity in infants, in contrast to adults, result in low levels of salivary SIgA antibody that preferentially bind CM of S. mitis but not PTCW. By utilizing isolated cell walls and membranes as sources of antigens for proteomics it may be possible to identify antigens common to oral streptococci and dissect the fine specificity of salivary SIgA antibodies induced by oral colonization by S. mitis.
Asunto(s)
Fraccionamiento Celular/métodos , Enterococcus faecalis/inmunología , Inmunoglobulina A Secretora/inmunología , Saliva/inmunología , Streptococcus milleri (Grupo)/inmunología , Adulto , Factores de Edad , Animales , Anticuerpos Antibacterianos , Sitios de Unión de Anticuerpos/inmunología , Enterococcus faecalis/crecimiento & desarrollo , Humanos , Factores Inmunológicos/inmunología , Lactante , Boca/microbiología , Conejos , Saliva/microbiología , Streptococcus milleri (Grupo)/crecimiento & desarrollo , Streptococcus milleri (Grupo)/metabolismoRESUMEN
Some oral anaerobes from the Streptococcus milleri strain group were found to secrete human specific hemolytic toxin, which was detected when bacteria were cultured in Todd-Hewitt broth and Brain Heart Infusion broth. The toxin elicited by the Streptococcus intermedius strain was partially fractionated by ammonium sulfate precipitation. Preincubation with glutathione or cysteine showed significant inhibiting effects; however, no effects were seen with dithiothreitol or beta-mercaptoethanol, and cholesterol was a weak inhibitor. Five kinds of protease inhibitor had no effect on the hemolytic activity, and rabbit preimmune and immune sera against the bacterial cells showed weak inhibition at a similar level. Digestion with trypsin, chymotrypsin, proteinase-K, subtilisin and pronase-P brought about a rise in activity, followed by a decrease during long-term incubation. Other enzymes tested showed no effects. Further, the presence of the intermedilysin gene in the portion with hemolytic activity was not identified by polymerase chain reaction.